Advances in Marine Biology, Volume 15

Advances in

MARINE BIOLOGY VOLUME 15

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Advances in

MARINE BIOLOGY VOLUME 15 Edited by

SIR FREDERICK S. RUSSELL Plymouth, England

and

SIR MAURICE YONGE Edinburgh, Scotland

Academic Press London New York

San Francisco 1978

A Subsidiary of Harcourt Brace Jovanovich, Publish-srs

ACADEMIC PRESS INC. (LONDON) LTD.

24-28 OVAL ROAD LONDON NW1 7DX

U.X. Edition published by ACADEMIC PRESS INC.

111

FIFTH AVENUE

NEW YORK, NEW YORK

Copyright

10003

0 1978 by Academic Press Inc. (London) Ltd.

All rights reserved

NO

p a r OF THIS BOOK MAY BE REPRODUCED IN ANY FORM BY PHOTOSTAT,

MICROFILM, OR ANY OTHER MEANS, WITHOUT WRITTEN PERMISSION FROM THE PUBLISHERS

Library of Congress Catalog Card Number: 63-14040 ISBN:

0-12-026115-4

PRINTED IN GREAT BRITAIN BY THE WEITEFRIARS PRESS LTD., LONDON A N D TONBRIDGE

CONTRIBUTORS TO VOLUME 15 CHRISTOPHER J. CORKETT,Dalhousie University, Halifax, Nova Scotia, Canada.

E. D. S. CORNER, The Laboratory, Marine Biological Association, Plymouth, England.

ANTHONY G. DAVIES,The Laboratory, Narine Biological Associa,tion, Plymouth, England. IANA. MCLAREN,Dalhousie University, Halifax, Nova Scotia, Canada.

M. R . REEVE,University of Miami, School of Marine and Atmospheric Science, Miami, Florida, U.S.A.

F. S. RUSSELL,Marine Biological Association, Citadel Hill, Plymonth, England.

M. A. WALTERS,University of Miami, Xchool of Marine and Atmospheric Science, Miami, Florida, U.S.A.

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CONTENTS CONTRIBUTORS TO

VOLUME 15

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The Biology of Pseudocalanus CHRISTOPHERJ. CORKETTAND IAN A. MCLAREN I. Introduction

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11. Systematics . . A. Nomenclature.. .. .. B. " Physiological " Species . . C. Variations in DNA Content D. Retrospects and Prospects . .

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VI. Excretion . . .. .. A. Nitrogen . . .. B. Phosphorus . . .. C. Retrospects and Prospects VII. Locomotion . . .. .. A. Routine Swimming . . B. Escape Reaction .. C. Retrospects and Prospects

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VIII. Nutrition .. .. .. .. .. .. A. Feeding Mechanism . . .. B. FoodEaten .. .. .. .. C. Feeding Rate . . .. .. .. .. D. Die1 Feeding Rhythms .. E. Assimilation . . .. .. .. F. Food Requirements for Sustenance G. Retrospects and Prospects . . .. IX. Reproduction .. .. .. .. A. Sex Ratio .. . . .. .. .. B. Oogenesis and Egg Laying . . C. Sperm and Spermatophore Production D. Mating .. .. .. .. E. Reproductive Rate . . .. .. F. Retrospects and Prospects . . ..

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.. .. X. Development and Growth . . .. .. .. A. Embryonic Development Rate .. .. .. B. Hatching .. .. .. .. C. Development Rate of Nauplii and Copepodids . . D. Longevity of Adults . . .. .. .. .. E. Body Size .. .. .. . . .. .. .. F. Body Composition and Weights . . .. G. Oil Storage . . .. .. .. .. .. H. Growth Rates. . .. .. .. .. .. I. Rate of Production of Egg Matter .. .. J. The " Balance Equation " and Growth Efficiencies K. Retrospects and Prospects . . .* .. ..

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XI. Life Cycles in Nature .. .. .. * . .. A. General Features, Terminology and Approaches

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B. Representative Life Cycles . . C. Retrospects and Prospects . .

.. .. XII. Vertical Migration . . A. Ontogenetic -Migrations .. B. Seasonal Migrations . . .. C. Die1 Migrations .. .. D. Retrospects and Prospects . . .. .. XIII. Production . . A. General Methods .. B. Production Estimates C. Retrospects and Prospects .. .. .. XIV. Parasites A. Dinoflagellates .. B. Gregarines . . .. C. Trematodes . . .. D. Nematodes . . .. E. Crustaceans . . .. F. Retrospects and Prospects XV.

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Role in the Food Web . . .. A. Effect on Phytoplankton . . B. Predators .. .. .. C. Significance in the Food Web D. Retrospects and Prospects . .

XVI . Acknowledgements . .

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Marine Biology and Human Affairs F. S. RUSSELL

I. Food from the Sea 11. Fish Farming

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111. Poisonous and Venomous Plants and Animals

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.. Physiological and Medical Aspects. . Pesticide . . .. .. .. Geology and Meteorology . . .. .. .. .. Pollution .. Conservation . . .. .. .. Man and the Marine Environment. .

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IV. Underwater Structures

V. Ship Design VI. VII. VIII. IX.

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Echo-sounding and Noise

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Review of

Nutritional Ecology of Ctenophores-A Recent Research

M. E. REEVEAND M. A. WALTER

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I. Introduction

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11. Feeding Mechanisms and Behavior .. .. .. 251 A. Feeding Mechanism and Behavior in Mnemiopsis 251 B. Comparison of Feeding Behavior in Other .. .. .. .. . . 259 Tentaculata. . C. Food of Tentaculata . . .. .. . . .. 263 D. Food and Feeding Behavior of Nuda . .. 265

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IV. Chemical Composition

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VII. Respiration and Excretion. .

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Conclusion

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Pollution Studies with Marine Plankton: Part 1. Petroleum Hydrocarbons and Related Compounds

E. D. S. CORNER

I. Introduction

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11. Hydrocarbon Levels in Sea Water .. .. A. Studies Primarily concerned with Alkanes. . B. " Dissolved " and Particulate Hydrocarbons C. Hydrocarbons in or near the Surface of the Sea D. Comprehensive Analyses .. .. ..

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111. Hydrocarbons in Plankton . . .. .. .. .. A. Phytoplankton .. .. .. .. . . B. Phytoplankton and Crude Oil as Sources of Hydro.. .. .. . . carbons in the Sea . . C. Zooplankton . . .. .. .. .. ..

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IV. Toxicity Studies with Phytoplankton .. .. .. 317 A. Studies using Crude Oils and their Water-soluble Fractions . . .. .. .. .. .. 319 B. Studies using Naphthalene . . .. .. . . 327 V. Mechanisms of Phytotoxicity

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VII. Fate of Hydrocarbons in Zooplankton . . .. .. A. Uptake and Release . . .. .. .. .. B. Quantitative Importance of the Dietary Pathway C. Long-term Exposure Experiments . . .. .. .. .. .. .. D. Metabolism .. .. .. E. Release of Hydrocarbons in Faecal Pellets. . VIII.

Toxicity Studies with Zooplankton .. .. .. .. .. .. .. .. A. Crude Oil .. B. W ater-soluble Hydrocarbons .. .. .. C. Possible Effects of Hydrocarbons on Reproduction by Zooplankton .. .. .. .. . . D. Summary and General Comments . . *. ..

IX. Conclusions . . .. A. Chemical Analyses B. Toxicity Studies C. Biochemical Work

X. Acknowledgements XI. References

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Pollution Studies with Marine Plankton: Part II. Heavy Metals

ANTHONY G. DAVIES

I. Introduction

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11. The Turnover of Heavy Metals by Phytoplankton .. A. The Kinetics and Mechanism of Metal Uptake by Phytoplankton .. .. .. .. . . B. The Effect of the Chemical Form of a Metal upon its Uptake by Phytoplankton .. .. .. C. The Role of Phytoplankton in the Biogeochemistry of Heavy Metals in the Sea .. ..

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111. Laboratory Studies of the Toxic Effects of Heavy Metals . . .. .. .. 398 upon Phytoplankton A. The Effects on the Growth of Phytoplankton . . 398 B. Synergism and Antagonism of Mixtures of Heavy Metals towards Phytoplankton . . .. .. 411 C. The Nature of Metal Toxicity in Phytoplankton . . 412

IV. Studies of the Toxic Effects of Heavy Metals upon Natural Populations of Phytoplankton . . .. 415 . . 416 A. The Effects on Primary Production Rates B. The Effects in Large Volume Sea Water Enclosures 419 V. Heavy Metal Concentrations in Natural Populations of Marine Phytoplankton .. .. .. .. 425

.. VI. The Turnover of Heavy Metals by Zooplankton . . A. Studies of Metal Fluxes through Zooplankton .. B. Food and Water as Sources of Metals for Uptake by Zooplankton. . .. .. .. .. .. C. The Effect of the Chemical Form of a Metal upon its Uptake by Zooplankton .. .. .. D. The Role of Zooplankton in the Biogeochemistry of Heavy Metals in the Sea . . .. .. .. VII. Laboratory Studies of the Toxic Effects of Heavy Metals upon Zooplankton . . .. .. .. .. A. The Effects on the Metabolic Activity of Zooplankton .. .. .. .. .. .. B. The Effects on the Feeding and Ingestion Rates of Zooplankton. . .. .. .. .. *. C. The Effects on the Growth and Development of Zooplankton. . .. .. .. .. .. D. The Effects on the Fecundity of Zooplankton .. E. The Effects on the Phototactic Response of Zooplankton .. *. .. .. .. .. F. The Effects on the Swimming Activity of Zooplankton .. .. .. .. .. .. G . The Combined Effects of Heavy Metals and Additional Environmental Stress upon Zooplankton

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Studies of the Toxic Effects of Heavy Metals upon Natural Populations of Zooplankton in Large Volume Sea Water Enclosures . . .. .. 457

IX. Heavy Metal Concentrations in Natural Populations of Marine Zooplankton .. .. .. . . 460

X. Conclusions

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Adv. mar. Biol., Vol. 16. 1978 pp. 1-231.

THE BIOLOGY OF PSEUDOCALANUS CHRISTOPHER J. CORKETTand IANA. MCLAREN Dalhousie University, Halifax, Nova Xcotia, Canada I. Introduction

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Reproduction .. .. A. SexRatio .. .. .. .. .. .. B. Oogenesis and Egg Laying . . .. .. .. C. Sperm and Spermatophore Production . . .. D. Mating . . .. .. . . .. .. . . E. Reproductive Rate .. .. .. .. F. Retrospects and Prospects . . .. .. .. Development and Growth . . .. . . .. .. A. Embryonic Development Rate .. .. .. R. Hatching .. .. .. .. .. .. C. Development Rate of Nauplii and Copepodids .. D. Longevity of Adults . . .. .. .. .. E. Body Size .. .. .. .. .. .. F. Body Composition and Weights .. .. G. Oil Storage .. .. .. .. .. .. H. Growth Rates . . .. .. .. .. .. I. Rate of Production of Egg Matter . . .. .. J. The ‘‘ Balance Equation ” and Growth Efficiencies .. K. Retrospects and Prospects .. .. .. .. Life Cycles in Nature .. .. .. .. A. General Features, Terminology and Approaches . . B. Representative Life Cycles . . .. .. .. .. .. .. C. Retrospects and Prospects .. .. .. Vertical Migration . . .. .. .. .. A. Ontogenetic Migrations . . .. .. .. .. B. Seasonal Migrations . . .. .. .. C. Die1 Migrations .. .. .. D. Retrospects and Prospects . . .. . . .. I .

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Production .. .. .. A. General Methods .. B. Production Estimates C. Retrospects and Prospects Parasites .. .. .. A. Dinoflagellates . . .. B. Gregarines .. . . C. Trematodes .. .. D. Nematodes .. .. E . Crustaceans .. .. F. Retrospects and Prospects Role A. B. C. D.

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in the Food Web .. . . Effect on Phytoplankton .. Predators .. .. .. Significance in the Food Web. . .. Retrospects and Prospects

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I. INTRODUCTION Over forty years ago, at a conference sponsored by the National Research Council of Canada, Russell (1934) put our subject into context: “ intensive study of the plankton in northern waters . . . supplemented

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by recent observations on the food of plankton-feeding fish have all pointed to the economic importance of a few species only. I n first rank can be placed Calanus Jinmarchicus, Temora longicornis, and Pseudocalanus elongatus . . , ” These three genera have all been extensively studied since, and Calanus has been admirably ‘(booked ” (Marshall and Orr, 1955). Perhaps because of its relatively large size, Calanus has been most favoured as an experimental animal and is much the best known copepod in a number of ways. Our knowledge of Pseudocalanus is somewhat complementary to that of Calanus. Out of an awareness, expressed even in the earliest copepod literature, of the extreme variability of size of PseudocaZanus in nature, has grown a rather precise set of (‘rules ” concerning its development, growth, and reproductive rates. Perhaps it can also be said that there has been more concern with the mean response to environmental variables in studies of Calanus and more interest in individual variation in studies of Pseudocalanus. The reader should be aware that we have generally avoided using papers in the vast copepod literature that make no direct reference to Pseudocalanua. This may disappoint readers who feel that a more complete or deeper account might have been inferred from systematic relationships. For example, the swimming of Calanus nauplii has been well described, and there is no reason to suppose that the morphologically very similar nauplii of Pseudocalanus would behave differently. However, since we can find no description of swimming of Pseudoculanus nauplii, we do not cover the subject. We have attempted to be analytical and synthetical where possible in our review, and do not simply summarize the observations and conclusions of other authors. Some readers may feel that on occasion we have selected or even abused the writings of others in the search for patterns and regularities. However, we have reserved our most personal assessments of research on Pseudocalanus for the sections in each subsection that we call ‘(retrospects and prospects ”. Therein we broadly assess what has been done and suggest promising (and unpromising) lines for future investigation. Some of our suggestions for future research may seem a little vague ; perhaps they have to be since real discovery is by nature unpredictable.

11. SYSTEMATICS A. Nomenclature 1. The Genus

The Genus was established by Boeck (1864) with the name Clausia, in honour of the late C. Claus. Later Boeck (1872) discovered that the

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name Glausia had been preoccupied by a parasitic copepod and therefore substituted the name Glausia with the new name Pseudocalanus. 2. Described species

(a) Pseudocalanus minutus (Kreryer, 1845) The first description of a species of the genus was the publication of a plate by Kreryer (1842-45), on which the animal was given the binomen Calanus minutus. No description of the plate was pubIished a t the same time, but since the plate was published prior to 1931 the plate and accompanying binomen are sufficient to describe a new species (I.C.Z.N.,Art. 16, a, vii). The original drawing is reproduced in Fig. 1 and is clearly a male copepodid V (see p. 30, Table I), although

FIQ. 1 . Calanus minutus Kreyer, 1845, the type species of Pseudocalanus. A male copepodid V. ( x 2.6 from original plate 41 in Kreyer, 1842-46.)

the diagram shows five thoracic segments and not the usual four. K r ~ y e r(1848) did publish, separate from the plate, a description of a male copepodid V of Calanus minutus in which he described the fifth thoracic segment as being rudimentary and free on its lateral and ventral sides. This evidence suggests that the fourth and fifth thoracic segments were incompletely fused or that the suture line of the fusion was still visible. This is not uncommon in Pseudocalanus and With (1915) illustrated copepodids V of both sexes showing small fifth thoracic segments. A problem arises since the date of publication of the binomen Calanus minutus is the date of publication of Kreryer’s plate, and we have been unable to obtain this exact date. It is known that the date must have been during or before 1845, when the last plates of this

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work were published (Sherborn and Woodward, 1901), and the date 1845 can therefore be used after the binomen (I.C.Z.N., Art. 21, f ) . According to de la Roquette (1842, p. 446), publication of the first plates had begun by 1842. We, therefore, cite both dates in our references except in purely nomenclatorial citations, K r ~ y e rpreserved some specimens which were examined by With (1915), who described them as “ belonging all to the penultimate stage (18 29) ”, presumably meaning one male copepodid V and two female copepodids V. However, Dr B. Frost (personal communication) has recently examined the same material and reports that it consists of three female copepodids V and one male copepodid IV. This appears t o exclude the possibility that the holotype is extant. I n view of other taxonomic problems to be documented below, great care should be taken in any future designation of a neotype.

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(b) Pseudocalanus elongatus (Boeck, 1864) P. elongatus was first given the binomen Clausia elongata by Boeck (1864) in a short description of adult males and females, with the observation that females were common and males rare in Christiana (Oslo) Fjord. Boeck made no reference to the earlier publications of K r ~ y e r(1842-45, 1848).

(c) Pseudocalanus clausii (Brady, 1865) Specimens of this species were originally found by Brady (1863) in the North Sea and erroneously ascribed by him to a quite different genus and species, Phadnna spinifera. Later Brady (1865) gave his specimens the new binomen Calanus Clausii, after the carcinologist C. Claus. (d) Pseudocalanus acuspes (Giesbrecht, 1 881) This species was originally described by Giesbrecht as Lucullus n. gen. acuspes, from the Bay of Kiel, and later given a very thorough description with excellent illustrations, by Giesbrecht (1882). (e) Pseudocalanus major G. 0. Sars, 1900 This species was described by Sars (1900) as “ so very resembling the type species [by this he meant P. elongatus] that I should have been very much inclined to regard it as only a large variety if both forms were not found together in the very same samples, without exhibiting any transitions ”. Sars (1900) did not refer to the publications of Krayer (1842-45, 1848), but does refer to Boeck (1864) and, implicitly, Boeck (1872),

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so that clearly he assumed that prior to his work the genus contained only P. elongatus (Boeck, 1864).

(f) Pseudocalanus gracilis G. 0. Sars, 1903 The classic work of Sars (1903) contains a description and figures of P. gracilis in the supplement at the end of the volume. He described the females of P. gracilis as being more slender than those of P. elongatus, as having a more conspicuously projecting frontal region and as having longer and narrower caudal rami. When females were placed on their side, the flexed first antenna was said to reach the end of the third urosome segment in P . gracilis, whereas in P . elongatus the first antenna reached only to the end of the genital segment. The male of P. gracilis was said by Sars ( 1 903) to resemble that of P. elongatus, the only distinction between them being the longer first antenna of the former. 3. Subsequent delimitations of described species

Pseudocalanus has been referred to under a variety of species names in the subsequent literature. Clearly the designation of at least some of the species has been less than satisfactory. We are concerned here with those papers that have attempted to clarify the status and characters of species. Much of the difficulty in subsequent work on the nomenclature of Pseudocalanus comes from the fact that Boeck, Brady, Giesbrecht, and G. 0. Sars did not mention the work of Krrayer in relation to the genus Pseudocalanus. Brady (1878) synonymizes his Calanus Clausii, 1865, with Pseudocalanus elongatus (Boeck, 1864). Giesbrecht (1882, addendum p. 167) admits the synonymy of Lucullus acuspes Giesbrecht, 1881, with Pseudocalanus elongatus (Boeck, 1864) and P. clausii (Brady, 1865). Giesbrecht (1882) says that he had not previously been aware of the identity of the species he described due to the poor original accounts given by Boeck and Brady. Giesbrecht also lists in his bibliography Kraryer’s (1848) text description of P. minutus, but not Krayer’s (1842-45) plate. Giesbrecht was thus clearly aware of Krrayer’s work, but probably did not consider the description to be detailed enough to be worthy of comments in connection with the genus Pseudocalanus. It seems inconceivable that Sars did not know of the work of his compatriot, H. Kraryer, but since Kraryer described an immature specimen, Sars may not have viewed it as Pseudocalanus. Clearly the species name minutus is available for a species of Pseudocalanus, and

THE BIOLOGY OF PSEUDOGALANUS

7

the question arises: is the form that Krayer described synonymous with P. elongatus (Boeck),P. major, G. 0. Sam, or P. gracilis, G. 0.Saw? With (1915) considered this question in detail and examined what he referred to as “Kraryer’s original specimens”, and was of the opinion that the three species described by Boeck and G. 0. Sars were synonymous, since he found transition specimens in shape of head, length of first antenna, and size. With did, however, state that Krayer’s specimens were in shape of the head most like P. gracilis, and were of middle size ’,. With (1915) therefore recognized only one species: Pseudocalanus minutus (Krcryer). Wiborg (1954) considered that G. 0. Sars was right in establishing three species and made use of the observations by With (1915) on Krcryer’s “ original specimens ” (see above) to designate P. gracilis as a synonym of P. minutus. He retained P. elongatus as a separate species and expressed some doubt about P. major, which he thought “may be an independent species or a large-sized P. elongatus”. He noted that P. minutus is normally larger than P. elongatus, but felt able to distinguish the two in samples from Norway, even when size showed considerable overlap, on the basis of body shape and length of second antennae. Although Brodskii (1948) indicated that two kinds of Pseudocalanus, differing in robustness of body, might be found in the Sea of Japan, he was content to follow With’s (1915) judgment of Sars’ species and to refer all his material to P. minutus. Later Brodskii (1950), in his major monograph on Calanoida of Soviet Far Eastern Seas and the Polar Basin, recognized three species : P. elongatus, P. major, and P. gracilis. In doing so he added to Sars’ (1900, 1903) criteria for separating the species, using relative lengths of the urosomes of adult females and the proportions of segments of the fifth legs of the adult males. However, in the synonomy of P. elongatus, Brodskii (1950) quotes Kraryer’s binomen, Calanus minutus, so that his three species should have been given as P. minutus, P. major, and P. gmcilis, according to the Law of Priority. Parran and Vervoort (1951) recognized three ‘‘ forms ” in one single species : Pseudocalanus minutus elongatus, P . minutus major, and P. minutus gracilis. Today, however, these “ forms ” should be considered as subspecies (I.C.Z.N., Art. 45, d, i, and Art. 45, e, i) and one of the three subspecies should be a nominate subspecies and have the same name as the species, i.e. Pseudocalnnms minutus minutus (I.C.Z.N., Art. 47, a). Fontaine (1955) synonymized all the described species under Pseudocalanus minutus (Krayer) and attributed size variations (some(‘

8

CHRISTOPHER J. CORKETT AND

IAN A. MCLAREN

times continuous, sometimes polymodal) in her material from northern Canada t o environmental influences. Kamshilov (1961) stressed the very great variation in size with no evidence of polymodalism, of Pseudocalanus in the White and Barents Seas and showed that the coefficient of variation (standard deviation/ mean) of size in his samples was twice that of Calanus (species not given). Furthermore, the ratio of cephalothorax length to length of urosome, which had been used (Brodskii, 1950) t o separate supposed species, had a continuous, unimodal distribution. He concluded that all his material should be referred to P. elongatus. However, here again, if this is done the name P. minutus has priority. Grice (1 962) stated that '' Pseudocalanus minutus was represented by two size groups ( P . minutus f. elongatus and f. gracilis) " in his collections from the Arctic Basin, but he did not routinely discriminate them. Cairns (1967) in his samples from the Canadian Arctic examined one criterion that had been used to define kinds of Pseudocalanus: there appeared t o be two groups of females, not completely separated by size, but showing a possible discontinuity, on a graph, of urosome length against cephalothorax length. The regression coefficients of urosome length on cephalothorax length were 0.520 for the small and 0.376 for the large ones. Within the wide continuous size range (about 1.0-1-4 mm in cephalothorax length) of the large animals, the length of the urosome relative to the cephalothorax decreased with size. Cairns did not attempt to refer his specimens to any species of the genus Pseudocalanus, and suggested that the large and small females might represent reproductively isolated forms or have resulted from different environmental conditions. Lacroix and Filteau (1971) believed that two " forms (per Farran and Vervoort, 1951), the small elongatus and the large major of Pseudocalanus minutus occur in the Baie-des-Chaleurs, off the Gulf of St. Lawrence. Adult females of the large form (cephalothorax mean 1.4 mm) were common in spring, but the small form (- 0.9-1.0 mm) predominated in spring and summer. The large form predominated as copepodid V (1.2-1.3 mm) in the deep, cooler waters throughout the summer. Enough examples have been given to indicate that problems of nomenclature and of delimitation of named species pervade the literature on Pseudocalanus. A formal systematic revision would involve the examination of much new material. Since we cannot do this here, we follow the practice of McLaren (1965) and refer only t o the generic name throughout this review on the biology of Pseudo))

-

THE BIOLOGY OF PSEUDOCALANUS

9

calanus. This is not t o deny the certainty that different kinds of Pseudocalanus exist, but that these kinds may be difficult t o accommodate either in the formalities of nomenclature or in prevailing concepts of species (see p. 11).

B.

"

Physiological '' species

Even if morphological differences among geographically isolated populations of Pseudocalanus are elusive, physiological differences occur that may signify reproductive isolation (cf. Carrillo B.-G. et al., 1974).

McLaren (1965, 1966) showed that the temperature response of adult female size and of embryonic development rate varied geographically. For example, extrapolation of the size-temperature relationship (Fig. 26) for Loch Striven, Scotland, to 0°C suggeststhat at this temperature (normal for a female from the Canadian Arctic) a monster larger than any known species of copepod would result. Of course this is hypothetical, since development would simply not take place a t this low temperature, and furthermore the size-temperature relationship may break down a t excessively low temperatures (see p. 122). Clearly, however, there are inherent differences between populations in these two parts of the world. Regional differences in development rate, for example, expressed as time to reach various stages (Fig. 25), are less pronounced. C. Va&ations in DNA content I n recent years a novel source of variations within and between populations of Pseudocalanus has been discovered. McLaren (1 965) described a large form of Pseudocalanus that coexists with a more abundant small form in Ogac Lake, a partially landlocked fiord on Baffin Island, northern Canada. The small form was believed t o be the same as the widespread Pseudoealanus of waters outside the fiord, the size of which had been reduced inside the fiord by elevated temperatures (see p. 117, Fig. 26). I n the large form embryonic duration (McLaren, 1966; see also p. 103, Fig. 22) and development times of older stages (McLaren, 1965 ; see also p. 113) are longer than those of the small form. McLaren (1965) speculated that the large form of Pseudocalanus was a polyploid. However, later work (McLaren et al., 1966) showed that both forms of Pseudocalanus from Ogac Lake contained the same chromosome number (n = IS), but that chromosomes in undivided

10

CIIRISTOPHER J. CORKETT AND IAN A. MCLAREN

eggs of the large form were much larger than those of the small form. The DNA content of nuclei a t the 32-cell stage was about seven times greater in the large form. This was attributed to polyteny, although today the whole question of repetitive or otherwise increased amounts of DNA tends t o be discussed in different terms. Woods (1969) added more information, demonstrating that the large forms in Ogac Lake and also in another landlocked fiord, Winton Bay on Baffin Island, were morphologically very similar, except in size, to the small forms and to the phenotypically larger forms of the cold seas outside. Female cephalothorax lengths of the large forms were respectively (means f S.E.) 1.09 f 0.040 mm and 1-16 f 0.044mm in Ogac Lake and Winton Bay, whereas the small forms were 0.85 & 0.009 mm and 0.84 f 0.007 mm. The small form is larger in cold waters outside the lakes, but its eggs are always smaller than those of the large form (see Table XVIII, and McLaren, 1965, his Fig. 1). Woods speculates on the adaptive meaning of these large forms. Noting that the effect of increased DNA per nucleus is probably an increase in cell size and a decrease in cell division rate, she suggests that this restores the normal size and cycle for an arctic population forced to exist in abnormally warm environments. Indeed, in Ogac Lake, McLaren (1969) showed that the small form may " waste '' much of its reproductive effort, since early broods matured and produced an unsuccessful second generation in summer. Under these conditions, there should be selective pressure for maturation later in the season, which would be thwarted by genetic exchange with the populations from outside the lakes, brought in by periodic high tides. Woods also notes arguments that larger size allows a greater range of food (see also p. 63). The large form during summer does show retarded maturation of the overwintered generation and slow progress of the new generation, compared with the small form (McLaren, 1969 ; Woods, 1969) thus restoring an essentially normal arctic life cycle (see p. 139). The large forms can be viewed as an " instant species ", almost certainly reproductively isolated from the coexisting small forms, in the manner of polyploid species among plants. It may be wondered if such discontinuous DNA variation is partly responsible for some of the sizepolymodalisms in Pseudocalanus noted in the literature. Clearly this raises nomenclatural and systematic problems : how many of the described species are of this sort? Can such variants, morphologically identical to widespread forms except in size, spring up independently among different populations, and should they be classed together on the basis of size? To what extent does such a size change allow rapid morphological divergence to occur!

THE BIOLOGY OF PSEUDOCALANUS

11

Recently the whole question of DNA variation in Pseudocalanus has become more complex. Hart and McLaren (1978) have shown that there is continuous variation in body size, egg size, and attendant embryonic duration in populations of Pseudocalanus from Halifax, Nova Scotia (p. 105). McLaren (1976b) showed that the size of adult females is strongly heritable (p. 123) and that size of adult females is strongly related to DNA content of somatic nuclei (p. 124). D. Retrospects and prospects What started out as a rather classical nomenclatural muddle has led t o a frontier ” problem in systematics. We feel that some of the nomenclatural confusion has been exacerbated by the attempts (e.g. Brodskii, 1950) to define species limits in terms of size and its correlates that are highly responsive to environmental influences and are also perhaps subjected to strong local differentiation under natural selection. We hope that the nomenclatural and systematic problems in Pseudocalanus will be cleared up by a revision of the genus that takes into account these aspects of variability. Further investigation is clearly needed on the role of quantitative DNA variation at the ecological, evolutionary, and molecular level. The DNA content of nuclei, presumably by mediating cell size and cell division rates (Woods, 1969),may act as a basis for the quantitative inheritance of body size and durations of developmental stages. This genetic basis for maintaining phenotypic diversity may be aided by size-assortative mating (see p. 84). Not only is such a mechanism quite extraordinary but its consequences may defy simple systematic analysis. Consider the possibility, given the strong heritabilities of size together with assortative mating, that disruptive selection could very quickly lead to two reproductively isolated populations where once there was one. Indeed, it is possible that ‘‘ biological species ” (sensu Mayr, 1963) could come and go rather quickly in this widespread genus of copepods. The situation in Pseudocalanus may find its counterpart in other groups of copepods, including the sibling species of Calanus, as mooted by McLaren et al. (1966).

111. DISTRIBUTIONAND ABUNDANCE

A. Geographical distribution 1. General Our survey of the distribution of Pseudocalanus is summarized in Fig. 2. Although well known in broad terms, its precise limits are

12

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

obscure in some places and its reported presence in some localities is dubious or in need of confirmation. Sewell (1948) collates virtually all earlier references to distributions of marine copepods, and it remains largely to update and correct his assessments. Sewell, like most previous and subsequent reviewers, concludes that Pseudocalanus is basically a neritic, northern genus, found in arctic seas extending southward along cooler coasts, and even beyond in deep waters.

FIQ.2 . World distribution of Pseudocnlnnus. Open-ocean boundaries in Atlantic after Edinburgh Oceanographic Laboratory (1973) and in Pacific after Omori (1965). Other sources in text. We consider this t,o represent the distribution of normally reproducing populations, although there are deep-wster records outside these limits, and animals may not commonly breed in the mid-Arctic Ocean.

I n considering the limits of the range of Pseudocalanus, Sewell cites records from as far south as Chesapeake Bay in the eastern U.S.A., from the North Atlantic Drift south of Iceland, from European waters as far south as Portugal, and from the Mediterranean. I n the North Pacific, he records it south t o Japan and Vancouver Island. I n addition, some anomalous records can be gleaned from Sewell’s review, and these must be examined.

THE BIOLOGY OF PSEUDOCALANUS

13

Sewell himself notes that supposed occurrences in the northern Gulf of Suez of Pseudocalanus and other North Atlantic forms are open to doubt. The samples in which these copepods occurred were from seawater taps draining tanks that were probably filled prior to passage of the ship through the Suez Canal. Sewell (p. 497) implies that Pseudocalanus reaches subantarctic or even antarctic waters through the deep Atlantic, but does not document this statement with references. Although he later (p. 499) lists the genus for deep Atlantic waters, he does not include it in his list (p. 513) of arctic or North Atlantic forms recorded from subantarctic or antarctic waters. Nor does it occur in the most extensive modern review of copepods of these waters (Vervoort, 1965). Sewell’s report of Pseudocalanus off western South America is based on the work of Wilson (1942). This very large work and a subsequent one by Wilson (1950) extended the distribution of Pseudocalanus over vast areas of the tropical and subtropical Pacific and Atlantic, far outside previously accepted limits. Some authors have corrected or expressed doubts about other records in Wilson’s (1942, 1950) lists. His records of Pseudocalanus seem to have been evaluated simply by being ignored in a number of subsequent publications on copepods of the waters surveyed by him. Contrary to Wilson’s claims, Pseudocalanus has not been found in extensive sampling of the California Current (Fleminger, 1967), off western South America (Bjornberg, 1973), in near-surface waters of the southern North Atlantic (Deevey, 1971), or in the tropical Pacific (Grice, 1961 ; Vinogradov and Voronina, 1963). A number of papers subsequent to Sewell (1948) give a more refined view of the distribution of Pseudocalanus. We cannot possibly consider more than a fraction of these, and confine our review to those that give a wide perspective or a more accurate assessment of the margins of its range.

2. Arctic Basin

Pseudocalanus is common in coastal arctic waters and has been recorded from many parts of the Arctic Ocean proper, generally in the upper 300 m (e.g. Dunbar and Harding, 1968). It is, however, evidently scarce and patchy in the more central parts of the basin, and has not been found in some surveys (e.g. Minoda, 1967). Harding (1966) found no subadults in his samples from the basin, and concludes that Pseudocalanus there is a n expatriate from surrounding neritic waters, especially the Chukchi Sea, where it is common (Grice, 1962).

14

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

3. North Atlantic and adjacent waters An excellent overview of North Atlantic distributions of adult Pseudocalanus has been constructed from the Continuous Plankton Recorder surveys (Fig. 2 18 in Edinburgh Oceanographic Laboratory, 1973). The marked abundance in upper waters off eastern Canada and western Europe contrasts with its general scarcity in the open Atlantic, where scattered records are shown southward to about 41"N. I n the inshore waters of eastern North America, Pseudocalanus has been noted as far south as Beaufort, North Carolina, a t latitude 34'40" (Pearse, 1936). However, this and a number of other identifications made by Wilson for Pearse's study must remain suspect, especially one reputed finding of Pseudocalanus innear-fresh water. Bowman (1971) states that Pseudocalanus does not occur south of Cape Hatteras, a zoogeographical boundary for many northern forms. It penetrates the Gulf of St. Lawrence to at least 60"20'W (Prdfontaine and Brunel, 1962). I n the eastern North Atlantic it can be common in the Bay of Biscay and off northwest Spain, but it is not listed for the waters off southwest Portugal (Vives, 1970). I t s distribution in the Baltic region has been recently summarized by Ackefors (1969a) and Arndt and Stein (1973). It penetrates the Gulfs of Bothnia and Finland in small numbers, but is evidently not found in shallow waters even in the northern extremities of the Baltic proper (Eriksson, 1973b).

4. Nediterranean and Black Xeas I n spite of the earlier accounts reviewed by Sewell (1948) the status of Pseudocalanus in the Mediterranean is uncertain a t best. Rose (in Trdgouboff and Rose, 1957) includes the genus in his taxonomic keys to Mediterranean copepods, but makes no mention of its occurrence in his text. Surveys and biological studies of copepods in the western Mediterranean by Gaudy (1962) and Vives (1967) fail to mention it or note its absence specifically. If it occurs a t all in the western Mediterranean, it must be as a rare expatriate from the Atlantic. I n the eastern Mediterranean, the thorough survey by Kimor and Wood (1975) failed to report it. However, Pseudocalanus does occur in the Adriatic. VuEeti6 (1957) states that " sporadic individuals '' occur all over the Adriatic, but Hure and Scotto di Carlo (1968, 1969) found it only in northern parts, where it was most common in May, although never dominant numerically. Recent work (Dr J. Hure, personal communication)

THE BIOLOGY O F PSEUDOCALANUS

15

indicates that it extends in some numbers down the coast of Italy, but not evidently beyond the Strait of Otranto. Pseudocalanus is also well known as a disjunct population in the Black Sea. Its distribution there is detailed by Afrikova (1975)) who maps its abundance by season and depth. 5. North Paci$c and adjacent waters

The best modern overview of the southern limits of distribution of PseudocaZanus in the North Pacific is supplied by Omori (1965, his Fig. 4). It is shown as most abundant in shallow waters of the Bering Sea and near the Aleutians, relatively abundant near the coasts of Hokkaido and southern British Columbia, less so in the open ocean in between. It was absent from some samples along the 43"N parallel between about 170"E and 175"W. On the Pacific coast of Japan, Pseudocalanus seems to be one of a number of boreal species that is not found west of the Bonin Ridge running south from the Tokyo region, although it occurs in Sagami Bay at the landward end of the ridge (Furuhashi, 1961). Furuhashi's southernmost records offshore are at 38"OO'N 145"23'E and (Furuhashi, 1966) a t 40"03'N 152'01'N. I n the Sea of Japan, Pseudocalanus occurs in the extreme southeast (Morioka, 1973) and off the coast of Korea (Mori, 1937). On the American coast, the southernmost reliable record appears to be at 38"52'N, just off the coast of northern California (Davis, 1949). 6 . Expatriates in deep water

The geographical limits outlined above can be considerably extended (and probably will continue to be) by scattered records from deep waters. These represent individuals carried from waters farther north, unable to rise to warmer surface waters, and presumably incapable of sustaining populations indefinitely. Ignoring problematical earlier records in Sewell (1948),we find records in deep water of two individuals off the Azores (Roe, 1972)) seven females a t 31"31'N 64"OO'W near Bermuda (Harding, 1972), and a southernmost recorded specimen a t 29'58% 22'58'W (Grice and Hulsemann, 1965). 7. Distribution in relation to water masses Copepod distribution is frequently discussed in relation to water masses or marine biogeographic zones. Pseudocalanus often figures as one of many " indicator " forms in such studies, only a few of which will be reviewed here. In the North Atlantic (e.g., Edinburgh Oceanographic Laboratory,

16

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

1973) Pseudocalanus is found in pure Atlantic as well as mixed arctic and Atlantic (subarctic, sensu Dunbar, 1947) waters. A sophisticated approach is found in Colebrook (1964), who used principal component analysis to group and classify copepods of the North Sea and North Atlantic. This results in an objective designation of " Para-Pseudocalanus " as a member of the intermediate group with respect to northsouth and neritic-oceanic gradients. Unfortunately, the lumping of Pseudocalanus with Paracalanus, a more southern form, makes the designation less useful. The southern limits of Pseudocalanus at about 42"N in the open waters of the western North Pacific (Omori, 1965) coincides quite well with the subarctic boundary, which is defined in strictly hydrographic terms (Dodimead et al., 1963). Pseudocalanus occurs in and is used in the definition of the North Pacijic temperate region of Brodskii (1956). 8. Distribution uith respect to distance offshore

Pseudocalanus is generally reckoned as a neritic copepod. This is very evident in maps of late-summer distribution in the Barents Sea (Zelikman, 1966, his Fig. 7), and on a grand scale in the North Atlantic surveys of the Continuous Plankton Recorder (Figs. 217, 218, in Edinburgh Oceanographic Laboratory, 1973). All the areas with the highest abundance of Pseudocalanus are within about 400 km of land. Nevertheless, Pseudocalanus does occur abundantly further offshore in northern extensions of the North Atlantic (Ostvedt, 1955). Motoda and Minoda (1974) refer to it as " typically oceanic " as opposed t o neritic in the Bering Sea. Possibly this is a matter of definition. It is at times most abundant in the central region of the Black Sea (Afrikova, 1975).

On a smaller scale, a number of authors have stated that Pseudocalanus is generally commoner away from the immediate vicinity of the coast. For example, Evans (1973) found that Pseudocalanus in 1969 was four times as common ten miles off the Northumberland coast as it was two miles offshore. However, Petipa et al. (1963) in a series of transects off the coast of the northern Black Sea found that regions of concentration varied, sometimes near shore and sometimes farther out. Furthermore, it may tend to become more abundant in enclosed bays than in the open waters outside. This is shown most clearly in a series of samples taken in summer 1960-62 from the Baie-des-Chaleurs, in the Gulf of St. Lawrence (Lacroix and Filteau, 1971). We conclude that Pseudocalanus is indeed predominantly a coastal form, but that the neritic-oceanic gradient is an unrefined one, allowing for many exceptions.

17

THE BIOLOGY OB P,SEUDOCALAhrUS

B. Abundance 1. General abundance

Pseudocalanus is not only widespread in northern seas, but is often said to be the most abundant form in many surveys of these waters. Perhaps the most impressive testimony to its numerical importance is from the long-term surveys with the Continuous Plankton Recorder around the British Isles (Fig. 3). Along with Calanus jnmarchicus (including C . helgolandicus, no doubt), Pseudocalanus (with a small admixture of Paracalanus) is found in virtually all the sampled areas. It is also numerically the most abundant form per sample. It ispossible, in our opinion, that Pseudocalanus is the most abundant metazoan in the world. 2.

'' Patches " and mms occurrences

Pseudocalanus may be found in '' patches ", many kilometres in diameter, that may be treated as dynamical and productive units (e.g. Thompson, 1976). This may be of considerable significance to A 1.6 0.4 02

B 0.01

0

0.05

0.1

0.5

I

5OloO 5

5

Pam-Reudacalanus rpp. Acartla a m .

Cantrapagas hamatus

Euchaob habas

Houromamma rabusta Pareuchadta narvefica Rhincalanus nasutus Plauramamma banalis Ploummamma adominah8 Phuramamma gmcilis Labidocero wllastoni Sapphirina app. Candacia armata Euchaeta acuta hfstridia /onfu Calanus minor Astidius armatus Anoma/acwa patsrsoni Calanus amcilis Contrapagas bmdy!'

i I

Plsummamma xiobias

).6 0.4 0.2

0

0.01

0.1 0.05

0.5

'

i 5

FIQ.3. Histograms of A, the proportion of sampled area (vicinity of British Isles) in which the species occurred, and B, the abundance of each species during 1948-66, as revealed by the Continuous Plankton Recorder Survey. (From Colebrook el al., 1961.) A.M.B.-15

3

18

CHRISTOPHER J. CORRETT AND IAN A. MCLAREN

fisheries (seep. 201). Fish (1936) interpreted such concentrations around the Gulf of Maine as " stocks " in the manner of fishery science. He felt that breeding occurred first in offshore areas and later inshore, where local concentrations were sustained by hydrographic circumstances from areas of higher production offshore. The number of stations involved makes some of his isopleths of abundance open to question. Soviet researchers have shown particular interest in such concentrations. Zelikman (1961) found mass occurrences of over 1oQ individuals per m3 near the mouth of the White Sea in midsummer 1956. Meshcheryakova (1964) describes less dense concentrations of Pseudocalanus and other copepods in the eastern Bering Sea. Explanations for this are vague, but a lack of coincidence with concentrations of Calanus is of interest. Kamshilov (1961) describes abrupt increases in abundance at boundaries of sharp temperature changes in surface waters on transects through the Barents and White Seas, ascribed to concentration by hydrographic forces. Zelickman and Golovkin (1972) agree that hydrographic forces are responsible for concentrations of Pseudocalanus and other zooplankters near bird colonies on Novaya Zemlya, but stress that the concentration is due to productivity, not " mechanical " consequences of hydrography. C. Temporal variations 1. Seasonality of occurrence Throughout its range, Pseudocalanus shows seasonal fluctuations in abundance in relation to primary production and other factors. Where life cycles are annual, it is clear that a numerical peak must occur during the season of reproduction. But even where more or less continuous generations occur, there may be marked seasonality in abundance. Some authors have attempted to discern large-scale patterns in this seasonality. Pavshtiks and Timokhina (1972) summarized the annual cycle of Pseudocalanus in the Norwegian Sea, showing that the summer peak of abundance occurs in late June in Atlantic waters, mid-July in mixed waters, and late July in the East Icelandic Current. Colebrook (1969) has systematized data from surveys with the Continuous Plankton Recorder by using the centre of gravity on the time axis of the area included under seasonal curves of abundance. Geographical variation in this statistic for Pseudocalanw (his Fig. 8, probably including some Paracalanus, although this was not stated) shows a reasonable pattern of early peaks in southern and coastal waters

THE BIOLOGY OF PSEUDOCALANUS

19

and later ones in northern and oceanic parts. Analysis of covariance shows that seasonality is correlated with temperatnre, but of course causes may be indirect. In the southern parts of its range, Pseudocalanus is a winter-spring form, and may disappear altogether from sampling areas during summer. Since this disappearance seems clearly related to high temperatures, i t is discussed separately (p. 24). 2. Year-to-year and long-term changes in abundance The whole subject of secular changes in the marine environment is of profound importance (Russell et al., 1971) and Pseudocalanus has figured in some of the discussions. 155C 965

-n E

oc

0

e"

594

36C 212

P .x 119 61

23

I

u +0 50 Ye0 r

FIG.4. Average numbers, per sample of 20 miles, o f Pseudooalanua (including Paracalanua) from the east-central North Sea in monthly periods from January 1948 to December 1972, taken by the Continuous Plankton Recorder. (From Glover et al., 1974.)

Pavshtiks and Timokhina (1972) tabulate a five-fold variation in production of Pseudocalanus (p. 187) in the Norwegian Sea in seven seasons between 1959 and 1969. This is vaguely related by them to the temperature regime and the timing of the spring maxima. Lacroix and Filteau (1971) found that Pseudocalanus averaged two and a half times as common in the Baie-des-Chaleurs, Canada, in 1962 compared with 1960. They suggest that a warm hydrographic

20

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

winter promoted strong vertical mixing and high production in the subsequent summer season in 1962. Meshcheryakova (1964) describes a roughly two-fold difference in abundance of Pseudocalanus and other forms in the eastern Bering Sea in 1958 and 1959. I n the former year, an earlier warming led to a strong diatom bloom in summer, which in turn encouraged strong representation of the copepods. Peterson and Miller (1975) found a marked reduction in abundance of Pseudocalanus off the coast of Oregon in 1971, evidently associated with reduced upwelling and warmer water than usual. I n the Black Sea, annual production of Pseudocalanus (p. 183) varied almost two and a half fold during the period 1960-66 (Greze et al., 1968.) The remarkable summary of long-term changes in the east-central North Sea by Glover et al. (1974) is summarized on Fig. 4 (whichincludes Paracalanus; G. A. Robinson, personal communication). These authors conclude that the long-term reduction in mean annual abundance is related to retardation in the time of the spring phytoplankton bloom, from late March to mid-April in the Atlantic off the British Isles, and from mid-March to mid-April in the North Sea. This has led to a reduced length in season of sustained production by the zooplankton from slightly more than seven months in the fifties to about six months in the early seventies. Underlying the biological trends is said to be a climatic trend involving the withdrawal of Atlantic influence from the North Sea,

D. Vertical distribution Pseudocalanus is generally found in the upper layers of the sea, although individuals have been taken as deep as 4 000-5 000 m (Grice and Hulsemann, 1965). Here we first outline briefly the main features of vertical distribution in the open sea, where the bottom may not set the deepest penetration. Then we consider inshore waters, where details of vertical distributions with respect to phydcal, chemical, and biological factors may be more evident than in the open sea. There is of course a dynamical aspect to this subject, which is dealt with at length elsewhere (Section XII). We refer throughout this section to daytime distributions of Pseudocalanus. The broad capabilities of Pseudocalanus are evident in vertical samples from deep, oceanic waters in various regions. I n the Norwegian Sea, most individuals occur above 50 m in spring, but below 1 000 m at other times of the year (Ostvedt, 1965). I n the southern Bering Sea,

TEE BIOLOGY OF PSEUDOCALANUS

21

&oda (1971) found that most animals were between 0 and 50 m in a series of samples taken from 28 May to 19 June 1962. Only a tiny fraction (average 0.1%) occurred in the deepest hauls (between 707 and 1 350 m). Over the deep Kuril-Kamchatka trench, 98% of the biomass of Pseudocalanus occurs in the upper 50 m, according to Arashkevich (1969). Dunbar and Harding (1968) found that almost all individuals were taken between 50 and 300 m under the ice of the Arctic Ocean ; at these depths, largely unmixed arctic waters are found. I n offshore regions of the Black Sea, which is of course anoxic in deeper waters, Pseudocalanus is found down to at least 200 m but is generally more common above 50 m (Afrikova, 1975). We conclude from these studies that Pseudocalanus is capable of living in very deep waters (unless prevented by lethal conditions, as in the Black Sea), but may only be evident if seasonal samples are taken. Studies of vertical distributions in shallower, inshore waters give more insights into the physical, chemical, and biological factors, that might control such distributions. Minoda and Osawa (1967) found that Pseudocalanus and other small copepods were concentrated by day at depths of the sonic scattering layers in the Okhotsk Sea in summer, 1963, and this coincided with the thermocline at the time. I n the Landsort Deep of the Baltic, Pseudocalanus was most abundant at 50-100 m, just below a thermal minimum, absent above 30 m, where temperature began to rise sharply, but present even down to 300-400 m, in spite of virtual absence of oxygen (Ackefors, 1966). At a shallower station south of Stockholm, Pseudocalanus always occurred below the thermocline at 20-30 m in spring and autumn, although a few were found near the surface in unstratified waters in winter (Ackefors, 1969b) ; salinities were low but varied only slightly with depth. I n a semi-landlocked bay on the island of Split, Yugoslavia (VuEetiE, 1961), Pseudocalanus occurred at the surface only in January and November, when the water column was almost isothermal and <13"C. Samples in May, June, and July showed it to be confined to deeper waters less than about 13°C. Carter (1965) and McLaren (1969) gave detailed accounts of the vertical distribution of all developmental stages in the strongly stratified waters of Tessiarsuk and Ogac Lake respectively, two landlocked bodies of seawater in northern Canada. Since much can be inferred from these studies about the ontogenetic and seasonal migrations of different stages, we discuss them later (pp. 158-163). It is sufficient to note that low salinities and low oxygen tensions respectively determined the upper and lower limits of distribution of Pseudocalanus. The work of Lee and Williamson (1975) in the Irish Sea also gives a

22

CHRISTOPHER J. CORKETT

m n I~LNA.

MOLAREN

wealth of information on the daytime vertical distributions of copepodids and adults (combined) at four stations a t several times of year, all accompanied by details of physical and chemical conditions in the water column. At the shallowest station (40 m) animals were generally commonest at mid-depths, whereas at another just slightly deeper (45 m) station they were more often found near the bottom. Both these stations had strong tidal currents and little thermal stratification. At a deeper station (100 m) there were single sharp peaks of concentration of three of the five sampling dates ; on each of these three dates there was distinct thermal stratification not present on the other two dates. The relative concentration at about 10 m on 22 January 1969 was more extreme than any other example depicted, and this concentration was in the middle of a sharp positive temperature gradient near the surface. Distributions at the deepest station (275 m) were remarkably uniform with depth at all times of year ; there was almost no thermal stratification. Lee and Williamson also depict the vertical distribution of NO,, NO,, PO,, and SiO,, which in some instances showed strong stratification when temperatures and salinities appeared uniform. Lee and Williamson single out two stations in March as showing chemical stratification in relation to copepod distribution. I n the shallowest station a high NO, value near the bottom is correlated by them with smaller numbers there. On the other hand, in the deepest station a sharp maximum of NO,, NO,, and PO, occurred a t 100 m, which according to Lee and Williamson coincided with a peak of Pseudocalanus; this peak appears to us to be unconvincing.

E. Xampling problems and microdistribution Three different problems may thwart accurate estimates of the mean numbers of copepods living in a sampled unit of water : (1) the sampling gear may allow some individuals to slip through the net meshes ; (2) individuals may avoid being captured by the apparatus ; and (3) individuals may be arranged in a clumped manner in the water, and subject to being undersampled or oversampled in any one catch. It is obvious that many studies of copepods including Pseudocalanus have badly undersampled the young stages. The work of Carter (1965), McLaren (1969) and others indicates that a mesh of No. 20 (apertures 76 pm) is adequate for all stages (eggs being of the order of 100 pm at least). A formal study of size selectivity achieved by sampling Pseudoculunus and other forms is offered by Sherman and Honey (1971). McLaren (1969) showed that a, 12.5 cm Clarke-Bumpus sampler towed vertically a t about 0.5 mls obtained on average about the same

TEE BIOLOGY OF PSE UDOUALAN US

23

number of adult females (mean 5% more, range 29% less to 34% more) as predicted from parallel hauls with a net 0.5 m in diameter. The difference between the two nets was significant only on three of seven occasions during the season. There was thus no good evidence that females can escape a net as small as 12.5 cm in diameter towed at this speed. The superiority of vertical net hauls in dealing with vertically stratified zooplankton, including Pseudocalanus, is confirmed by sampling in enclosed experimental systems (Lawson and Grice, 1977). Of greater interest is the possibility of real horizontal variations in microdistribution. The parallel vertical net hauls of McLaren (1969) described above gave weak evidence that clumping on a horizontal scale of a few centimetres or metres did not occur in Ogac Lake,a peculiarly stable system hydrographically. Barnes and Marshall (1951) used a rapidly taken series of pump samples of about 6 1 each to test the distributions of various forms at 10 m off Millport, Scotland. In one series, nauplii of Pseudocalanus showed significantly nonrandom clumping, whereas nauplii of a number of other copepods did not show any such tendency. Barnes and Marshall suggest that such small scale clumping may result from the fact that Pseudocalanus, unlike the other species, carries its eggs until they hatch as nauplii. Recently Smith et al. (1976) have demonstrated marked aggregation of adult Pseudocalanus and other copepods in shallow waters off the coast of Oregon. They found that concentrations varied three- to ten-fold over horizontal scales of 30 to a few hundred metres. This variation was attributed to physical processes, largely related to vertical displacements of marked depth-peaks of concentration by internal waves in this near-shore setting. Presumably vertical net hauls, as opposed to the pump samples from fixed depths used by Smith et at., would have shown less variability on a horizontal scale.

F. Physical-chemical limits to distribution 1. Temperature

Psezldocalanus often tolerates great temperature variations throughout its range. However, temperature clearly is involved directly or indirectly in setting its limits of distribution. Although Kolosova (1975) concludes, from extensive field sampling, that Pseudocalanus in the White Sea has a “temperature preference” of 2-8”C, this may reflect a wide variety of hydrological and biological causes rather than any debilitating effects of extreme temperatures. The limits for existence presumably exceed those for reproduction and development. McLaren (1966) lists for a number of localities experi-

24

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

mental temperatures at which successful embryonic development occurred. A t 0°C a t least some eggs hatched in all localities. The highest temperatures listed are 7.1 and 7~9°Cfor two localities in the Canadian Arctic, 12-0°C for Halifax, Nova Scotia; 13-1"C for Woods Hole, Massachusetts ; and 14.9OC for Loch Striven, Scotland. Corkett and Zillioux (1975) hatched eggs from Plymouth at 16-2"C. Thompson (1976) found that individuals from the southern North Sea could be reared at 3.7"C only up to NV, whereas those at 5-0"C could reach maturity. We shall indicate later (p. 108), that development below 8°C may have been in other ways abnormal. It is expected that local populations will be adapted to local temperature extremes, but it is of some interest to consider the absolute thermal limits of Pseudocalanus in nature. Deevey (1960a) graphs the extreme range " as -1.8' to 24.5"C and the " breeding range " as -1.0" to 19°C. However, these ranges must be looked at more critically. There is no doubt that Pseudocalanus exists and must a t times breed in the arctic a t the temperature of freezing of full sea water (ca. -1-8"C). The upper limit is less certain. Sampling techniques must take into account the fact that Pseudocalams may occupy the cooler parts of stratified water columns. The record of a few individuals at 20-21OC in the unstratijied Cape Cod Canal (Anraku, 1964a) must reflect its real potential for existence there. The occurrence (noted as " rare ") a t 23°C in the Japan Sea (Morioka, 1973) also seems to have taken full account of vertical stratification of temperature (the species otherwise was found only at or below 13°C). I n the Black Sea, where surface temperatures in open water probably exceed those found in any other part of the range of Pseudocalanus, it stays in cooler depths in summer and autumn, but Porumb (1973)says that it almost disappears when surface waters warm to 23-26.5"C. We are unable to find any satisfactory upper temperature limit for breeding of Pseudocalanus in nature. Deevey (1960a) found eggs in samples from Delaware Bay in June, but only surface temperatures are given, and we have no indication that eggs were viable. 66

2. Salinity There is limited laboratory information on the adaptability of Pseudocalanus to extreme salinities. McLaren (1966) found that mortality during embryonic development was equally high at salinities of 25.7 and 32-3%, for animals from Ogac Lake, Baffin Island, a brackish-water environment. McLaren et al. (1968) concluded that mortality of developing embryos was significantly higher only at the lowest experimental salinity of 2 5 ~ 2 %for ~ animals from full strength

THE BIOLOGY OF PSEUDOCALANUS

25

sea water near Halifax, Nova Scotia. This seems to be weak evidence for local adaptation to low salinities of Ogac Lake. Certainly Pseudocalanus in nature is quite euryhaline or, properly " euryhaline-marine " (Jeffries, 1962). It is known from quite saline waters (37%,) in the Adriatic (VuEetid, 1961), but presumably does not have to contend with much greater hypersalinity. However, it is often found in highly freshened waters. Kuznetsov (1973) found that Pseudocalanus avoided the freshened (6-80/103)upper 6 m of the Laptev Sea in October. The lowest estimate of the limiting salinity appears to be that of Hessle and Vallin (1934), who state that it occurs at 4-5%, in the Baltic. However, this estimate may not have taken full account of stratification. The very accurate vertical sampling in the Baltic by Ackefors (1969b) revealed 6 ~ 0 %as ~ the lowest salinity where Pseudocalanus occurred. This is the lowest reliable estimate of which we are aware. Ackefors (1969b) and others have noted that Pseudocalanus is found in such low salinities only when the water is cold, in the depths in summer and near the surface only in winter. Ackefors also suggests that it does not spawn in the shallow waters near Asko, near Stockholm, where salinity does not exceed 7.5%,. 3. Oxygen

Pseudocalanus lives routinely in the Baltic at depths where the oxygen concentration is usually less than 2 ml OJl, and sometimes very near zero (Ackefors, 1969a). The lowest oxygen concentration where Pseudocalanus has occurred can be inferred from Ackefors (1966), who found some individuals between 400 and 500 m in the Baltic in August, when oxygen below 300 m was 1 ml/l and only 0.04 ml/l a t 440 m. A few individuals were taken at 31 m in Ogac Lake, Baffin Island, by McLaren (1969). At this depth oxygen was unmeasureable, but the very strong stratification on a small depth scale makes this record less reliable than those of Ackefors. Clearly, however, Pseudocalanus is capable of withstanding virtual anoxia. 4. Pollutants Studies on the effects of pollutants on marine organisms are becoming commonplace, and a few published experimental studies deal with Pseudocalanus. Grice et al. (1973) found that acid waste sufficient to reduce seawater pH to 6.7 (control 7.8) gave 30% mortalities of adults in 24 h. Lower pH gave lOOyo mortalities. They concluded that these effects were unlikely to be important in nature.

26

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Lee et al. (1977), using large polyethylene enclosures in the sea, found that initial fuel oil concentrations of about 40 p.p.b. (major non-volatile components) above the control concentration, had no measureable effect on zooplankton populations that were 44-91 % Pseudocalanus. Gibson and Grice (1977), again using large polyethylene enclosures in which Pseudocalanus was the predominant copepod, added concentrations of copper. At the highest level of 50 pg Cull Pseudocalanm populations fell to 50% of their original levels at a rate 2.5-3 times faster than did controls. Reeve et aZ. (1977) studied rates of ingestion and production of faecal pellets by P~eudocaZ~n~s removed from these enclosures. Some depression in these rates when water samples from the enclosures were used was due to the reduction of food organisms by effects of enclosure or copper, so that rates determined on standard food (from outside the enclosures) give a more reliable indicator of effects of copper as a stress on the copepods. Ingestion rates on the standard food by control copepods (mean 14-2 pm3/cop./day) were below those that we believe to be “saturation” levels (see p. 68). There was no clear effect of copper by day 2, but by day 4 both ingestion rates and faecal production were reduced. By day 9, when populations in the enclosures had been severely depressed, these rates among animals exposed to 5 pg Cull were about half those of controls. I n these enclosures, there was a shift to smaller phytoplankton species. Steele and Frost (1977) speculate from their simulation models involving Pseudoca1anu.s (see p. 209) that this shift in phytoplankton size could have resulted because of depletion of their grazers, largely Pseudocalanus.

G. Retrospects and prospects Although the overall distribution of Pseudocalanm is well documented, we lack knowledge of its precise southern limits in offshore waters in particular. It would also be useful to have more refined information on the status of populations at the fringes of its range: whether they occur year-round or seasonally, in shallow or deep water, breeding or not. Pseudocalanus clearly adapts to a wide range of physical and chemical conditions, and it should not be assumed that its geographical limits are set solely by physiological tolerances. We shall show later (Sections IX and X) that temperature has profound effects on size, development rate and reproductive rate, and therefore on population dynamics. It may be that extreme conditions limit a population’s resistance to

THE BIOLOGY O F PSEUDOCALANUS

27

predation or ability to compete with other forms before the limits of physiological tolerance are reached. Lee and Williamson (1975) quite rightly point out that from their work selected examples of vertical distributions . . . could be quoted as supporting nearly all the conflicting statements that have been in previous publications on the vertical distribution of P . elongatus ”. Yet vertical distribution in relation to temperature stratification often figure in such publications, with animals seeming to concentrate in, over, or under thermoclines. This seems to suggest that they may choose ” not to enter warmer or colder waters even when this is well within physiological tolerances. Of course temperature is very often correlated with other variables : with salinity gradients, although these often seem slight by comparison ; with food, which may accumulate in or near thermoclines; and indeed with almost anything that varies vertically. We shall later discuss the possible significance of temperature in vertical migration (pp. 175-179). Long-term studies of Pseudocalanus and other marine organisms may tell us much about natural and man-caused changes in the biosphere (see Glover et al., 1974). The experiments discussed here deal largly with acute responses of Pseudocalanm to pollutants. Eriksson (1973a)has suggested that sex ratios of Pseudocalanus may be distorted in coastal waters by accelerated mortality of females. Steele and Frost (1977) have deduced from their simulation models that stresses like pollution may in general tend to favour small plant cells and therefore smaller copepods like Pseudocalanus. Although speculative, these arguments illustrate the kind of population information that may be required to understand future changes in distribution and abundance of P8eudocalan.u~. ‘I

‘I

IV. MORPHOLOGY Pseudocalanus is typical of most crustaceans in that after hatching at an early stage of development it adds successively new segments and appendages. Pseudocalanus hatches as a nauplius, which is the most immature larval form among the arthropods. There are six naupliar stages (abbreviated N I-N VI) followed by six copepodid stages (abbreviated c! I--C VI), of which the last (C VI) is the adult. It is possible to divide the body from C I onwards into three regions or tagmata according to two different kinds of schemes. The first of these is based on theoretical considerations and we use the three terms,

28

CHRISTOPHER J. CORKETT AND IAN A. MULAJ3EN

head, thorax and abdomen for all stages. The position of the border between thorax and abdomen is arbitrary, and is discussed later. The second scheme is practical and is based on the observable shape of the body. We use for the three tagmata of this scheme the terms cephalosome, metasome and urosome.

1

N1

NIP

NY

FIQ.6. Characters of distal segment of first antenna and posterior end of body used to identify Pseudocalanus nauplii. (From Ogilvie, 1963.)

THE BIOLOGY OF PSE UDOCALAN US

29

A. Embryo The embryology of Pseudocalanus has not been investigated, but Corkett (1966) photographed the external features of eggs of Pseudo&nus. After Marshall and Orr (1955) the following stages of Pseudocalanw eggs were observed and photographed ; two-cell stage, segmentation-cavity stage, gastrulation stage. This is followed by a period when no detailed structure is visible until the developing nauplius can be seen, which occasionally twitches prior to hatching. The egg membranes are described in a later section (p. 107).

B. Nauplii The naupliar stages I1 to VI of Pseudocalanus are illustrated by Oberg (1906). N I of Oberg (1906) is really N I1 (Ogilvie, 1963). The newly hatched nauplius (N I) is illustrated by Corkett (1968). Keys to the identification of Pseudocalanus nauplii are given by Ogilvie (1953) and Faber (1966). The most useful character for the identification of nauplii is the arrangement of the caudal armature as seen from a ventral view. Figure 5 gives a summary of the characters used in the identification of the six naupliar stages of Pseudocalanus. Ogilvie (1953) expresses some of these characters in tables useful for discriminating different genera and species. The first three naupliar stages contain only three pairs of appendages, but in N IV the posterior portion of the body has lengthened and the rudiment of the first maxilla has appeared (Oberg, 1906). By the time N VI is reached there are rudiments of thoracic appendages. The appearance of rudiments of segments in N IV, N V, and N VI means they are sensu strict0 metanauplii (Snodgrass, 1956), but the term metanauplius is not normally used and Pseudocalanus is universally referred to as having six naupliar stages.

C. Copepodids (CI-C V ) 1. Structure

The basic structure of copepodids (along with adults) is summarized in Table I. The cephalosome is the observable large, anterior division of the body ; it is not divided into observable segments. The metasome consists of those observable segments between the cephalosome and the main body articulation with the urosome. The urosome consists of the narrow observable segments behind the main body articulation.

30

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

TABLEI. NUMBERS AND ARRANGEMENTS OB SEUMENTS AND SWIMMINU FEETIN Pseudocalanus (modifiod after Corkett, 1968)

Stage

sex

I I1 I11

-

IV

? d ?

-

-

IV

V

s

V

9 d

VI (Ad.) VI (Ad.) ~

No. NO. No. No. No. observable observable observable observable pairs of thoracic abdominal meta-some urosome swimming segments segments segments segments feet 4b 5b 6 5

5 5 5 3” 4

l b

l b

1 2 2 3 3 3c 4

2 3 4 4 5 4

5 4

5=

~~~~~~~

Fifth highly modified. After Oberg (1906). Plus genital segment (see text p. 34).

The basic structure of the five immature copepodid stages are given in Table 1, and we do not repeat all this information below. Corkett (1968, his Table I) used the term “segments thoracique libres” instead of metasome. 2. C I

This stage has three metasome and two urosome segments as in all copepod species (Gurney, 1942). Oberg (1 906) states that in the early copepodid stages the small anterior segment of the urosome was separated from the metasome during moult and therefore belongs to the thorax. We therefore place the border between thorax and abdomen between the first and second urosome segment (Table I). C I has two pairs of swimming feet and the rudiments of the third (Oberg, 1906). 3. C I I We consider (as in C I ) that the border between the thorax and abdomen lies between the first and second urosome segment (Oberg, 1906). C I1 has the rudiments of an additional fourth pair of swimming feet. 4. C I I I

It is not clear where the border between the thorax and abdomen lies (either at the main body articulation or between the first and second

THE BIOLOQY OF PSEUDOCALANUS

31

urosome segment), but the latter is likely by analogy with C I , C 11, C V 9, and C VI.

6. C I V Sexual dimorphism appears at this stage. The fifth pair of swimming feet in the male is small and uniramous (Kraefft, 1910). The fourth metasome segment is the result of the fusion of the fourth and fifth thoracic segments, but this fusion is occasionally incomplete. It is again not clear where the border between the thorax and abdomen lies (see above) : probably between the fist and second urosome segments. 6.

CV

As above (C IV) the fifth pair of swimming feet in the male is small and uniramous (Kraefft, 1910). Here again the fourth metasome segment is the result of the fusion of the fourth and fifth thoracic segments with occasional cases of incomplete fusion as in the C V 8 illustrated in Fig. 1. From the side views of the female urosome of C V in Kraefft (1910) it seems clear to us that the border between thorax and abdomen lies between the first two urosome segments, which are swollen at this stage prior to fusion to form the genital segment in C VI. When young female stages of Pseudocalanus are parasitized, sometimes this fusion does not take place (see p. 194).

D. Adults (C V I ) 1. Arrangement of somites and appendages into tagmata

Within the arthropods somites are grouped into tagmata, such as head, thorax, and abdomen. Use of such a term does not necessarily mean that a structure by this name contains the same homologous somites in different groups of Crustacea. I n the adult of Pseudocalanus the first thoracic segment is fused with the head (unlike all previous copepodid stages) to form the cephalosome; this leaves three metasome segments (Table I). The theoretical and practical arrangements of somites and the names for tagmata useful for adult Pseudocalanus are given in Table I1 and some are illustrated in Fig. 6. Column 1 in Table I1 shows the basic theoretical number of somites which are homologous to those of other Crustacea (Borradaile et al., 1961, their Table 7). I n column 2 of Table I1 we show the theoretical distribution of somites into the three previously mentioned tagmata. We adopt the position of Owre and Foyo (1967) and consider the maxillipeds as being the last pair of head appendages and for convenience consider the

TABLE11. THEORETICAL AND OBSERVABLE ARRANGEMENTS OF SOMITES,TAGMATA, AND APPENDAGES IN ADULTPseudocalanus 1

Theoretical somites 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 Telson

2

3

Theoretical arrangement of somites into tagmata 1 2 3 4 5 6 1 2 3 4 5 6 1 2 3 Anal segment

Head

Thorax

I J

Abdomen

4

Observable tagmala in male.

Appendages

First ant,enna Second antenna Mandible First maxilla Second maxilla Maxilliped 1st swimming foot 2nd swimming foot 3rd swimming foot 4th swimming foot 5th swimming foot Genital operculiim ($2)

I

Cephalosome

5

G

Observable tagmata in female

Cephalosome Cephalothorax

J

Metasome fused

Metasome

genital segment

genital segment Urosome

lurosome

Jfused

33

THE BIOLOGY OF PSEUDOCAL-ANUS

First antenna Second antenna Mandible First maxilla Maxii I iped First swimming foot thoracic somite Second swimming foot thoracic somite Fourth and fifth thoracic sornites

Third swimming foot Fourth swimming foot Genital operculum

Genital segment (sixth thoracic somite and first abdominal somite) Second abdominal somite Third abdominal somite

Fro. 6. Diagrammatic representation of an adult female P8eudocatanus (see Table I1 and text). The abdominal and thoracic somites are numbered in theoretical sequence and the anterior observable tagma is the cephalosome. (After Sam, 1903, and Owre and Foyo, 1967.)

somite bearing the first antenna as being the first somite, thus giving rise to six head somites. We further adopt the position of Owre and Foyo (1967) and consider the next six somites as belonging to the thorax. Owre and Foyo (1967) state that the abdomen, as defined by them, contains " one to five somites " ; this statement is misleading as there are only four possible somites behind the thorax as they define it. We

34

CHRISTOPHER J. CORKETT AND L4N A. MCLAREN

therefore consider the abdomen as containing three somites followed by the telson or anal segment. Column 3 shows the well-known sequence of appendages. Columns 4 and 5 show the practical subdivisions of the body for both sexes in Pseudocalanus according to the system of Owre and Foyo (1967). The fourth and fifth thoracic somites are fused to form the third metasome segment. In the male the genital segment corresponds to the sixth thoracic somite only. I n the female the genital segment is formed by the fusion of the sixth thoracic somite and first abdominal somite. The genital operculum of the genital segment represents rudiments of the sixth thoracic appendages. Finally a useful division is the cephalothorax (column 6) which contains all the main body apart from the urosome and is that part of the body measured in biometric studies (cephalothorax length). 2. Appendages

(a) Structure With the exception of the first antenna the appendages of the Crustacea may be reduced to one or other of two types : the biramous limb, which in copepods includes the second antenna, mandible, maxilliped and thoracic legs, and the phyllopodium, to which belong the first and second maxillae. For the purpose of this review the two branches of the biramous limbs will be referred to as an internal endopod and an external exopod. On the phyllopodium type of limb projections occur and these will be referred to as endites when internal and exites when external. A more detailed account of the structure of copepod limbs is given by Gurney (1931). (b) First antenna The first antenna is a structure sui generis. It does not belong to the biramous or phyllopodium type and is not comparable in detail with the structure of any other crustacean limbs (Borradaile et al., 1961). The first antenna in the Calanoida never has more than 25 segments, and this number is frequently reduced by fusion of segments 1 and 2, 8 and 9 and more often 24 and 25 (Gurney, 1931). Griffiths and Frost (1976) illustrate the first antenna of male and female Pseudomlanus with the female first antenna possessing 23 free segments of which the first and second, eighth and ninth appear fused (Fig. 7B). The male first antenna has further fusions of segments especially at the base where there are well developed aesthetes (Fig. 7A). The aesthetes are chemo-sensoryin function and are probably used to detect

THE BIOLOGY OF PSE UDOCALANUS

36

C

'A

t

en

t ex

'

rl exp

FIQ.7. Appendages of adult Pseudocalamzca. A, first antenna of 3. B, first antenna of 9. C, second antenna of $2. D, mandible of 9. E, first maxilla of 9. F, second maxilla of 9. G, second maxilla of 6. H, maxilliped of 9. I, first left thoracic (swimming) leg of ?. J, second left thoracic (swimming)leg of ?. K, fifth pair of thoracic legs of 3. Symbols are: a aesthetes; b blade; en endite; end endopod; ex exite; exp exopod; 11 left leg; rl right leg; s stout seta; sb serrated blade; t teeth. (A and B from Griffiths and Frost, 1976; C, D, E and H from Schnack, 1976; F from Marshall and Om, 1966; G, I, J and K from Sam, 1903.)

36

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

a pheromone produced by freshly moulted females prior to mating (Pa 84).

(c) Second antenna The second antenna contains an exopod and endopod with well developed terminal fans of setae (Fig. 7C). The second antenna is used by Pseudocalanus in locomotion (see p. 5 2 ) and in filter feeding (see p. 54). (d) Mandible The mandible consists of a blade used for mastication of the food (the morphology of the blade is considered in connection with feeding mechanism, see p. 57). On the mandibular blade is situated a biramous palp (Fig. 7D) which is used with the second antenna to create water currents used in locomotion and feeding. (e) Pirst maxilla The first maxilla is a flattened phyllopodiuin type of appendage with a well developed endite and exite that play a part in feeding (Fig. 7E). The long flexible setae on the exite help maintain the feeding current (see p. 54) and the teeth on the endite (or gnathobase) are used to comb food from the setae and setules of the second maxilla before the food is passed to the mouth. (f) Second maxilla The second maxilla (Fig. 7F) is the appendage used to filter food from the feeding current. This phyllopodinm type of appendage has a number of endites with well developed setae on which are situated setules. The arrangement of setae and setules and their use in filter feeding is considered later (p. 54) as is the possible use of the terminal " stout seta " (p. 58). The second maxilla in the adult male is vestigial (Fig. 7G) showing that filter feeding cannot take place in the adult of this sex. (g) Maxilliped The maxilliped consists of a single endopod (Fig. 7H) with well developed setae. The setation is not reduced, unlike that found in carnivorous copepods (Anraku and Omori, 1963 ; Arashkevich, 1969 ; Schnack, 1975). Rotation of the maxillipeds is used to enhance the feeding eddies (see p. 54).

THE BIOLOGY OF PSEUDOCA LA N V S

37

(h) Thoracic appendages The first four thoracic appendages or swimming feet are flattened biramous appendages (Fig. 71, J) probably used in the escape reaction to avoid predators (see p. 53). It is possible the shape of these appendages are responsible for the name “ copepod ” ( K W T ~ ,oar ; TOVS, foot), a name first used by Milne-Edwards in 1830 (Russell, 1934). The exopod on the second, third and fourth swimming feet of the female has a terminal serrated blade (Fig. 7J). The adult female has no fifth thoracic legs but the adult male has a highly modified fifth pair of legs consisting of a three-segmented right leg, and a five-segmented left. Sam’ illustration (Fig. 7K) hints at fusion of two segments in the first basal segment of the right leg.

E . Retrospects and prospects Although its external anatomy can be satisfactorily described in general terms, little is known of details that might serve to separate different kinds of Pseudocalanus (see Section 11). Furthermore, virtually nothing is known (except by inference) about the internal structure of Pseudocalanus. For purposes of description and exposition, we have found it useful to compare theoretical and practical arrangements of the series of segments and appendages (Table 11). There is still considerable confusion in published accounts of such series, and we hope our approach will find more general use among copepodologists and others. Future work on the morphology of Pseudocabnus in relation to function should certainly include studies of the fine structure of the first antenna1 aesthetes with which males are so richly endowed. It would also be useful to have insights into the ways in which the fifth legs of the males function in mating.

V. RESPIRATION Respiration of Pseudocalanus has been quite extensively documented, generally as one of a number of forms in comparative studies. The ultimate aim of such studies has generally been to supply one component of the “ balance equation ” of growth or production (see p. 181). Respiration is measured as oxygen consumption. Little appears to have been done using more biochemical approaches to metabolic rate, although Muhammad (1965) has demonstrated a strong correlation between oxygen consumption and succinic dehydrogenase activity in a variety of mixed zooplankton, including Pseudocalanus.

38

CHRISTOPHER J. CORKETT AND IAN A. MCLBREN

A. Factors injuencing rate of oxygen consumption Our survey of the determinants of respiration rate deals with variables that are expected to be important in nature. Little appears to have been done on certain more methodological questions. Pavlova (1975) has shown that degree of crowding may be important in Pseudocalanus. Rates of 0, consumption (means & 95% c.1. in pg/cop./h) by uncrowded (3-4 cop./l6 ml) adults were 0.225 f 0.032 for males and 0.190 f 0.223 for females; for crowded (10-12 cop./l5 ml) adults, they were 0.092 & 0.030 and 0.086 f 0.037 respectively. Generally speaking, however, authors have standardized such matters for comparisons between experiments. 1. Body size Conover (1959) was evidently the first to include PseudocaZanzcs in a relationship between rate of oxygen consumption (R, in p1 . O,/cop./ day) and body weight (W, in mg dry wt) for a number of copepod species: 0.856 log W. Log R = 2.068 The relationship for four experiments using adult female Pseudocabnus alone, with a length range of 0-80-0.91 mm, at 20°C (probably an excessive temperature for the animaIs from Southampton, England), gave a regression coefficient of 0.433 which was less reliable than the value of 0.856 for all species (see above). Because weight of an organism may be variously made up of metabolically inactive material, such as stored oil, ripe ova (as opposed to rapidly growing oocytes) or gut contents, cephalothorax length might be a better predictor of respiration rate of like-shaped animals. Raymont (1959) assumed that respiration followed a " surface law ", and that it would be proportional to the square of length, and therefore corrected respiration rates of copepods accordingly. However, within the size range of Pseudocalanus used by him (adults of both sexes 0-94-1.13 mm) no correlation of respiration rate and length was noted. Conover (1959) fitted a relationship of respiration rate (R, as above) to cephalothorax length (L, in mm) for a variety of copepods: log R = 0.353 2.713 log L. The less reliable regression coefficient for PseucEocalanus alone was 1.645. We do not have data for respiration of nauplii and copepodids of Pseudocalanus from which to derive an intraspecific relationship, and such experiments as conducted by Conover (1959) demonstrate that i t is difficult to use adults or copepodids of a narrow size range to derive an accurate relationship between respiration and body size of

+

+

39

THE BIOLOGY :OF PSEUDOCALANUS

Pseudocalanus. It is better to use a general, interspecific relationship, making use of a wide size range of animals. Probably the best available interspecific relationship to date is that fitted by Ikeda (1974) to " boreal " zooplankton, including Pseudocalanus: log R = 1.437 0.783 log W. Again R is pl O,/cop./day and W is dry w t in mg. We can transform this for cephalothorax length (L in mm) by using the length-weight relationship given on p. 127. log R = 2.850 log L - 0.070. The mean habitat temperature for the " boreal " species in this relationship was 8.6"C and, although Ikeda (1974) did not attempt to correct for experimental temperature, most of his respiration rates were determined at temperatures close to this habitat temperature. The literature provides a few estimates for temperatures near 8.6"C of respiration rates of Pseudocalanus of known weights or lengths (Table 111). Agreement with the rates predicted from Ikeda's equation is quite good, with the overall mean of predicted values about 5% higher than that observed. Polyakova and Perueva (1976) give another series of respiration values a t 8.5 and 10°C. Unfortunately, they give total lengths and wet weights of their animals. Using the conversion factor of dry wt = 0.15 wet wt, which they use to transform dry weights in Conover (1969) t o

+

TABLE111. OBSERVEDRESPIRATION RATESOF Pseudocalanus AND THOSE (1 974) EQUATION FOR " BOREAL " ZOOPLANKTON

PREDICTED FROM IEJZDA'S

Experimental temperature ___

"0

Size

mm

Source pg dry wt

29.6a 26-7a 14.Sa 13.7= 13.5 13.3 12.0

Obeerved

Predicted

1.61

1.73 1.61 1.00 0.96 0.94 0.94 0.86

8 8 8 8 6-8 6.8 11.2 10

0.98

0.41 0.55 0.94 0.79 1.12 1.56

10

0.87

0.74

N

N

N

Respiration, p~o,lcop.lday

Estimated from graph.

1.25

0.80

0.67

Anraku (1964b) Anraku (1964b) Anraku (1964b) Anraku (1964b) Ikeda (1974) Ikeda (1974) Ikeda (1970) Marshall and Om (1966) Marshall and On: (1966)

40

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

wet weights tabulated by them, we find that their observed values of R in p1 O,/cop./day (ten estimates) average 22% lower than those predicted from Ikeda's equation. 2. Temperature

Evidently only Anraku (196413) has systematically examined effects of temperature on respiration rates of Pseudocalanus (Fig. 8). Animals were kept at experimental temperatures for 24 h before 0, consumption was measured. Clearly temperature has the usual positive

Temperature ("C)

Fro. 8. Respiration at different seasons of adult female Pseudocalanus from near Cape Cod, Massachusetts, expressed per copepod and per unit dry weight. Approximate habitat temperatures are shown as H on each curve in the righthand graph. (After Anraku, 1964b.)

effect a t the lower end of the temperature scale, and use of rateldry wt appears to give more comparable results. It is worth noting, as well, that Raymont's (1959) estimates for both sexes of Pseudocalanus from Woods Hole a t 15"in April-May (mean 2.45 p1 O,/cop./day) were similar to those found by Anraku in the same locality and season some years later (ca. 1.9 p1 O,/cop./day, from Fig. 8). However, Raymont's copepods averaged 0.99 mm in length, or about 11-5 pg dry wt (from equation p. 127), so that respiration on a per weight basis was about 200 pl O,/mgldryLwt/day, which is much in excess of that found by Anraku (ca. 75 p1 O,/mg dry wt/day, from Fig. 8). The dry weights of the copepods used by Anraku can be deduced from Fig. 8 to be about 27 pg in May, which seems excessively large for lean animals (p. 127), and they may have had much stored oil. For these reasons we believe that

THE BIOLOGY OF PSEUDOCALANUS

41

Anraku’s results can only be used in a relative way for comparisons of temperature response. There are obvious seasonal differences in temperature response (Fig. 8). I n the cool months (February and December) rates were lower at 22.5” than at 15°C) so that the animals were beyond the “ optimum ’) a t the highest temperature. On the other hand, rates at the lower temperatures were inversely related to habitat temperature (indicated by ‘‘ H ” in Fig. 8) ; that is, copepods taken in cooler waters were better able to maintain high respiration rates at cold temperatures in the laboratory. Anraku describes such seasonal differences in terms of lower Qlo (ca. 1.3 in Feb. and 1.7 in Dec., v. 3.7 in Aug.) for copepods from the cooler months, but this is merely descriptive of the higher rates at lower temperatures in the monotonic parts of the curves on Fig. 8. The differences between seasons may have resulted from longterm acclimation (i.e., reversible changes in physiology or activity that take longer than the 24-h adjustment period used by Anraku). Differences could also have resulted from permanent developmental differences between cold-water and warm-water generations (in the manner of size, see p. 116), or even from genetic differences (cf. Bradley, 1975).

Whatever the cause of seasonal differences, we may agree with Anraku (196413) that his results “indicate an adaptation to cold” by Pseudocalanus. This may lead to maintenance of similar respiration rates in nature regardless of season. Anraku tabulates habitat temperatures of 7-1, 14.1, and 6.7”C for animals collected in May, August, and September respectively, and we assume 2°C for February. At these temperatures the weight-specific respiration rates were quite similar (- 38-60 pl O,/dry wt/day ; Fig. 8). The falling off a t high temperatures of respiration for animals collected in the cooler months may be more pathological than physiological, and is of little interest in the context of natural conditions. Anraku (196413) noted that animals died or behaved abnormally a t 22.5”C even in August, and that Pseudocalanus disappeared from the Woods Hole region when temperatures were above 20°C (see p. 24). Raymont (1959) found, in experiments conducted on the adults of both sexes, an elevated respiration in Pseudocabnus from Woods Hole in April-May in one experiment a t 20°C compared with the mean rate of several experiments a t 15°C. However, he noted that repeated runs showed a drop in the rate at 20°C. Conover (1959) estimated that at 20°C adult female Pseudocalanus from Southampton respired 196-256 (mean 234) p1 O,/mg dry wt/day, much in excess of the rate for Woods Hole (Fig. 8). Conover’s experiments were short-term (4-8 h) and his

f

42

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

animals may not have become physiologically damaged while those of Anraku were 24 h in duration. However, Conover’s results may also have had little relevance for natural conditions, since habitat temperatures a t the time of his study were said to be 11.5OC. 3. Light Although light could influence the activity and therefore oxygen consumption of copepods, only Pavlova (1975) appears to have examined its effect on adult Pseudocalanus from the Black Sea. She standardized animal sizes and degree of crowding for experiments being compared. Rates of oxygen consumption (means f 95% c.1. in pg/cop./h) by males at 3 cop./l5 ml were 0.101 f 0.078 in the dark and 0.450 f 0.351 in diffuse light ; for males at 4 cop./l5 ml, rates were 0.360 & 0.181 in the dark and 0.710 f 0.425 in diffuse light; for females a t 4 cop./l5 ml, rates were 0.190 f 0.118 in the dark and 0.490 f 0.380 in diffuse light. Thus it appears that respiration is higher (and more variable) in the light. The animals may simply be more active, trying to “escape” light conditions that obtain by day near the surface (see Section XII). Pavlova (1975) found that respiration rates of females suspended by day in containers at depths of 50-100 m in the sea were similar to rates in the dark in the laboratory. Unfortunately, containers were larger, animals more crowded, and temperatures considerably lower than those of laboratory experiments, so we do not feel that her results give a meaningful indication of the natural rates obtaining under very low light levels. B. Respiration and food requirements Anraku (1964b), Marshall and Orr (1966), and Hargrave and Geen (1970) have calculated minimum food requirements or proportions of food used for respiration by Pseudocalanus. Some of these experiments were with feeding animals, others with “ starved ” animals, and it may be noted parenthetically that Raymont (1959) found no significant difference between the results of two experiments with unfed and seven with fed Pseudocalnnus. I n view of the variability of conditions and results, we do not feel it worthwhile to list the food requirements calculated by the above authors, nor to convert other individual respiration estimates. Furthermore, we feel that caIculations that use RQ values for carbohydrate, fat, and protein separately or in some chosen mixture give an unwarranted air of precision. Instead, we shall here assume an RQ of 1, so that 1 pg of dry body tissue or food requires 1 pl of 0, to be

THE BIOLOGY OF PSEUDOCALANUS

43

metabolized. If we take Ikeda's (1974) formula (p. 39) as an adequate predictor of 0, consumption, and assume a seasonal temperature compensation (as implied in Fig. 8) then minimal food (F)expressed as a percentage of body weight (W in mg) can be given by : log F = 0.437 - 0.217 log W.

An adult copepod of 10 pg might thus need about 7% of its weight daily as food to compensate for respiration, while a nauplius of 1 pg might need about 12%.

C . Retrospects and prospects No doubt much has been left undone in the study of respiration of Pseudocalanus. Nothing appears to be known of the effects of salinity, oxygen tension, and other such variables. Studies of the effects of food concentration deserve expanding, and the effects of temperature and light have not been fully explored. We feel that the use of dry weight as a measure of size has made it difficult to compare results from different seasons and localities, and make a plea for use of a better measure of metabolic size, perhaps length. However, whatever measurements might be made, the experimental violence done to such tiny animals by crowding them into small volumes of sea water and perhaps forcing them to be abnormally active may make any results somewhat unnatural. Our conclusion that respiration might involve expenditures of the order of 10% of body weight each day might not be worth refining through further respiration measurements. It is possible that weight losses can be determined more directly in starving animals, or perhaps by reductions in the size of the oil store (see p. 129). If the strategy of respiration studies is to forge links between food availability, growth, and production, then this can probably be done more directly without use of a " balance-equation " approach (see p. 132).

VI. EXCRETION Excretion has been extensively studied in zooplankton, including copepods. Although excretion may be of interest as a phenomenon in its own right, there have been two main strategies in such studies : (1) rates of excretion supply one component in the " balance equation " of growth or production, and (2) excretion supplies nutrients to phytoplankton. The elements of interest in excretion are nitrogen and phosphorus.

44

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

A. Nitrogen Although amino acids and especially urea are produced by copepods, the current view (Conover, in press; Smith, 1975; Ikeda, 1974) is that NH, is the predominant form of excreted nitrogen by non-feeding copepods. Only NH, excretion has been measured in Pseudocalanus. Butler et al. (1969) and Ikeda (1974) have made a few measurements, but Christiansen (1968) has made extensive analyses of NH, excretion by Pseu,docaZanus from Bras d’Or Lake, a landlocked arm of the Atlantic in Nova Scotia. Some of the factors influencing NH, excretion are of interest in the context of experimental design, but of little significance in nature. We will detail some of these methodological matters first, since Christiansen’s (1968) work is not readily available. 1. Methodology

Christiansen (1 968) determined concentrations of NH, colorometrically, standardizing to a salinity of 25%,. It is important to realize that, unlike for respiration experiments, food conditions during the experiment are of crucial importance. It is expected that a copepod that is feeding or has a gut full of food would excrete a t a higher rate than a copepod that was not fed and had an empty gut, both because of increased metabolic production of waste products and because the methods used to measure excretion may include the leaching of unassimilated nutrients from faeces. Christiansen (1968) placed actively swimming animals in unfiltered sea water (containing any food present in nature) in 140 or 450 ml bottles during the experimental period when excretion rates were measured. Ikeda (1974) kept his animals in natural raw sea water for one day prior to the experiment and then placed animals in filtered sea water during the experimental period. He even removed faecal pellets produced during the experimental period in an effort to measure basal levels of excretion. Size of bottle was found by Christiansen to have no significant effect on excretion rate separate from the effect of concentration of copepods (see below). Experimental and control bottles were placed on plankton wheels in the dark at controlled temperatures. Experiments with antibiotics and mixtures of copepodids of Temora longicornis and Pseudocalanus indicated no significant effect of bacteria in experiments lasting up to 14 h a t 2°C. Christiansen concludes that corrections for bacterial uptake of NH, are unimportant in short-term experiments, a t least. Crowding of animals had a powerful effect on reducing production of NH,. Unspecified mixtures of copepodids of T . longicornis and

THE BIOLOGY OF PSEUDOCALANUS

45

Pseudocalanus were used to demonstrate this. Copepods were placed in 450 ml containers at concentrations from 50 to 600 per container.

This crowding effect ’’ is most likely an artifact of experimental “

conditions and is discussed in more detail below. The log-log regression given by Christiansen for the relationship had a regression coefficient of -1.04 which was close to unity (P< 0-001). A regression coefficient of unity would mean an inverse proportional relationship which clearly cannot apply to very low concentrations of copepods since it would imply indehitely high excretion rates. Excretion rate also declined more or less exponentially with time in experiments with mixed copepodids in unfiltered seawater, so that rates were about five times as high at 2 h after the commencement of experiments compared with 12 h later. Although Christiansen ( 1968) suggested that quiescence and perhaps metabolic shifts would explain reductions in excretion rates, both when the copepods were crowded and when experiments were conducted over longer periods of time, he also concluded that food may become depleted in the darkened bottles. The importance of behavioural quiescence was not supported by his demonstration that excretion rates of C V Pseudocalanus (20 cop./l30 ml) were unaffected by different amounts of time animals were kept prior to the conduct of experiments. We conclude that experiments with fed animals under crowded conditions and of long duration are likely to give reduced excretion rates due to depletion of food. We can therefore readily agree with Christiansen that short term (2-4 h) experiments with uncrowded (<150 cop./J) copepods of the size of Pseudocalanus might be necessary to evoke excretion rates realistic for natural environmental conditions. 2. Effect of food concentration The inference that food shortage might be involved in long-term experiments or crowded conditions is reinforced by Christiansen’s (1968) direct experiments. Using mixed natural food in nutrientenriched seawater, he showed that excretion rate by C V Pseudocalanus (25 cop./l30 ml for 4 h at IOOC) in pg NH,-N/cop./h were 0.046 & 0.006 and 0.068 f 0.005 (means & 95% c.1.) at food levels of about 39 and 93 pg N/1 respectively.

3. Effect of body size It has been widely demonstrated that the smaller zooplantonic species and individuals within species have generally higher weightspecific excretion rates. Christiansen (1 968) demonstrated that this effect of body weight was marked among several species of copepods

46

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

and for several stages of Psezldocalanus. The interspecific and intraspecific relationships were very similar (Fig. 9). The regression coefficients of near unity imply that excretion rate per individual is virtually the same for a tiny nauplius (about 0.6 pg in Fig. 9) and a full-grown

1\ NP-PI

‘Ooo

-

\

,““““a

sirnilis

\

ti 1-u

I

-

log E = O,066

20

t-

‘\

\

\

”V,V,,”D

finmarchicus

G 0.001

0-01 Mg dry w t . ( W )

u (

FIQ.9. Weight-specific excretion rate for adults of different species of copepods (open circles, broken line) and for different stages of Pseudocalanw (closed circles, unbroken lines). (After Christiansen, 1968, who included in his analysis the point for CalanusJinnaarchicus from Corner ct al., 1966.)

adult (ca. 14 pg): about 0.68 and 0.86 pg NH,-N/individual/day respectively. This is hardly likely to be a “natural” state of affairs. We ~ as similar suggest that this anomalous finding of Christiansen ; t well results in Corner et al. (1965, their Table 4) are related to the “crowding effect” described earlier (p. 45). The crowding effect was inversely related to excretion and gave a log-log regression coefficient close to unity, like the effect of body weight. The effect of crowding together numbers of animals of a given biomass may have the same effect in

THE BIOLOGY OB PA'EUDOCALANUS

47

reducing excretion rate (through depletion of food or whatever mechanisms) as occurs when the biomass of individuals increases and the numbers are held constant, sts in Fig. 9. Rather than attempting to correct for the crowding effect in the relationship between excretion rate and body size, we turn to the work of Ikeda (1974), who fitted the following expression to non-feeding " boreal " zooplankton (terms as in Fig. 9) : log E = 0.182

- 0.21 log W.

This expression (which includes values for Pseudocalanus) gives an excretion rate for a 14 pg animal (full-grown adult) of 0.052 pg NH,N/individual/day and for a 0.6 pg animal (nauplius) of 0.004 pg NH,N/individual/day. Since Ikeda (1974) measured excretion by unfed animals we would expect these values to be lower than those calculated from Christiansen (Fig. 9). More important, however, is the fact that the excretion rate calculated from Ikeda (1974) for the biomass of a nauplius is an order of magnitude smaller than that calculated for an adult. This seems to us to be a more realistic result although we are fully aware that it has yet to be demonstrated that extrapolation may be made from the interspecific relationship of Ikeda to the intraspecific relationship implied in the above calculations. 4. EfSect

of temperature

Unfortunately, Christiansen (1968) once again used mixtures of copepodids of Temora longicornis and Pseudocalanus to test effects of temperature (T, in "C) on excretion rates (E, in pg NH,-N/cop./day) a t varied densities (C, in No./450 ml) : log E = -1.59 - 0.585 log C

+ 0.0137 T.

Effects of crowding (C) and temperature (T) were highly significant ( P < 0.001) and the use of a multiple regression enables the effect of temperature on the release of nitrogen to be determined after the effect of crowding has been removed. The regression coefficient of 0.0137 implies that, whatever the density, a 10°C rise in temperature will increase the excretion by 37%, i.e. a low Qlo of 1.37. Support for this low Qlo comes from the work of Ikeda (1974) who gave measurements of excretion rates by non-feeding Pseudocalanus in two experiments a t about 7°C (4-3 and 5.0 pg NH,-N/mg dry wt/day) and one experiment at 12°C (4.1 pg), again implying a low Qlo. Nival et al. (1974) also suggest that a low Qlo of excretion rate for non-feeding copepods is characteristic.

48

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

5. Effect of salinity Christiansen (1968) again used mixtures of copepodids of T . longicornis and Pseudocalanus at 10°C to test the effect of salinity on excretion rates at varied densities. Christiansen gives a multiple regression in which both crowding ( P < 0.001) and salinity (P< 0.01) are highly significant. Unlike temperature the effect of salinity was found to be quite pronounced, doubling the rate of excretion between lo%, and 32%,. Excretion in the more euryhaline Acartia tonsa was not significantly affected by salinity. Presumably the acute effect shown by Christiansen could have some importance in highly stratified waters, but acclimation or adaptation may well occur in persistently brackish waters occupied by Pseudocalanus. 6. Effect of oxygen concentration

As already noted (p. 25), Pseudocalanus can be found at very low oxygen concentrations in nature. Christiansen ( 1 968) carried out 4-h experiments at 10°C with C V Pseudocalanus, 20 cop./l30 ml a t 20 different oxygen concentrations between 1.3 and 6.9 ml 02/l. The effect of excretion rate was we11 fitted by a negative exponential expression (exponent -0.3) such that there was an approximate doubling of the rate at 1 ml O,/I compared with that near 0, saturation. N

7. Nitrogen requirements

We suggest elsewhere that a value of 7% N might be found in lean ” Pseudocalanus and discuss the possibility that even higher values in Christiansen (1968) may include gut contents (p. 126). Christiansen’s equations (Fig. 9) suggest that a feeding adult animal of 10 pg dry w t might excrete about 0.8 pg of NH,-N daily (and perhaps more as organic N). If we assume that its food is also 7% N by dry weight, this implies a production of NH,-N in excreta (and possibly leached from faeces) in excess of the body nitrogen per day. This is a higher value than any listed by Corner and Davies (1971, Table VIII). However, it may be reasonable, as rations of up to 140% of body weight per day have been observed (Paffenhbfer and Harris, 1976, see p. 63). Ikeda’s (1974) formula for non-feeding animals gives a better indication of basal requirements, indicating that an adult Pseudocalanus (10 pg) might excrete about 0.040 pg NH,-N/day, or about 5.6% of its N content (if N is 7% of dry wt). This estimate agrees with that of Butler et al. (1969), who found that “ mixed small copepods ” (in fact over 90% C I V and C V Pseudocalanus) in filtered seawater “

49

T H E BIOLOQY O F PSEUDOCALANUS

excreted (mean and 95% c.1.) 3.98 & 0.61 pg NH,-N/mg dry wtlday. Mean dry weight was 11.7 pg, and contained 7.8% N, so that this rate represents about 5% of body nitrogen per day. B . Phosphorus Hargrave (1966) and Hargrave and Geen (1968)present information on phosphorus excretion by Pseudocalanus from Bras d’Or Lake, Nova Scotia. Both dissolved inorganic phosphorus (DIP) and dissolved organic phosphorus (DOP) were measured. Again we deal briefly with methodology to the extent that it may influence whether the estimates being sought are representative of those occurring in nature. As was the case for excretion of NH,, food conditions in these experiments are important and again may be implicated (although the evidence is less complete)in reducing DIP under crowded conditions and in experiments of long duration. Unfortunately, the authors did not consider Pseudocalanua in all their experiments. 1. Methodology Hargrave (1966) showed that both DIP and DOP declined significantly in brackish water without zooplankton held for 24 h in the dark. Thus it is not surprising that both DIP and DOP increased more in laboratory experiments with copepods when antibiotics were added, which they were routinely. About 67% of dissolved phosphorus was DOP in one set of experiments with C V and adult Pseudocalanus described by Hargrave and Geen (1968). Thus measurements of DIP alone (which was the measurement usually made in experiments) should be corrected if they are to be used as estimates of rate of release of total dissolved phosphorus. Copepodids of Temora longicornis and Pseudocalanus in experiments with unfiltered seawater were observed to concentrate in the lower half of unstirred experimental bottles and produced significantly less (.P < 0.05) DIP than did those in stirred bottles (Hargrave and Geen, 1968). However, no direct measurements of the effect of crowding were made. Hargrave (1966) also found significant declines in production of DIP after about 10 h in unfiltered seawater by a mixture of copepodids of T . longicornis and Pseuducalanua. However, the high temperatures (18-20°C) in the experiment could have been a factor. 2. Effect of food concentration

Mixed copepodids of T . longicornis and Pseudocalanus released significantly less DIP in membrane-filtered seawater than in natural sea A.M.B.--15

4

50

CHRISTOPHER J. CORKETT A N D IAN A. MCLAREN

water at in situ temperatures. Means, 95% c.I., and ranges in pg DIP/cop./day were : 0.0075 f 0.0007 (0-0030-0.0160) and 0.0210 f 0.0055 (0.0111-0.0551) respectively (data from Hargrave, 1966). The former may be taken as a non-feeding excretion rate. Hargrave also showed that release by copepodids of Pseudocalanus of DIP and DOP was reduced by passage of the medium through filters of different pore sizes smaller than 3-5 pm. These presumably removed food which can be used by Pseudocalanus (see p. 63). Copepodid concentrations (650-1 000/1) and temperatures (17-1 9°C) were rather high in these experiments. 3. Effect of body weight

Hargrave and Geen (1968) include Pseudocalanus in a graph of rate of DIP excretion (E, in pg DIP/mg dry wt/day) as a function of body size (W, in mg dry wt) for nine species which can be written from Hargrave (1966) as : log E

=

-0.663 - 0.296 log W.

The authors do not state whether filtered water was used or not, but the rate predicted from this equation for a 10 pg Pseudocalanus is 0.0085 pg DIP/cop./day, which is close to the non-feeding rate (0.0075 pg DIP/cop./day) noted above. Certainly the larger animals (Mysis, Gummarus, Crungon) in their regression could not have been feeding much from the sea water. We take it that the regression illustrated by Hargrave and Geen represents some sort of physiological relationship uninfluenced by much faecal production. TO support this, Hargrave (1966) shows a close proportionality between oxygen uptake and phosphoros excretion by seven of the nine species used in the regression of DIP excretion on size. 4. Effect of salinity

No significant correlation was found between excretion rate and salinity of a mixture of copepods, including Pseudocalanus, from Bras d’Or Lake (Hargrave, 1966). However, details of the experiment and results are not given. 5 . Phosphorus requirements Hargrave ( 1966) conducted experiments on non-feeding copepodids

or Pseudocalanus with empty guts. By including direct estimates of loss of body P, he calculated that 18.2% of total body phosphorus could be released as faecal particulate P per day. I n addition the copepodids released some 7.7% of total body phosphorus per day in dissolved form

THE BIOLOGY OF PSEUDOCALANUS

51

(DIP and DOP). This estimate agrees with that of Butler et al. (1969) who calculated the excretion of total P in membrane filtered water, They found that " mixed small zooplankton " (almost all Pseudocalanus) excreted (mean f 95% c.1.) 0.77 & 0.07 pg P/mg dry wt/day. Their animals weighed about 11.7 pg each and were 0.61% P , so that about 11 % of their body P was excreted daily. The regression of D I P on body weight suggests that higher values may be attained by feeding Pseudocalanus. Assuming (from the interspecific relationship of Hargrave and Geen; see p. 50) that a 10 pg Paeudoealanus, containing 0 . 6 1 ~ 0P, produces 0.0085 pg D I P per day, this will be about 14% of the body Plday. This could be as little as onethird of the total dissolved P produced, since DOP was not measured (see methodology, p. 49). C. Retrospects and prospects The evidence seems clear in these studies on Pseudocalanus that " starvation rates of excretion are much lower than excretion rates found during feeding. Experimental and technical difficulties of working with a tiny animal under fed conditions make it unlikely, in our opinion, that precise estimates of excretion rates can be obtained that are representative of those found in nature. Moreover, the role of such an animal in replenishing nutrients probably depends as much on system qualities of the animal's environment-the amounts and kinds of food available-as on the character of the excreting animal. If basal excretion estimates are required it is probably better for purposes of calculation t o assume general functions of the sort derived by Ikeda (1974) relating body size, temperature, and excretion rate of different species, rather than t o try and estimate such rates directly with such a small animal as Pseudocalunus. While there have been attempts t o use excretion rates in balance equations of growth of copepods (e.g., Corner and Davies, 1971), the most that can be said for the rates available for Pseudocalanus is that they are compatible with feeding rates and with the direct and accurate estimates of growth rate that can be made (see p. 132). Altogether, we conclude that further detailed studies of excretion in Pseudocalanus might be of intrinsic interest, but would have little strategic value for biological oceanography. ')

VII. LOCOMOTION A. Routine swimming Gauld (1966) examined several genera of calanoids, including Pseudocalanus, and observed that they usually swim in a smooth,

62

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

gliding fashion, using principally the second antennae for propulsion, in the manner described by Lowndes (1935). Urry (1964) used stroboscopic illumination to determine that the second antennae of adult Pseudocalanus vibrated at 1 890-2 580 cycles/min, a rate considerably higher than the 1 200 cycles/min reported for Calanus by Lowndes (1935). The movement of the second antennae in copepods is complicated (Gauld, 1966),but evidently the endopod provides propulsion when the exopod is in a recovery stroke and vice versa, so that continuous propulsion is maintained. Giesbrecht (1882) illustrates the second antennae of both male and female Pseudocalanus. The setae of the male

f?

I

1

J I cm FIQ.10. Swimming excursions of adult female Pseudocalanus during seven-second paths. The sequences of positions (each straight portion an excursion) from which the scatter diagrams were based are shown on a reduced scale on the right. Each excursion (dot on graph) is plotted as an absolute value, and the average horizontal and vertical components of movement are given by dark bars on the axes. (After Anderson, 1974.)

are longer than those of the female and completely setulated, suggesting a more efficient swimming potential, which is probably used in hunting for a mate during the short life of an adult male. I n addition to the second antennae, the mandibular palps are involved. The palps rotate a t their base, presumably to reduce resistance on the recovery stroke, since Gauld (1966) states that the effective thrust must be mainly during the backward stroke of the palps. The action of these two pairs of appendages gives the slow gliding motion and in producing this forward motion also creates a broad, U-shaped vortex or swimming swirl (this should not be confused with the feeding current, p. 54).

THE BIOLOGY OF PSEUDOCALANUS

53

Cushing (1959) uses a calculated 24-h swimming rate for Pseudocalanus of about 0.2 cmlsec but this is based on slender data. Anderson ( 1974) has analysed the routine swimming of Pseudocalanus in spatial terms. He observed that, although it occasionally swims in loops (see mating, p. 84, for a possible explanation), its undisturbed movement is usually similar t o the " hop-and-sink " motion described by Bainbridge (1952) for Calanus. Periods of vertical ascent often alternate with periods of passive sinking. I n photographic records of this motion, this appears as " J "-shaped paths (Fig. 10). Anderson analysed photographs of swimming Pseudocalanus t o measure the vertical and horizontal component of each straight excursion during seven-second paths. The horizontal displacement was assumed t o be equally distributed in all directions. The distribution of the vertical and horizontal components of velocity is summarized on Fig. 10. The vertical component of 0.061 & 0.010 cm/s considerably exceeds the horizontal component of 0.031 f 0.006 cm/s (means f 95% c.1.). This confirms in a quantitative way the " hop-and-sink " motion.

B. Escape reaction As Gauld ( 1 966) pointed out, most calanoid copepods, including Pseudocalanus, can make large leaps or jerks t o avoid contact or capture. Rapid movement makes it difficult t o see exactly what limb movements are involved, but there seems little doubt that the swimming feet in particular are brought into play. Although Anderson (1974) does not describe an escape reaction for Pseudocalanus he postulates that its small-scale motion, doubling back on itself vertically, makes it less vulnerable t o predation by ctenophores than are some of the other species of copepods he studied.

C. Retrospects and prospects Slow swimming is reasonably well understood, but it would be useful to clarify the mechanism used in the escape reaction. Perhaps more important would be a hydrodynamic study of the sort that have tentatively been developed for other species of copepods. Certainly the work of Anderson (1974) implies that the routine swimming rates could be readily documented, and it would be of some value in connection with " energy-balance " approaches t o growth and production (p. 181) t o estimate the energetic costs of such motion.

54

CHRISTOPHER J. CORRETT AND IAN A. MCLAREN

VIII.

UTRITION

A. Feediny mechanism 1. Filter feeding (a) Feeding current Esterley (1916) and Cannon (1928) have given an account of the feeding current in Calanus and Gauld (1966) confirmed that these observations on the way particles are filtered and ingested in Calanus also apply t o Pseudocalanus. The feeding current is primarily a pair of eddies flowing round the bases of the maxillipeds. The pair of eddies are formed from the swimming swirl or vortex and are enhanced by the rotation of the maxilliped and by the movement of the f i s t maxilla. The exite on the first maxilla carries long flexible setae (Fig. 7E) which suck water forward through the setae of the second maxilla and then sweep it away outwards.

(b) Filtration The setae and setules on the second maxilla (Fig. 7F) are used to filter food organisms from the feeding current. The second maxilla in female Pseudocalanus has been studied in detail by Schnack (1975). We are interested in three basic measurements : (1) length of setae, (2) length of setules, and (3) distance between setules; and how these measurements vary for: (a) different setae along the maxilla, (b) different setules along a given sets, and (c) different seasons of the year. The length of the setae along the second maxilla showed no general trend to increase or decrease, but varied in length from about 90 pm t o 110 pm with a few smaller setae of 50 pm (Schnack, 1975). The length of the setules varies both among and on setae and Schnack (1975, her Appendix B) showed that setule length varied from 3.0 to 9.0 pm on a few selected setae. TABLEIV. THE DISTANCE BETWEEN SETULES ON THE SETAEOF THE SECOND MAXILLAO F ADULT FEMALE Pseudocalanus. (From ScEinack, 1975.) Position tip tip base base

Month December 1970 July 1971 December 1970 July 1971

Average distan,ce between setules ( p m )

95% c.1.

8.2 7.0 4.0 3.0

7.9-8.5 6'7-7.2 3.8-4.1 2.8-3.1

55

THE BIOLOGY O F PSEUDOCALANUS

The distance between setules varied both for different setae along the second maxilla and for different positions along given setae. At the base of the appendage the setules on the seta were closer together than a t the tip (Table IV). Seasonal variation is also encountered and the average distance between setules in July is significantly smaller than that found in December (Table IV). We suggest that this is related to seasonal variation in body size in response t o temperature (p. 116). Schnack (1975) considered the minimum distance between setules in female Pseudocalanus to be 3.0 pm (95% c.1. 2.8-3.1) which was the smallest value in the seven species she studied, leading her to conclude that Pseudocalanus was specialized for filter feeding and therefore essentially a herbivore. The detailed measurements on the second maxilla are not easily used t o reconstruct an effective mesh size t o determine the efficiency of removal of small food organisms, since the mesh size must vary with the distance between setae. This distance must vary with position of the second maxillae, presumably under control of the animal. However, the means of distances between setules on setae from about 3 to 8 pm (Table IV), do suggest that Pseudocalanus would be more efficient in filtering particles larger than about 8 pm. Poulet (1977) tabulates differences in number of setae, size and vibration rates of feeding appendages of adult female and C I11 animals. He also calculates the hypothetical volume swept by the stroke of these appendages. Naturally enough, relative mesh size (number of setae per appendage length) is smaller, vibration rate of appendages faster, and the hypothetical volume swept by each appendage is smaller among younger animals. Although these data do not give direct estimates of minimal sizes of potential food (see above) they show clearly that younger animals could secure smaller food particles more efficiently. Once food particles have been filtered from the feeding current by the second maxilla the particles are combed off the setae and setules of the second maxilla by the teeth on the endite of the first maxilla (Fig. 7E) and passed t o the mouth. (c) Mastication Before food particles are passed through the mouth into the oesophagus they undergo mastication by the mandibular blades. Anraliu and Omori (1963) found that filter-feeding copepods had short and rounded teeth whereas predatory copepods had teeth that were few and pointed. Petipa (1975), in a recent study of the origin and classification of feeding types of calanoid copepods, suggests that the Pseudocalanus type ” can be derived by reduction of the masticatory function ((

B FIG.11. Scanning electron micrographs of the right adult female mandible of Pseudocnlanus. A, chewing surface. B, detail of dorsal teeth. Symbols are; c1-c4, central teeth; cli, diastema; d,-d,, dorsal teeth; v,, ventral tooth. (From Sullivao et al., 1976.)

THE BIOLOGY OF PSh'UDOCALANUS

57

from the " Calanus type ", and can lead by further reduction to a " Paracalanus type '' of feeding. Earlier Itoh (1970) tried to quantify such differences by calculating an " edge index " ( E L ) which took into account the length and distance between teeth relative to the total width of the mandible blade. Schnack (1975) using Itoh's E.I. (with some modification) found she could identify three groups of copepods : (1) E.I. < 500, continuous filtering herbivorous copepods; (2) E.I. 500-900, omnivorous copepods ; (3) E.I. > 900, carnivorous copepods. Schnack (1975) found Pseudocalanus females had an E.I. of 365 and they were placed with Paracalanus parvus females (E.I. of 265) in the first category. Sullivan et al. (1975) examined the teeth on the mandibular blades of eleven species of copepod in detail using a scanning electron microscope. They confirmed the work of Beklemishev (1 954a) that the teeth had siliceous crowns set in a chitinous mandibular blade. They also showed that the right and left blades were not identical. The sharp projections on the teeth of one mandible fit into the grooves on the teeth of the opposite mandible, suggesting a cracking rather than a grinding action. I n the herbivorous copepods such teeth would be well suited for breaking diatoms, and Sullivan et al. (1975) comment that " possession of glass teeth for eating food in glass cases is surely one of the lyrical symmetries of nature ". Based on the work of Beklemishev (1954a),Sullivan et al. (1975) have divided the teeth into three groups. Starting with the ventral edge of the blade and progressing to the dorsal edge these groups are : ventral, central, and dorsal. The right mandibular blade of PseudocaZanus has a single large ventral tooth separated by a diastema from the rest (Fig. 11A). The left mandibular blade of Pseudocalanus was not studied but based on work on other species of copepods, the left blade probably has two ventral teeth into which the single ventral tooth on the right blade would fit. The ventral tooth on the right blade of Pseudocalanus is followed by four central teeth (Fig. 11A) which are followed by three dorsal teeth of a complex nature (Fig. 11B) and Sullivan et al. (1975) suggest that these dorsal teeth can " tightly grip food of a softer nature ". 2. Peeding on large particles Cushing (1955) states :

" I have Been a Psedocalanus take a Biddulphia sinensis almost as big as itself, break it and filter off some of the contents ". This is an important observation because it suggests that, despite evidence given above that Pseudocalanus is basically a

58

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

continuous filter feeder it is, nevertheless, able to feed by another mechanism, evidently involving selection of larger particles. Gauld (1964) suggests that larger particles are captured by Pseudocalanus as in other copepods, by use of a sweeping movement of the second maxillae. He also pointed out that the second maxilla of Pseudocalanus has at its tip a long stout sets (Fig. 7F) which ‘‘ would be a very effective seizing and holding weapon ”. The maxillipeds may be involved in this feeding process as well since they are known to play an important part in the grasping of food by carnivores such as Euchaeta norvegica (Gauld, 1964) and Oithona similis (Schnack, 1975).

B. Food eaten 1. Xpecies eaten in nature

Pseudocalanus is capable of ingesting a wide variety of food. Corkett (1966) has shown that individuals in the laboratory will take in fine sand and produce faecal pellets and Schnack (1975) finds “ sand ” (fine mineral particles) in guts in nature. Pavlovskaya and Pechen’Finenko (1975) have shown that Pseudocalanus in the laboratory will ingest and assimilate algal detritus, humus particles, and even melanin. These and other such experiments are reviewed later, but they tell us nothing about the food available t o and chosen by animals in nature. Until recently, the gut contents of wild-caught Pseudocalanus had not been extensively studied. Lebour (1922) concluded that diatoms formed the main food, especially Coscinodiscus and Thalassiosira. Flagellates were also thought to be eaten although indistinguishable in the guts. Later, Lebour ( 1 923) found coccoliths in guts, as evidence that coccolithophorids are eaten. Marshall (1949) examined guts of over 100 live Pseudocalanus from off Millport, Scotland, and found that 73 were empty or had indistinguishable remains, 18 had diatoms, 4 had radiolarians, 1 had a flagellate, and 10 had crustacean remains. Although the crustacean remains show that Pseudocalanus can be carnivorous (and we have found that females will eat their own nauplii) the morphological evidence of the previous section and other studies in nature (below) imply that they are overwhelmingly herbivorous. Beklemishev (195413) stresses this, and found only the diatoms Chaetoceros and Thalassiosira in guts of adult females from the Bering Sea. Schnack (1 975) presents a great amount of information on food and feeding of copepods off Kiel Bay, near the mouth of the Baltic. She determined the kinds and amounts of food in guts of adult female Pseudocalanus taken in November and December 1970, and in July,

THE BIOLOGY OB PSEUDOCALAh’US

59

September and October 1971. These contents were compared with food in the water column, sampled by a net with mesh of 54 pm and by plankton pump which sampled particles too small to be caught by the net. She found that the spiny diatoms Chaetoceros spp. were almost uneaten. Xkeletonema costatum was common in the water but scarce in guts, possibly due t o its fragility. I n general, she concludes that Pseudocalanus is a herbivore, that individuals favour small food species, that they remove only a fraction of the range available in the plankton, that they favoured centric over pennate diatoms, and that small forms like Exuviaella baltica and Heterocapsa triquetra were a t times important. These conclusions do not take into account the possibility of local concentrations of food or copepods in the water column (which would complicate conclusions about selection of one sort of food or another), and they are unsupported by statistical analysis. Zagorodnyaya (1974) has produced by far the most detailed assessment of food in guts of Pseudocalanus from samples taken in the Black Sea off Sevastopol in January and March 1973. She identified and tabulated some 50 food species, predominantly diatoms and dinoflagellates, with some chrysophytes. No animal food was found. The three most regular species were the dinoflagellate Exuviaella cordata, the diatom Cyclotella caspia and the chrysophyte Coccolithus huxleyi. Diatoms were selected in excess of their proportionate representation in the water column, and the reverse was generally true of dinoflagellates (depth distributions of copepods and food species were taken into account). However, she concludes that species selection was generally based on size. This and other aspects of Zagorodnyaya’s important work are dealt with elsewhere. Feeding experiments conducted in the laboratory using natural seawater and its contained food give some indication of the food species used in nature, even with the reduced options in time and space. Curl and McLeod (1961) give anecdotal observations on the selective removal of Skeletonema sp. from stored plankton samples by Pseudocalanus and Acartia tonsa. Parsons et al. (1967) also noted use of Xkeletonema in more formal experiments. Geen and Hargrave (1966) found that Pseudocalanus did not eat many of the long-spined Ceratium sp. nor the spiny, chain-forming diatom Chaetoceros, but rather favoured larger flagellates in natural seawater samples. Parsons et al. (1967) also indicated that Pseudocalanus was unable to derive much value from samples of a bloom of Chaetoceros socialis and C. debilis. In contrast, Poulet (1974) observed chains of Chaetoceros sp. being devoured by females kept in unfiltered seawater. A species by species account of the food of Pseudocalanus in nature

60

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

could in principle become almost endless. A number of the above authors have indicated that size is a chief determinant of suitability of a food species. This subject is dealt with next. 2. Xixe of food

Clearly there must be some lower limit to the size of food particles that can be filtered and some upper limit to the size of organisms that Pseudocalanus can “ handle ”. Observations on the maxilla, show that. the smallest distance between setules on the setae is 3 pm (p. 5 5 ) and individuals can evidently contend with food almost as large as themselves (p. 57). However, much effort has been expended on the search for size-selective feeding within these limits. As shall be seen the evidence for size-selective feeding by filter-feeding animals is by no means conclusive. Evidently the first formal consideration of size-selective feeding by Pseudocalanus was by Hargrave and Geen (1970). They showed that filtration rate was significantly higher for cells about 10 pm in maximum diameter than for cells of about 0.5, 5, and 7 pm (about 12 v. 4-5 ml/cop./day respectively). Their results are marred by being based on experiments with a mixture of C I V and C V of Pseudocalanus and Temora longicornis feeding in water from which particles > 35 pm had been removed by filtration. Zagorodnyaya’s (1974) analysis of gut contents of Pseudocalanus in the Black Sea is especially valuable for its information on several stages of copepodids and on adult females (Table V). Counts and measurements are based on unspecified “ standard methods ”, and unfortunately there is no clear indication in her paper that the percentage values in Table V are for cell numbers or total cell volumes, although

-

TABLEV. PERCENTAGE REPRESENTATION OF DIFFERENT SIZE GROUPS OF UNICELLULAR ALGAEIN THE GUTS OF Pseudocalanus AND IN THE WATER COLUMN. (After Zagorodnyaya, 1974.) Size group

(rm)

< 10 10-20 20-30 30-40 40-50 >50

January

March ~

CIIICIV 43 50 7 -

_ _

59 34 6 1

_ _

CV 38 37 19 1 1

Ad. Q Water 16 38 31 8 5 2

25 41 23 4 3 4

C IIf C I V

~___

50 7 36 7 -

-

53 21 21 3 2

C V 61 16 13 8 1 1

A d . ? Water 54 15 20 8 2 1

14 66 17 1.6 1-4

THE BIOLOGY OF PSE U DOCA LAN U S

61

one can infer the former. Formal statistical analyses of the data as presented in Table V cannot be made, but they support Zagorodynyaya’s conclusions that the copepods consumed mostly cells < 10 pm (which were especially Cyclotella caspia and Coccolithus huxleyi) and that older stages and adults were more competent with larger cells. Extensive studies by Poulet (1973, 1974, 1976, 1977), and Poulet and Chanut (1 975) have advanced the whole subject of grazing by copepods to new levels of sophistication. The recurrent theme in Poulet’s work is size-selective feeding. For each grazing experiment, Poulet placed 50 or 100 lively adult female Pseudocalanus in a liter beaker containing seawater screened through a 160 pm mesh. Duplicate containers filled with screened seawater served as controls. All experiments were carried out for 19-20 h a t temperatures close t o those of the water from which the sampies were removed. The experimental and control samples were analysed with an electronic particle counter set for particles between 1.58 and 114 pm (earlier work) and up t o 144 pm (later experiments). This produced data on concentrations (volume in p.p.m.) versus particle diameter (spherical equivalent). Poulet (1973) studied grazing in samples from 5 m a t five stations in a transect from the head of Bedford Basin, a highly enriched environment, to beyond the entrance of Halifax Harbour, Nova Xcotia. He also used a vertical series of samples from five depths between 0-60 m from the middle of Bedford Basin. From his analyses of the changes in the concentration-size distribution of particles after grazing, he concluded that the copepods were well able t o consume particles from 1.58-114 pm (although not readily if < 4 pm). Poulet (1973) found, however, that grazing occurred on smaller particles (< 25 pm) when these were more than about half of the total concentration and occurred on larger particles when these were equally concentrated as or more concentrated than the smalIer ones. That is, “ the heterogeneity of particle distribution in time and space can be overcome by copepods by shifting their grazing pressure from one size of food t o another ”. Poulet (1974) expanded his work to a two-year study using water samples from 5 m in Bedford Basin. He described the seasonal cycles of particles in six size categories : 1.6-3-6 pm, 4-0-9-0 pm, 10-1-22-6 pm, 254-57.Opm, and 64-1 14 (or 64-144) pm. The seasonality of abundance of particles in nature is of course a property of the environment he studied, and is not reviewed here. Against this background of seasonal availability, Poulet studied possible selection of particles using electivity indices for all but the smallest of the above six size categories. I n agreement with his earlier conclusions (Poulet, 1973), he argued that

62

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

" in general, and within certain limits, Pseudocalanus minutus consumed those particles that were present in the greatest concentration ',.. This evidently need not be understood as indicating selection of those that are most concentra,ted. Poulet (1974) is somewhat equivocal on this question. Although he applies no statistical tests, he discusses seasonal variations in electivity indices a t some length, in this sense endowing the indices with " reality ". Finally, however, he concludes that Pseudocalanus is " unselective in its feeding. There is, however, no doubt that electivity varies with particle size. The numbers of positive and negative electivity indices can be estimated for each of his five particle-size groups from Poulet's (1973) text and his Fig. 5 . The indices were positive in la%, 33%, SOY0, 68%) and 59% of experiments for the smallest through largest particle size categories respectively (heterogeneity x2 = 45.3, d.f. 4, P < 0.01). This would seem to suggest that smaller particles ( < l o pm) are less readily removed by the animals. Poulet also presents a series of graphs with regressions of rate of consumption against particle concentration, which we summarize in Table VI. He concludes that " the highest ')

TABLEVI. REGRESSIONS OF FOOD CONSUMPTIONBY ADULTFEMALE Pseudocalanus ON PARTICLE CONCENTRATION (After Poitlet, 1974.)

Particle size

y=a+bx a

PS 1.6-3.8 4.0-9.0 10.1-22.6 25.4-57.0 64.0- 144.0

__

n

60 80 89 84 73

-0.413 - 0'093 0.365 0.430 0.341

r

b 0.214 0.707 0.991 0.932 0.636

0.17 0.54 0.63 0.83 0.62

c

z = particle concentration (In of mg/l). y = food consumption (In of mg/h/cop. x

Given as log in original, but In meant, and with corrigendum from author (regression and correlation coefficients reversed in original).

[correlation] coefficients were computed in size group 3 and 4, showing a good feeding response towards particles in the 10 to 50 pm range ". This conclusion cannot be inferred from correlation coefficients. More interesting questions might be asked using analysis of covariance, but this cannot be done from the data given. However, the elevations ( a ) of the regressions for the two smallest particle sizes (1.6-3.6 pm and 4.0-9.0 pm) in Table V I are lower than for larger particles. Except for the smallest particles, for which the variance of estimate must be very

THE BIOLOGY O F PSEUDOCALANUJY

63

large (r not significant a t P = 0.05), the slopes (b in Table VI) are similar for each regression. The value of a for particles < 4 pm is about -0.4, and for particles > 10 pm is about 0.4. The difference suggests that the smallest particles are retained about half as efficiently 2) as are larger ones. (i.e., eO.8 Poulet and Chanut (1975) come t o somewhat different conclusions by using two non-parametric tests t o detect possible differences in the size-frequency distributions of particles in diets and in controls. The more sensitive Kolmogorov-Smirnov test for the maximum difference between the cumulative frequency curves in controls and diets showed significant differences ( P < 0.05) in 16 of 42 experiments. However, in each of these experiments the difference was due t o the increase in particles over control level, either substantiaIly (5-25% over control) or grouped in a narrow size-range. Poulet and Chanut conclude that statistically significant examples of apparent selective feeding are due to the formation of smaller particles from larger ones by the activities of the copepods-for example by the breakup of chains of diatoms. Certainly this may partly explain the low electivity of the smallest particles and the differences in the regressions of consumption rate on particle concentration between large and small particles. The general conclusion t o be reached from Poulet’s earlier work does not altogether agree with the observations of Zagorodnyaya (1974) who found that Pseudocalanus favoured cells < 10 pm (Table V). Her experiments cannot be considered as well controlled as Poulet’s, but her observations that the diets of older copepodids and adults may include larger food particles is an amplification rather than a contradiction of Poulet’s work. It seems that Pseudocalanus may make use indiscriminately of a wide range of food, but that the upper limit of that range may increase with body size. We have given anatomical evidence (p. 5 5 ) that seasonally smaller females may be able t o filter smaller particles. Finally, in his most recent paper, Poulet (1977) concludes that copepodids (C I-C IV, mostly C 111) consume food particles
64

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

humus particles, melanin) to varying degrees by Pavlovskaya and Pechen’-Fenenko (1975). Since a substantial fraction of particulate organic matter in coastal waters is non-living, the work of Poulet (1976) adds an important refinement to his studies of particle grazing by Pseudocalanus. Using measurements of adenosine triphosphate (ATP) he estimated the proportions of living and non-living particulate carbon in control and grazed samples of seawater between 29 February, 1972, and 24 January, 1973. (Other features of his grazing experiments are noted in the previous section.) The living fraction in the sea (control samples) varied from 7 to 42%, with a mean of only 21% of total particulate carbon. He estimated that on average about 29% of the ingested carbon was living. Poulet found that selectivity [sic] indices for the living fraction were positive in 56% of experiments. However, a sign test and a runs test indicated that the selectivity values for living and non-living particles did not differ. Poulet concludes that “ seasonal variation of the body size and the number of eggs laid per female , . . give indirect evidence that the assimilation of a diet composed mainly of non-living particles occurs and is channelled into growth and reproduction ”. However, food probably has little effect on body size (p. 122), and there is in fact no direct indication that the non-living fraction is either assimilated or nutritious.

C. Feeding rate The aim of observations and experiments on Pseudocalanus as a consumer of food is presumably to establish for individuals of a given stage, sex, and size, the rate of removal of food of a particular kind at a particular density in the water. Temperature and perhaps salinity can be assumed to be important in determining this rate. Such data tell us nothing about the assimilation of food nor of its subsequent utilization. Although they are in principle interconvertible, rates of food consumption by copepods have traditionally been expressed in two ways: ( 1 ) as Jiltering rate and (2) as feeding rate, or, more precisely, ingestion rate. Filtering rate measures volume of water swept clear of food (or other particles). The dimensions of filtering rate are volume of water per animal per unit time, and the expression of food consumption in this manner has a hopeful ring ; it was once believed that copepods could be treated as automata that swept the water at rates independent of the character and abundance of the food therein. This contention

65

THE BIOLOGY OF PSEUDOGALANUS

has to be demonstrated, and some support for this view is found in the work reviewed in the previous sections. We consider, however, the expression of food consumption in terms of filtering rate to be a needlessly abstract way to express the intake of food and prefer to use ingestion rates. Ingestion rates can be expressed in a number of ways. Although often given as rates per individual or as rates per unit of body weight, rates given as a percentage of body weight (wet, dry, carbon, etc., as appropriate to the technique) are perhaps the most useful. The appropriate time unit is one 24-h period (we will follow the convention of using ‘‘ per day ”) in a priori recognition of the possibility of die1 rhythms. We refer to the total ingestion per day as the daily ration. The vast literature on consumption of food by copepods (reviewed recently by Marshall, 1973) is full of observations and experiments that do not seem to supply useful information on any of the above topics ; we make no attempt to be exhaustive in our review of matters other than those specified above as they relate to Pseudocalanw. 1. Effect of food concentration

Much of the earlier literature on feeding by copepods argued or assumed that filtration rate was independent of concentration, and this was proposed for Pseudocalanus as well (Gauld, 1951). The extensive experiments by Marshall and Orr (1966), using uptake of 32Pfrom algal cells, allow some inferences to be made about the suitability of certain food species. However, their variable and very low estimates of filtering rates give no information on effects of food density. They used small concentrations of cells, and losses of 32Pin faeces and excretion were not accounted for. The first indication of the seemingly obvious possibility that Pseudocalanus could be satiated a t high food concentrations comes from the work of Anraku ( 1 9 6 4 ~ ) . He showed that females in (evidently) 6-h experiments did not ingest more than about 5 x lo4 cells/day of the diatom Thalassiosiru Jluviatilis or filter more than about 40 ml/day of the medium (Fig. 12A). Urry (1965) found that rate of production of faecal pellets diminished but was not asymptotic a t concentrations of lo6 cells/ml of the flagellate Isochrysis galbana. Corkett (1966) extended these studies to higher concentrations. Although rate of faecal pellet production cannot be used to estimate feeding rates, it indicates the shape of response to density of two food species (Fig. 12D, E). Corkett also found that at concentrations of I . galbana less than 4 x lo4, faecal pellets were about A.M.B.-15

5

66

CHRISTOPHER J. COREETT AND IBN A. MCLAREN

half as long but the same width (and therefore about half the volume) of those pellets produced at higher concentrations. Parsons et al. (1967) estimated concentrations of food organisms in control and grazed samples of seawater with an electronic particle counter, and found that ingestion rate levelled off at high concentra. tions (Fig. 12B). Furthermore, they found that feeding virtually ceased

p " 0

3

I

L

2

l

3

4

5

2

Celts x103/rnl

4

5

G

10

I I5

20

Concentration (pm3 x 106/rnl)

50

Cells x

ro3/rnl

119 C / (

FIU.12. Evidence of saturation of feeding rates of Pseudocalanue. A, ingestion rate of the diatom !z'halassioaira Juuiatilk (after Anraku, 1964~). B, ingestion rate of mixed natural phytoplankton (after Parsons et a2.. 1967). C, ingestion rate of II mixture of the diatom flkeletonema sp. and '' p-flagellates " (after Parsons et al., 1969). D, number of fmcal pellets produced with the diatom Laudetia boreal& as food (after Corkett, 1966). E, number of faecal pellets with the chrysomonad 180chry8i8 galbana (after Corkett, 1966). F, ingestion rate (mean of all stages) of Pseudo. caEanua feeding on Thatassiosira rotula as a function of nominal food densities (after Paffenhofer and Harris, 1976).

in Pseudocalanus and other species at low food concentrations. Accordingly, they felt that daily ration r could be given as a function of food density p by the modified Ivlev equation : r = R [1-e-k(P-Pd]. Here R is the maximum (asymptotic) ration, p o is the level a t which feeding ceases, and k is the proportionality constant applying to the particuIar feeding species or food type. Unfortunately, the alga Chaetoceros s p . that predominated in their seawater samples was thought to have been almost uneaten, so the relationship between food density and ingestion rate is open to question. Parsons et al. (1969) confirmed the utility of their modified Ivlev equation in describing feeding rate of Pseudocalanus (with a small

THE BIOLOQY OF PSEUDOCALANUS

67

admixture of Oithona sp.) on food consisting of a mixture of Skeletonemu sp. and “ p-flagellates ” (Fig. 12C). Frost (1974) graphs ingestion rate of cells of Thalassiosira JEuviatilis by Pseudocalanus as a function of cell density. He shows an abrupt threshold of ingestion rate of about 1 400 cells/cop./h beginning at a cell density of about 1 100 cells/ml. Because he does not give data points on his graph, we do not include it in Fig. 12. A later paper (Robertson and Frost, 1977), also without data for Psezdocalanus, gives a maximum daily ration of 1-13 times body weight in carbon. We turn to the work of Paffenhafer and Harris (1976) for the most complete study of ingestion rates as a function of food density. They showed that filtering rate and ingestion rate were roughly proportional to body weight between stages C I and C V, so that the amount ingested per unit body weight at any given food level is constant for these stages. We use their nominal values of food density (actual values deviated little) in Fig. 12F. We have choaen to fit by eye more-or-less asymptotic curves to the examples of satiated feeding in Fig. 12, although we are aware of alternatives (e.g. Frost, 1974). A lower limit of food density a t which feeding ceases, as indicated in the work of Parsons et al. (1967, 1969) is not always evident in Fig. 12. Because the relationships have been assumed to be asymptotic, we have chosen to estimate the levels of food supply that lead to 90% of the maximal ingestion rate. We have used original estimates of cell volume and carbon content where given. The size and carbon content of T. Jlzlviatilis are from means in Mullin et al. (1966). Corkett and McLaren (1969) give the volume of Isochrysis galbana. Corkett and Urry (1968) give disc diameter of Lauderia borealis, whose volume is estimated from published illustrations by assuming that discs are twice as broad as high. Carbon contents for these two species are estimated from the carbon/volume regression in Mullin et al. (1966). There are startling variations in the results of this comparison (Table VII). The very high value for L. borealis is almost certainly due to the fact that it did not stay well in suspension in the static cultures used by Corkett (1966). The motile I . galbum is probably too small (4-8 pm) to be efficiently filtered; the work of Poulet (p. 63) suggests that it might be removed at about half the rate applicable for larger cells. Although Chuetoceros appears to be saturating a t high levels, it was hardly ingested at all. Even between the two species of Thalassiosira there appear to be differences (Frost’s estimate, cited above, for T.jluviatilis gives an abrupt threshold level at about 1 100 cella/ml or 0.21 pg C/ml). Clearly all these experiments give evidence

68

ORRISTOPHER J. CORKETT AND IAN A. MCLAREN

TABLEVII. LABORATORY ESTIMATES OF NEAR-SATURATION FOOD CONCENTRATIONS FOR Pseudocalanus. (From Fig. 12 ; see text.)

Food species

Thalassiosirafluviatilia Isochrysis galbana Lauderia borealis Chaetoceros sp. Skeletonema sp. and " p-flagellates " Thalassiosira rotula

Cell volume

Phytoplankton concentration at 90% of rnazimum ingestion rate

pm3

cells/ml

pm3/m1

pLgClml

1370 100 27 000 -

1700 160000 4300 -

3.9 x 106 16.0 x loG 116.0 X 10" 5-2 X lo6

0.32 2.72 5.14 0.27

-

-

11.5 X lo6 1.1 x 106

1.09 0.09

for satiation of feeding rates, but tell us little about the food levels at which this may be achieved in nature. Paffenhofer and Harris (1976) argue that their food densities match those found in nature, and that their use of large, rotating culture-vessels contributed to their low estimates of satiation food densities using T . rotula. It is also possible to calculate the maximal rates of ingestion from the experiments summarized above. Table VIII assumes the same TABLE VIII. ESTIMATES OF NEAR-SATURATION INGESTION RATESBY Pseudocalanus. (From Fig. 12 and Table VII.)

Daily ration Food species llm3

Thalassiosirafluviatilis Chaetoceros sp. SkeZetonerna sp. and " p-flagellates " Thalamiosira rotula

110 x lo6 0.8 x 106 40 x 40G 195 X loG

PFLS

c

8.14 0.04

3.8 16-1

conversion factors used in Table VII. The estimate for Chaetoceros sp. is clearly not comparable with those for the other food species, and supports the contention of Parson's et al. (1967) that it is hardly eaten because of its spines; the maximum ingestion rate is clearly not set by satisfaction of " hunger ". Although the estimates of ingestion rates in Table V I I I otherwise show considerable variation, this is not as extreme as the differences among saturation levels of food supply (see Table VII). It should be noted that the high estimates of ingestion

THE BIOLOGY OF PSEUDOCALANUS

69

rates (up to 140% of body weight per day, see Fig. 12F) of Paffenhofer and Harris (1976) are for growing copepodids, and the rest are based on adult females. There is clear evidence of high variability in feeding rates of individuals. I n a small series of week-long experiments with one or two copepods per vessel, Corkett (1966) found that one female consumed as much as 330 x lo6 pm3 Isochrysis/day for a week, whereas another took as little as 60 x 106 pm3/day. Similarly, the experiments of Delalo (1964) indicate that individuals varied greatly (see p. 73). We can now consider the relationship between food density and feeding rate in natural seawater as found in the two-year study by Poulet (1974). We have shown (Table V I ) that the regression coefficients of logarithms of ingestion rates of particle concentrations are somewhat less than unity when each size group of particles is considered separately. This is a suggestion that saturation might occur at higher food levels. However, the overall regression of ingestion rate (Y in mg X 10-4/h) on food concentration (X in mg/l) is given by Poulet (1974, with his corrigendum and correction as in our Table VI) as: In Y = 1.29 In X -0.33 ( r = 0.80, d.f. 93). The value of r after transformation to z and calculation of S.E., implies that the regression coefficient (1.29) is significantly greater than 1.0. Thus there is no evidence for levelling off of feeding rate with increased concentration of particles : quite the contrary. This non-proportionality is probably an artifact of grouping particles of different sizes, since smaller particles, for which regression elevations were lower, were also present in lower average concentrations (Poulet, 1974, his Fig. 9). In his later study of feeding by copepodids (Poulet, 1977) there is a weak positive correlation (weighted r = 0.61, d.f.8, p 0-06, from his Table 1) between feeding rate and monthly mean food concentrations. There is also no evident leveling off of rates at higher concentrations. The particle concentrations measured by Poulet (1974) averaged 5.2 p.p.m. or 0.27 pg C/ml, using his conversion factor. I n spring they frequently exceeded 10 p.p.m. or 0.52 pg C/ml. I n his study of copepodid feeding, Poulet (1977) tabulates food concentrations averaging 3.80 p.p.m. (0.20 pg C/ml). These values are close to or exceed those Bhat have been found to saturate feeding rates in some laboratory experiments (Table VII), especially the critical ones of Paffenhofer and Harris (1976). An explanation might be sought in the fact that on average some 79% of the particulate carbon in Poulet’s (1976) experiments was non-living. It this is relatively non-assimilable or otherwise unnutritious (contrary to Poulet’s suppositions) the copepods might continue filtering a t maximal rates in spite of very high food concenN

70

CHRISTOPHER J. COREETT AND IAN A. MOLAREN

trations. This, however, does not explain the low ingestion rates exhibited by the copepods in Poulet's experiments (see p. 72). 2. Effect of temperature

Although temperature certainly has a profound effect on the growth and development of Pseudocalartus (Section X), its effect on feeding rate has apparently been examined only by Anraku (1964b). His results are somewhat difficult to interpret. It should be borne in mind that only five females were used to determine each experimental point in Fig. 13. Anraku (1964b) does not report the concentrations of 50

L

1141181111 11111 2 16 20 :

' 0

Temperature ("C)

FIG.13. Filtering rates at different seasons of adult female Pseudocalunua from near Cape Cod, Massachusetts. Approximate habitat temperatures, H, are shown on each curve of the right-handgraph. (After Anraku, 1964b.)

Thlassiosira Jluviatilis he used, and it is worth noting that filtering rates were in other experiments about 30 ml/cop./day a t 1 000 cells/ml and only about 10 ml/cop./day a t 5 000 cells/ml (Fig. 12A). Furthermore, dry weight may be an inappropriate measure of filtering capacity, especially since it includes '' inert " materials like stored oil. Although the effects of temperature have by no means been fully explored in Anraku's experiments, it seems possible to conclude that high temperatures, even 15"C, depress feeding rates. The feeding rates per unit weight a t " in situ " temperatures (after Anraku, 1964b) are interpolated in Fig. 13B. There is a suggestion that the " optimum " rate lies near these habitat temperatures. The fact that filtering rate in August at 15°C is almost as high as that a t 8°C might be taken as a hint of acclimation to warm summer temperatures. However, there remains a great difference between rates at habitat temperatures, suggesting that no seasonal adjustment has occurred. However, let us suppose for the purpose of exposition that the size of

THE BIOLOGY OF PA’EUDOCALANUS

71

the copepods were as indicated in the length-temperature relationship for nearby Long Island Sound (Fig. 26, p. 118): 1-00 mm in May, 0.98 mm in December, and 0.81 mm in August. Assume further that the filtering capacity of a copepod is proportional to the square of its length: 1.00, 0.96, and 0.66 respectively for the above months. It can be seen that the similar filtering rates per copepod in May and December and the muchlower rate per copepod in August (Fig. 13A)make sense in terms of this approximation of filtering capacity. We do not offer this as a serious analysis, but as a plea for more thoughtful considerations of copepod size in design of such experiments. 3. Daily rationns in nature We have given several estimates of the maximal daily ingestion by copepods in the laboratory (Table VIII). Here we will summarize attempts to estimate the daily ration of Pseudocalanus in nature. Zagorodnyaya (1974) derived a mean daily index of fullness of guts (asa percentage of body volume) from observations on sampIes collected over 24-h periods in the Black Sea during January and March, 1973. Translating these indices into amounts of food consumed during 24 h required a measure of the rate of passage of food through the guts of the copepods. Zagorodnyays quotes a symposium paper by Pavlovskaya and Pechen’-Finenko (see also the same authors, 1975, for more details), who carried out experiments to determine the time required for clearance of radioactive food (Gynmodinium kowalevskii) from guts of adult females. Pavlovskaya and Peehen’-Finenko (1975) conclude that food passes through the gut at 8°C in 30 min. This seems to be somewhat arbitrary : their Fig. 2 indicates that almost all the radioactivity was evacuated from guts a t 20 min, and they choose to use an asymptotic curve to describe the series of points, which allows no real estimate of time for complete evacuation. However, their estimate (20 or 30 min) compares closely with direct observations by Delalo (1964) that identifiable remains of Prorocentrum micans appeared in faeces on average about 40 min after commencement of feeding. Using 0.5 h as an estimate of time for passage through the gut the daily ration of Pseudocalanus was calculated by Zagorodnyaya (1974) from mean daily indices of fullness: 24 multiplied by (index of fullness)/O-5. No trace of food was found in adult males. Table I X indicates that C V and adult females in March had the highest daily rations. These stages showed most marked diurnal vertical migration, and Zagorodnyaya’s work is considered later in that context (p. 165;). Poulet (1974) summarized his extensive experiments on grazing by female Pseudocalanus on natural food particles as monthly estimates

72

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

TABLEIX. DAILYRATION OF BLACK SEAPseudocalanus. (From Zagorodnyaya, 1974.) Ration,

yo of wet body weight

Month

GIV

CV

Ad.?

7.1

6.0

2.6

5.0

7.0 11.8

5.7 12.1

CIII January March

of mean food concentrations and mean daily rations. Rations were also expressed as a percentage of body weight, assuming that each adult female weighed 6.7 pg carbon (in a later paper, Poulet, 1976, he uses a value of 7.14 pg C). I n fact, if Pseudocalanus near Halifax vary seasonally as they do in other localities (Fig. 26, p. 118), a three- ta four-fold seasonal variation in body weight (from the relationship on p. 127) may be involved. Bearing in mind these possible sources oj error, the work of Poulet (1974, his Table 1) implies an essentially lineal relationship between food supply and ration. I n his study of feeding bj copepodids, Poulet (1977) shows (his Table 1) that they ingested ar average ration of 0.18 pgC/cop./day using his estimate of body weigh1 for copepodids (3-26 pg dry wt) and assuming them to be 50% carbor (see p. 126), we estimate that they consumed on average only 11% of their body weight per day (seasonal range, 3-16%). The estimates of daily rations as a percentage of body weight in nature of 2.6-12.1% (Table I X ) by Zagorodnyaya (1974) and of 2 . 2 4 5 5 % by Poulet (1975, 1977) are in general much smaller than those that can be gained at saturation feeding levels in laboratory experiments (Table VIII, assuming body weight of 6.7 pg C, as for Poulet’s work, above). The estimate by Poulet (1976) that only an average of 21% of the food of Pseudocabnus is living does nothing to explain this discrepancy. Non-living food is unlikely to be more nutritious than living food, and thus be equally satisfying as living food at lower densities. I n conclusion, there remain major questions about the amount of food consumed by Pseudocalanus in nature. These can only be resolved after a consideration of the amounts of food required by Pseudocalanus.

D. Die1 feeding rhythm It is probable that vertical migrations of Pseudocalanus routinely take it into depths where food is scarce, and some authors have assumed in their calculations that feeding only occurs at night (McLaren, 1963;

73

THE BIOLOGY OF PSE UDOCALAN US

Parsons et al., 1969). It is of interest to know if there is a feeding rhythm that is independent of this behaviour ;the evidence is equivocal. Anraku’s (1964c) 53-h feeding experiments revealed initial “acclimation ” of rates, but no evidence of die1 rhythmicity. Sharp changes could conceivably have occurred during the night, when no observations were made, but the amount of food eaten by morning was about as predicted from the rates prevailing during the previous day. Lock and McLaren (1970) found food present in guts of Pseudoculanus at all times of day and night (artificial light cycle) in rearing experiments. These experiments were with animals that had spent varying amounts

‘\ \#

\\ \\

/P\

1 ;z--el-*..

0a700

1100

:a..--

..*-_..--

1500

1900

Time of day

Fro. 14. Number of cells of tho dinoflagcllate Prorocentrum micans eaten by recently captured adult female Psedocalanus during 4 hour intervals. (After Delalo, 1964.)

of time in the laboratory. Possibly any natural rhythm had been lost. Delalo (1964) attacked the question directly. He placed freshly caught individuals in 15-20 ml receptacles and fed them the large (40-50 pm) dinoflagellate Prorocentrum micans at 200-400 cells/ml. The medium was changed every 4 h and the number of tests (indigestable) of P. micans were counted in the preserved faecal pellets. Strong feeding peaks were evident (Fig. 14). He found considerable individual variation among females in intensity of feeding, but peak times coincided in all individuals in February and in 3 of the 4 in March (when one peaked 1900-2300 h). Delalo compared mean consumption during the period of 6 h on either side of the peak time with that during the remaining 12 h and found significant differences by t-test in both February and March. Although this test is formally questionable in view of the small samples and inhomogeneity of variances, the coincidence of peaks is in itself clear evidence of cycling (9 out of 10 ;binomial

74

UHRISTOPHER J. CORKETT AND IAN A. MOLAREN

expectation 115, P < 0.01). Delalo also attempts to analyse the difference between the February and March curves using x2 (method unclear), and concludes that the difference is non-significant. However, the fact that all four March peaks are later than all six February peaks is significant (runs test, P < 0.05). Delalo allowed one individual in March to continue feeding for a further 24 h, and found that the feeding rhythm persisted, although at reduced amplitude (maximum 34 cells v. 81 cells consumed, at 1500-1900 h rather than 1900-2300 h, and total intake during the 24 h 143 v. 192 cells, estimated from his Fig. 2). Although it is unwise to

L t

‘ 80 OOE v1

f

.-

40

3

20

-

Jan.

c

0 1200

2000

0400

1200

l i m e of sample FIG.15. Observations on fullness of guts of copepodid and adult stages of Pseurlocalanua captured at various times of day in the upper 150 m of the Black Sea. (After Zagorodnyaya, 1974.)

read too much into the performance of one individual, there is thus a suggestion that behaviour changed after a period in the laboratory. Delalo speculates about the relationship of his results to feeding rhythms of vertical migration. This subject has been examined extensively by Zagorodnyaya (1974, 1975), using samples taken in January and March (dates not given) in the northern Black Sea. She dissected out guts of copepodids and adults (20 of each stage in each sample where possible) to estimate volumes of compact food masses. For present purposes it is enough to illustrate the clear evidence for rhythmic feeding in nature (Fig. 15). Most recently, Mackas and Bohrer (1 976) have used a fluorometric technique for detecting chlorophyll in gut contents to illustrate a moderate die1 rhythmicity in the intake of food by Pseudocalanus in waters near Halifax, Nova Scotia.

I

THE BIOLOGY OW PSEUDOCALANUS

75

Our general conclusion is that Pseudocalanus may show strong diel feeding rhythms in nature that can perhaps only be demonstrated in freshly caught individuals. Certainly short-term observations on feeding behaviour and feeding rates must a priori be considered suspect. Prudence might also suggest that only freshly caught individuals should be used if natural rates are being sought. E. Assimilation The only attempts to measure assimilation by Pseudocalanus are evidently those of Pavlovskaya and Pechen’-Finenko (1975), who worked with C V and adult females (numbers in each experiment unspecified) from the Black Sea. These authors used a wide variety of radiocarbon-labelled foods-living dinoflagellates, diatoms, and infusoria, freshly prepared detritus of single-celled and multicellular algae, humus of plant origin, and melanin. One set of experiments was run in triplicate at 8°C for 8-17 h, during different times of the 24-h period to accommodate possible diel irregularities. The amounts of food assimilated were measured by radioactivity of the copepods, which were dried for weighing after their guts were evacuated of unlabelled food. The amount of food assimilated by the copepods was very low in these experiments : maximally 3% of body weight in the case of the diatom Skeletonema costatum, followed by melanin (1.8%). The large dinoflagellate Peridinium trochideum, detritus from the multicellular alga Cystoseira barbata and humus of plant origin were all taken very little: 0-1-0.2% of body weight during the experimental period. By using 15-min exposures to labelled food, collecting all faeces, and measuring labelled CO, as respiration, Pavlovskaya and Pechen’Finenko were able to estimate assimilation efficiencies (percentage of ingested food appearing in bodies or respired). They used corrections for changes in feeding rates during the 24-h period in order to express results in percentages of copepod body weight per day, but the relative distributions of ingested food among faeces, body, and respiration are unaffected by this mode of presentation (Table X). Clearly there are marked differences in the amounts ingested of the different foods and the extent to which they are assimilated and used in respiration. Much of the assimilated food appeared to be lost in respiration during the experimental period. The two dinoflagellates were evidently relatively undigestible, while detritus of Ulva regida was little ingested. Other results by Pavlovskaya and Peched-Finenko are expressed by them only as assimilation eficiencies: Uronemu

76

CHRISTOPHER J . CORKETT AND IAN A. MCLAREN

TABLEX. ASSIMILATION OF FOOD BY C V AND ADULTFEMALE Pseudocalanus. (After Pavlovskaya and Pechen’-Finenko, 1975.)

yo of body wtjday Kind of food

Gymnodinium kowalevskii Peridinium trochoideum Detritus, Platymonas viridis Detritus, Ulva rigida

Unassimilated Respired (infaeces) (inCO,) 8.86 17.36 8-19 1.69

4.90 4.59 10.32 2.23

Remaining Assimilation in body eficiency (%) 0.57 0.85 9-26 0.45

38 24 71 61

marinum 68% ; detritus, Cystoseira barbata 53% ; humus 82%; melanin 7 3 yo. Thus, with the exception of the two species of dinoflagellates (Table X), most foods were digested relatively efficiently. However, 8s noted above, some of these were not readily ingested (detritus of the two multicellular algae, plant humus). It may be concluded from this work, that ingested food is generally assimilated with high efficiency ; but, in the absence of statistical analysis, it is not possible to conclude that the efficiencies ranging from 53 to 82% really differ significantly. Furthermore, the rather startling difference in the distribution of assimilated matter between the body and respiration “ pools ” (Table X) raises questions about the validity of the approaches and techniques (cf. Conover and Francis, 1973). Finally, in view of the enormous variability in rates of ingestion of food by Pseudocalanus shown in previous sections, minor differences in assimilability may be of little consequence.

F. Food requirements for sustenance 1. Amount of food The amount of food required by Pseudocalanus, like the amount consumed, is a matter of definition. Later sections consider the food requirement for growth (p. 133). Here the concern is with food requirement for sustenance of the adult and is measured by the survival of adult female Pseudocalanus kept in the laboratory. Urry (1965) demonstrated, as a prelude to determining food requirements for survival, that starved females from the overwintered generation off Plymouth (see p. 145) survived longer than the females from the f i s t spring generation. Corkett and Urry (1968) demonstrated that growth medium (Erdschreiber medium) alone could not be

77

THE BIOLOGY OF PSEUDOCALANUS

used as a food, and that varying culture volumes between 10 ml (1 copepod present) and 500 ml (2 copepods present) had no effect on survival if food supply was adequate. Corkett and Urry (1 968) showed that antibiotics did not prolong life of starved animals, but that small amounts of penicillin and streptomycin increased survival when the copepods were fed. Large amounts of antibiotics (more than 100 i.u. of each per ml) were clearly toxic. Antibiotics were not used by Urry (1965) and Corkett and Urry (1968) in their tests of food requirements, but the adults were placed in fresh culture medium every week. Two algal species have been tested a t varied concentrations. Isochrysis galbana at 30 000 cellslml promotes an almost full length of life (Table XI), although this level may supply only 60% of the food that Pseudocalanus is capable of ingesting (see Fig. 12E). Corkett and Urry (1968) found that much smaller quantities of the large diatom Lauderia borealis are needed-500 cells/ml being sufficient (Table XI) which is a level of only about 30% of that which can be ingested (see Fig. 12D). TABLEXI. SURVIVALOF ADULTFEMALE Pseudocalanua AT VARYINGFOOD CONCENTRATIONS. (From Urry, 1965, and Corkett and Urry, 1968.) Algal species Lauderia borealis

N o . of copepods

Concentration (cells/ml)

18 18 18 18

0 500 2 000 4 000

Isochrysis galbana

Average life (days) 14 41 53 46

N o . of copepods 20 20 20

20 20

Concentration (cells/ml) 0 1000 5 000 30 000 100 000

Average life (days) 17 23 26 58 83

2 . Quality of food Urry (1964, 1965) tested the quality of a number of species of

algae in sustaining adult females. Each species was fed to a group of copepods (taken from the same samples from nature) at 30 000 cells/ml and compared with a control group of copepods fed Isochrysis galbana at 30 000 cellslml (Table XII). Clearly a wide variety of food species permit the survival of adult females. It is of interest to look at those species that seem nutritionally inadequate (Table XII). Urry (1964, 1965) does not explain the

78

UHRISTOPHER J. UORKETT AND IAN A. MOLAREN

inadequacies of Nitzschia gotlandica or Hemiselmis virescens. He does suggest that, although Gymnodinium veneJicum has been known to produce poisonous exocrines, his cultures had evidently lost their toxicity. TABLEXII. SURVIVAL OF ADULTFEMALE Pseudocalanus EXPRESSED AS A PROPORTION OF THE CONTROL GROUP. (After Urry, 1964, 1965.) Rood apeciesa Diatoms Phaeodactylum tricornutum Nitzsohia gotlandica Dinoflagellates Cryrnnodkium veneficum a. vitiligo Arnphidinium sp . Cryptophyte Hemiaelmis virescens Chlorophytes Chlorella stigmatophora Dunaliella tertiolecta Chlamydomonaa coccoides Chlarnydomow sp. Chrysophyte Dicrateria inornata

Relative length of life

N o. of copepods

0.93 0.37

48 40

1.14 0.59 0-51

48 61 20

0.68

25

0.57 0-61 0.98 0.89

48 20 20

<0.36

~~

a Both food species and control (Zsochlyais galbantc) at 30 000 cells/ml, replenished weekly.

G . vitiligo is not known to be toxic, but Amphidinium spp. have this reputation. Some Chlorella spp. are toxic, and Urry (1965) showed that Pseudocalanus survived poorly (0.56 of control) when fed Isochrysis galbana in a cell-free extract of C. stigmatophora. Urry also found undigested cells of this species in the faeces. No such toxicity effects were noted in the other relatively unsuitable chlorophyte, Dunaliella tertiobcta. Corner and Cowey (1968) compared amino acid contents of the chrysophyte Dicrateria inorwta with that of the diatom Phaeodactylum tricornutum, taking (from Urry, 1965) the former as a " poor " and the latter as a " good " food. There was little difference in amino acid contents, and we suggest that the former is probably " poor " because of its small size (3-6 pm) compared to the Isochrysis control ( P 8 pm).

THE BIOLOGY OF PSEUDOUALANUS

79

Although these experiments on food quality are of some intrinsic interest, they probably tell us little of the nutritional problems faced by Pseudocalanw in nature. None of the toxic or unsuitable species are widespread or dominant members of the open-water coastal plankton. PseUd0calanu.s may more often face food shortage, or refractory or unnutritional non-living food, than with unsuitable food species within the suitable size-range. G. Retrospects and Prospects We are left somewhat unsatisfied by published work on feeding of Pseudocalanus and, by inference, of other copepods. We are uncertain whether the low selection of small ((10 pm) particles is because of poor sieving ” or a result of creation of small particles from large. It is possible that seasonal differences in the temperature response of feeding rate would not be found if a more appropriate measure of size of individuals were used. We are not really sure whether Pseudocalanw clears its gut in half an hour, twenty minutes, or even less. However, the most serious questions seem to us to concern the great variability of estimates of feeding rates, whether calculated from fullness of guts in nature, from rates of removal of natural food in seawater samples, or from removal of defined food species in laboratory experiments. The work of Paffenhafer and Harris (1976) stands apart in revealing that very high feeding rates at low, natural food concentrations can be shown by copepods reared in the laboratory. We think it is quite probable that wild-caught animals (generally adult females) may fail under most conditions to show meaningful feeding rates. More attention should be given to residual die1 rhythms. Experiments should clearly be conducted with uncrowded animals, preferably with some sort of mixing of the medium. Consideration should be given to the character of individuals in grazing experiments; are they fat or lean, recently moulted or not, gravid or infertile, or are they even in some sort of resting stage? Recently Conover (1978) has made the bold suggestion, using examples of unsaturated feeding rates shown by Poulet (p. 69) for Pseudo&nw, that copepods adapt in nature by producing more digestive enzymes in response to increased food levels. Clearly this ability could underline the great variability shown by Pseudocalanw under various experimental conditions, and we look forward to further explorations of Conover’s hypothesis. We will show later that maximal, temperature-dependent rates of ‘I

80

CHRISTOPHER J. CORKETT AND

IAN A.

MCLAREN

reproduction (p. 98), development of stages (p. 156) and production (p. 189) may occur during a substantial part of the year in temperate waters. Further, it seems that Pseudocalanus can suspend development and enter an overwintering, “resting” phase even when food is plentiful (p. 157). Studies of food choice and feeding rates, plagued with inaccuracies and uncertainties, may be irrelevant when Pseudocalanus is so seldom food-limited in nature. Altogether, we conclude that more extensive studies of nutrition in Pseudocalanus may not be the most revealing approach to finding out how it “ works ” in nature.

IX. REPRODUCTION A. Sex ratio Although sex ratio is also an aspect of development or demography and could be considered in the section on development and growth or life cycles, we include it here because of its clear relationship to reproduction. Sex ratios of adult Pseudocalanus, like those of many other copepods, are strongly distorted in favour of females. Many authors give sex ratios in relation to localities and seasons, and some speculate on the differences. It would not be profitable to quote many of these, since it is quite clear that the shortage of adult males in nature is due to their restricted life span relative to that of females, and not to markedly unbalanced primary sex ratios. We will quote some of the evidence for this conclusion from field samples and laboratory studies. The short life of adult males is most obvious in seasonal samples from high latitudes. I n the far north, adult males may appear in numbers only around the time when overwintered copepodids begin to mature and then evidently disappear, whereas adult females may be found through the year (e.g. Ussing, 1938; Grainger, 1959). Ostvedt (1955) found that males reached maturity and fertilized females in the depths below 600 m in the Norwegian Sea, but failed to ascend with the females to the surface in spring. A number of authors have shown that males and females in plankton samples are roughly equal in abundance among C IV and C V, which show secondary sex characters (e.g. Ussing, 1938 ; Bogorov, 1939 ; Marshall, 1949 ; Fontaine, 1955 ; McLaren, 1969). The small deviations from unitary sex ratios that are discussed by some of these authors are probably due to different rates of development of males and females in these copepodid stages (McLaren, 1969). Laboratory evidence indicates clearly that adult males have shorter lives than females (p. 114), but rearing experiments are somewhat equivocal. Thompson (1976) used I . galbana and reared 87 adult

THE BIOLOGY OF PSEUDOCALANUS

81

females and 29 adult males at a range of temperatures ; no trend in sex ratio was evident in relation to temperature. On the other hand, Paffenhijfer and Harris (1976)obtained 73 adult females and 74 adult males on a diet of Thalassiosira rotula. There thus appears to be some trophic factor in determining adult sex ratio, but whether this operated in these experiments through differential mortality of males or through primary sex determination is unknown. We think it very unlikely that a trophic factor was responsible for the different sex ratios in samples from the North Sea and White Sea as speculated by Mednikov (1961). The diatom T . rotula is also better than the flagellate I . galbana in efficiency of feeding (Table VII), and for growth and development (p.113). We consider that the short life of adult males is related to the fact that females are fertilized only once during their lifetime, shortly after they mature (see p. 83). Under these circumstances early reproductive efficiency of the males is important, even a t the possible cost of a long life.

B. Oogenesis and egg laying The oogenetic cycle of Pseudoculunus has not been studied, but the superficial appearance of developing eggs has been described by Corkett (1966),who distinguished three stages (Fig. 16). The immature stage containing small developing eggs in the ovary, but not in the oviduct, is found in C III-C V, and in newly matured adult females (Fig. 16A, B). The semi-ripe stage contains some large eggs that may extrude into, but not fill, the oviduct and its diverticula (Fig. l6C). The eggs in the ripe stage (Fig. 16D) may fill the oviduct and its large anterior diverticula. After laying, the animal reverts to a semi-ripe condition. Corkett also found a few females with a degenerating ovary; these females were more abundant at the beginning of the year and perhaps came from the overwintered generation. Egg laying has not been described, but as in Calanus (Marshall and Orr, 1952)the eggs are extruded in a soft condition and become round and firm only after they are attached to the genital segment. The structure of egg masses and sacs and their membranes is considered elsewhere (p. 8 5 ) . C. Sperm and spevmatophore production

The spermatogenetic cycle has not been described, nor is it known how many spermatophores can be produced by a male during its short

a2

CIIRISTOPHER J. CORKETT AND IAN A. MCLAREN

life (see p. 114), although a single male has been known to fertilize up to three females in the laboratory (Hart and McLaren, 1978). Giesbrecht’s (1882) illustration of a spermatophore shows suggestion of layering of the masses of sperms within (Fig. 17A), and Sixth thoracic segment

A

Fiith thoracic segment

B

G e n h opercufum

ovary

7iverticulum

Egg mass

Left oviduct

FIQ.16. Development of the ovary of Pseudocalartus. A, C V. B, adult female with immature ovary. C, adult female With semi-ripe OVV. D, adult female with ripe ovary. (After Corkett, 1966.)

Hirschfeld (1974) indicated that the tightly packed sperms are arranged in a helical manner. She counted about 360 sperms in an acetocarmine squash of one such spermatophore. We will see that these would be enough to fertilize the eggs produced by a large (1.1 mm or so) female

THE BIOLOGY OF PSEUDOCALANUS

83

A B Fro. 17. Spermatophores of Pseudocalanua. A, spermatophore extruded from mele gonopore. B, unusually large number of spermatophores attached to a single (abnormal, see text) female. (From Giesbreoht, 1882.)

during her lifetime: about 35 eggs per clutch (Fig. 19A) for a total of 10 clutches or so (p, 93).

D. Mating Some authors have commented on the fact that the elongated spermatophores are rarely found in nature. Adult males are also shortlived (see p. 80). These facts in themselves suggest that mating is normally a once-in-a-lifetime act for each female. Giesbrecht’s (1882) report of up to 70 spermatophores attached to single females is extraordinary. Giesbrecht’s illustration (Fig. 17B) is of interest in indicating what appears to be a rudimentary fifth leg. This can be

84

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

associated with parasitic sterilization or intersexuality (p. 194) and possibly the mechanism of mate attraction (see below) could be badly distorted. Unlike some species (e.g., Eurytemora herdmani ; McLaren, 1976a) Pseudocalanus seems difficult to mate in the laboratory. I n one experiment about four C V females were allowed in the presence of a single adult male in 54 individual 125 ml bottles with adequate food; mating success was less than 10% (Hart and McLaren, 1978). Increasing size of culture vessels beyond 100 ml seemed to have no effect on mating success (Hirschfeld, 1974). Hart and McLaren (1978) found that there was evidence for size-assortative mating ; i.e. large males chose large females and vice versa, when given a choice in the laboratory. The evident sexual fastidiousness of Pseudocalanus may be in part related to this size-assortative mating. Some of the events leading up to mating have recently been clarified by Griffiths and Frost (1976). They found that males placed in filtered seawater with recently matured females exhibited swimming behaviour similar to that described for other species by Katona (1973) : rapid zig-zags alternating with slow figure-eights and loops. Ten experimental beakers, with five males in each, were subjected to 20 min with maleconditioned, 20 min with female-conditioned, and 20 min with control (freshly filtered) water. The frequency of occurrence of the three kinds of male behaviour is given in Table XIII. Clearly these elaborate patterns of swimming have a sexual connotation and are probably evoked by substances produced by freshly moulted females. TABLEXIII. FREQUENCY OF OCCURRENCE OF DIFFERENT MALE BEHAVIOURS IN DIFFERENTLY CONDITIONEDWATER. (After Griffiths and Frost, 1976.) Conditioned water Behaviour zig-zag looping figure -eight

Male

Female

Control

1 5

10 10

4

4

8

3

1

Griffiths and Frost obtained further evidence for such a pheromone in Calanus by demonstrating that there was a significant accumulation of radioactivity in adult males exposed to seawater conditioned by labelled males or females. They showed that this radioactivity was concentrated around the positions of aesthetes on the antennules of

THE BIOLOGY OF PSEUDOCALANUS

85

Calanus males. Their drawing of the first antenna of the Pseudocalanus male (Fig. 7A) shows a rich endowment of aesthetes. Griffiths and Frost suggest that the function of the elaborate swimming behaviour of males may be to bring widely spaced individuals together. However, we can find no published observations on the way in which the male approaches and attaches the spermatophore to the female, presumably using the remarkable fifth legs (Fig. 7K) during what must bc a very ephemeral coupling.

E. Reproductive rate 1. General pattern of egg production

The following terminology (Corkett and McLaren, 1969) is used throughout this review. A true egg sac is a group of eggs clearly surrounded by an outer membrane and attached to a female. An egg mass is a similar group of eggs attached 60 a female, such eggs adhering to one another by the outer egg membrane, with no further surrounding membrane. The general term clutch. is used for the eggs contained within a sac or mass. The eggs are generally carried by the female until they hatch, although the clutches may in some populations under some circumstances break up before hatching (Corkett and Zillioux, 1975). When clutches break up they often leave a mass of eggs attached to the female and the rest of the eggs become detached singly or in groups. We assume that eggs are always laid in sacs which can be recognized by the symmetry imparted to them by the outer membrane. I n plankton hauls these egg sacs are often disrupted and leave only a fcw eggs (i.e. masses) clinging to the genital segment. It is not possible to establish patterns of egg production in nature as observations have to be made over a period of time on individual females. We believe that the normal potential production of eggs by females (not reduced by food limitation) is illustrated by the examples in Fig. 18. These individual females, captured as adults in nature, were among those producing the largest number of egg sacs in their respective experiments, and perhaps had only recently been fertilized in nature. The reproductive life of these females can be divided into two phases. First there is a reproductive period of sustained production of clutches of more-or-less similar size (horizontal lines in Fig. 18). This may be followed by a period during which clutches are smaller or even entirely inviable (eggs disintegrate soon after being extruded and were presumably infertile). It is not known if this apparent infertility is due

86

CHRISTOPHER J. CORKETT AND IAN A. MULAREN

t o shortages or inadequacies of stored sperm or to physiological shortcomings of the females. Death in any case may follow after a quite long, essentially post-reproductive period (after horizontal lines, Fig. 18).

Reproductive rate per female can be analysed in terms of five variables : (a) the number of eggs in clutches ; (b) the rate of production of clutches; (c) the total number of clutches produced during the reproductive period ; (d) length of post-reproductive period ; and (e) the proportion of eggs in these. clutches that hatch as nauplii. 20-•

..

Fertile clutch o Inferti18 sac t Death of female

10 -..*,-I

$30;

.

I

Time (doys)

FIQ.18. Production of successive clutches in the laboratory by female Pseudocalanw captured in nature. A, female from off Plymouth, England, kept at 844OC in excess laochrysis galbuna, replenished weekly (after Corkett and Zillioux, 1976). B and C, females from off Halifax, Nova Scotia, kept at 6-7"C, at 3 X lo4 cells/ml (B) and 3 x 105 cells/ml (C) of I . gulbanu replenished weekly (after Corkett and McLaren, 1969).

(a) Number of eggs in clutches Figure 18 indicates that, although there is little evidence for trends during the full reproductive period of females, the size of individual clutches may be quite variable. There is some suggestion in Fig. 18 (and in other examples depicted by Corkett and McLaren, 1969) that exceptionally small clutches are often followed by exceptionally large ones, and vice versa. Perhaps this involves withholding eggs between oogenetic cycles. Although a variety of explanations have been offered for clutch size in Pseudocalanus, the importance of female size was first explicitly recognized by Marshall (1949). McLaren (1963) showed that her data from Loch Striven on number of eggs (E)as a function of cephalothorax length (L, in mm) were closely described by E = 19.36.LB.g4.

87

THE BIOLOGY OF PSEUDOCALANUS

Subsequently McLaren (1965) and Corkett and McLaren (1969) concluded that this same function described egg numbers in unbroken sacs from Long Island Sound, Nova Scotia, and the Canadian arctic, provided local difference in egg size were taken into account. Reanalysis of these data shows that clutch volume is indeed more closely related to length than is the number of eggs in the clutch (Fig. 19). C

0 O

Loch Striven, Scollond

n

0 Halifax.Nova Scatia

A Ogac Lake,Baffin lslond eFoxe Basin OSouthwes?Baffin Island Long Island Sound *Unpam Bay

A

0.0

A

em

/

0

E = 0.0201L449

m

*

Cephalolhorox length(L1 in mm

FIQ.19. Clutch sizes in number (A) and calculated volumes (B) as functions of cephalothorax length of adult female Pseudoculunw. (After McLaren, 1966, and Corkett and MoLaren, 1969.)

88

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

The relationship between clutch volume and length is significantly greater than cubic (95% c.1. of the exponent in Fig. 19B are 3.44-4.94). This relationship can be compared with that describing body weight as a function of length, which is also significantly greater than cubic (exponent 3.40-3.88, see p. 127). Although an upper limit to clutch volume may be imposed by space within the body, as suggested by McLaren (1965), the fact that mean clutch volume in a female is smaller than the largest clutch volumes (Fig. 18A, C) suggests that some other adaptive control is operating. Perhaps sizes of clutches are controlled due to hydrodynamic restrictions required for swimming, especially for escape. Corkett and McLaren (1969) found that mean numbers of eggs in a sac were uninfluenced by wide variations of food supply (Fig. 20). Five females kept by them a t an even lower food supply of 3 000 cells 1. galbanalml failed to produce any sacs in 5-7 days. The coincidence of large clutches and spring diatom outbursts (Marshall, 1949) is a consequence of large female size in the first new generation developed at spring temperatures (see Section XI for many examples). Corkett and Zillioux (1975) found that temperatures had no effect on clutch size of females captured off Plymouth, but that those produced at higher temperatures tended to release more of their eggs prior to hatching times (Table XIV). It seems possible to conclude from the above work that mean clutch size during the reproductive period of the female is determined by her size, and is influenced little if a t all by circumstances surrounding her during this time. Next we examine work on Pseudocslanus against this conclusion. E F F E C T S OF TENPERATURE ON EGGPRODUCTION B Y Psetl,doca~anus FROM OFF .PLYMOUTH. (After Corkett and Zillioux, 1975.)

TABLEXIV.

1.3 4.0 8.4

16.2

2 7 13 7

12.3 11.3 & 2.2 13.8 f 2.1 11.5 -J= 1.7

4 5 12 42

0-7 1.1 2.3 3.4

24.5 35-4 38.2 36.9

11-38 9-57 7-88 3-71

THE BIOLOUY OF PSEUDOCALANUS

89

Paffenhofer and Harris (1976) measured reproduction performance of a number of females reared at 1 2 O C . Average clutch size was 16-26, but there was variation within and between experiments. Apparently they included small clutches produced late in life in determining mean clutch sizes. This is perhaps reflected in the smaller mean clutch sizes in an experiment started with 10 " older '' females. Since no data are given on the sizes of individual females, it is not possible to explore their observations further. Thompson (1976) found in extensive experiments that females from North Sea stock produced on average about half as many " viable eggs " per clutch as might be expected from the relationship indicated in Fig. 19A. Her estimates are actually based on counts of nauplii, and will be considered later (p. 94). Sazhina (1971) gives a clutch size of 27 for Pseudocalanus from the Black Sea. Elsewhere (Sazhina, 1968, 1974) she lists values of 27-30 eggs at ZO"C, 16 a t 15-17"c, and 9-11 at 18-22°C. The trend within this (elevated) temperature range is interesting, but we are given no details on sizes of fernales (tabulated simply as 1.2 mm) or the conditions of capture or rearing. We conclude by emphasizing the importance of specifying body size (and egg size if possible) in any experiments or analyses on egg production by Pseudocalunus. fb) Rate of production of clwtches A female obviously cannot produce a new sac while one is being carried on her genital segment. Even if the egg sac is dropped prematurely, a new one does not usually appear until nauplii in the detached sac are hatching; Corkett and Zillioux (1975) noted a single exception. The rate of production of sacs is determined by the time taken for embryos to develop to hatching and by the time between hatching of one clutch and appearance of the next. Embryonic duration is controlled by temperature but time between clutches can be subject to other influences, of which food supply can be assumed to be important. In a high food supply, a new sac can appear within an hour of hatching of the previous one. This lapse of time was determined precisely for a few females from Halifax, Nova Scotia by Corkett and McLaren (1969) and can also be estimated from the numbers of sacs produced by females at various food concentrations (Corkett and McLaren, 1969, their Fig. 2). To estimate this time, it is assumed that the first and last sac during the reproductive period (i.e. at the ends of the horizontal lines in Fig. 18) were discovered a t random with respect

90

UHXISTOPHER J. OORKETT AND IAN A. MULAREN

to their stage of embryonic development. The proper estimate of number of clutches during this period is then one less than the number observed. Each of these clutches can be assumed to have taken about 4.7 days at the given experimental temperature of 6 7 ° C (see Fig. 22). The remaining time can be allotted to periods between clutches. Evidently little time is lost between clutches at high food densities, TABLEXV. ESTIMATES OF TIMESBETWEEN HATCHING OF A CLUTCHAND APPEARANCE OF A NEW ONE IN Pseudocalanus. (After Corkett and McLaren, 1969 ; see text.)

Days between clutches Isochrysis galbanaa celEs/ml

Number of females

Range

0.6 0.8

0.3-0.8 04-4.1 0.1-2.4 1.0-5.1 3.2-6.1

3 9 3 6 3

600 000 300 000 150 000 30 000 15 000 ~

Mean.

1.2

3.7 4.6

~~~~

Food level replenished weekly.

although a few individuals may show longer delays than most (Table XV). The average time between hatching of successive egg sacs is about 20% longer than the time of embryonic duration when food is adequate (>30 000 cells/ml, replenished weekly ; Table XV). Production rate of clutches begins to drop at levels below 150 000 cells/ml, but probably not much above 30 000 cells/ml, when Iaochryaia galbana is

p

z

015,000 Cclls/ml

5-

mo 30,000

Cellr/ml

~ I S 0 . 0 0 0Celb/ml

91

THE BIOLOGY OF PSEUDOCALANUS

used. It is interesting to note that a t only 15 000 cells/ml the number of eggs in a clutch is not reduced (Fig. 20). Now we may examine other, less extensive estimates of rate of production of clutches against the observations given above. Sazhina (1971) assumed an interval of 3-5 days and later (Sazhina, 1974) 3-5 days between successive clutches for Pseudocdanus from the Black Sea. She did not consider possible effects of temperature on the interval but these estimates refer to 8-10°C (see Fig. 22). Corkett and Zillioux (1975) derive an estimate of the maximal rate of production of sacs for well fed animals from Plymouth by dividing 12-6 (the mean number of eggs in a sac) by the time taken for development to hatching at their four experimental temperatures. They found that the observed rates of egg production were only about 50-70y0 of these maximal rates (compared with 83% for Halifax females, above). However, they based their estimates on rate of egg production during the whole laboratory life of the female, including the post-reproductive period. The extensive work of Thompson (1976) allows us to estimate the time between hatching and sac formation for seven females reared and mated in the laboratory. Here the reproductive period was explicitly designated, as time of appearance of last clutch was noted. It is possible, however, that females could have slowed down rate of production of sacs before this in some cases (cf. Fig. 18). From Thompson's data, we estimate that there was generally a mean lapse of a day or so between clutches of females (Table XVI). Longer periods occurred TABLEXVI. PARAMETERS OF REPRODUCTION BY SEVENFEMALE Pseudocalanue REAREDIN THE LABORATORY. (After Thompson, 1976.)

Temp. "G

Est.a time to hatching

Total no. of sacs

6.1 8.6 10.8 13.5 13.5 14.5 14.5

5.2 4.0 3.2 2.5 2.5 2-3 2-3

7 10 5 16 6 16 3

Eat. time Obs. time Obs. time Obs.b between first to carrying of postmating to first clutch, laet clutch, clutch,es, reproduction day8 days time, days dW8 4 3 4 16 16 2 2

43 40 24 54 16 44 6

1.9 0.5 2-8 1.3 0.7 0.6 0.2

From Fig. 22. Time between appearance of last clutoh and death of female.

38 10 3 70 64 67 41

92

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

between clutches of the female at 10*8"C,which died shortly after her fifth clutch appeared and may have been abnormal. Excluding this female, the times between appearance of successive clutches averaged about 28% longer than the times for development to hatching of eggs of the various temperatures. This is a good match for the 20% estimate for females from Halifax (above). Paffenhofer and Harris (1976) tabulate the means in four experiments of number of sacs produced by females, numbers of eggs per sac, and periods of nauplii production in days. If the last are to be interpreted as mean times between hatching of first eggs and hatching of last, then an estimate of times between successive clutches can be made, as above, by subtracting one sac from the mean number produced, I n the one experiment with Thalassiosira rotula as food, four clutches (five minus one) were produced in 14.33 days. It is assumed that this experiment was carried out at 12-5'C like their rearing experiments and that embryonic duration is about 2.8 days (Fig. 22). This implies a mean period between appearance of successive clutches about 24% longer than the time for development to hatching. I n three other experiments using Peridinium trochoideum, (concentration?)the mean delays between hatching of one clutch and appearance of the next were 3.3, 4.7 and 6-0 days. This seems to confirm the conclusion of Paffenhofer and Harris (1976) that P. trochoideum is a poor food for Pseudocalanus. From all these sources, we conclude that successive clutches of well fed, fully reproductive females should appear at intervals equal t o about 1.25 multiplied by the duration of embryonic development for the given temperatures. The lapse of time between mating and the first clutch has been estimated for seven laboratory-reared females by Thompson (Table XVI). If we exclude the two females at 13.5"C as abnormally retarded, the first clutch in Pseudocalanus may appear in about the time taken t o hatch a clutch. Since the female is fertilized very soon after moulting to adulthood (see p. 83), a period between maturation and first clutch similar to that between hatching of successive clutches might be assumed for demographic or productivity studies. (c) Total number of clutches We have shown that females in the laboratory produce a number of normal or complete egg sacs, and then may produce smaller or infertile ones during an essentially post-reproductive period (Fig. 18). Among females captured in nature, some will have already

THE BIOLOGY OF PSEUDOCALANUS

93

expended part of their reproductive potential. Those females producing maximal numbers of egg sacs are more revealing of reproductive potential. Of 33 females whose reproductive histories are depicted by Corkett and McLaren (1969), eight produced between 8 and 11 full-sized clutches and between 0 and 3 small, late ones. This strongly suggests that the normal reproductive potential for females off Nova Scotia is about 10 successive clutches. Sazhina (1971) suggests that Pseudocalanus in the Black Sea may produce 21 clutches, but this estimate is based on an assumed interval of 3.5 days (embryonic duration at 10°C) and an unsupported estimate of 75 days for the length of the reproductive period. The largest number of clutches produced by an individual captured off Plymouth was 9 (see Fig. 18A), all of which hatched successfully (Corkett and Zillioux, 1975). Laboratory raised and fertilized females can, in principle, give more precise estimates of potential number of egg sacs. Results of three such experiments are tabulated by Paffenhofer and Harris (1976). I n one experiment using Peridinium trochoideum as food, 15 females produced an average of only 2.27 (range 2-4) egg sacs. I n another experiment three females produced only one, two, and two sacs respectively. This is good reason (along with the delays between sacs noted on p. 90) for thinking that P. trochoideum is an unsuitable food. I n another experiment using Thalassiosira rotula as food, six females produced an average of five clutches (range 2-9), which is lower than might be expected from the maximal performances of wild-caught females. Paffenhofer and Harris state that unfertilized eggs were colIected when females were about to finish egg production, but it is possible that some of their females did not fulfil their reproductive lives. The experiments were carried out in bowls, in which T . rotula may not have stayed in suspension. Another experiment with 12 females in a rotating beaker prolonged the period of reproduction, but no data are given on total numbers of clutches. The seven females fertilized in the laboratory in Thompson's (1976) study (Table XVI) produced an average of 8.9 egg sacs, close to the potential indicated for wild females. The variation in number of sacs in her experiments may be " natural ", whether related to premature death (female at l0-S"C)or to premature infertility (females at 13.5, 14-5OC). We are given no information on whether the late clutches of the females producing 15 and 16 sacs were full sized. For purposes of calculating demographic or production parameters, it may be perfectly acceptable to assume a potential of 8-10 clutches. I n fact, observed or assumed mortality rates will ensure that very few

94

CHRISTOPHER J. CORKETT AND IAN A. MOLAREN

females will reach this potential, or food may become seasonally scarce long before this potential is reached. (d) The length of the post-reproductive period Evidently females can live for some time after reproduction has essentially ceased. We have suggested above that the eight out of 33 females that carried 8-11 successive complete clutches in the experiments of Corkett and McLaren (1969), had been captured soon after maturity. The mean reproductive span of these eight females was 51 days, and they lived an average 41 days beyond this. The seven females reared and mated in the laboratory by Thompson (1976) had mean reproductive periods (mating to last clutch) of 39 days and post-reproductive periods of 40 days (TableXVI). Elsewhere we suggest that the potential length of life may be set physiologically by temperature (p. 115), and the post-reproductive period would accordingly be shorter in females that have been thwarted from sustained reproduction by food shortages. However, for females producing a full complement of about ten clutches at a maximal temperaturedependent rate, the evidence suggests that an essentially postreproductive period can occupy the second half of adult life; however, given natural mortality rates this post-reproductive life is unlikely to be expressed in nature (but see work of Martens, 1975, discussed on p. 208). (e) Proportion of eggs hatching Corkett and McLaren (1969) and Paffenhofer and Harris (1976) found that infertile or otherwise unviable eggs are normally produced only when females approach the end of their reproductive lives. Death of some embryos at other times does occur, and might be expected among females confined to small experimental vessels. It would seem unlikely that infertility or embryonic death are frequent in natural populations. Therefore, it is a little surprising that Thompson (1 976) found that the rate of production of ‘‘ viable eggs ” by females from the North Sea was considerably lower than might be expected. Thompson gives estimates of average numbers of “ viable eggs ” per sac for a very large sample of females of known cephalothorax lengths. Those between 0.72 and 0.78 mm produced on average 5.1 such eggs and those 1.00-1.12 mm on average 9.3 eggs, or about half the mean number carried in sacs of females from Nova Scotia or Scotland (Fig. 19A). Her estimates of “ viable eggs ” for females mated in the laboratory and for females captured seasonally are also substantially below expected levels.

THE BIOLOGY OF PSEUDOCALANUS

95

Thompson’s counts of “viable eggs” were based on counts of nauplii found on periodic examination of females. We have found (unpublished observations) that females may eat some and on occasion all their nauplii, often very soon after they have hatched. This is presumably the major source of discrepancy between Thompson’s results and those summarized in Fig. 19. A useful study of viability of eggs of Psezcdocalanus could be made with containers adequately large for females to oxygenate their eggs by swimming, and by removal of sacs shortly before they hatch, so that no eggs are lost. 2. Theoretical rates of reproduction

Embryonic durations can be established accurately, and we have suggested that the time between successive clutches (or hatchings) might be about 25% longer than these durations when food is adequate. We have also shown that numbers of eggs in a clutch can be predicted for local populations, and that inviable eggs can probably be ignored. Thus estimating the maximal potential rates of reproduction per individual female might seem straightforward : simply divide the observed or calculated mean egg number in clutches by 1-25 multiplied by embryonic duration at the given temperature. This is in effect what is suggested by Sazhina (1974). However, this needs further refinement, as some females may not have produced their fist clutches and others may be post-reproductive. I n populations with continuous, overlapping generations the relative abundance of fully reproductive females will depend on the age structure of the adult female population. If mortality rates or recruitment of new adults are high, the pre-breeding females will be relatively common. Post-reproductive females will be more prevalent if mortality or recruitment rates are low. We have suggested for purposes of approximation that the prereproductive phase might on average last as long as the interval between appearance of successive clutches and that the post-reproductive phase might occupy the second half of a female’s life. We have used these approximations to estimate the proportion of females that might be fully reproductive in populations of various age structures. For simplicity we assume that the number in each successive age-group is a constant fraction of the preceding one (which is the same as assuming that constant mortality and recruitment rates have prevailed for some time). For a series of such populations, in which the oldest reproducing females (tenth clutch) ranged between 1% and 50% as common as the newly matured ones, elementary analysis shows that reproducing females constitute between 59% and 73% of the population.

96

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

We suggest that it might be assumed that an average of 65% of females in populations with continuous, overlapping generations are fully reproductive. It is on this basis along with the assumption that 25% of fully reproductive females will be between clutches a t any given time that we have calculated the potential rates of reproduction depicted in Fig. 21.

Temperature PC)

FIG.21. Average number of eggs produced per day by populations of food-satiated females of various lengths as functions of temperature. Unbroken curves are for females from off Halifax, Nova Scotia, and broken curves for females from southwest B a 6 n Island (see Fig. 19).

3. Reproductive rates in nature Although there are a number of published observations on the number of eggs borne by females in nature, these generally fail to account for inevitable disruption of sacs and masses in preserved samples. To ensure accurate estimates, free and attached eggs must be counted in samples taken with suitably fine-meshed nets. There are numbers of estimates of egg numbers in the literature for which no such information on methods is given. Reproductive rates have been estimated for one population in which egg production was highly discontinuous. From counts of adults, nauplii and free and attached eggs in successive fine-mesh samples from Ogac Lake, Baffin Island, McLaren (1969) estimated that clutch size was 19.6 f 5.0 (mean and 95% confidence limits from his Table 2). The wide confidence limits for this estimate from 14 " broods '' (eggs or young from roughly synchronous clutches ; see p. 135) are not surprising considering the many possible sources of error, but the mean

97

THE BIOLOGY OF PSEUDOCALANUS

estimate is gratifyingly close to the expected clutch size of about 18 eggs for females of about 0.85 mm (mean for the season from McLaren's Fig. 6) as suggested in Fig. 19A. McLaren (1969) also estimated that the time between successive broods was approximately a week, which is about as expected at the temperature of 2-7°C that prevailed in the

2

4

6

8

10

12

14

Temperature ( T I in "C

FIG.22. Embryonic durations of Pseudocalanus from three localities. BBlehrQdelr's temperature function for Halifax animals from McLaren et a2. (1969). Function with b = -2.06 fitted to data in McLaren (1966) for the large form from Ogac Lake and to data for North Sea animals from Thompson (1976).

upper waters of Ogac Lake at the time of his studies (i.e. 1.25 x 4-5to 8 days, the times taken for embryonic development a t 7 O C and 2°C; Fig. 22, Table XVIII). After two or three successive broods in early summer in Ogac Lake, the food supply diminished rapidly, and the consequence was evidently not simply reduced reproductive rate, but A.Y.B.-15

6

98

CHRISTOPHER J. CORKETT AND IAN A. MULAREN

virtual cessation of reproduction, and finally death of the overwintered females (see p. 138). I n continuously breeding populations, it should be possible to use egg counts (free and attached) and temperatures t o estimate reproductive rates, in the manner suggested by Edmondson et 02. (1962). This has not evidently been done for Pseudocalanue. However, Marshall (1949) lists total numbers of free and attached eggs in fine-meshed samples taken weekly from Loch Striven, Scotland, along with mean numbers of eggs on those females that were carrying full, unbroken egg sacs. This allows us to estimate the proportion of females that were carrying egg sacs at the time of sampling. This has been done for the purpose of understanding life cycles (see later, p. 143). On average about 40% of females in the population bore egg sacs during the season of sustained reproduction, in March through August (Fig. 29, p. 142). From laboratory evidence (previous section), we have estimated that about 65% of females should be in full reproductive condition and that a further 25% of these should be between clutches, so that we expect that some 50% of females should be bearing egg sacs in any given sample. The slightly lower estimate from animals from Loch Striven suggests that the pre-reproductive period or the period between clutches has been underestimated in laboratory experiments, or possibly that predation takes a toll of eggs from prematurely broken sacs. Preliminary observations by Corkett (1 969) suggest that egg numbers may at times indicate near maximal reproductive rates for a population of females in which there are no pre-reproductive or postreproductive individuals (i.e. 1.25 multiplied by the observed number of eggs per female in Table XVII, close to the predicted numbers on April TABLEXVII. PREDICTED AND OBSERVED NUMBERS OF Ecas PER FEMALE IN BEDFORD BASIN,NEAR HALIFAX, NOVA SCOTIA,DURING 1969. (After Corkett, 1969.)

Dale

25 March 3 April 9 April 15 April a

Number of eggs per female Predicteda 0bse~ved 15.2 18.5 19.5 24.5

6.0 13.9 14.6 9.5

Based on mean size of 99, from Fig. 19A. Determined as by Corkett and McLaren (1969).

Relative size of oil sacb (mean f 95% c.1.) 0.01 f 0.003 0.02 & 0.006. 0.03 f 0.005 0.04 f 0.008

99

THE BIOLOGY OF PSEUDOCALANUS

3 and 9). However, on March 25 and April 15, observed numbers were less than expected even for a population composed in part of nonreproductive females (i.e. 25% non-reproductive on average, as estimated in previous section). On March 25 some females had no visible stored oil, and 88% of oil sacs were less than 0.02 in relative volume (Table XVII). However, on April 15 oil sacs were relatively large and 17% of females had oil sacs less than 0.02 in relative volume. This suggests that the presence of stored oil in these females reflected a level of food in excess of that required to support maximal reproductive rates. TABLEXVIII. PARAMETERS OF B~LEHRADEK’S TEMPERATURE FUNCTION, WITH b = -2.05, DESCRIBINQ EMBRYONIC DURATIONS OF Pseudocalanw. (Data from McLaren, 1966; McLaren et al. 1969; and, for the North Sea, from Thompson, 1976.)

Locality Halifax, N.S. Woods Hole, Mass. Frobisher, N.W.T. Ogac Lake, N.W.T. (small form) Ogac Lake, N.W.T. (large) Millport, Scotland North Sea

Mean egg diameter pm f.95% c.l.

Temp. at time of sampling

Parameters a

a

121.6 & 1.8 127-4 & 3.7 130.4 &- 3.3

0.1 -0.7 - 1.7

2 144 2 312 2 280

- 13.40 - 13.87 -13.84

108.5 & 2.3

4.6

2 105

- 13.00

155.3 & 2.6 123.6 f 1-8 -

4-6 8.9 1o a

3 467 2 290 1 845

-13.17 -13.63 -11.45

Approximate annual mean temperature of southern North Sea; not given in Thompson (1976).

F . Retrospects and prospects A number of mysteries remain concerning reproduction by Pseudocalanus. The nature of the mating act and the reasons for low mating success in the laboratory are unknown. The reasons for unbalanced sex ratios in some laboratory rearings (Thompson, 1976) are obscure. Perhaps qualitatively or quantitatively inadequate food promotes a tendency for a greater proportion of females to be produced. This strategy could be adaptive for Pseudocalanus, since females live longer (p. 114) and, even ifthey mature at a time when foodis inadequate for their young, they need not ‘‘ waste ” all their reproductive efforts then.

100

CFLRISTOPHER J. CORKETT AND IAN A. MULAREN

Because of the evidently simple rules governing clutch size and clutch frequency in Pseudocalanus, control of its potential reproductive rate is well understood. Evidently these potential rates may at times apply in nature. We need more studies from nature to investigate this supposition using frequent sampling with fine-meshed nets, like that of Marshall (1949). Possible indicators of maximal reproductive rate such as presence of adult males a t a certain frequency or sizes of the oil store in adult females, should be explored. Pseudocalanus has been noted as having a generally lower reproductive rate than that found in some other common copepod genera such as Acartia, Centropages and Temora (Corkett and Zillioux, 1975; Dagg, 1977). After demonstrating that Pseudocalanus females (unlike those of Acartia tonsa and Centropages typicus) did not show reduced reproductive rate when fed intermittantly (12h/day), Dagg (1977) concluded that Pseudocalanus is adapted to patchy and irregular food availability. Since we have already shown (p. 73) that Pseudocalanus has a marked die1 feeding rhythm, Dagg’s laboratory feeding schedule may not have been altogether appropriate as a model of such patchiness. However, clearly the restrained reproductive rate of Pseudocalanus does require more exploration in the comparative manner begun by Dagg.

X. DEVELOPMENT AND GROWTH Readers will find this section of our review perhaps the most difficult and condensed, but we believe that it is through the observations and analyses outlined here that the role of Pseudocalanus as an important producer in the food web of northern seas may be best understood. It is possible to describe rates of development in terms of the moult into successively different morphological stages (Egg, N I-C VI). Growth is a conceptually separate phenomenon from development, and indeed some authors have expressed growth rates of copepods directly as a continuous increase in weight (wet or dry, or of some element). However, if the weight of particular stages can be determined, then growth rates can be estimated by using the times taken to reach these stages. We therefore pay considerable attention in this review to the determinants of development rates and body sizes as separate processes. We will show that temperature is of profound significance in determining development rates and sizes, and therefore growth rates, of Pseudocalanus. I n considering these matters, we make some use of BBlehrBdek’s (1935) temperature function. Empirical justification for the use of this function has been given by McLaren (1963). We use it in

THE BIOLOGY OF PSEUDOCALAN US

101

this review partly because it has already been used in publications about Pseudocaianus, and partly because of the need of some analytical function for interpolation and prediction. Since the use of BBlehrAdek’s function has recently been subjected to some criticism (Bottrell, 1975), we describe its properties briefly here. BBlehrBdek’s equation is one of several three-parameter equations that can adequately describe responses of physiological rates to temperature. The development time (D) in days of a development stage or stages (eggs, N I-C VI), of Pseudocalanus is represented as a function of temperature (T) in “C by: D = a(T-u)b, where a , u and b are fitted parameters. On a linear plot the constant u (often called the biological zero since it is the theoretical temperature a t which development time is infinite) describes the position or origin on the temperature scale. D = a when T = 1 u and therefore the constant a is the theoretical development time one degree above u ; log a is also the intercept on the y-axis of a plot of the log of development time against the log (T-u), the parameter b is the slope of this line. The parameter b in this equation can be assigned a constant value. The assignment of a constant value to b is arbitrary; however, as pointed out by McLaren et ai. (1969) when b is made a constant then a for embryonic duration among closely related animals is related to egg size. Furthermore, they show that a is strongly correlated with habitat temperature among more distantly related copepods.

+

A. Embryonic development rate 1. Zffects of temperature Figure 22 shows that BBlehrBdek’s temperature functions with

b = -2.05 (from McLaren et al., 1969) describes quite well the differences between three geographically separate populations of Pseudocalanus. The wide 95% confidence limits for means of North Sea animals in Fig. 22 generally exceed the ranges in development times for individual clutches given by McLaren (1966) and McLaren et al. (1969). These wide intervals and the anomalous duration at 8.6”C probably result from less frequent experimental observations (Dr B. Thompson, personal communication). However, this also illustrates the usefulness of having a general temperature function, like BBlehrQdek’s, for interpolation with such data. The assumed value of b = -2-05 (see above) has been used to fit the parameters of BBlehrAdek’s function to development times of different populations of Peeudocalanzcs (Table XVIII). Clearly, the great

102

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

difference of the large form from Ogac Lake (Table XVIII, Fig. 22) from all other populations is its proportionately slower development at all temperatures. If curvature ( b ) is held constant, the position on the Celsius scale (x)of its temperature response does not differ palpably from that of the small form, but the value of a is substantially greater (Table XVIII). The regression of a on egg size for the populations of Pseudocalanus given in Table XVIII, even when the large form from Ogac Lake is excluded, is significant at p = 0.05. McLaren et al. (1969) showed that with b = -2-05 the parameter u for 11 species of copepods from the arctic to the tropics was related to average environmental temperature and suggested that u may be used in this manner to indicate temperature adaptation. The regression of u on the temperature at time of sampling for populations of Pseudocalanus given in Table XVIII is not significant (p 0.09). Other estimates of development time in the literature (Marshall, 1963; Katona and Moodie, 1969; Sazhina, 1971, 1974) are not sdEciently accurate (usually given to the nearest whole day) to express possible regional differences. The earlier work of McLaren (1965) gives estimates of the relative times taken to hatch at different temperatures of embryos of Pseudocdanus that had undergone partial development, and has been superseded by the subsequent work cited above.

-

2. Effects of salinity

McLaren et al. (1968) found that the effect of salinity on development times of Pseudocalanus from Halifax, Nova Scotia, was slight. Only in one of several experiments was development significantly slower at abnormally high salinities (about 9% slower at 39x0 than at 29-35%,). Survival of embryos, however, may be affected by salinities (see p. 24). 3. Xeasonal and short-term temperature acclimation

Recent work (Hart and McLaren, 1978) shows that embryonic duration of Pseudocalanus varies to some extent with the temperature experienced by females. The seasonal differences in development time are given in Table XIX. If one considers the females collected in April t o be cold acclimated and those collected in October to be warm acclimated then one would expect that development in the laboratory at 10°C and 6°C would be relatively shorter for the April animals than the October ones. This does not occur and Hart and McLaren (1978) conclude that the seasonal differences in development times can not be attributed to temperature acclimation, but is possibly explained by the larger size of females and their eggs in the colder season (see p. 105).

103

THE BIOLOUY OF PSEUDOUALANUS

Hart and McLaren (1978) also looked a t short-term acclimation by keeping females a t lO"C, 6°C and 2°C during the period of oogenesis. They found that eggs hatched slightly but significantly later at 10°C when females had been kept at 2°C or 6°C. Thus short-term differences in development appear to be attributable to temperature acclimation unlike the seasonal differences noted on Table XIX. TABLEXIX. SEASONAL VARIATIONS IN BODYSIZEAND EWRYONIC DURATION OF Pseudocalanw FROB HALIFAX, NOVASCOTIA. (From Hart and McLttren, 1978) Month of collection of females April October

A~~Tox. Mean cephalothorax Temp. at time of length i n females (mm) collection 2 13

1.12 0.86

Mean development times i n hours (f 95% c.1.)

10°C 82.0 f 2.3 73.4 f 1.3

6°C 125.0 f 1-7 117.0 f 1-6

4. Effects of body and egg size The constant a in Brjlehrtidek's equation is the theoretical development time one degree above a (p. 101). It must be stressed that under no circumstances do we visualize actual development of Pseudocalanus taking place either at a (biological zero) or one degree above u. However, it is of interest that, when b is given the constant value of -2.05, then a is positively correlated with egg diameter amongst geographically different populations of Pseudocalanus (p. 102, Table XVIII). Thus, as suggested by McLaren (1965) and McLaren et al. (1969), by accounting for local differences in temperature through fitted values of a, the fitted values of a reflect the effects of egg size. The effect of size is of course most clearly shown by the embryonic duration of the large form from Ogac Lake (Fig. 22). If Bi5lehr&dek's equation is used with the values for the parameters given in Table XVIII for the large and small forms of Pseudocalanus from Ogac Lake, then it can be shown that the ratio of the embryonic duration of the large to small form is about 1.6:1 ab any temperature between 0 and 10°C. This ratio is more similar to that of egg diameters than to that of egg volumes (-3:l). McLaren (1 966) points out that proportionality of development time and egg diameters might be expected if control were through surface : volume restrictions. Corkett and McLaren (1969) give a positive correlation between egg diameter and cephalothorax length for the Halifax, N.S., population

104

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

IA

**

I

80

I

* A

. +

I

.

**

4 March April

70

Odune

* *

July

*oci. 7

Aua.

0.15

0.14

E E

v

0.13

. v

m

4 4

Fro. 23. Relations botween embryonic duration at 10°C (A) and egg diameter (R) with size of adult females. (After Hart and McLaren, 1978.)

THE BIOLOGY OF PSEUDOCALANUS

105

and recently Hart and McLaren (1978) have investigated this relationship in detail for samples taken from March to October (Fig. 23). The;y showed that within the local Halifax population embryonic duration at 10°C (Fig. 23A) and egg diameter (Fig. 23B) are strongly correlated with cephalothorax length. It might be expected that egg size by itself would be responsible for differences in development times through, for example, the surface :volume ratio decreasing with increasing diameter (McLaren, 19GG). Hart and McLaren (1978) present evidence that shows that factors other

FIG.24. Hatching of Pseudocalanus. Note the bulging of the inner membrane, followed by the crumpling of the outer membrane, and finally the rupture of the inner mombrane, which is left protruding from the outer membrane. (From Marshall and Om, 1964.)

than egg size alone determine development time. They showed (using partial regression analysis) that female size had a significant effect on embryonic duration independent of egg size and that embryonic duration was significantly correlated with size of male parent as well. In addition they found that individual egg size within a clutch had no effect on the development time of that individual egg. I n the previous section we discussed seasonal and short-term acclimation and we now see how such acclimation may be affected by size. Since size of female P ~ e u ~ ~ ~changes u l u seasonally ~ ~ ~ ~ near Halifax (Table XIX) and elsewhere (p. l l G ) , seasonal differences in embryonic development rate can be partly attributed to such changes.

'"1

A

t

"1 501

D=u(T+ 13.40)-2'05

Ad.?, u=19350\

t\

m

\

\:

c III, u- I1890

\

crn.u=12850*

\

\

.

= 9190

"r 2ot

x

IO

O

'*\

2

4

06

8

10

12 l Temperature ("C)

FIQ.26. Times between hatching and the beginning of various stages of Paeudocalanw from (A) new Halifax, Nova Scotia (each point an individual, after McLaren, 1974) and from (B) the North Sea (open circles are means, closed circles individuals, after Thompson, 1978).

BBlehr6dek'e funQtiOIIf3 aa for embryonic duration (see Fig. 22). exoept for differences in a for each stage.

THE BIOLOGY OF PSEUDOCALANUS

107

In conclusion Hart and McLaren (1978) have shown that short-term temperature acclimation affects embryonic duration in Pseudocalanus slightly, and in the expected compensatory way, but that seasonal differences can be attributed to environmentally and perhaps genetically (p. 124) determined differences in sizes of parents and eggs.

B. Hatching Hatching in Pseudocalanus has been described by Marshall and Orr (1954). The embryo is surrounded by two membranes and during hatching the outer one splits, the inner one bulges out, increasing in volume t o about twice that of the embryo, and eventually the nauplius struggles and breaks out through the inner membrane (Fig. 24). The discarded inner membrane may remain attached to the outer (Marshall and Orr, 1954) or the inner membrane may become completely detached from the outer during hatching (Corkett, 1968, who gives a photograph of a newly hatched nauplius, a hatching one, and two embryos).

C. Development rate of nauplii and copepodids The phyaiological and hormone control of development rate (i.e. moulting rate) of some Crustacea is reasonably well understood. However, the subject has barely been explored in Copepoda (Carlisle and Pitman, 1961) and evidently not at all in Pseudocalanus. Of particular interest would be information on physiological or hormonal causes of suspended development during the winter at middle and a t higher latitudes (see p. 157). However, for many purposes it is of greater interest to enquire into the two obvious controls of development rates in nature-temperature and food supply-which are the substance of the following sections. 1. Eflect of temperature

Although Pseudocalanus has been reared in a number of laboratories, the effect of temperature on times taken to reach various postembryonic stages of development has been examined only in populations from Nova Scotia (Corkett and McLaren, 1970 ;McLaren, 1974) and the southern North Sea (Thompson, 1976). These researchers used excess amounts of food and argue that the development rates of the copepods were maximal at the various temperatures. Their results are summarized in Fig. 25. Corkett and McLaren (1970) supposed that time taken to reach any given stage might be the s-me multiple of embryonic

108

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

duration a t any given temperature. Assuming that B6lehrBdek's function applies with a = 13.4 and b = -2.05 (i.e. the same as for embryonic duration in Fig. 22), this is the same as assuming that the temperature responses of times to develop to older stages can be adequately described by differences in a alone. This can be amplified as follows. If for stage I,

D,

= a, (T

- a)b

and for stage 11,

:.

D,, = a,,(T - a)b A t any temperature T

DIP,, = %/a11 if b and cc are common to all stages. Figure 25A strongly supports this view for Pseudocalanus. Thompson (1976) estimated average times (by planimetric integration of numbers versus time) taken to reach various stages from samples taken at regular intervals from large populations. She did not separate males and females in these estimates. I n spite of methodological differences, her results (Fig. 25B) are very similar to those for Pseudocalanus from Nova Scotia. Here, too, the times taken to reach various stages are well described by the equation for embryonic duration (see Fig. 22), with changes only in the proportionality constant, a. However, there is an awkward " step " in the relationship for older stages at about 8°C. This suggests that the animals at colder temperatures were somehow thwarted from more rapid development. Possibly there was some abnormality of response in the laboratory cultures a t these rather low temperatures for this North Sea population. Additional evidence for this assertion comes from the fact that Thompson (1976) was unable to obtain successful hatching of eggs a t temperatures below 3.7"C, or successful rearing of stages above N V at 3.7"C. Table XX gives estimates of the relative amounts of time required to reach successive stages of development, expressed as multiples of the time for embryonic duration; that is, the a of Bglehrhdek's function for each stage, calculated as in Fig. 25, is divided by the a for embryonic duration from Fig. 22. Because of the possibly abnormal retardation of development below 8°C among North Sea animals (see above), we have calculated for use in Table XX the a values for successive stages only for data from higher temperatures in Thompson (1976). This gives estimates of a for the older stages that are 3-5-4.8% smaller than those in Pig. 25B.

THE BIOLOQY OF PSEUDOCALANUS

109

OF THE RELATIVE TIMES TAKEN TABLExx. ESTIMATES BY Pseudocalanus TO REACH VARIOUS STAGES. (Data from Fig. 25.)

Beginning of stage N I1 N 111 N IV NV N VI CI

c I1 c I11 c IV cv c VI

a

Nova Scotia

North Seaa 0.18 0.55

4.29

-

5-99

-

9.03

2.01 2.82 3.51

3.91 5-14 6-14 7.12 8.14 9.46

Multiples of embryonic duration only for data >S"C (see text).

The estimates for Pseudocalanus from Nova Scotia and the North Sea, of the relative times taken to reach various stages are very similar (Table XX). Among all nauplii, N I11 (the first feeding stage, see p. 112) has the longest duration (1.5 embryonic durations). All copepodid stages have roughly the same relative durations. It is probably sufficiently accurate for some purposes to assume that each copepodid stage has about the same duration as embryonic duration a t any temperature ; from Table XX the five stages average 0.95 embryonic durations (i.e. (9.03-4+29)/5)in the Nova Scotia animals, and 1-11 (i.e. (9.46-3-91)/5) in those from the North Sea. We believe that the estimates in Table XX, or perhaps the approximations noted above, can be used to predict the duration of developmental stages of Pseudocalanus in conditions of excess food, provided embryonic durations are known for at least two temperatures (the minimum required to fit BGlehrBdek's function with b assumed to be constant at -2.05). Since embryonic duration, ignoring the large form of PseudocaEanus, differs little among regions (Table XVIII), especially at higher temperatures (Fig. 22), we propose that estimates for older stages from different geographical localities should not differ much from those shown in Fig. 25. It is possible to examine this proposition using scattered estimates for other localities. I n using published estimates, the possibility has to be considered that food shortages, disease and other such factors can prolong development times beyond those possible a t the given temperatures. Indications of

110

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

such retardation might be the occurrence of a wide range in development times, or of some " stragglers " that moult long after the others. The first recorded laboratory rearings were those off Plymouth by Crawshay (1915) who found that adulthood was reached 35-40 days after hatching at 12.3"C. Although the food supply used by him cannot be assessed, it was probably inadequate for maximal development. Katona and Moodie (1969) succeeded in rearing large numbers of Pseudocalanus over perhaps four generations in large-volume containers a t 15°C. They obtained information on developmental durations only from eight smaller vessels of 250 ml volume, in which the progeny of individual females were reared. These gave estimates considerably longer and more spread out than those in Fig. 25B: 16-25 days for the time from hatching to C I and 30-38 days to adulthood. Numbers and sexes are unspecified. Although Katona and Moodie give no information on feeding procedures in the small vessels, they indicate that the rearing medium in the large-volume containers was renewed only every two weeks. Algal food (mostly Platymonas sp. and Isochrysis galbana) was added to these containers to keep cell concentrations at 100000/ml, but altogether the conditions of rearing do not seem to have been as optimal as those used by Corkett and McLaren (1969) and Thompson (1 976). Corkett (1970) reported on the successful rearing of three individuals from Halifax, N.S. beyond C I from among those reared to this stage by Corkett and McLaren (1969). Although the fastest individual developed at precisely the rate predicted from Fig. 25A (26 days to adulthood), the other two lagged considerably, so that mean time to maturity was 35 days. The food medium was added to but not changed every few days as by McLaren (1974). We assume that the retarded individuals were not developing at a physiologically maximal rate. The large number of experiments conducted on Pseudocalanus off the island of Sylt in the North Sea by Paffenhbfer and Harris (1976) were designed to test the effects of food concentration on growth rates (see p. 130). They used large-volume, rotating containers, and changed food frequently. Within the range of food concentrations used by them, there was little or no evidence of retardation. I n eight experiments, the times between hatching and C I were 10.5-12.5 (mean 11.4) days, and between hatching and 50% adult were 24-32 (mean 25.3) days. If we assume that embryonic duration for animals off the island of Sylt is the same as for the North Sea animals studied by Thompson (see Fig. 22), then the times estimated by Paffenhbfer and Harris are precisely as predicted from Table XX : 10-7-11.8 days for hatching to

THE

nIoLom OF

PSEUDOCALANUS

111

C I and 24-8-26.0 days for hatching to adulthood (the range representing both Nova Scotia and North Sea ''multipliers" respectively). Sazhina (1968) reared individuals of the Black Sea population of Pseudocalanus to maturity in the laboratory at 8-10°C. Details of experimental procedures and numbers of animals involved are not given, and estimates are presented only to the nearest day. However, results conform quite well to those summarized in Fig. 25. Durations for nauplii in days were: N I (l), N I I (2), N I I I (3), N I V ( 3 4 9 , N V (3-4), N VI (2-3) for a total naupliar period of 14-18 days. The lower value is probably a better indication of potential rates. For copepodids, the times were: C I (2), C I1 (2), C I11 (3), C I V (5) and C V (5). Lengthening of duration in later stages (contrary to Table XX) indicates to us that these stages were not developing a t maximal rate. Nevertheless, the lower estimate of the range 3 P 3 9 days given by Sazhina as time from hatching to maturity agrees with mean estimates for other populations (Fig. 25). Sazhina (1974) gives duration of the naupliar stages (presumably equivalent to appearance of C I ) for animals from the Black Sea as 14-18 days at 8-10°C (a summary from Sazhina, 1968) and 1 6 1 9 days at 11-13°C ; the times at the higher temperatures suggest that development was retarded (see Fig. 25). For " Mediterranean " animals (no locality given, but presumably the Adriatic) she gives 12.3 days at 11-13"C, very close to times in Pig. 25. There are no details on numbers of copepod or conditions of rearing, except that green algae were used in the Black Sea experiments and mixed food in the " Mediterranean '' ones. However, we predict with some confidence that rates applying elsewhere (Fig. 25) will also apply to Black Sea and Adriatic populations. Andreeva ( 1 976a) determined the relationship between temperature and the duration of C I from the Sea of Japan as about 12 days a t 3.5"C and 4 days a t 15°C. These times are much longer than predicted on the assumption that stages take about the time required for embryonic duration (about 7 days at 3.5"C and 2.2 days at 15°C in the North Sea ; Fig. 22). However the ratio of times a t 3.5"C and 15°C is appropriate (3-0 in the Sea of Japan, 3.2 in the North Sea). Andreeva's (1976a) plot of individual durations shows considerable scatter (the range averaging about & 40% of the mean durations between 10" and 15OC). The animals may have been experiencing physiological or feeding problems that retarded their development rates. I n an earlier note, Andreeva (1975) gives 2-3 days for embryonic duration, 12-13 days for the naupliar period, and 23-25 days for the copepodid period, but does not state the experimental temperature involved. The first two are reason-

112

CIIBISTOPHER J. CORKETT AND IAN A. MCLAREN

able for temperatures of ca. 10-12"C (see Fig. 22, 25)) but the last shows clear evidence of retardation (see Table XX). 2. Effects of food supply

We have indicated above that inadequate food may retard development in rearing experiments. Differences in quality of food have evidently not been looked into, but therc is much information on effects of amounts of food. Thompson (1976) found that nauplii reach N I11 but do not develop further if starved. From her estimates of stage duration at the eleven temperatures shown in Fig. 25B, it can be estimated that the relative times (i.e. as units of embryonic duration) taken to reach N I1 and N I11 were 0.22 and 0.53-almost precisely as for the fed N I1 and N I11 in Table XX. Her estimates of duration of starved N I11 are invalidated by the death of all of them in this stage. We conclude (with Thompson) that food is unnecessary to sustain maximal development rates during the first two nauplius stages and that N I11 may be the first feeding stage. Corkett and McLaren (1970) found no retardation in individuals reared a t 11 6 ° C in concentrations of Isochrysis galbana replenished weekly at 1.5 x lo5, 3 x lo5, or 6 x lo5 cells/ml. Mean time from hatching to C I was 10.6 days and range of individual times was 9.612.5 days. However, a t 3 x lo4 cells/ml, the results were complex. I n two dishes with three nauplii each, three individuals reached C I in 11.5, 12.6 and 14.5 days. The rest remained in nauplius stages for 25 days, and then were fed abundant food (6 x lo5 cells/ml) and reached C I after about nine days. Nine days is close to the number of days that would be predicted to be taken if these retarded nauplii were stopped a t N I I I (9.6 days between beginning of N I I I and C I at ll*5OC, from Fig. 25B). These results suggest that 3 x lo4 cells/ml of Isochrysis galbana replenished weekly is a critical, threshold level for sustaining development rate. It has been shown (p. 90) that this is also a critical level for maximal production of eggs and will be shown (p. 129) that this is the level at which fat is laid down. This level of I . galbana, a t the time of replenishment, is about 500 pgC/I. calculated from Table V I I and may be effectively about half this level in view of probable filtering inefficiencies of such small cells (see p. 63). Paffenhofer and Harris (1976) examined effects of food supply extensively and rigorously. As noted above (p. 110) there was little evidence of developmental retardation at any food concentrations used by them. At (nominal) food concentrations of 5 6 2 0 0 pgC/l., the ranges of times for individual experiments were 10.5-11.5 days to reach C I

THE BIOLOGY OF PSEUDOCALANUS

113

and 24-26 days to reach adulthood. At a nominal food concentration of 25 pgC/l. (actually about 30 pgC/l.) C I was reached on average in 10.5 days. However, at the same nominal food concentration (actually about 21-27 pgC/l.) subsequent development was retarded slightly, and time t o 50% adult was 29 days. It appears that this level of food is about a t the threshold for sustained development. Since the experiments of Paffenhofer and Harris involved the diatom Tholassiosira rotula in rotating culture vessels, the lower estimate of this critical level compared with the results using the smaller I . galbana (above) is probably more nearly natural. Paffenhofer and Harris’ (1976) work is of great importance in demonstrating that development times are little affected by food concentrations comparable with those occurring in nature. They report that algal concentrations near the island of Sylt ranged from 68-780 pgC/l. (annual mean 192), and refer to comparable values for total particulate C for the northern North Sea. They conclude: (‘This suggests that all developmental stages of the neritic P . elongatus are able t o adapt t o a range of food concentrations, neither mortality nor generation time being markedly affected by low food concentrations” . 3. Genetic variation of development rates Differences in development rate between populations of Pseudocalanus may have a genetic component. McLaren (1965) concluded from field samples that the small form from Ogac Lake developed at about 0.42 stages per day compared with 0.28 stages per day for the large form. This ratio of development rates (1.5 :1) is about the sarne as the ratio of embryonic durations (1.6 :1, p. 103) determined in the laboratory for the large and small populations of Pseudocalanus from Ogac Lake. These differences in development rates are clearly genetical. I n eggs a t the 32-cell stage the DNA content was about seven times greater in the large form than found in the small form (McLaren et al., 1966). Woods (1969) speculates that this larger amount of DNA is itself responsible for the slower development rate and that this slower development rate serves t o restore a life cycle more suitable for the environments of Ogac Lake (p. 139), and we have in fact referred earlier to the large form of Pseudocalanus as a possible ‘(instant species ” (p. 10). Hart and McLaren (1 978) found that embryonic duration was significantly related t o size of males chosen for extreme sizes from a sample taken near Halifax, Nova Scotia. Since the male could only contribute

114

CHRISTOPHER J. CORKETT AND IAN A. MOLAREN

genetically, Hart and McLaren conclude that development rate, like size (see p. 124) was heritable among these animals.

D. Longevity of adults We have already indicated (p. 93) that female Pseudocalanus may potentially produce of the order of ten successive clutches of eggs over a period of time determined by temperature, and that they might live for an equal post-reproductive period. However, in nature it is unlikely that maximal potential longevity is often attained, and post-reproductive life is inconsequential for demography and production since neither growth nor egg production takes place. The brief review here may, however, be of some intrinsic interest. The maximum realized length of life in nature is over two years in the high arctic, where adult females may live for some months at least (p. 137). Adult males, as already noted (p. SO), are scarce in nature because of their shorter lives. I n Crawshay's (1915) pioneer work, one adult female survived for 121 days in the laboratory at 12.3"C. The laboratory work of Urry (1965) and Corkett and Urry (1968) on effects of quantity and quality of food on mean survival of adult females has already been discussed (p. 177). However, as their experimental animals were captured in nature, the maximum, rather than the mean might be a better reflection of their potential longevity. Accordingly, we have estimated from Urry (1964) the age of the longest lived individual in each of 21 experiments at 10 5 1 ° C in which food was considered to be adequate for sustained existence (i.e. Isochrysis at 30 000 cells/ml or more replenished weekly, and food species that gave comparable survivorship, Table XII). The mean of these 21 estimates is 107 days (range 84-144). Because different numbers of copepods were used to initiate these experiments, confidence intervals cannot properly be estimated. The eight wild-caught females considered by Corkett and McLaren (1969) to have fulfilled their reproductive potential of 8 or more clutches (see p. 93) lived on average 92 days (range 75-103) at 6-7". The seven females reared and mated in the laboratory by Thompson (1976) lived on average 79 days (range 31-140) after mating, which occurred soon after maturity (Table XVI). The most that can be said from these scattered results is that adult female Pseudocalanus are capable of living for more than 100 days at quite elevated temperatures. Males, by contrast, were found by Urry (1964) to live for only 15 days on average, compared with 33 days for females when both were given 30 000 cells/ml Isochry& galbana. Mean survival for starved

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THE BIOLOQY OF PSEUDOCALANUS

females in several experiments ranged from 17 to 19 days indicating that males evidently gained no sustenance from I . galbana. Probably because of small numbers, there was no evident effect of temperature on longevity of the seven females reared by Thompson (Table XVI). However, a trend is evident (Table XXI) for starved adult females as reported by Corkett and Urry (1968). The very short survivals a t 19.5-21°C are surely pathological, but those at lower temperatures may reflect normal physiological processes that may also apply to the determination of potential length of life of feeding adult females. TABLEXXI. LONGEVITY OF STARVED FEMALE Pseudocalanus IN TEE LABORATORY. (After Corkett and Urry, 1968.) No. of copepods 91 11

13 7

Temperature “C 5-7 10-17.5 11-19.5 19-5-21

Mean longevity (days) 71 35 24 4

Recently, Dagg (1977) compared survival over a short period of ten starved and ten continuously fed (superabundant Gonyaulax tamarensis) female Pseudocalanus and found no difference (six alive after sixteen days). However, these numbers seem rather small on which to base his conclusion that Pseudocalanus “can withstand rather low periods of total starvation, suggesting that they are capable of metabolically removing themselves from changes in the food environment.”

E . Body size Pseudocalanus shows a great deal of variation in body size. Cephalothorax lengths of individual adult females have been recorded as small as 0.67 mm (Carter, 1965) and as large as 1.8-1.9 mm (Lacroix and Pilteau, 1971, their Fig. 4). We believe that explanations for this variability are now largely complete. Therefore we attempt to review only a fraction of the large literature that refers to seasonal and local variations in size of Pseudocalanus with its many speculations. Rather, we will attempt to demonstrate the propositions that size is affected by temperature, but not directly by food supply, and that there is marked genetic variation in size, both within and between populations. The systematic significance of this variability has already been referred to (p. 11).

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

1. Effects of temperature on size (a) Inferences f r o m nature The earlier literature on size variations of Pseudocalanus in nature is admirably reviewed and analysed by Deevey (1960b). She found high negative correlations ( r < -0.6) between length of adult females and temperature in waters with annual temperature ranges of 14°C or more. Conversely, correlations with temperature were smaller, but high positive correlations ( r > 0.5) were found with various measures of phytoplankton during the month prior to sampling where waters had an annual temperature range of 13°C or less. Where possible, she came to these conclusions using partial correlations. Deevey (1960b) concluded that these correlations imply causality : that the relative effects of temperature and food on seasonal variations in length depends on the extent of the annual temperature range and the quantity of phytoplankton present. McLaren (1963) argued that the association between size and food supply was indirect, operating through the effect of food shortage in retarding development (p. 112). I n all localities from which data were available, Deevey (1960b) found that food supply was negatively correlated with temperature-partly a consequence of spring maxima in phytoplankton. This means that size might be most closely correlated with immediate temperatures when they are low since food would allow rapid development then, and this form of dependence cannot be eliminated by partial correlation. I n order t o stress the sufficiency of temperature as an explanation of seasonal differences in size, we will first discuss some examples from nature in which Deevey (1960b) and others have implicated food as well. Loch Striven, Scotland (Marshall, 1949; Marshall et ab., 1934) has a fairly narrow annual temperature range (6O-l3"C, means for water column, see Fig. 29). I n 1933 the major diatom bloom occurred in late March to early April, with smaller bloom in May, July and late summer. Few diatoms were found in winter. Adult female Pseudocalanus showed small size in winter, an abrupt increase to maximal size in April, and then a gradual decrease during summer (see p. 143, Fig. 29). Size and temperature are strongly and negatively correlated between April and autumn, but the persistence of small animals during the cold, diatom-poor winter reduces the correlation with temperature, and introduces the strong partial correlation between size and food during the previous month, as calculated by Deevey (1960b). However, the original papers make it clear that these small overwintering adults had been developing from about mid-July of the previous year, whereas the large adults in April were born beginning in February (see also

THE BIOLOGY OF PSEUDOCALANUS

117

p. 143). McLaren (1963) used such information from Marshall (1949) and Marshall et al. (1934) to plot mean size of adult females on each collection date against estimated mean temperatures during their lifetimes. The results (Fig. 26) indicate that temperature is a sufficient explanation for size variations. Similar arguments, taking into account small overwintering copepodids and adults from the previous warm autumn, can be used to explain the reduced correlation between size and temperature at time of sampling in other mid-latitude areas with narrow temperature ranges considered by Deevey (1960b); for example, the North Sea (see p. 147), the English Channel (see p. 146) and Norway (see p. 151). I n high latitudes, the protracted life histories of Pseudocalanus may even lead to spurious positive correlations between size and temperature at time of sampling. Thus Ussing (1938) found in East Greenland that reproduction during the phytoplankton maximum in June gave large young copepodids in July which developed (probably in deeper water) to large, overwintering C V by September (see p. 141). Reproduction in summer, when food is scarcer and temperature slightly higher, gave smaller copepodids that did not moult into C V until the following spring. There was little size variation in the subsequent adults. Clearly temperature at the time of capture would be a poor indicator of size under these conditions, with large C V occurring during the warm summer and smaller ones in the cold spring. But it cannot be concluded, as Ussing (1938) and Deevey (1960b) did, that food is directly responsible for the observed size differences. Similar interpretations can be applied to size variations described in other northern localities (see also p. 141) by Digby (1954), Fontaine (1955), Grainger (1959) and Cairns (1967). However, in some of these localities there is bimodalism of size that might also reflect genetically different size forms (see p. 123). Studies of two landlocked populations in northern Canada are free from effects of larger or smaller individuals being recruited from outside the study areas. Carter (1965) found that size of adult females varied little in Tessiarsuk, Labrador. From depth distributions and temperature profiles it seems that this population was subjected to a narrow range of temperatures during the season. A group of small, rapidly disappearing females a t the beginning of the summer may have largely or completely matured in the warmer waters of the previous autumn (see p. 140). McLaren (1969) described seasonal variations in sizes of all copepodids and adults in the three basins of Ogac Lake, Baffin Island. By detailed analysis of phytoplankton, temperatures and depth distributions of the copepods, he concluded that size variation

118

CHRISTOPHER J. CORKETT AND LBN A. MCLAREN

could be attributed to temperature during development. A paradoxical recovery of size of young copepodids during later summer when food was scarce and temperatures rising he suggested might be due to relatively more rapid development of smaller individuals or to their disproportionately high mortality. McLaren's (1969) study stresses the complexity of interpretation that is required to explain size variations in high latitudes, even in a relatively " controlled " situation. All in all, size varies rather narrowly within most of these highlatitude areas, and variations between localities with different thermal

Temperature ("CI

FIG.26. Relationships between female size and environmental temperatures in various localities. Points for Canadian arctic are (see text): 1, Tanquary Fiord; 2, Foxe Basin; 3, Ungava Bay; 4, Tessiarsuk; 6, Ogac Lake. (After McLaren, 1966, with additions for the Canadian arctic from Cairns, 1967, and Carter, 1966.)

regimes can be more revealing (Fig. 26). The points for Foxe Basin (2 in Fig. 26), Ungava Bay (3) and Ogac Lake (5) have been used and explained previously (McLaren, 1965). That for Tessiarsuk (4) is from Carter (1965), and represents the mean size of females on 10 and 20 September (estimated from his Fig. 5), since these developed during summer (his Fig. 4) at depths around and below 10 m (his Fig. 7) where temperatures can be estimated (his Fig. 3). The point (1) for Tanquary Fiord represents mean size in 1963 and 1964 combined from Cairns (1967, his Fig. 5) and assumes that small seasonal temperature variations near the surface are unimportant relative to the almost uniform temperatures of -1"C throughout the season between 20 and 100 m (his Fig. 1). Given these approximations and assumptions, Fig.

THE BIOLOGY OF PSEUDOCALANUS

119

26 indicates that there may be uniform temperature response of populations of Pseudocalanus throughout the Canadian north. As noted above, Deevey (1960b) concluded that temperature is all-important when its seasonal range is wide. McLaren (1963, 1965) chose to depict her size-temperature relationship for Long Island Sound (Fig. 26). Here the copepod populations might be relatively “ enclosed ”. A ten-fold variation in chlorophyll had no statistically significant effect on size, and the water is almost unstratified during the growing season, so that mean temperatures are more reliable. Unfortunately, there appears only limited information on seasonal size variations of the otherwise extensively studied populations of Pseudocalanus in the Black Sea. Kovalev (1967) lists mean total (?) lengths for animals from several depths for the months of February (1.17-1.19 mm), August (1.09-1.12 mm) and September (1.08-1.09 mm), which suggest that seasonal size variations in this population may be rather narrow. A later note (Kovalev, 1968) indicates a range of 1-00-1.23 mm in the Black Sea and 0-8P1.04 in the Adriatic. The relationships in Fig. 26 together seem to be overwhelming evidence from nature that temperature is the primary environmental determinant of body size of Pseudocalanus. They also reveal clear regional variations in response. McLaren ( 1965) fitted BBlehrAdek’s temperature functions to the data (with fewer points for northern Canada) in Fig. 26. This is not necessary to describe the obvious adaptation of each temperature-response curve to the regional thermal r6gime. Adaptation is hardly surprising ; otherwise monstrously large copepods might occur in cold northern waters, and exceedingly small ones in warmer regions. It is also interesting that the same range in mean body size seems to be maintained in all three geographical regions in Fig. 26.

(b) Evidence from the laboratory Only recently has the effect of temperature on body size been confumed in experiments. A hint of the effect is first found in the work of Katona and Moodie (1969), who collected females off Plymouth when the surface temperature was 86OC. These produced laboratory cultures that went through about four generations in the laboratory at 15OC. Means of cephalothorax lengths are given as 1.01 f 0-05mm in the females from nature and ranged from 0.84 f 0-02 to 0.89 f 0.004 in successive samples from the laboratory stock (the & values are said to be S.D., but are very small and are presumably S.E.). Without data for other temperatures the reduced size of females in the laboratory cannot be assigned with certainty to temperature.

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CHRISTOPHER J. CORKETT AND IAN A. MULAREN

A series of experiments were carried out with C I11 captured near Halifax, Nova Scotia, and reared to maturity at constant temperatures of about 5, 8 and 12°C and varying temperatures of 8-12", with a mean of 10°C (Lock and McLaren, 1970). The results showed the expected inverse relationship between female size and temperature, and no significant response of male size. The founding populations of C I I I differed in mean size, so that the effect of temperature is best expressed as percentage increments of size between stage I11 and adult (Table XXII). Although the size difference of the females at the temperature extremes proved to be highly significant, confidence intervals cannot be set for the percentage increments, as the animals were not reared individually. McLaren (1974) added to these results by rearing animals to C I11 in the laboratory. Somewhat surprisingly, he found no effect of temperature. Mean cephalothorax lengths (k95% c.1.) were 0.60 f 0.04 mm a t 4.2"C, 0.60 f 0.02 mm a t 7-3"C and 0.59 -& 0.02 mm at 11*7"C. TABLEXXII. MEAN SIZEINCREMENTS OF Pseudocalanus C I11 FROM HALIFAX, NOVASCOTIA, REARED TO ADULTHOOD AT VARIOUS TEMPERATURES. (After Lock and McLaren, 1970.)

yo size increment to ad. $2

Mean size C 111 mm 0.55 0.63 a

5°C

8°C

10°C"

60 59

52 51

45 49

12°C 43

45

12 h at 8" and 12 h at 12" each 24 hour.

Thompson (1976) carried out extensive experiments to determine the effects of temperature on size. She presents her data as tables with mean lengths, S.D. and numbers, and graphs the means without regressions. We have chosen to present her results by fitting regressions to her means (weighted by n ) for each temperature (Fig. 27, insert). This allows us to express the relative effect of temperature as the percentage decrement in length (mean calculated for all temperatures) for each degree Celsius. This in turn allows comparisons between the effects of temperature on each stage (Fig. 27). A striking pattern emerges from Thompson's results. If two groups of copepods were reared a t 10" and 11°C respectively, the latter should average about 1%longer at N V . However, if they are reared further t o

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THE BIOLOGY OF PSEUDOCALANUS

C I, they might differ little in length. This suggests that during the interval between NV and C I, temperature may have a positive effect on size. After C 111, the effect is clearly negative again. However, the confidence intervals for C V and adult animals are very wide. This is I I I I

II

, I I I

II

I I

I

I I I I I

I I I f I I

-5.0 4

6

8

10

12

( I

I

I I I I

I

I I I II

1; I I

14 I0

OC

I

I I I

I I I

N I N I I N I I I N I P N Y N P I CI

CII

CIIl C E C Y Ad.

Fra. 27. Effect of temperature on body size of North Sea Psewlocalanus reared in the laboratory (data from Thompson, 1976). The unbroken lines are based on linear regressions of lengths on all experimental temperatures. The broken lines are based only on linear regressions of lengths on temperatures above 7.3"C, as shown in the insert for C 111-adult (cephalothorax lengths, sexes combined). Overlap of the 95% c.1. with a length decrement of 0% indicates non-significance of the temperature effect.

not a consequence of small numbers of older stages ; the same numbers of individuals of each copepodid stage were measured at each temperature. Rather, it is due to the erratic effects of low temperatures on size of these older stages, as clearly shown on the insert of Fig. 27.

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CSIRISTOPRER J. OORKETT AND L4X A. MULAREN

The older stages at the three lowest temperatures in Thompson's experiments also seemed to develop more slowly than expected (Fig. 25B),and we have suggested (p. 108) that temperatures below 8°C might have caused abnormalities in the laboratory. Therefore we have fitted regressions to the means for higher temperatures, as indicated on the insert of Fig. 27. The regression coefficients,expressed as percentage length decrements per "C, show a more persistent and significant pattern of increase among older stages when only temperatures above 8" are used. Thompson's results, as expressed in Fig. 27, are compatible with those of McLaren (1974) in showing the small effect of temperature on size of C I11 as compared with adults (see above). However, they do not confirm the conclusions by Lock and McLaren (1970) that adult males were little affected by temperature. From tabulated results in Thompson (1976), the percentage decrement in length per "C is calculated as 3.43 & 1.86% for adult females and 2.92 f 1.54% for males (the 95% confidence limits are calculated for regressions on temperatures above 8"C, as for Fig. 27). There is some resemblance between the regression for adults from the southern North Sea and that for females from Lock Striven (cf. Fig. 26, Fig. 27 insert). 2. Effects of food on size

We have already suggested (p. 116) that the association between body size and food supply in nature is indirect : that food shortages may retard development and therefore diminish the correlation between size and current temperatures, but that body size is not ultimately affected by this shortage. Certainly the association between temperature and size in the field leaves little room for a further effect of food. McLaren (1963) showed from data in Deevey (1960b) that a ten-fold variation in chlorophyll (as an index of food) did not correlate with the deviations from the size-temperature curve for Pseudocalanus in Long Island Sound. Food was kept a t very high levels in the experiments of Lock and McLaren (1970), McLaren (1974) and Thompson (1976), ao that any effect of food supply was not evident. We have suggested (p. 110) that animals from Plymouth reared by Katona and Moodie, which were retarded in reaching maturity, may have been short of food. However, their size (female cephalothorax lengths 0.84-0.89 mm) was similar to that predicted for females from the southern North Sea (0.83 mm, from regression for females a t temperature > 8°C.) The extensive experiments of Paffenhofer and Harris (1976) give excellent evidence on the effects of food on size. They give sizes only

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THE BIOLOGY OF PSEUDOOALANUS

as ash-free dry weights, which are not directly comparable with the cephalothorax lengths considered above. It is clear that wide variations in food supply have little influence on ash-free dry weight, except for males a t the lowest concentration of food (Table XXIII). The great importance of these results (as for their results on development times described on p. 113) is the demonstration that food has such a small effect on body size at food concentrations a t the lower range of possible food supplies in nature. TABLEXXIII. WEIGHTS OF Pseudocalanus ADULTSREAREDFROM NORTH SEA POPULATIONS. (After Paffenhofer and Harris, 1976.)

Nominal food concentration (pg (711)

No. experiments Mean pg ash-free dry wt Mean 68 pg ash-free dry wt a

25

50

1

2

16.0 3.1

100

200

4

1

18.3 (14*0-22.5)a 22.1 (19.0-23.7) 9.5 (7.0-12.0)

11.3 (8.4-13.9)

17.5 13.2

Ranges in parentheses.

3. Genetic variation in body size Differences in the size-temperature relationship between populations of Pseudocalanus (e.g. Fig. 26) are more marked than the differences in developmenctemperature relationships (e.g. Fig. 25). The differences in size (p. 9) between the large and small forms of Ogac Lake and Winton Bay are also genetic, and McLaren (1965) suggested that the differences might be due to cell size, rather than cell number. Woods (1969) enlarged on this possibility and argued from the literature that " an increase in the amount of DNA should result in an increase in the cell volume but a decrease in the metabolic and division rates". However, Pseudocalanus also shows a great deal of continuous size variation within samples from any given locality (see Figs 29-33). Some of this variation is doubtless due to the occurrence of animals that have been exposed to different temperatures during maturation. Recent evidence (McLaren, 1976b) indicates that size is also markedly heritable. McLaren reared at 10°C femaIe offspring of families from 9 males each mated to 2-4 females. This enabled him to estimate by analysis of variance the contributions of males and females to the cephalothorax length of adult female offspring (see Falconer, 1960). McLaren found the heritability (h2)of cephalothorax length of adult

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

females at 10°C was 0.93 (significantly different from 0,P OeOl), based on male parents. This is extraordinarily high, and indicates that size (which we have already shown to be little influenced by food supply) under uniform conditions a t 10°C is almost entirely genetically determined. Most interestingly, the female contribution to size of her offspring (which is normally greater than that of the male in most such experiments, because of non-genetic maternal effects) was very small and not significantly different from zero. McLaren (1976b) suggests that this result is related to size-assortative mating. A further development of our understanding of the continuous variation in size of Pseudocalanus near Halifax, Nova Scotia, comes from the recent demonstration (McLaren,1976b) that size of females in the smaller range (ca. 0.8-1.05 mm in Fig. 23) is strongly correlated with cellular DNA contents. This suggests that the continuous variation iii body size is a result of continuous variations in cell size, caused by the continuous variation in nucleus size. N

F. Body composition and weights The composition of the marine plankton has been extensively studied, sometimes with little obvious application to problems of growth, development and production. Much of the literature on copepods is reviewed by Ikeda ( 1 974), but there is little information on Pseudocalanus. Here we will review what is known, and will focus ontjhat which is useful in describing the size and growth of the animals. This approach leads into a consideration of oil storage in the next section. 1. Wet and dry weights Wet weights are probably unsatisfactory as a measure of size of Pseudocalanus. Marshall and Orr (1966) give a range of estimates of from 50% to 80.7% water (mean 72.3%) for 15 lots of C IV, C V, and adult female animals. Nakai (1955) lists a value of 87.7% water for a very large lot of unstaged animals. Ikeda ( 1 970) gives a mean wet weight of 40 pg and a mean dry weight of 12 pg (i.e. 70% water) for one lot of 80 adult females and 8 adult males. Harris and Paffenhofer (1976) used ash-free dry weights as a measure of size in their experiments with Pseudocalanus and state that the average animal was 12.8y0 ash. Ilceda (1970) found that the ash-free weight of the above-mentioned lot of Pseudocalanus was 9.66 pg/animetl (i.e. ash was 20y0 of total dry weight). Nakai (1955) found that ash was only 2.3% of dry weight of his samples, and Laurence (1976) obtained a value (mean S.D.) of 8.50 f.0.11%.

THE BIOLOGY OF PSEUDOCALANUS

125

2. Calori$c content Martens (1975; see also Kraneis and Martens, 1975) has measured

calorific content of Pseudocalunus. He expresses his results in a regression on carbon (dry wt in pg) as: cal = 0.023 0.0067 x carbon. Assuming that the relationship is actually a proportional one, 1 pg C = 0.011 cal. This agrees well with an estimate of 4.6 cal/mg dry wt (at 50% carbon, giving 1 pg C = 0.009 cal) in Greze (1970) and with a mean (& S.D.) of 5-07 5 0.18 cal/mg dry wt (5-54 & 0.20 cal/mg ashfree dry wt) in Laurence (1976).

+

3. Lipid and protein

Substantial work has been done on the lipid and protein fractions of copepods, but only the work of Nakai (1955) seems to have dealt with Pseudocalanus. For a large number of animals he calculated that fat was 17.3% of dry weight and protein 71.5%. This estimate of fat content cannot be taken as average for purposes of calculation. The variability of lipid (in the " oil sac ") is implicit in some of the following sections. Nothing seems to be known of the qualitative aspects of lipids in Pseudocalanus although other copepods have been extensively studied. The composition of body proteins of Pseudocalunus has evidently not been studied. However, Jeffries and Alzara (1970) have assayed free amino acids in this and other copepods, largely in the context of environmental salinities. They detected all the standard amino acids except cystine and ornithine. Glycine was about one-third and proline about one-fifth of the total, and taurine, alanine and arginine were also well represented. The total free amino acid (about 550 pmoles/g dry weight) was about as expected for marine fornis, but the predominance of the two above-mentioned amino acids gives Pseudocalanus the highest index of " biochemical dominance " and the lowest index of " biochemical diversity " of the six species studied by Jeffries and Alzara. They believe that this is related to its euryhalinity and general adaptability (p. 2 5 ) . 4. Carbon, nitrogen, phosphorus, hydrogen and silicon

Some elements have been used as measures of size, biomass and condition " in studies of growth, production and excretion of copepods. Carbon was estimated by a wet-ashing method to be 49% by regression on individuals of dry weight between 3 and 30 pg by McLaren (1969). These were formalin-preserved animals and had lost "

126

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

much of their lipid. Two lots of unpreserved animals from Ogac Lake (where little lipid is found in the copepods) deviated little from the overall regression. Ikeda (1974) gives estimates of 45-1-46.5% C for four samples of individuals (stages not given) taken in May and June 1971, off Japan and weighing 10.7-16.4 pg in dry weight. Another four samples of C V taken in September 1970, weighed 25.5-27-9 pg/ individual and were 63.3-66-7 % C. Ikeda’s samples were fresh-frozen and analysed by gas chromatography. The very high C values in the September sample were presumably attributable to high lipid content in large oil sacs. Indeed Ikeda illustrates an individual stage V copepodid from the September sample showing a very large oil sac. Martens (1975) gives a regression of carbon (C) on dry weight (W) as: C = 2.27 0-187 x W (expressed in the same weight units). However, such a regression might more properly be fitted through the origin. His mean estimate is 36% carbon, with wide variations. To convert Pseudocalanus dry weights to carbon, Paffenhofer and Harris (1976) use conversion factors of 30% for N I-C I, 34% for C I-C 111, and 37% for C 111-adult. However, they later (Harris and Paffenhtifer, 1976) make it clear that these estimates are based on the much larger Calanus helgolandicus from California. We suggest that dry weights of Pseudocalanus when they are not very lipid-rich are about 50% carbon. I n calculations of growth and production, it may be better to consider the highly mobile lipid carbon separately. Ikeda (1974) determined that nitrogen was 11.4-12-4% of dry body weight of Pseudocnlanus in MayJune and only 6.4-7.2% in September. The low C :N ratio in September is presumably due to high fat content (see above). Christiansen (1968) reported higher N contents for NV-VI (dry w t 0.60 pg, 15% N), CI-I11 (2.73 pg, 27% N), C V (9.75 pg, 28% N) and adults (14-42 pg, 20% N). These animals, unlike those analysed by Ikeda, had been feeding in the laboratory. We have shown (p. 45) that feeding animals produce substantially more NH,, and the high N values in Christiansen’s animals may in part represent gut contents. Martens (1975) gives a regression of nitrogen (N) on dry weight (W) as : N = 0.477 0.027 x W (expressed in the same weight units). The average N content was 6.5% of dry weight. Butler et al. (1969) reported that N was 7.8% of dry body weight in “ mixed small copepods ”, which were in fact almost all C IV and C V Pseudocalanus. We suggest that a figure of 7% N for ‘‘ average ” unfed Pseudocalanus might be used. Hargrave (1966) lists phosphorus contents for starved animals as 0.58 pg for copepodids and 0.01 for nauplii; this seems high. Butler

+

+

THE BIOLOQY OF PSEUDOCALANUS

127

et aZ. (1969)give the only estimate known to us of P as a per cent of body weight of Pseudocalanus (see above qualification) : 0-61% of dry body weight. Hydrogen content wa8 estimated by Ikeda (1974),who found that, as might be expected, it varied with carbon. He found C:H ratios of 6.5-6-9:l in samples from MayJune and 6-6-6.9:l in September, evidently unaffected by amount of fat present. Nakai (1955) lists SiO, as constituting 0.04% of dry weight of Pseudocalanus. Although they have been studied in the marine plankton, we can find no information on other elements in Pseudocalanus. 5. Weight-length relationships

A number of authors have listed weights or other measures of body size for various purposes. We believe that cephalothorax length is often the most useful measure of size, not only because it is easier to determine, but because it is not subject to variations due to food contents, oil storage and preservation effects. Given this variability, it may be better for some purposes to calculate weights from a general weight-length regression. Krylov (1968) determined the weight-length relationship of a variety of formalin-preserved copepods (unfortunately total length in mm and wet weight in mg). He found that K in the expression, w t = K (length)3was roughly the same for all copepodid stages of a species. He gives two estimates of this constant for Pseudocalanus : 364 for White Sea animals, and 336 for Black Sea animals. Robertson (1968)gives dry weight to length relationship for seven samples of " Para-Pseudocalanus " in which the calculated exponent (2-13) is substantially less than the above. The size range of his samples (means 0.70-0.91 mm) was probably too small for an accurate determination. McLaren (1969) showed that the exponent in the weight-length relationship was greater than 3.0 (from his original data, 95% c.1. 3.40-3-88)for a much greater size range of formalin-preserved specimens (C 111, C IV, CV, adult 3 and adult $2) from the Canadian arctic : dry wt in pg = 11.9 (cephalothorax length in mm).3*64 The greater-than-cubic exponent is also implicit in observations that large Pseudocalanus are relatively wider (McLaren, 1965). We suggest that the above formula is much the most reliable available. It applies to " lean " animals, and may be a good indicator of weight for various metabolic contexts. However, there is no doubt

128

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

that unpreserved animals may be heavier than predicted by this regression. The mean weights of four lots of preserved animals from Plymouth (Conover, 1959) averaged only 7% higher than predicted by the above equation from their lengths. Weights of 13 unpreserved samples from Loch Striven (Marshall and Orr, 1966) average fully 67% larger than predicted. We suggest that the difference is due to fat contents. 6. Weight of eggs Adults do not grow (although females may increase their store of oil ; see next section), but females do produce eggs. Two lots of preserved eggs from southwest Baffin Island gave almost identical dry weights of 0.30 and 0.31 pg per egg (McLaren, 1969). Clutches from this locality averaged about 70 eggs with a total volume of about 0.65 mm3 (Fig. 19B). This allows us to write a general relationship for all localities from Fig. 19B : Dry weight of clutch in pg

=

6.35 (cephslothorax length in mm).4*1s

G. Oil storage The oil sac in Psezcdocalanus in nature can vary greatly in size from non-existence t o a condition where it almost fills the body (see illustration in Ikeda, 1974). Some laboratory studies have been made on possible sources of variation. Lock (1968) carried out rearing experiments on C I11 animals captured in nature, in an effort to determine the effects of temperature on size (seep. 120). He also measured the oil sacs in animals maturing in one experiment. He approximated the size of oil sacs by making models of them on a scale of 100 :1with modelling clay, and determining volumes by displacement. More oil was stored at low temperatures (Table XXIV), a t which temperatures females, but not males, were also larger in body size (p. 120). Because of great variability among individuals, this effect is only significant (p < 0.05) among males. The size of oil sacs is significantly smaller (p < 0.01 at 5 and 10°C) in eggbearing females, suggesting that other (unfertilized) females had not used their oil store in oogenesis. The size of oil sacs in males and females a t alternating temperatures of 8-12°C (mean 10°C) was larger than for animals raised at 8" and 12", but because of great variability, the effect fell short of significant. Even so, there is a hint in these results that some sort of energy bonus may accrue from alternating temperatures.

129

THE BIOLOGY OF PSEUDOCALANUS TABLE

XXIV.

OIL STORAGE IN Pseudocalanus ADULTSREARED FROM TAKEN IN NATURE.(From Lock, 1068.)

CIII

Mean size of oil sac in p 3 >: l o 0 f 95% c.1. (no. measured) Temperature "C 5 8 1On 12 a

Males 7 . 3 & 1.69 (18) 4.3 f 1.75 (10) 4.8 1.42 (25) 1.6 & 0.91 (16)

Females without eggs

Females with eggs

9.2 f 3.08 ( 7 )

4.6 f 3.90 (8) 1-7 f 1-06 (16) 3.6 f 2.59 (13) 2.0 f 1.33 (12)

-

11.6 f 2.41 ( 7 )

-

8" for 12 h, 1 2 O for 12 h each 24 hour during rearing.

Corkett and McLaren (1969) found that oil sacs of adult females in the laboratory were more-or-less constant in size a t given food levels during the period of egg production. Generally the oil sacs became smaller and sometimes even disappeared altogether before death. Experiments with a small number of females suggested that oil sacs did not increase much in size when food concentrations (Isochrgsis galbana replenished weekly) were increased beyond 1 x lo5 cells/mI, but that they were very small or absent at food concentrations of 3 x lo4 cells/ml or less. As we have shown elsewhere, at these low food concentrations eggs are not produced at the maximal rate (p. go), and development rates may be retarded (p. 112). Paffenh6fer and Harris (1976) noted well-developed oil sacs in all copepodids and adult stages at food concentrations of 50, 100 and 200 pgC/l., and by inference not in those reared at 25 pgC/l. At 25 pgC/l. time to adulthood was slightly longer (p. 113) and adult males weighed less (p. 123), perhaps due to reduced oil content. Observations of occurrence and sizes of oil sacs in Pseudocalanus in nature may thus be of great importance, for we believe that the presence of an oil sac in copepodids (except when they are overwintering and perhaps " resting '' in some sense) indicates that development is proceeding a t a temperature-determined rate. This has to be verified. Similarly, we believe that oil sacs in adult females may mean that they are producing eggs at maximal, temperature-dependent rates (cf. p. 98).

H. Growth rates Growth rates are implicit in the previously described observations on development rate, body size, and length-weight relationships. For example, for animals from Halifax the length of C I11 animals and A.Y.B.-16

7

130

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

the length increment to adult females has been given on p. 120 and in Table XXII. We assume that each copepodid stage takes the same amount of time as does embryonic development a t each given temperature (p. 109, Fig. 22). From these times and the estimated weights (equation, p. 127) of the C I11 and adult animals, the instantaneous coefficient of increase (k)of dry weight on a daily basis (t is time in days between stages) can be calculated from the weight of the earlier stage (W,) and weight of the later stage (W,) from : W, = Woekt. Then, 100(ek-1) gives growth rates as percentages. of body weight per day which is convenient for comparison with the practice (which we have followed) of giving rations, respiration losses, etc., in this form. For Pseudocalanus C I I I to adult female from Halifax, these are then approximately 7, 10, 12, 14, 15 and 17% of body weight per day at 0", 3", 6", So, 10" and 12.5". The same procedure can be used for Pseudocalanus (sexes combined) from the southern North Sea, using data from Thompson (1976). The regressions for length (L) on temperature (T) between 8 and 14°C are from Fig. 27 (insert): L = 0.4270-00264(T-11-74) for C I ; L = 0-577-0*00336(T-ll.74) for C 111; L = 0.900-0-3452 (T-11-74) for adults. Using the same procedure as for the Halifax animals, growth rates for C I to C I11 are estimated at 16, 19, 25 and 32% ,at So, lo", 12.5" and 15OC respectively. For the period between C I11 and adulthood (both sexes combined), the estimates are 16, 18, 18 and 17% respectively at the same temperatures. It can be seen that growth rates increase with temperature among older copepodids in the Halifax population, but not in the North Sea animals. This is because the negative effect of temperature on size is especially marked in the North Sea animals. Clearly, different populations will exhibit quite different responses of growth rate to temperature, depending largely on the size-temperature relationship. Paffenhafer and Harris (1976) estimated instantaneous growth rates a t 12.5OC directly as increase in ash-free dry weights of copepods. They indicate that little fat is included in these estimates. We have already discussed their observations that food had no effect on development rate, except at the lowest concentrations used by them (p. 113), and that low food levels had little effect on weight of adult females, but more on adult males (p. 123).Table XXV summarizes their findings on effects of food on growth rates, converted from instantaneous rates to percentages of body weight per day. The manner in which they evaluated naupliar weights is not evident, as young nauplii may not feed (p. 112) and may lose weight during early stages. The low growth rates between C I and adult a t a food level of 25 pgC/l. are probably related to the smaller size of adult males and slightly retarded develop-

THE BIOLOGY OF PSEUDOCALANUS

131

ment rates a t this level (Paffenhofer and Harris, 1976). The high values for C I-C I11 and low values for C III-adult at food levels around 50 pgC/l. are interpreted by Harris and Paffenhsfer (1976) to mean that older copepodids are less able to secure rations a t this low food level. However, it is possible (since the rates for C I-C I11 are unusually high) that experimental error is involved. At any rate, the extreme rates at 50 pgC/l. tend to cancel out, giving overall rates of 18-23% growth of body weight per day between C I and adult. There is thus excellent agreement between growth rates at 12.5" given by Paffenhofer and Harris (1976) and those estimated above from the work of Thompson (1976) for animals from a nearby part of the North Sea. We therefore feel that laboratory or field estimates of the relationship between size and temperature in any given locality are all that is required (given the relative invariance geographically of development rate in relation to temperature) to calculate potential growth rates in nature when the food supply is adequate.

I. Rate of prodwetion of egg matter Adult males may not feed (p. 114), but adult females certainly do. Yet they cannot grow in the usual sense after the final moult to adulthood. Paffenhofer and Harris (1976) and Harris and Paffenhofer (1976) make a curious attempt to calculate growth rates of adults from " 50% adult to full adult ". This may compound the effects of larger adults maturing later, oogenesis and possibly fat deposition, and the very small estimates of growth rate do not appear to be very useful. The most significant use of food by females is surely in egg production. It is possible to determine the potential rate of production of egg matter in the same terms that we have used for growth of body dry weight. As an example, we use Thompson's'(1976) data, from which cephalothorax length (L) of an adult female (note that the regression in our Fig. 27 is for adults of both sexes) is given as a function of temperature (T), when greater than S"C, by:

L

= 0.939

- 0.0322 (T - 11.57).

Dry weight of adult femalesis calculated from the expression on page 127 and weight of their clutches from formula on page 92. Time between clutches is assumed (see p. 128) to be 1-25 multiplied by the duration of clutches (Fig. 22). This allows us to express amount of egg matter produced per day as a percentage of female weight, which is determined by the given temperature. These values are lo%, 12%, 14% and 16% per day at 8, 10, 12.5 and 15°C respectively. These estimates for the

132

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

southern North Sea are thus not far short of those for growth rate of older stages given in the previous section. Since eggs may be more calorific than most body tissues, egg production might be nearly equivalent to growth of younger stages.

J. The ‘‘ balance equation and growth efliciencies ”

There have been a number of attempts with zooplanktonic species to determine growth rates and the various efficiencies involved by estimating values of the components in the “ balance equation ” of growth (review in Conover, in press). The general equation can be written : G=I-E-U-R where G is growth rate, I ingestion rate, E egestion rate (faecal production), U excretion rate and R respiration, all in the same units. Gross and net growth efficiencies are respectively the proportions of I and of I - E that appear as G, all in the same units for a particular time period. These efficiencies are sometimes calculated by use of information on chemical composition, respiration, excretion and population data. The only such calculations known to us for Pseudocalanus are in Christiansen (1968). He used nitrogen contents and excretion rates to estimate a gross growth efficiency of 74y0and a net growth efficiency of 12.3% for the interval N V to adult for animals in Bras d’Or Lake, Nova Scotia. The values are at best approximate, since some components were calculated by difference from outmoded observations. We have described assimilation efficiencies (I-E),/I, averaging about 65% in Pseudocalanus (p. 76). Respiration losses (R) have been estimated to amount to about 10% of body weight per day (p. 43). As Conover (in press) points out, we seem to know least about the significance of excretion (U) in terms of the balance equation of growth. Dissolved organic N and P might be represented by an equivalent loss in body weight, but even NH, excretion involves some loss that is not expressed in respiration. It has been suggested that about 5% of body N (p. 49) and about 10% of body P (p. 51) might be required each day by Pseudocalanus for maintenance. Perhaps 213 of the P might represent organic loss (p. 49) and a small amount of N (not measured in Pseudocalanus) likewise. A figure of 10% of body weight for U is probably generous. The work of Paffenhofer and Harris (1976) showed that older stages of Psewdocalanw may grow (Table XXV) at about 10% to 17%

133

THE BIOLOGY OF PSEUDOGALANUS

TABLEXXV. EFFECTSOF FOODSUPPLY(Thalassiosira rotula) RATESOF Pseudocalanus FROM THE NORTHSEA. (After Paffenhofer and Harris, 1976.) Growth as N o . of experiments 1 2 4 1

Food

cone.=

ON

GROWTH

yo body wt/day

(pgC/Z.)

Hatching to CI

C I to end c III

End C I I I to adult

25 50

20 15 16-17 20

12 30-46 22-37 19

11 4-7 17-21 17

100 200

Nominal levels; actual levels deviated somewhat.

of body weight per day at ingestion rates (Fig. l2F) of 60-140% per day. Entering these values as G and I along with the above estimates for E, R and U, we can see that the balance equation may in fact be balanced only at the lower ingestion rate of 60% of body weight per day. Harris and Paffenhofer (1976) have fortunately estimated gross growth efficiencies directly from measurements of ash-free dry weights of accumulated rations and of growth in the same units during various intervals between hatching and adulthood (Table XXVI). Paffenhofer and Harris (1976) concluded that daily ingestion rates continued to increase at food levels above those at which growth rates did not increase further. This should lead to reduced growth efficiencies (probably due to reduced assimilation efficiency) at higher food levels. Although Harris and Paffenhofer (1976) found that the regressions of growth efficiencies for various developmental stages on food concentrations were all negative, none was significantly different from zero a t the 5% level. However, since the estimates of daily ration were based on large numbers of measurements, the reduced efficiencies implied in the work of Paffenhofer and Harris (1976) might on analysis prove significant. I n other words, we suggest that the lack of significance between growth efficiencies and rations in Harris and Paffenhofer (1976) results from unnecessary grouping into reduced sample sizes. At any rate, the results in Table XXVI may be taken to imply gross growth efficiencies of the order of 25% at food levels that are comparable to those occurring routinely in coastal waters. I n order to grow at rates of 10-20% per day, or produce egg matter at about the same rates, a copepod must thus consume at least 4 0 4 0 % of its weight per day. Most laboratory studies of feeding rate have not revealed such high values for daily rations (p. 72).

134

CHRISTOPHER J. CORKETT AXD IAN A. MCLAREN

TABLEXXVI. GROSS GROWTHEFFICIENCIES OF Pseudocalanus. (After Harris and Paffenhofer, 1976.)

Nominal food level WCP. Thalassiosira rotula 25 50 100 200

Number

Mean growth eficiencies

of

experiments

N I-C I

C I-C 111

C 111-Adult

1 2 5 2

24.7 25.8 24.4 25.6

39.7 24-7 24.8 25.8

21.5 27.1 20.1 14.5

K. Retrospects and prospects It is we think possible that more is known about the growth and development of Pseudocalanus than of any other copepod. There have been a number of themes in the work that we have reviewed that are only now coming into a coherent focus. First of all, the work on Pseudocalanus stresses the great importance of temperature in the control of growth and development of planktonic animals. I n the zooplankton literature in general, there has often been more stress on food and feeding and a tendency to carry out elaborate experiments a t single temperatures, instead of attempting to determine the temperature functions of growth processes. Secondly, as already noted (p. 133), we believe that most estimates of feeding rates in the laboratory are too low to support observed growth rates, such as those found by PaffenhOfex and Harris (1976). We will later argue (p. 155) that Pseudocalanus can develop and grow a t temperature-dependent rates in nature, indicating that no shortage of food occurs during large portions of the year. Thirdly, we have shown that approaches to questions of growth and development through the " balance equation ", admittedly somewhat desultory in the case of Pseudocalanus, are outclassed by the direct approaches of Paffenhofer and Harris (1976). Respiration, excretion, assimilation and grazing are all of interest in their own right, but are unlikely to give a very accurate appraisal of the growth status of the animal in nature. Finally, we suggest that the conclusion that Psezcdocalanus frequently grows at maximal, physiological rates in nature may be true of other important copepods as well. We believe that use of temperature functions of size and development rate and perhaps simple transforma-

THE BIOLOGY OF PSEUDOCALANUS

135

tions between species based on size (Corkett and McLaren, 1970), or perhaps DNA content (McLaren et al., 1966), will lead to powerful, predictive techniques for the future.

XI. LIFE CYCLESIN NATURE A. General features, terminology and approaches We have learned enough about the biology of Pseudocalanus from laboratory studies to know that even the most obscure or complicated life cycles must have some general features. It will help to review these before considering examples from nature. The nauplius emerges after an embryonic duration that is controlled by temperature (p. 101). The individual may develop after a temperature-dependent time up to N I11without food (p. 112), but any further development cannot occur without food. Above a certain threshold level of food supply, development will proceed at a temperature-dependent rate. If food is sustained, each copepodid stage may take roughly the same amount of time that is required for embryonic duration at the given temperature (p. 109). Mating must occur shortly after the females moult into adulthood (p. 83) and her first eggs may appear after moulting on average about the time taken for embryonic duration (p. 92). The female carries sacs or masses of eggs that constitute a clutch of eggs. If food is sustained, females may in theory produce up to ten or so clutches (p. 93), but this number is probably seldom achieved even in continuously breeding populations, because of natural mortality. A new clutch can appear after the previous one hatches, and after a further lapse of on average about 25% of the time taken for embryonic duration a t the given temperature (p. 93). Each clutch gives rise to a brood of young and all the broods produced by a female belong to the same generation (sometimes wrongly called a brood in the plankton literature). Although the time between hatching and appearance of first eggs has been called generation time in the literature on zooplankton, including Pseudocalanm (e.g. Paffenhsfer and Harris, 1976), this is incorrect. The true length of a generation is technically difficult to calculate, and we will find no use for it in this account. Nevertheless, the appearance of successive generations can often be recognized, as we shall see. We distinguish the productive season as that time of the year when growth, development and reproduction are sustained. Pseudocalanus suspends development during winter at high latitudes and may dis-

136

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

appear, at least from inshore waters, during summer in the southern parts of its range. We shall present evidence that animals may store much oil and " voluntarily" suspend development in summer or autumn. We refer to these animals as being in " resting stages " although we do not imply that they give up all activities. Because of such periods of rest, the productive season tends to start with a population of animals that are largely at the same stage of development, generally late copepodids or adults, so that the initial and even subsequent generations can be more-or-less synchronous. We shall show that at high latitudes where development is slow this synchrony of life cycles allows us to identify, not only generations, but also more-or-less synchronous broods within these generations. Such a nearly simultaneous spawning gives a cohort (a generation or even a single brood) of animals that can be identified in sequential samples. Although attempts have been made in the literature on Pseudocalanus to infer life cycles from counts of various stages in temperate waters, this is much more difficult. Females may spawn a series of broods that nre unlikely to be synchronous, so that it is not possible to follow a single cohort of offspring through successive developmental stages. However, we can take advantage of two facts : (1) even when younger stages develop rapidly and are ephemeral, adult femsles are long-lived (p. 114) ;and (2) the size of adult females is strongly affected by seasonal differencesin temperature (p. 11 6). Thus where generations are more-orless synchronous and successive, there should be periods when female size is stable, followed by periods of rapid change in size, followed by periods of renewed stability at a new size. We shall see that this is precisely what is observed when samples are taken with sufficient frequency. Most attempts to describe life cycles have been based on samples taken in the open sea, where exchange of water and populations with different histories makes interpretation more difficult. Only Fish (1936), working in the Gulf of Maine, has attempted to follow life cycles while tracing presumed movements of populations in a, region. Studies in semi-enclosedbodies of water (Marshall, 1949 ; Carter, 1965 ;McLaren, 1969) have produced the most detailed and accurate information, giving us insights into life cycles elsewhere.

B. Representative life cycles 1. Tanquary Fiord, Ellesmere Island The most extreme environment in which Pseudocalanw has been studied in detail is Tanquary Fiord, at 81'N in the high Canadian arctic.

137

THE BIOLOGY OF PSE UDOCALAN US

Here Cairns (1967) took a series of samples with fine-meshed nets in 1964 and estimated the relative abundance of stages (Fig. 28A). At the beginning of the season, in late May, at least some adult females had already reproduced, but the young were “ stalled ” in NIII (which can be reached without food). After the first week of June, wasting of the snow-cover over 2-5 m of ice allowed light to penetrate the water, and development of these nauplii commenced.

.

...

EGGS-=

.

.

I

-

I

I

~

’

. ,

I

I

I

I

U

Tessiarsuk

,

NO/m2 -0

10

May

20,000

20

I June

July

Aug.

Sept.

FIG.28. Life cyclos of Pscudocnlanws in three localities in northern Canada as shown in the development of stages in samples from successive dates. Cohorts are traced as G,B,, where n is the generation number and m is the number of a brood within this generation. (A, after Cairns, 1967; B, after McLaren, 1969; C, after Carter, 1965.)

At the same time, a group centered on C I11 began to develop. Later in the season, a second mode of nauplii appeared, but did not gain much development before the end of the seaaon. We suggest that these two modes of nauplii represent respectively two groups of more-or-less synchronous clutches produced by the female population, which was clearly dying out by the end of the season. We have accordingly labelled these cohorts as G,B, and G,B, (as first and second broods of generation 2). We agree entirely with Cairns (1967) that the frequency distribution of stages can only be interpreted as reflecting a two-year cycle in this

138

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

population of Pseudocalunus. The oldest animals (Go) represent a generation that was born two summers previously. The mode of copepodids advancing from C I11 to C V during the summer (GI) represents a generation that was born in the previous summer. The older mode of the current generation (G,B,) advanced to C I I by the end of the summer. Each of these generations need only advance one more stage to achieve the level of development found at the beginning of the 1964 season. The younger animals (G,B,) presumably would die out. Although only coarser-net ( # 6) samples were available to Cairns from 1963, the patterns among copepodids were similar to those of 1964: adults and a mode a t stage I11 were present a t the beginning of the season, and modes at C V and C I1 a t the end. Cairns also noted that a few small individuals disappeared from C IV and C V largely before mid-June and appeared as small adults (mean ca. 1 mm) among the large ones (mean ca. 1.2 mm) thereafter. He suggests two possible explanations. The small animals may represent a basically annual form, perhaps genetically distinct (see p. 113). Or, they may represent the persistence of a limited number of individuals from the second seasonal brood of two years previously (i.e. a putative GOB,). Surface waters warmed from about -1°C on 12 May to +l"C on 25 August 1964. Although this does not seem to be a large difference, the size-temperature response in these cold waters is very steep (Fig.' 26), and could readily explain the difference in sizes. 2. Ogac Lake, B a B n Island

The most detailed study of Pseudocalanus in a high-latitude setting was made by McLaren (1969), who was able to follow life cycles of isolated populations in three basins of Ogac Lake, a landlocked fiord on Baffin Island a t 63"N. The cycle in the innermost basin in 1957 is summarized in Fig. 28B. The overwintering Go matured rapidly in mid-June, large numbers of eggs (G,B,) appeared on 18 June, and it can be inferred that G,B, largely died out in summer, and that GIBl formed the basis of the overwintering generation. McLaren (1969) concluded from size-frequency distributions that almost all adult females in early August were Go, and that they were largely replaced by G,Bl in mid-August, so nauplii in late August were members of a new generation, as indicated in Fig. 28B. Perhaps some of G,Bl participated in the overwintering group. Life cycles in the middle and outer basins of Ogac Lake in 1957 were also basically annual (McLaren, 1969). Because it is more productive than the inner basin, G,B, in the middle basin largely matured by

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late August, and GIB, reached older copepodid stages. The outer basin is more productive than the middle or inner basins, but colder, so that development of three broods was sustained, although slow, during summer. I n late August, a tidal incursion greatly reduced the population, and in mid-September the potentially overwintering older copepodids were derived from G,B, in the outer basin. The pattern in the middle basin in 1962 was similar to that of 1957, except that a third brood, G,B, was sustained and reached late naupliar stages by mid-August. McLaren (1969) showed that the frequencies of individuals in designated broods were about as expected if each of the females in the same samples had produced a clutch of a size determined by their mean body size (see p. 96). That is, each brood did indeed represent a more-or-less synchronous production of full-sized clutches by the entire adult female population. The time between broods was about as expected from the prevailing temperatures in the lake (see p. 97). The observations thus indicate that reproductive rate was maximal during the short period in early summer when food was above some threshold, but was negligible thereafter. Evidence from an experiment with fertilized polyethylene columns suspended in the middle basin in 1962 showed that production of G, was dependent on the size and abundance of females of Go, not on the amount of food that was present during the productive season (McLaren, 1969). I n the fertilized columns, GIBl and G,B, completely matured and GIB, reached late copepodid stages by early August, when a massive G,Bl appeared before termination of the experiment. The appearance of second summer generations in all basins of Ogac Lake, and especially in the fertilized column, indicates that the basically annual life cycle in these Pseudocalanus from high latitude need not be intrinsically controlled. That is, there is not some obligatory resting or overwintering stage a t the end of summer. I n the warm waters of Ogac Lake, the tendency for unseasonable and wasteful maturation and reproduction at the end of summer may be more pronounced than it is in colder waters of the seas outside. Woods (1969) suggested that the large forms in Ogac Lake and in the similar Winton Bay represent evolutionary attempts to restore normal arctic size and slower development rates in these unusually warm waters. I n support of this, she showed that two broods of the previous summer were represented in the overwintered generation of the large form in Ogac Lake in 1962, and that G,B, of the large form only reached young copepodid stages by early August, whereas (see above) G,B, of the small form had matured by this time. This means that the earliest broods of the large

140

CHRISTOPHER J. CORRETT AND IAN A, MCLAREN

form (and therefore those produced before overwintered females have suffered further mortality) are the most successful ones. 3. Tessiarsuk, Labrador Carter (1965) studied the life cycle of Pseudocalanus in Tessiarsuk, a landlocked fiord at 56'30" in northern Labrador. Here, as in Ogac

Lake, there were advantages of sampling an isolated marine population. At the beginning of the 1961 sampling season in the outer basin of the lake, the first brood of young had already developed to a mode at C I (Fig. 28C). Adult females (Go)during early summer were distinctly bimodal in size (means of about 0.75 and 0.95 mm in Carter's Fig. 5). The smaller females had presumably matured in the warmer waters late in the previous summer, whereas the larger ones had overwintered as copepodids and matured in the colder waters of spring and early summer. The disappearance of the small females after the beginning of August presumably signals the replacement of Go by newly matured individuals of G,B,, but not before Go had produced a second burst of clutches, G,B,. The young in September are clearly a new generation, G2BI. Samples taken by Carter (1965) in the following spring, on 10 April 1962, showed much reduced populations, with modes at C I11 and N 111. This suggests that most of G,B, had died out, and that the founding generation in 1962 originated largely in G,B, o i the previous summer, as in Ogac Lake. The presence of only a single size group of large (m. 0.95 mm) mature females on 19 May also indicates that all of them had matured from copepodids in the cool waters of spring. Carter suggested that copepodids in the lake in spring 1962 were probably remnants of G,B,, which would imply a basically semiannual cycle. Our analysis implies that G,B,, already " stalled " as nauplii in early September, died out during winter. The life cycle in the somewhat isolated inner basin of Tessiarsuk was very similar to that depicted above, except that GIBz was slightly more difficult to discern in midsummer, possibly due to sampling errors in this larger, deeper basin. The frequency distributions of stages at the end of the 1961 season and a t the beginning of 1962 were almost identical to those found in the outer basin. The life cycle in Anaktalik Bay, outside Tessiarsuk, did not show the same clarity as those in the lake. Both early and late naupliar maxima appeared, but whether these represented successive broods or generations cannot be discerned. A marked scarcity of older copepodids and adults, except at the beginning of 1962, suggested to Carter that most adults may have matured and spawned outside Anaktalik Bay.

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4. Other arctic local~ties No other populations from arctic waters have been studied in the detailed way of those from Tanquary Fiord, Ogac Lake and Tessiarsuk. However, the few other studies in general conform to one another. Grainger (1965) indicates that breeding and development of Pseudocalanzts in the Arctic Ocean were unknown, and we know of no subsequent studies. Ussing (1938) and Jespersen (1939) found that the overwintering stock in fiords of East Greenland matured and began to breed in April and May, with egg-bearing females present until August. Most of G, reached C IV in late summer and overwintered in this stage. A few matured to produce nauplii of G,, probably unsuccessful, in autumn. Digby (1954) found that the cycle in Scoresby Sound, East Greenland, was basically annual small late copepodids giving rise to small adult females in spring, and younger copepodids developing in the cold waters to larger adults in early summer. Some of these large adults live through another winter, thus having an essentially 14-year cycle. A few individuals spawned early in the year may have matured between September and November and have been responsible for nauplii at that time, this G, being probably unsuccessful. Grainger (1959), working in a comparably high-arctic locality in Foxe Basin, northern Canada, found that adult males were abundant only in March through May, and adult females (and their nauplii) were common from April to September. The predominant overwintering stages were C I V and C V. Although Grainger concludes that the cycle was basically annual, he suggests that part of the population may have taken 14 or 2 years to mature. His argument is based in part on the occurrence of a size-bimodalism in C V females, but not clearly in any other stage. The larger size mode occurred between late September and early February, and disappeared at the time of appearance of substantial numbers of adults in the samples. Grainger (1959) interprets size differences (probably not correctly) in terms of food supplies, but we can only suppose that the large C V females represent a stock that had developed in colder water, perhaps elsewhere. Fontaine (1955) studied Pseudocalanus life cycles in the subarctic (sensu Dunbar, 1947) waters of Ungava Bay, northern Quebec. She concluded that the cycle was basically annual, but that a small portion reached maturity and spawned during late summer. 5. Loch Striven, Scotland

The magnificent survey by Marshall (1949) is by far the most thorough and revealing study of Pseudocalanus life cycles from tem-

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

perate latitudes. It is based on weekly vertical hauls from 40 fathoms with a fine-meshed (200 mesh/inch) net. Here, the most useful means of delimiting generations is by size changes in successive samples of

1.4

-I

mn

L I

I.2

1.a

0.e L

0.6

I Jan.

1 March

I

1 May

I

I

July

I

I Sept.

I

F I ~29. . Relative abundance of C IV and egg-bearing adult female Pseudocalanus, and length-frequency distributions of adult females in samples from Loch Striven, Scotland, with temperature data. Frequency scales of histograms varied and some samples left out for clarity. Generations designated as G,. (Data from Marshall, 1949, and Marshall et al., 1934.)

adult females. By reanalysing her data and expressing them in different ways, we have come to conclusions that extend or are a t slight variance with her own. Figure 29 summarizes much of what we have to say. As Marshall herself concludes, it seems clear that the adults (Go)at

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the beginning of January trace their origin from warmer waters in the previous autumn. Some 50% of the overwintered animals at this time were in C V, and the slight size increase of adult females in subsequent weeks is a consequence of maturation of these small C V animals at the cooler temperature obtaining in late winter. By 20 February the proportion of egg-bearing females showed a sharp increase, and this proportion was generally maintained until early August. We can define the productive season in terms of sustained high egg production as late February to early August. The appearance of the new G, spawned and developed in cold waters is dramatic, and shows little overlap with the females of the overwintered generation. Subsequent generations of adult females were spawned in increasingly warmer waters, and this is expressed in diminished size through the season. If successive generations retain any of the synchrony evident in the appearance of G,, this should be expressed as a series of persistent size modes, representing the females of the current generation. Between these periods of persistent size, there should be periods of rapid change of size, as females of the new generation replace those of the previous one. This is precisely what is observed in Marshall’s (1949, her Table X) data. Provided a suitable spacing of samples is chosen (there is not room for all her samples in our Fig. 29), the succession of generations appears very convincing. I n each case, we have included the first and last samples in which a particular dominant size mode is expressed. We infer that during the productive season in Loch Striven, there were six successive generations (Gl-G,) of Pseudocalanw. Although useful in suggesting maximum reproductive rates in nature (p. 98), the information on proportion of egg-bearing females (Fig. 29, top) offers little insight into the possible succession of generations. There is perhaps a hint of reproductive decline among old females of Go in mid-March and G, in late April. However, the proportion of C I V in the samples supports our conclusion that six generations of adults were produced. Marshall (1949, her Fig. l ) shows abundance of each stage as a percentage of the total numbers of animals. However, this method of expression is influenced by the highly variable mortality of younger stages and the persistence of adult females, which do not pass through in a “ wave ” of abundance. We have thought it better to express abundance of C IV as a percentage of total copepodids (Fig. 29, top). If the population were at equilibrium and if no mortality occurred during copepodid stages, then C I V on average should constitute 20% of all copepodid stages. Clearly neither of these situations applies and C IV is generally scarcer at the beginning

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CIXRISTOPHER J. CORKETT AND IAN A. MCLAREN

of the season. However, as the season progresses, and particularly after G,, there is clearly an accumulation of C IV, and we agree entirely with Marshall that these animals have suspended development to form an overwintering stock (Go); at the same time some animals clearly continue to undergo normal development. These animals are evident as a series of peaks, through the season, of C IV. Only in the case of G, is there any ambiguity in these peaks, and all of them fall, as expected, shortly before or around the time of appearance of the new size modes of females. TABLEXXVII. OBSERVEDAND PREDICTED TIMESOF APPEARANCE OF SUCCESSIVE GENERATIONS AND BODYSIZESOF ADULTFEMALE Pseudocalanua IN LOCHSTRIVEN, SCOTLAND.(See text.)

@en. no.

Mean temp “C

Dates

Times between generations (days)

Cephalothorax length (mm)

Observed Predicted 0 bserwed Predicted

G, G2

G, G, G, G,

< 20 Feb-4

April

4 April-15 May 15May-12 June 12 June-3 July 3 July-24 July 24Jdy-14A~g

6-5 8.5 12 12.5 14 15.5

>42

48

42 28 21 21 21

38 28 27 24 22

1-14 1.04 0.99 0.94 0.83 0.78

1.12 1-03 0.89 0.88 0.81 0-75

We now proceed to indicate that the successive generations of animals in Loch Striven were developing at a rate that was temperaturedependent. The first appearances of successive generations are shown in Fig. 29, as are temperatures during the developmental periods of each generation. We assume that successive generations spend their lives nearer the surface than the bottom. This is true of earlier generations and at least of young stages in later generations (Marshall, 1949, her Fig. 15). The times between appearances of generations from the data in Fig. 29 are influenced first by the basically weekly sampling schedule carried out by Marshall during much of the season. Second, the f i s t appearance of a generation of adult females represents the << vanguard ”, not the bulk of such females. In calculating predicted times between generations (Table XXVII), we have used the mean estimate for the period from hatching to adulthood from Thompson’s (1976) experimental work for the southern North Sea. As can be seen (Fig. 25B) “ vanguard ” females in the laboratory may mature sub-

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stantially earlier than average ones, so we ignore the short additional time required for embryonic duration. We have used the regression of size of adult females as a function of temperature from Thompson (1976), ignoring the evidently abnormal size response below 8", and extrapolating for lower and higher temperatures (p. 122). The concordance of observed and predicted generation lengths and sizes is remarkably good (Table XXVII). We take this analysis as a strong indication that part of the population of Pseudocalanus in Loch Striven was undergoing growth and development a t a maximal, foodsatiated rate between late February and late July. The prolonged absence of females of G, into September and the drop in reproductive rates after early August suggest that growth and development were thwarted by food shortages late in the season (although Marshall, 1949, registered abundant diatoms and flagellates then). The new mode of females on 2 October would be " G, " but presumably at least some joined the overwintering stock for the subsequent year, so we designate them as Go. One fascinating aspect of the situation in Loch Striven as revealed by Marshall is the co-occurrence of part of the population that develops a t maximal rates, and part that enters some sort of resting phase. We shall later suggest that this is commonplace among temperate populations of Pseudocalanus. 6. Plymouth, England

Digby (1950) analysed vertical hauls with fine meshed nets from a station five miles off Plymouth, England, in water about 50 m deep (Fig. 30). At the beginning of the season adults of Go were small. Digby interprets the mode of larger adults appearing in late February as harbingers of the new generation, but the fact that they are not as large as subsequent females indicates that they matured from small overwintered copepodids (cf. Loch Striven, above), and we designate them as part of Go. Large adults of GI were taken on 3 April and 28 April. The appearance of G, on 14 May, 41 days after 3 April, seems a little behind schedule (expected ca. 36 days a t ca. 9"C, see Fig. 25B), but large numbers of nauplii on 8 May (adults not measured then) indicate that G, appeared a few days prior to 14 May. The distinct drop in size on 2 June appears to represent G3, on schedule (expected ca. 32 days a t 1Crll"C). Digby considered that the small peak of nauplii in late June represented the beginning of G,, but we would interpret them as nauplii of G,, whose adults may be evident then and more clearly in early July, Digby acknowledged that " abundance of later stages and percentages present a rather confused picture " and

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

that " the generations become more merged ", but he concludes from abundance peaks that three generations were spawned between late June and early October. Abundance peaks (particularly those in which many stages seem to peak simultaneously) are unreliable and size of adult females reveals little, presumably because of relatively small temperature change during August to October (Fig. 30, bottom) and because of advective influences in this open-channel sampling site. Naupliar numbers had dropped to low levels in September, and C I V

FIG.30. Size-frequency distributions of adult female Pseudocalanw and interpretation as generations (G,,),in samples from off Plymouth, England, with temperature data. (After Digby, 1950.)

and C V began to outnumber C 1-111 in late September, comprising fully 85% of copepodids on 9 October and 91% on 23 October. Digby interprets this preponderance of late copepodids as a peak in a developmental sequence of the last generation of the season, and some may have matured, judging from the small drop in size of females in late October. However, since Pseudocalanus almost disappeared thereafter, we suggest that these late copepodids represent an accumulation of overwintering stages, perhaps better represented in deeper waters farther offshore. I n conclusion, we suggest that the productive season at Plymouth

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extended from late February until a t least the end of October, with overwintering stages accumulating during this month. Up to early July the succession of generations was about as predicted from prevailing temperatures. If we assume that subsequent generations were not thwarted by food shortage, and able to develop at a temperaturedependent rate, at a mean temperature between early July and late October of about 16"C, a further five generations could have been spawned, for a total of nine generations during the productive season. 7. Northumberland coast, England

Evans (1977) made a thorough study of life cycles and production of copepods at a station five miles off the Northumberland coast. He concludesthat size of Pseudocalanus is strongly affected by temperature, but argues that statistically significant differences between size distributions are the sine qua non for separating generations. He delimited the following sequence of generations between February 1971 and July 1972 :

. . JFM)(AM)(J)(JAS)(O)(NDJFM)(A)(M)(JJ ... I n each case, the brackets delimit samples of females for the given months whose mean sizes differed (P < 0.05) from those immediately preceding and following. The appearance in 1971 and 1972 of a supposed new generation in April coincides with the timing in Loch Striven (Fig. 29) and off Plymouth (Fig. 30). Examination of Evans' data (his Fig. 10) indicates that the population sampled in April 1971 was indeed G , (along with May sample, largest mean length of season with small 96% c.1.). However, the sample of April 1972 probably included overlapping generations of Go and G , as females were intermediate in mean size between those of March and May, and had the highest 95% c.1. of any sample during the two years (cf. similar situations in Fig. 32, below). The completeness and accuracy of subsequent statistical delimitations of generations is questionable for four reasons. (1) samples were monthly, which may have allowed some generations to be unsampled (cf. G, in Fig. 29): (2) normal variate statistics may be inappropriate for distinguishing some generations with non-normal size-frequency distributions : (3) advective influences off the Northumberland coast (as off Plymouth) could confuse the pattern. This would appear to be the only possible explanation for a sharp and significant drop in size in October 1971 at a time of falling temperature (Evans, 1977 his Fig. 3, 10): (4) as Evans himself notes, extra generations may have occurred

Z

J

3

I

'I I

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at times when sea temperatures were relatively stable. For example, mean temperatures for 1971 were 11.4, 12.3 and 12.4" respectively in July, August and September (Evans, 1977, his Fig. 3). Evans uses a temperature function (from Corkett, 1970) for age of maturity that leads to an overestimate of the time required (see p. 110). He concludes, using the temperature function, that seven generations, rather than the five distinguished by him, could occur at a temperaturedependent rate during the year. We estimate that a mean temperature of around 10°C (from his Fig. 3) could have allowed about seven generations (from Fig. 25B) to develop during the period between early April a
8. West coast of Sweden Adler and Jespersen (1920) tabulated data on mean temperatures and mean lengths of female Pseudocatanus from the Kattegatt and somewhat further offshore in the nearby North Sea. These data amply show the response to temperature (see Deevey, 1960b), but cannot be used to discriminate generations. Eriksson (1973a) has recently depicted histograms of size variation of female Pseudocatanus during 1968-1970 in the Kattegatt off the coast of Sweden. Samples were in general too widely spaced in time for delimitation of generations, and considerable size variation occurred on some dates, possibly due to the wide temperature variations in this strongly stratified and hydrographically complex region (Fig. 31, bottom). However, the data are especially interesting in showing differences in the appearance of the first new generations each year. I n 1969, the sequence was similar to those in Loch Striven (Fig. 29), and off Plymouth (Fig. 30), with large individuals of the overwintering generation appearing in mid-April, along with the first of G,, which became strongly represented by the end of the month. I n 1970 the same general sequence occurred, except that individuals of Go were somewhat larger at the end of 1969 and therefore into 1970. However, in 1968 the females in mid-February were as large as those of GI in subsequent years. Pseudocalanus was also evidently more abundant at this time of year in 1968 than in the following two years (Eriksson, 1973a, his Fig. 4). It seems unlikely that these animals could have matured in the warmer waters of the previous autumn, and we have designated them as G,. No information is given by Eriksson (19738) on conditions during the preceding months that could have encouraged such st winter generation.

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9. Coast of Norway

From variations in size and composition of samples of Pseudocalanus from Msre (62'52") on the coast of Norway, near Bergen, Ruud (1929) concluded that there were three main '' spawnings " (abundance of eggs and nauplii),one in March, a second in May, and a third near the end of July. I n the absence of size-frequency distributions of females, detailed assessment is not possible. I n Oslo Fjord (Wiborg, 1940), Pseudocalanus is said to spawn all the year round. Breeding was most intense in February to May, June to July, and August to September, but it is not possible to infer any sequence of generations from the available data. Overwintering in C IV and C V was common only in the innermost parts of the fiord. I n NordBsvatn, a semi-landlocked fiord near Bergen, a succession of spring-summer generations probably occurred (Wiborg, 1944), but this cannot be inferred in the absence of size-frequency distributions of females. Older copepodids overwintered in deeper water. Wiborg (1954) studied life cycles of Pseudocalanus in greater detail from samples taken in 1949-1950 from three localities off the Norwegian coast, giving information on size-frequency distributions of adult females. Here, as in the North Sea area, interpretation is complicated by the passage of water through the sampling areas. He estimated that water might be transported from the southernmost sampling site (Sognesjeen, 61'04") to the next one up the coast (Ona, at 62"54'N), in about a month. However, populations in the vicinity of Eggum, at 68'08'N off the Lofoten Islands, are presumably quite isolated from the southern ones. I n all three localities, the f i s t main spawning period tabulated by Wiborg occurred between late February and the beginning of April. He concluded that there were three subsequent main spawning periods in each locality in both years, a t various times in MayJune, JulySeptember, and September-November. Wiborg interprets size variations of adult females (his Fig. 66, and Table 51) partly in terms of population origins, but size variations certainly follow the pattern shown by populations farther south. At the more southerly Norwegian stations, distinctly larger individuals of GI appeared on 28 March 1950 a t Sognesjsen, 28 March 1949 and 17 April 1950 a t Ona, and on 23 April 1949 and 28 April 1950 at Eggum. There is a suggestion in these and subsequent samples that generations in the more northerly Eggum were some weeks behind the other two localities, but samples were too infrequent for detailed analysis, and the transport of animals may give a misleading picture.

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We illustrate the sequence of samples a t Eggum because of its interest as a northerly locality with a long productive season (Fig. 32). Samples during 1949 were taken from 190-200 m up to 50 m (except on 5 January when the whole water column from 195 m was sampled). Thus we have chosen to use estimates of mean temperatures for depths 50-200 m (from isopleths, Wiborg, 1954, his Fig. 7) as a measure of the temperatures experienced by the developing animals (Fig. 32, bottom). Although Go, G, and G, seem quite distinct and in keeping with the pattern in, for example, Loch Striven (Pig. 29),the lapse of time between the appearance of G, and G, seems too short (from some time between 8 and 23 April to 21 May, or perhaps 40 days at most) for a, generation

FIO.32. Size-frequencydistributions of adult female Pseudoculunus and interpretations as generations (G,) in samples from Eggum, northern Norway. (After Wiborg, 1954.)

to develop a t a mean temperature of ca. 6"C, which should take more than 50 days (see Fig. 25). Possibly this is a consequence of transport northward of populations that were slightly more advanced farther south. Although G, seems clear in late June, size did not diminish further in August. This could be interpreted to mean that no generation was produced in the interval but it can be noted that temperature changed very little during this period, rising a bit more steeply in autumn when size showed a further decrease. Certainly the sustained presence of nauplii in all three localities, with a distinct increase in autumn, suggests that populations could have been developing at maximal rates during this period. At a mean temperature of about 7"C, two generations of adults could have been produced between late July and late September, for a total of four or five for the season.

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Wiborg (1954) distinguished between P. elongatus, which name he believed applied to the common coastal form. ‘‘ having 3-5 annual spawnings ’’ (more according to us), and P. minutus (syn. P. gracilie, see p. 7) which he believed applied to the offshore, northern form of the Norwegian Sea (see below). He found some of the latter in his samples, distinguished by their shape and larger size, and states that “ it seems likely that P. minutus in Norwegian coast waters has only one spawning a year, in March-April”. Recently Davis (1976) has examined the “overwintering strategies” of Pseudocalanus and other copepods in some Norwegian fiords and sounds in the Troms0 area. He confirmed that between early November and late February C IV and C V were abundant, with a scattering of C 111,but that only a few adult females and no adult males were to be found. 10. Norwegian Sea The study by Ostvedt (1955) in the Norwegian Sea a t 66”N and 02”E depicts a life cycle of Pseudocalanus in sharp contrast to those of neritic waters farther south or west. Here there is a single annual spawning (egg-bearing females late April to late June), and a pronounced seasonal migration from and to the depths (see p. 162). Almost the entire population is below 1 000 m as C V between July and April. These become mature and females are fertilized and rise to the surface layers, where the new generation is spawned. These descend as C V to repeat the annual cycle. Soviet workers (Pavshtiks and Timokhina, 1972) have studied seasonal differences in the timing of the season of production in the Norwegian Sea, being essentially earlier south and west and late north and east. They illustrate (their Pig. 3) seasonal peaks of numerical abundance of Pseudocalanus in June in Atlantic waters, early July in mixed waters, and in late July in the East-Icelandic Current. However, no information is given on the timing of its major spawnings. 11. Baie-des-Chaleurs,Gulf of St. Lawrence Lacroix and Filteau (1971) did not use fine enough nets to capture

younger stages, and probably began sampling too late in the season (May-June) to detect overwintering generations. However, their sizefrequency distributions of their ‘‘ small form ” changed during the season in a way that could reflect turnover of several generations. Early in the season, a group of large females (cephalothorax means 14-1-5 mm) was present, but these largely or entirely disappeared as the season progressed. On the other hand, large C V (means 1.2-

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1.3 mm) occurred throughout the summer in deeper, colder water. Lacroix and Filteau interpret these as a population of another " form ", major (see p. 8), possibly having an annual cycle. Parallels with Wiborg's (1954) distinction between life cycles of presumed P. elongatus and P. minutus (see p. 152) may be noted. 12. Gulf of Maine and Bay of Pundy Fish (1936) made extensive studies of abundance of all stages of

Pseudocalanus (grouped as ova, early and late nauplii, early and late copepodids, and adults) in the waters of the Gulf of Maine and Bay of Fundy. To some degree the account is confused by the use of small numbers of samples over wide areas to depict " stocks ". However, some such scheme is necessary in view of the open circulation of waters in the Gulf of Maine. Fish concluded that the succession of generations was clearest in the stock that spawned first (i.e., G o ) in the western coastal region of the Gulf in March-April. He found advanced nauplii in late April and adults (GI) in late May. Temperatures at this time in this area were probably of the order of 3 4 ° C just below the surface (Bigelow, 1927), so that a month for this period of development is as expected (see Fig. 25A). A maximum of late nauplii (of G,) in the latter part of June, ova and young nauplii (G3)on July 30, and late nauplii (G3)on August 15, are all attributed to this western Gulf stock in various parts of the region. Eggs at the entrance of the Bay of Fundy in September 15-16 were attributed to G,. Although neither the details of the life cycle nor the mean temperatures during development can be very accurately determined, it seems likely that more generations could have occurred between late May and mid-September than the two supposed by Fish. 13. Delaware Bay

Studies on the American east coast south of Cape Cod by Fish (1925), and by Deevey (1952, 1956), are inadequate for details of life

histories of Pseudocalanus. However, they do amply confirm the fact that Pseudocalanus is essentially a, winter-spring form in this southern extremity of its range. It is also of some interest to note from Deevey (1952) that adult males occurred in samples throughout the season (January-August) in Block Island Sound. Also, neither there nor in Long Island Sound (Deevey, 1956) was there any evidence for a preponderance of a " resting '' stage in summer. Deevey ( 1960a) depicts size-frequency distributions of females during 1931, 1932 and 1933 from samples taken off the mouth of Delaware Bay. Sampling was most frequent during the first part of

154

CHRISTOPHER J. CORRETT AND IAN A. MULAREN

1931 (Fig. 33). Deevey suggested that there were winter, spring and early summer ,generations in these waters, but we infer that there were more. We assume that the large individuals on 10 February were later recruits to GI developed in colder waters. The appearance of three generations in 121 days between 22 January and 25 May is reasonable for a mean temperature of about 6°C (ca. 45 days per generation, from Fig. 25). The shortening of the period between generations up to G , is also reasonable, but it is affected in part by the low frequency of

FIa. 33. Size-frequency distributions of adult female Pseudocalanus and interpretations as generations (G,) in samples from outside Delaware Bay, eastern U.S.A. (After Deevey, 1960a.)

sampling. I n the period between 25 May and 23 July, three generations could have developed a t the high temperatures (i.e. up to G,). However, the resemblance in size between females from 22 July and those on 10 December, when Pseudoculanw reappeared in samples after a summer and autumn of absence, suggests that animals in July had already suspended development for the summer season, to reappear as Go of the subsequent season. We suggest that an origin of these Go animals in G6 (as in Fig. 33) is more probable than an origin in G,. In 1933, animals in November were much larger than those of July, indicating to Deevey that an earlier autumn generation had been produced in that year. 14. Black and Adriatic Seas

Extensive studies of Pseudocalanus in the Black Sea have been made by Soviet and Roumanian investigators, but there appear to be

THE 'BIOLOQYOF PSEUDOCALANUS

155

no data on size-frequency distributions of females, from which generations might be inferred. Porumb (1971, 1972) tabulates much information on the relative and absolute abundance of all stages of Pseudocalanzcs, including eggs. Although it is present in all seasons, reproduction is more intense in winter and spring, when its biomass is 5-6 times as great as in summer or autumn. Reduced abundance during the season of high reproduction is attributed to predation. Altogether, it is not possible to use Porumb's numerical data to follow generations. Sazhina (1971)has assumed that development rate during the period from October to March in the Black Sea is not food limited. Based on her laboratory estimates (Sazhina, 1968) of age of maturity at 34-39 days at 8-lO"C, she suggests that five generations are produced in a 180-day period (October through March). Although measurements on Pseudocalanus from the Adriatic are available in some works (e.g. Friichtl, 1920) these are inadequate for use in delineating life-cycles. Recent work (e.g. Hure and Scotto di Carlo, 1969) confirms that Pseudocalanus is a winter-spring form. VuEetid (1957) found egg-bearing females in the landlocked Malo Jezero from January to July. 15. Sea of Japan Andreeva (1975, 1976b)suggests in preliminary reports that Pseudocalanus in the northern Sea of Japan shows year-round reproductive activity, with one generation in spring, two in summer, and one in autumn. Reproductive peaks about 50 days apart in summer are said to match experimental generation lengths (egg to egg) of about 45 days, but as we suggest elsewhere (p. l l l ) , her experiments probably overestimated the copepodid period substantially.

C. Retrospects and prospects Although populations of Pseudocalanus in the far north are clearly short of food for much of the year, the evidence indicates to us that those of temperate waters during the productive season are not food limited, but develop, grow and reproduce at temperature-dependent rates. This conclusion matches the laboratory observations by Paffenhofer and Harris (1976), who found maximal development and growth rates a t food levels that are generally present in nature (see p. 113). I n Table XXVIII, we summarize theoretically possible numbers of generations of adult females that might be successively detectable in the plankton during productive seasons of various lengths at various seasonal temperature means. These are given to the nearest number of generations, calculated from the times to reach maturity

156

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

TABLEXXVIII.

APPROXIMATE

NUMBERS OF GENERATIONS OF Pseudocalanus FOODLIMITED IN NATURE

EXPECTED WHEN NOT

Seasonal mean temperature ("C)

Length of productive season (months) 4

5

6

7

8 ~

6 8 10 12

3 3 4

3 3 5

5

6

3

4

4 6 7

5 6 8

~-

5 6 8 9

(mean of the two localities in Fig. 25). Presumably very long productive seasons do not occur in the far north or at the southern extremities of the range, so that Pseudocalanus might be expected to have about 3-7 generations per year (plus an overwintering or oversummering generation) wherever it occurs in temperate waters. As yet few attempts have been made to go beyond descriptive accounts of life histories of Pseudocalanus (or other marine copepods) in order to deal with population dynamics. McLaren (1969; see also p. 96) showed that abundance of younger stages in Ogac Lake, Baffin Island, was related to abundance and body sizes of adults when food was sufficiently abundant. A paper by Lishev and Freimane (1970) seems to contain the beginnings of an analytical and predictive extension of such relationships, but their collecting techniques are not given clearly and their analysis is marred by questionable reasoning in places. They begin with the unlikely suggestion that Pseudocalanus is basically monocyclic (i.e. one generation per year) in Baltic waters. They then regress the abundance of young stages during various months in the period from 1959 to 1965 on the abundance of younger, (or parental) stages during earlier months. First, they show that the abundance of nauplii in May of various years was positively related to the mean abundance of adult females and males in February and May ; this seems reasonable, although the proper relationship would presumably be proportionality of nauplii with adult females. Next, they show that the abundance of C I V in August samples is positively related to the abundance of C I, C I1 and C I11 combined in May. They assume that the C I V in August actually derive from the earlier young stages. This leads them to propose that the negative deviation of the observed regression line from the line of identity is a measure of intervening mortality. It is highly probable that generations occurred between May and August, but the observed correlation of abundances between seasons is none the less interesting. A strong correlation between abundance of adults

THE BIOLOQY OF PSEUDOCALANVS

157

in October and copepodids in August is less interesting than are similar correlations between abundances of various stages in February and abundances of younger stages in the previous October (presumably no new generations had occurred during this winter period). Lishev and Freimane (1970) also attempt to show effects of temperature on development rates using the relative abundances of younger stages as functions of temperature, but their arguments are certainly erroneous where not incomprehensible. However, in spite of some difficulties with their work, it is of clear importance in showing the strong autocorrelations that may occur within and even between generations of Pseudocalanus. As they recognize, this may have great predictive value, and we urge that their approaches be tried by others. Some interesting problems about life cycles of Pseudocalanus remain to be explored. We do not altogether agree with Heinrich (1962) in classifying Pseudocalanus as a copepod that will breed all year round, depending only on a sustained crop of phytoplankton. The work of Marshall (1949) and others seems to suggest that Pseudocalanus may “voluntarily” suspend development before the end of the productive season. We speculate that Pseudocalanus may have a lifehistory strategy that involves a choice between entering a resting stage as the usual end of the productive season approaches and going on to mature and reproduce. Perhaps those individuals that have accumulated enough stored oil are somehow triggered into ceasing further development (at C IV-V in Loch Striven). Those that, for any reason, have been unable to store enough oil at a critical stage may go on to produce another generation of individuals, some of which may be successful in preparing for overwintering. It may be noted that this speculation implies that the best-fed individuals are in terms of organic matter the least productive during the latter part of the season. This may seem contrary to intuition in terms of production ecology, but is perfectly in keeping with the evolutionary viewpoint that organisms are not “ interested ” in maximizing production, but in perpetuating their own kind. Another problem that needs to be explored is the possibility that different forms (species) of Pseudocalanus have intrinsically different life cycles as suggested by Wiborg (1954) and Lacroix and Filteau (1971). Another explanation is that large, oil-rich individuals in deeper water during summer in temperate waters may have simply developed to an overwintering stage in cooler, deeper waters. Certainly, animals of high latitudes, although basically annual or biennial, do not always seem to be able to “ prevent ” themselves from untimely maturation and reproduction in abnormally warm waters, such as

168

CHRISTOPHER J. UORKETT BND IAN A. MCLBREN

Ogac Lake, and autumn maturation occurs among some individuals even in the colder arctic and subarctic waters of Greenland a,ndUngava Bay. This suggests to us that these northern animals, given enough food, could have several successful generations in a year, without necessarily suspending development. We suggest that further studies of " landlocked " populations in various parts of the world would be revealing, as would experiments on the character of overwintering stages.

XII. VERTICALMIGRATION Elsewhere we have discussed vertical distribution of Pseudocalanus in the sea during the daytime (p. 120). I n this section we are concerned with the dynamic aspect of vertical migration. Die1 vertical migrations have attracted most interest, but Psezdocalanus shows longer term migrations as well and we designate these as seasonal and ontogenetic migrations. Ontogenetic migrations are defined below, but it is important to note here that we limit our definition of seasonal migration to migration t o the depths that clearly involves some sort of resting or overwintering phase in the population (p. 157). A. Ontogenetic migrations By ontogenetic migration, we mean the change in position of individuals in the water column (as observed by night or day) as they develop. Of course, individuals cannot be followed through extended time, and we must infer such movements from the distributions of stages a t any given time or over a period of time. The tendency for younger stages of Pseudocalanus to occur nearer the surface has been noted a t least as early as Kraefft (1910), and by numerous authors since (e.g. Ackefors, 1969a, b ; Marshall, 1949; Wiborg, 1940, 1954). Exceptions have occasionally been noted (e.g. Minoda, 1971). There are few details in most of these studies, but observations from two stratified, landlocked arms of the sea in northern Canada (Carter, 1965; McLaren, 1969) and from the Black Sea (Afrikova, 1975) are quite revealing. Carter (1965) illustrates vertical distributions of all stages in the inhabitable upper 30 m (oxygen depleted in deeper water) of Tessiarsuk, in northern Labrador, on 2 July, 2 August and 26 August 1961. On each occasion there were proportionately more egg-bearing individuals among deeper females and earlier nauplii were found slightly deeper on average than were later ones. A pattern of upward swimming after

THE BIOLOQY OF PSEUDOCALANUS

159

hatching is implied. Older stages were found generally with maxima in progressively deeper water, sometimes several successive stages showing the same patterns of distribution. A marked bimodalism in depth distribution of some stages is of interest. On 9 July there were peaks of all stages at or just below the sharp thermocline and halocline at 4-6 m, and deeper maxima of C 1-111 and adult females between 10 and 15 m. There were similar dual maxima on 3 August in C 111-IV, CVd, and adult females. There is a suggestion on 9 July of descent among older nauplii, which would have been members of second brood (G,B,, see Fig. ZSC). Perhaps the deeper individuals among C 1-111 in early July and C 111-V represent later-spawned individuals, some from Temperature ("C)

06

7

8

9

Sample depths

10

20

30 %.Salinity

FIG.34. Vertical distributiom of P.sedocalanua in the middle basin of Ogac Lake, B a f f i Island, around mid-day on 21 August, 1967. (After McLaren, 1969.)

the second brood of the season, while those staying near the surface were earlier in origin. The bimodal depth distributions among adult females may also reflect a dual origin, for during early summer two size modes representing autumn and spring maturation (see p. 140) were present. Carter (1965) also depicts detailed depth distributions in Anaktilik Bay, outside Tessiarsuk. I n the bay, where there was little stratification, older stages were found deeper, as expected, but there were no sharp depth-maxima. A series of samples from Ogac Lake, a landlocked fiord on Baffin Island, were particularly revealing (Fig. 34). The pattern can be interpreted as ascent to the surface by young nauplii, followed by descent beginning a t C 111. Here again (as in Tessiarsuk, above) it is possible that the two groups in C I11 represent two different origins, in

160

CHRISTOPHER J. CORKETT AND IAN A. MOLAREN

-A,---

-!.

/-----

lo-! ( ! \

-!

1

,.‘

--

-\

20- ‘.,Chlor.

\.

-

-

#

30

I

*

I

-*

,’

.‘

,-7-.-. I

I

I

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suggests that the ontogenetically descending animals were “ seeking ” colder waters and were loathe to penetrate the temperature peak at 15 m. The pattern of depth distribution in Ogac Lake in mid-August 1962 was entirely different (Fig. 35). The peak of abundance of all

THE BIOLOGY O F PSEUDOCALAA'US

161

stages near the surface was related to a persistent peak of phytoplankton abundance just below the halocline (the chlorophyll peak was reduced by mid-August, as seen on Fig. 35; see McLaren, 1969). The deeper maxima of Pseudocalanus were also found at higher food levels. Females in the deepest samples in 1962 showed a bimodal size distribution, the smaller ones probably representing animals that had recently descended from the concentration in warmer, near-surface waters (McLaren, 1969). This suggests that descent can be a sudden " decision " at any stage. Afrikova (1975) depicts percentage distributions of all stages (eggs, NI-V grouped, N V I and each copepodid stage separately, adult females and adult males) in seven strata down to 200 m in three regions (northwestern, central, eastern) of the Black Sea in February, April, August and November. It is difficult to generalize from this massive survey without data on time of day, hydrographical and phytoplankton conditions, but a few observations on possible ontogenetic movements can be made. A t times the peaks of depth distributions of all stages coincided, so that any movements must have taken place within the 25 or 50 m strata. At other times, adult females, ova, and sometimes NI-V showed primary or secondary peaks in deep water that were not exhibited by N VI so that an upward migration of young stages is implied. Copepodid stages generally occupied mid-depths, with adults often showing clear primary or secondary peaks in deeper water in most regions and most seasons. This suggests that adults may migrate deeper (we will show this below, p. 166, to be true of adult diel migrants in the Black Sea). An exception was in April in the northwestern part of the sea, when females were most abundant near the surface, whereas younger stages were deeper. We offer the following tentative conclusions about the nature of ontogenetic vertical migrations by Pseudocalanus. (1) Younger stages are generally found nearer the surface. (2) Animals at the same stage, but of different developmental histories, may show different depth preferences. (3) The avoidance of warm, near-surface temperatures by, in particular, older stages may be involved in their descent.

.

B Seasonal migrations Many authors have noted the tendency of copepod populations to be found in deeper water as the productive season (as defined in p. 135) progresses. Althoughall stages may be involved, often the descending population may have developed to one or a small number of stages. Among those in more recent studies who mention the phenomeA.X.B.--15

8

162

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

non in different populations of Pseudocalanus, VuEetM (1961), Carter (1965), Ackefors (1969b) and Porumb (1971) have suggested that gradual seasonal descent is related to withdrawal from warming of the surface waters. However, others have described a migration t o the depths that clearly involved some sort of resting, overwintering phase in the population, and it is this phenomenon that we wish to designate as seasonal migration. Marshall (1949) found that all stages in Loch Striven, Scotland, were mostly above 10 m from late January until late June. Most were below 10 m during July, but nauplii, and C I and C I1 were still above 10 m in August. Although she invoked warming of the surface waters (see Fig. 29) and possible water movements to explain parts of this pattern, Marshall clearly ascribes the descent of the major part of the population after the end of June to the preponderance of the overwintering stage C IV (later becoming C V). Ussing (1938) and Digby (1954) describe a similar seasonal sinking of stages between C I1 and adult in the fiords of East Greenland. They overwinter and gradually develop at depths below 50 m. I n the shallower Foxe Basin, in arctic Canada, there was no evidence for such a partial disappearance of the stock in winter (Grainger, 1959). Kovalev (1967) indicates that copepodids and females are absent from suface water of the Black Sea in summer, but that there are peaks at 75-100 m in both summer and winter. The more extensive work of Afrikova (1975) shows a strong secondary peak of abundance in deep water of all copepodid stages in the central part of the Black Sea in August 1951. No such peak is evident in samples from November 1954, so that the August distribution might be taken as an example of seasonal descent. However, in February 1956, in the northwestern part of the sea, all copepodid stages were most abundant in the deepest layers, with secondary peaks in shallower waters, ao that the pattern of descent is not clear. We cannot determine if there is a seasonal resting stage in these Black Sea animals. A profound seasonal migration in the Norwegian Sea has been described by Bstvedt (1955) and confirmed by Hansen (1960). The overwintering animals below 600 m (mostly 1 000-2 000 m) consisted almost all of C V. A small number of C IV stayed above 600 m. When the ascent began in March, a large portion of the mature animals had already matured below 600 m, and few adult males reached the surface layers. By June, C V began to predominate, and by July most of these had moved into deep water. Although only a few studies have demonstrated it, we believe that the seasonal descent of " resting " overwintering stages may be the rule

THE BIOLOGY , O F PSEUDOCALANUS

163

among Pseudocalanus populations in temperate waters. McLareii (1963) argued in support of conteiitions by some authors that this behaviour allows animals to conserve their energy stores in the cool depths. McLaren (1974) stressed the fecundity advantage froin large size in colder waters (p. 116) when life cycles are seasonally interrupted. In the Norwegian Sea, the animals migrate beyond 600 m, where there is a sharp temperature change, to depths that are 6-8OC cooler than the surface a t any season (0stvedt, 1955). I n inshore waters, the surface may become colder than the deeper waters by the time animals return to the surface to commence breeding (Marshall, 1949). We shall demonstrate in the next section (p. 171) that these “resting ” stages may forego diel vertical migration into warmer waters in summer.

C. Die1 migrations Before proceeding further we must clearly distinguish between the operation o f ” and the “ significance of ” vertical migration. Hardy (1956) writing about the phenomena of vertical migration reported, “ It would be nice and tidy if one could just pin down the operating factor to some such relatively simple thing as the movement up and down of a particular light-intensity-although it would in no way tell us what was the actual reason, in terms of advantage to th,e animal, for such a reaction being evolved ’’ (our emphasis). I n this review, in addition to purelydescribing vertical migration, we will largely concentrate on aspects that relate to the “ significance of ” rather than the “ operation of ”, since relatively little study has been made on the physiological control of vertical migration by Pseudocalanus. “

1. Observations

Although there are numerous published observations on vertical distributions of Pseudocalanus, few show differences between night and day. Here we will stress studies that are based on close spacing in depth and time, give insights into behaviour of different stages, or offer additional data on the physical, chemical and biological circumstances found in the water column at the times of sampling. I n addition to describing diel migration, we will bring the reader’s attention to conditions in the water column (temperature, food, predators) and to differences among seasons and among stages, all of which have some bearing on the possible adaptive significance of migration described later. Evidently the first sketchy observations on vertical migration of

164

CHRISTOPHER J. CORKETT AND IAN A. MULAREN

I""

+ t I L m & +I

1

150

1

I

I+-

..,-

75:

-

*+ + 1

1

1

1

Time of doy

FIG.36. Vertical distributions of Pseudocalanwr in the Black Sea at various times, during a 24 hour period in March 1973. (Data for C I, C 11, and adult 3 from Zagorodnyaya, 1974, and for C I11 adult 9, including unbroken and broken arrow8 for presumed strong and weak migrants, after Zagorodnyaya, 1975.)

Pseudocalanus were made by Savage (1926), Bigelow (1926) only implying that it probably occurred in this copepod. Savage found greatest abundance near the bottom by day and fairly even distribution in the water column at night, but supplied few details. Migration in the Black Sea was described by Nikitine (1929) and ;t number of authors subsequently. Porumb (1971) studied migrations

THE BIOLOGY OF PSEUDOCALANUS

166

into the top 5 or 10 m, and found that animals did not enter these waters when temperatures rose above 23°C. The most thorough study of vertical migration of Pseudocalanus in the Black Sea (Zagorodnyaya, 1974, 1975) also gives the most striking examples of the phenomenon of which we are aware (Fig. 36). Samples are taken on unspecified dates in March 1973, about 40 miles off the coast near Sevastopol. Unfortunately, Zagorodnyaya gives no details on methods of capture, population sizes or physical-chemical conditions in the water column. Although she states that temperature of all strata a t the time was about 8" (which temperature she uses later in assuming a speed of passage of food through the guts of the copepods), there may have been some thermal stratification at this time (see Petipa et al., 1963). No data are given on vertical distributions of copepods in another series from January 1973, although migration was said by Zagorodnyaya to be weak. Clearly, vertical migration was marked in older stages a t least, although doubtfully in adult males (Fig. 36) whose scarcity may have produced sampling problems (see unlikely distribution a t 1900 h). Zagorodnyaya (1975) concluded tentatively from inspection of the distributions that there were two classes of migrants : strong and weak. For example, the almost complete compression of adult females from the upper layers at 0300 into the layer at 50-75 m at 0700, indicates that these upper-layer animals moved faster and farther than those that moved, at most, from 50-75 m to 75-100 m. We include her arrows, tentatively interpreting the movements of these supposed weak and strong migrants (Fig. 36). By using this grouping into strong and weak migrants, in ways which she does not make altogether clear, Zagorodnyaya (1975) estimated the residence times of individuals in different depth layers. Using observations of the gut contents of copepods in each layer and experiments on passage of food through the guts (see p. 71), she was able to estimate daily rations for weak migrants, strong migrants, and for possible non-migrants. Accepting the existence of these classes of migrants in March, she showed that strong migrants in stages C I11 to adult female consumed more than weak migrants which in turn consumed more than supposed non-migrants. She also analysed gut contents of animals in January and concluded that the differences between weak migrants and strong migrants during that month were very slight, evidently because migration was only of small amplitude. Zagorodnyaya (1974) showed from observations on fullness of guts (see discussion, p. 74) that almost all food was secured at night. The actual rations she calculated (maximum about 12% of weh &/day by strongly migrant C V) would not be enough to satisfy the daily needs of

166

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

growing copepods (see p. 133). However, the relative differences in daily ration among classes of migrants are of great interest, since they imply that strong migrants either needed more food or could obtain more food, both of which may have energetic implications. Zagorodnyaya and Svetlichnyi ( 1976) demonstrated diel changes in specific gravity of individual Pseudocalanus associated with these diel rhythms of feeding and migration : adult females, but not non-feeding adult males, showed a lower specific gravity at night when their guts were mostly full. Afrikova ( 1976) analysed vertical distributions of Pseudocalanzcs from samples taken at P 5 h intervals over three 24 hour periods in June 1959 from the west-central Black Sea. The net samples were from 0-10 m, 10-25 m, 25-50 m, 50-75 m and 75-100 m, presumably (not stated) by stratified oblique or vertical hauIs. Her Fig. 2 (date?) illustrates a marked thermocline from ca. 8°C to ca. 16°C between 25 m and 10 m with near isothermy below and above these depths. There was a more gradual halocline from ca. 21%, to ca. 18%, between 75 m and 50 m. Afrikova, in ways not made fully explicit, calculated mean depths of occurrence for each stage at each time during the three 24 hour series. These mean depths (her Table 1) do not match the peak depths for grouped stages on her Figs 1 and 2 (which show continuous curves of biomass against depth rather than histograms of numbers in each sampled layer). We have difficulties in interpreting some of her results, and we feel that the amplitudes and rates of migration for each stage as tabulated by her must be strongly dependent on the discrete sampling intervals used for depths and time. However, the general pattern of peaks in her Figs 1 and 2 seems to be fairly consistent within stages between sampling dates. Because Afrikova (1976, her Fig. 1) grouped eggs (which cannot possibly migrate unless attached to females) with N 1-111, we cannot interpret movements of these youngest nauplii. Older ones (N IV-VI) are shown as having moved up by day in one sampling series, but generally as peaking in the 25-50 m layer at all times of the day and night during the other two series. Peaks of both C 1-111 and C IV-V are shown as generally between the 10-25 m and 25-50 m layers (i.e. evidently at around 25 m, just below the thermocline). The older animals (C IV-V) were much more concentrated in the upper part of the 10-25 m layer (i.e. in the warmest water available) by night. Adults (sexes combined, but presumably largely females) were much deeper by day, with peaks shown at the boundary of the 50-75 m and 75-100 m layers. At night, peaks were generally between the 25-50 m and 50-75 m layers (one peak shown between the 10-25 m and 25-50 m

THE BIOLOGY OF PSE UDOCALAN US

167

layers). Adults thus showed the greatest amplitude of migration, but this did not evidently carry them substantially into the thermocline ; this contrasts with the above-described pattern for older copepodids. Afrikova does indicate (without giving data) that adult males in one 24 hour series rose in evening (2000--2100 h) to 0-10 m. From dissection of guts, Afrikova (1976) concluded that most feeding by copepodids and adult females took place in the 10-25 m or 25-50 m layers, but that some food was secured in deeper waters also. She states that migration by Pseudocnlanus in general was less abrupt nnd the diel feeding rhythm less marked than for Calanus kelgolandicus in the same waters. She also concluded that temperatures and salinities had little influence on the amplitude of migration. However, as we have indicated in the previous paragraph, the response to temperature does appear to vary with stage : young individuals being generally resident at all times at cool mid-depths; older copepodids moving from middepths into warm, near-surface waters at night ; and adults living deep by day, migrating farthest up to mid-depths, but not generally accompanying the older copepodids into the thermocline. I n the Adriatic region, VuEetii: (1961) found that animals in Vcliko Jezero (a coastal saltwater lake) occurred near the surface in Janiiary and deep in summer. However, her limited samples revealed no evidence of diel vertical migration during either season. Bogorov ( 1946) found that Pseudocalanus remained constantly in shallow waters in the Barents Sea during summer, but found a weak movement toward the surface in localities in the White Sea farther south. Bogorov interpreted these patterns in terms of the 24 hour daylight obtaining during summer a t the higher latitude. However, we note that surface waters a t these high latitudes would be warmer farther south, especially by Iate summer. McLaren (1969) took daytime and night-time samples from another arctic location, Ogac Lake on Baffin Island (Fig. 35). Here, in spite of complete darkness at night on 14-15 August 1962, and in a highly stratified water column, there was very little, if any, vertical movement of any stage. Slight apparent differences in the distributions (Fig. 35) can be attributed to sampling errors and the differences in depths sampled by night and day. What is of interest is the strong concentration, night and day, at depths where chlorophyll concentrations were highest (the near-surface peak in chlorophyll had been more pronounced in previous weeks ; McLaren, 1969). Development rate at this time of year in 1962 (McLaren, 1969, his Fig. 2 ; see also Fig. 28B in this work) was very slow, and possibly the animals were obliged to feed throughout the 24 hours. There was strong migration of immature Sugittu elegam (making up the mobile

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

part of the predator carbon biomass in Fig. 35). It is of interest that the deeper peaks of predator concentration and the night-time peak of migrant predator concentration coincided with copepod concentrations. Some studies in the eastern North Atlantic and adjacent waters have supplied little information on diel vertical migration relating to physicalchemical conditions (e.g. Savage, 1931 ; Malikowski and Ciszewski, 1962). Hansen (1951) found that adult female Pseudocalanus did not migrate through marked temperature-salinity gradients at 9-1 3 m in waters near Oslo in late June 1947. Adult males were less migratory, and we infer that younger stages (said by Hansen to occur a t 0-50 m) were above or were able to migrate through the gradient, but details are not given. The most detailed study in European waters has been made by Lee and Williamson (1975) who observed vertical distributions and movements of Pseudocalanus (adults and copepodids combined) in the Irish Sea. Although they depict vertical distributions of temperature salinity, 0,, SiO,, NO,, NO, and PO,, we doubt that most of these have any influence on vertical movements of copepods (contrary t o their suggestions ; see also our comments in p. 22), and we show only temperature in Fig. 37. On every date, there appears to have been some vertical migration. However, there was no single pattern with respect to timing of the light cycle. The upward movement well before sunset on 4 May 1969 was particularly striking, as was the lag after sunrise on 4 July 1970, and the presence near the surface a t all times on 21-22 October 1970. These observations are contrary to the usual notion that migrants avoid the surface during daylight hours. Migration appears to have been least coherent on 21-22 October 1970, when the waters were the least stratified thermally of the five dates. I n general, the distributions give the impression that only parts of the populations moved towards the surface, and on 11-12 August 1970, it appears possible that another part might actually have moved down. Wide spacing of the horizontal plankton hauls, the rather strong currents in the Irish Sea, and the grouping of all older stages in the counts by Lee and Williamson, probably preclude closer analysis of the data in Fig. 37. I n the Pacific and adjacent waters, Japanese and Soviet workers have investigated diel migrations of Pseudocalanus, but only some studies refer to conditions in the water column. Minoda and Osawa ( 1967) found Pseudocalanus and other copepods concentrated near the bottom of the thermocline, which coincided with a daytime sonic scattering layer in the Okhotsk Sea in early August. Their widely spaced hauls, above, within, and below the scattering layer, gave no

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evidence of diel migration of Pseudocalanus. The most extensive studies are in Minoda (197 l),who found relatively uniform distributions in the upper 50-200 m during May, June and July in the seas east of Kamchatka. We interpret his Figs 13 and 14 as showing increase at the surface by night at two stations, a decrease at two stations, and no change at three stations. Thermal stratification was generally weak in the upper 50 m.

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The most complete set of observations of diel vertical migrations of Pseudocalrinus anywhere have been made by Lock (1968) in the relatively enclosed Bedford Basin, near Halifax, Nova Scotia. He paid particular attention to precise depth controls, using horizontal hauls with quantitative Clarke-Bumpus plankton samplers. The rather coarse-mesh net (variously clogged) led to variable results for the youngest copepodid stages, and we review here only the depth distributions of stages C 111-V and adults (Fig. 38). Lock separated the sexes of C IV-V, and although we group them in Fig. 38 we at times

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discuss sexual differences below. Phytoplankton cell counts do not include very small forms or flagellates, which may predominate in Bedford Basin (Poulet, 1974). I n early May, C 111-V appeared to be concentrated just below the thermocline at mid-day. Migration to the surface was strong in C I11 and C IV, which were found near the surface well before sunset (see similar situation early in the season in Fig. 37). Among the C IV at the surface in late afternoon 80% were males, although overall in the water column 52% were males. Female C I V thus appeared to spend more time in the cooler depths. Stage C V only occurred near the surface in the midnight sample, and sexes behaved similarly. The evidence for migration of adults (both sexes) is weak but there does appear to have been some increase in shallow waters at night. Animals rising above 10 m would have experienced somewhat warmer waters, but the counts of phytoplankton cells suggest that little variation in food occurred in the upper 35 m or so. In mid-July, the pattern was rather different (Fig. 38). Migration in C I11 appeared weak, with perhaps only a small part of the population involved, in the middle of the night. The non-migration in C IV is very striking. This stage (see scale differences in Fig. 38) greatly dominated the population a t the time, and was found concentrated in deep water throughout the 24 hours, with only a small hint of increase at the 6 m level around midnight. Clearly these are " resting ", potentially overwintering animals, similar to those that have been documented in Loch Striven, Scotland (see p. 144). The change in behaviour in C Vis equally striking. Although part of the population may have stayed in deep water, there was a clear movement to shallow water commencing before sunset, and a movement away from the shallows commencing before sunrise. We suggest that, although some C V may also have been resting, some were active migrating members of a developing generation, like their counterparts in Loch Striven. Both sexes of C V were involved in the peaks nearer the surface at night, but only males exhibited this peak in the early morning, thus staying longer in the warm water. Adult females also clearly moved upward to the surface at night, perhaps not quite as far or for quite as long as did C V. Adult males showed little if any migration. It can be seen from the lower panels of Fig. 38 that migrants in July generally did not enter the very warm surface waters. Those a t 6 m (early night peak of C V) were a t about 12"C, where phytoplankton was also evidently very concentrated. Those at 12 m (peak of femaIes shortly after sunset) were at about 7OC at a concentration of cells about two-thirds the maximum value at 6 m.

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I n mid-December the evidence for migration among young stages was equivocal. It appears possible that the bulk of the population stayed deep during the 24 hours, and that another part stayed in shallower waters, some approaching the surface near sunset. The situation among adults is intriguing. There was a clear concentration, especially of adult females, at the surface for a short period after sunset. The water column was almost isothermal and phytoplankton evidently scarce a t the time. Lock (1968) also graphed means and variances of sizes of adult females for times of day between which depth distributions changed most rapidly (lower panels of Fig. 38). Although he does not supply significance tests for most of the differences between depths and times, he does state that at least 100 females were measured from each timedepth sample. The interested reader may be able to estimate the approximate significance of the differences between means by seeing where they differ by more than d ( v a r . length)/2/100. By these crude methods deeper animals can be shown to have been generaIly significantly larger on average than those from shallower depths. I n a few instances, significant differences occurred between mean sizes at the same depth at different times of day. Also, P tests (ratios of variances) imply that animals were more variable at certain times of day a t the same depths. We do not describe such differences in detail or attempt to explain them here, but simply offer them as indications that animals of different sizes within the same stage may have different migratory habits. 2. Control by light

Although other factors may influence the depths occupied by Pseudocalanus (e.g. pressure, see Digby, 1967),the " operating factor " of diel migration has generally been considered to be related to diel light cycles. Little detailed work has been done on the relationship between diel migrations and diel light levels in Pseudocalanus, but some observations have been made, particularly in relation to solar eclipses, and these we outline below. Two studies have been made involving the response of Pseudocalanus to solar eclipses. Skud (1968) sampled quantitatively 0, 10 and 20 m at, before, during and after a total eclipse in late afternoon (totality 1745 h) of 20 July 1963, on the coast of Maine. Temperature was about 11.3" at 0 m and 9.3"C at 20 m. Pseudocalanus was one of two species of copepods out of seven believed by Skud to exhibit some response to the diminished light. His graph indicates that Pseudocalanus did not occur in surface samples, so his evidence for upward movement

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must be based on changes in the middle depth, at 10 m. On July 20 only a few per cent of the population were at 10 m before the eclipse, compared with some 30% during and after totality. However, his graph also shows that on the previous day (when animals in general occurred higher in the water column, possibly because it was less bright), 40% of the population was a t 10 m before, 70% during, and 55% after the time of totality on the following day. Such variations seem to us to make interpretation difficult. Sherman and Honey (1970)made a study of the vertical distributions of Pseudocalanus a t 0, 10 and 30 m in relation to the near-total eclipse a t Boothbay Harbour, Maine, on 7 March 1970. The water column a t the time was almost isothermal a t 1-6-2-0"C. On the day before the eclipse, adults of both sexes were generally concentrated near the bottom, rising to the surface in late afternoon, and evidently leaving it after sunset, reminiscent of the situation in December in Fig. 38. An odd concentration at 10 m occurred in late morning of both days, but the upward movement at totality of both sexes, especially males, seemed quite clear. It is not surprising that Pseudocalanus should respond to decreased light but observations in the previous section suggest that the response is greatly modified by seasons and circumstances. The rise to the surface well in advance of sunset early in the season in the Irish Sea (Fig. 37) and Bedford Basin (Fig. 38) is especially intriguing, as is the complete lack of response of some stages at some times in some conditions of the water column. I n the literature in general a common response to light is said to involve upward movement with change of light at both dusk and dawn, with randomization and net downward movement during darkness. There is no evidence of this dusk and dawn rise in the observations on Pseudocalanus in the Irish Sea (Fig. 37) and Bedford Basin (Fig. 38); there was rather at times an asymmetry in the nocturnal response, with more animals near the surface at dusk than at dawn. 3. The adaptive value of die1 vertical migration Many explanations have been posed for the significance of the pervasive phenomenon of vertical migration of zooplankton. Whatever the explanation, it is axiomatic that migrant behaviour must generally confer some selective advantage to individuals. I n asking why animals migrate, it is assumed that there must be some '' cost " involved as well. Evidently Afrikova (in a 1972 symposium proceedings quoted by Klyashtorin and Yarzhombek, 1973, but unavailable to us) has estimated from loss of stored oil that the energy consumed by a migrant Pseudocalanus would add little to its " routine " metabolism. This is in

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accord with other recent work on energetic costs of vertically migrating Pseudocalanus (Svetlichnyi, et al. 1977) based on calculations of work required to overcome hydrographic resistance and gravity. As is so often true of animals, there is probably more than one explanation for a complex behaviour. Furthermore, explanations involving selective advantage in evolutionary time may be difficult or impossible to test. The best we can do is to see if one or another explanation appears to conform better with observations made at some times in some places. We propose here to examine three hypotheses as possible explanations of die1 vertical migration by Pseudocalanus. The first of these, the predation hypothesis, has not been developed in the literature as a specific explanation for vertical migration of Pseudocalanus. However, it is so widely accepted and seems so plausible that we feel it should be included with reference to observations on Pseudocalanus. The second, energy-bonus hypothesis was developed as part of an essentially demographic hypothesis with specific reference to Pseudocalanus by McLaren (1963). Later, McLaren (1974) found the energy-bonus to be unnecessary in the development of the demographic hypothesis. However, the energy-bonus part of the original (McLaren, 1963) hypothesis stands on its own, and we feel justified in including it with reference to observations on Pseudocalanus. The third, demographic hypothesis, was clearly developed with Pseudocalanus as a model (McLaren, 1974). I n examining these three hypotheses we will again bring the reader’s attention to conditions in the water column (temperature, food, predators) that are of consequence in these hypotheses. (a) The predation hypothesis McLaren (1963, 1974) agreed that, of a number of long-standing explanations for vertical migration, avoidance of visual predators during the day was most plausible and perhaps the chief reason for migration at night, rather than by day, even if migration had some other primary advantage. Of course, the observations on Pseudocalanus show that in general it does leave the surface by day, which could indeed help in avoiding visual predators. However, this does not always seem to be a sufficient explanation of its migratory behaviour. For example, why do young stages (which are possibly most vulnerable to predation) tend to remain near the surface (e.g. Fig. 34) and be less migratory (Fig. 36)? Why are adult males, admittedly short-lived even without possible predation, generally so much less migratory (Fig. 36 ; 16-17 July in Fig. 38)? Why do animals approach the surface at times long before sunset (May,

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Figs 37 and 38), or stay there after sunrise (3-4 July, Fig. 37), or spend the entire daylight period in well lit surface waters (p. 167 ; October, Fig. 37), if such behaviour would expose them to visual predators? I n Ogac Lake non-migrant Pseudocalanus may contend with their chief predator Sagitta elegans (presumably a non-visual predator), migrating toward the surface a t night (Fig. 35). Pearre (1973) also showed that migrating S. elegans in Bedford Basin fed on Pseudocalanus most heavily at night. A case might be made in these situations for the evolution of descent by Pseudocalanus at night to avoid predation. All the above observations seem to pose problems for the predation hypothesis of vertical migration. Nevertheless, the advantages of avoiding visual predators seem so plausible, that we believe that predation must play some role in the diel migration of Pseudocalanus, but that it is at times over-ridden by other factors. (b) The energy-bonus hypothesis McLaren (1963) offered an original analysis of the way in which vertical migration in thermally stratified waters might be advantageous to the individual migrant because of enhanced fecundity (by which he meant clutch size in the case Pseudocalanus). He began his arguments with the widely held notions that animals in deeper, cooler water could conserve energy and that this energy gain could be advantageous for a number of reasons. This had been offered as an explanation especially for the seasonal retreat from warm surface waters by overwintering stages (see p. 162). McLaren (1963) assumed that Pseudocalanus could secure all the food it required by night nearer the surface waters, where food would usually be more abundant and more rapidly ingested at the generally higher temperatures. This food would then be metabolized with less respiratory loss in deeper, cooler waters, thus conferring an energy bonus that would be partly lost by staying in the warm surface waters. We have seen that Pseudocalanus may be satiated by food levels that are well within the range found in nature (p. 68), and that it may show marked diel rhythms in feeding (p. 72). We have also shown that, in populations where migration surely occurs, animals are capable of developing at maximal, temperature dependent rates (p. 144). Altogether, the evidence we have reviewed confirms the premise that Pseudocalanus can at times secure an adequate daily ration for maximal growth rates by feeding during only part of the 24 h period. Even if energetic costs of migration are small (p. 173), the energybonus hypothesis predicts that migration would not be advantageous in an isothermal water column. McLaren (1963) reviewed a wide

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literature on different species suggesting that migration was generally weaker in unstratified waters. Certainly some migration occurs in Pseudocalanus in unstratified waters. However, our review suggests that migration was weakest in two seasonal series (Figs 37, 38) in autumn and winter when waters are isothermal. Observations in December in Bedford Basin (Fig. 38) seem to suggest that adults in particular “sampled” the surface water at dusk and, finding them unsuitable (no warmer?), returned to deeper water. We have also recounted other evidence that vertical distribution and migration of Pseudocalanus is in general strongly affected by temperature. McLaren (1963) suggested that the energy bonus of migrants might appear in accumulated oil for overwintering in copepods. The work of Lock (1968) indicated, if not conclusively, that animals reared in alternating warm temperatures by day and cold temperatures by night laid down more oil (p. 128). However, animals observed in a similar experiment seemed to have food in their guts at all times during the 24 h period suggesting to Lock and McLaren (1970) that a diel feeding rhythm might not have been involved. As McLaren (1963) pointed out, the complete cessation of migration by resting animals in the overwintering stock (see C IV on July 16-17, Fig. 38) might be viewed as an extension of the energetic gains of diel migration : these animals by ceasing migration altogether conserve energy at much lower temperatures. Altogether, there appears to be some support in observations of Pseudocalanus that part-time residence in deeper, cooler water may be metabolically advantageous for diel migrants when food is sufficiently concentrated. (c) The demographic hypothesis From the review of the effects of temperature on development rates and sizes of zooplankters, including Pseudocalanus, McLaren (1963) proposed that residence of developing females by day in deeper, cooler waters would increase adult size, and therefore fecundity, over non-migrant individuals in warmer surface waters. There was little difficulty in demonstrating an advantage for animals where life cycles were set by seasonality. For example, female Pseudocabnus in the surface waters of the Norwegian Sea would become larger and ultimately have large clutches if they migrated daily to cooler waters, before descending as C IV-C V to overwinter in the cold depths (see p. 152). Since they would not mature untilthe following spring, the slight retardation of development caused by diel migration to cooler waters during summer would be inconsequential.

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However, the situation is different for populations that develop continuously. McLaren (1963) showed that increased clutch size in these continuously developing individuals would not compensate for the retarded maturity at low temperatures. I n these, the potential rate of increase (i.e. assuming no mortality) of Pseudocalanus was positively related to temperature. Therefore, he devised a model using von Bertalanffy 's well known growth equation to suggest a mechanism whereby increased " anabolism '' in warmer water and reduced " catabolism " in colder waters (i.e. the energy bonus of the previous hypothesis), could enhance the body size and therefore clutch size of females beyond that expected at the mean temperatures during development. I n this way, McLaren deduced that there would be an important demographic advantage to migrants in potential rate of increase. Lock and McLaren (1970), in a laboratory test of part of McLaren's (1963) ideas (see p. 120), showed that no suchenhancement of female size occurred when animals were raised from C I11 to adulthood in an alternating temperature cycle, For this reason, among others, McLaren (1974) revised the demographic hypothesis as it applied to Pseudocalanus in particular. First of all, he noted that his earlier arguments were based on false premises : the selective advantage (and evolutionary fixation) of a trait depends, not on the potential rate of increase (i.e. no mortality) of its bearers, but on the realized rates of increase in the face of natural mortality. His subsequent model of the demographic advantages of die1 vertical migration (McLaren, 1974) does away with the necessity for incorporating theoretical metabolic effects. The essence of his model can be described, with references to data and observations in this review, as follows. 1. Young animals gain little size advantage from living in cool waters (p. 120, Fig. 27), so that development (McLaren assumed up to C I11 for calculations) in warm surface waters would have little effect on ultimate body size but would accelerate maturity. 2. Older stages gain a size advantage at low temperatures (Table XXII, Fig. 27) which can be translated into increased clutch size (Fig. 19A), potentially sustained for ten successive clutches (p. 93). He therefore assumed that migration occurred between the beginning of C I11 and adulthood, to give this advantage in clutch size. 3. He then assumed that there were two classes, migrant and nonmigrant, of animals between C I11 and adulthood. 4. He assumed that both non-migrants living near the warmer surface or migrants spending part of the time in deeper, cooler water were food-satiated and developed at maximal, temperature-dependent

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rates (as in Fig. 25), and that adult females produced eggs at maximal, temperature-dependent rates (as in Fig. 21). 5. He assumed that the non-migrant populations living constantly near the surface were kept in equilibrium, with no increase or decrease of numbers of any stage over one or more generations. 6 . Under these assumptions, when the same constant mortality rates were applied to all stages, both migrant and non-migrant classes, the hypothetical migrant class decreased relative to the numerically constant non-migrant one. That is, the increased clutch size of migrants failed t o compensate for the added mortality resulting from delayed maturity. 7. With the further assumption that the mortality rate in early life (hatching to beginning of C 111) was greater than that during later life (C I11 to adults and their carried eggs), he was able to show that the hypothetical migrant population could increase relative to the nonmigrant one. That is, when mortality rates were relatively small among older animals, the advantage gained through increased clutch size by migrating for part of the day into colder waters was greater than the disadvantage from mortality prior to the delayed maturity. The advantage for the migrant over the non-migrant was greater, for a given temperature decrement experienced by the migrants, when surface waters were cooler; e.g. the advantage for a 5°C decrement would be greater if the difference was between 5 and 10°C than between 10 and 15°C. McLaren (1974) finally indicated that details and some of the assumptions of his model could be relaxed, and concluded “that whatever other values it has, vertical migration in thermally stratified waters may offer important demographic advantages to migrants whose fecundity is increased by development in low temperatures.” We can examine this conclusion against observations of Pseudocalanus. Clearly it must be understood as applying only to females. The fact that young stages of Pseudocalanus in general are less migratory or nonmigratory and often live near the warm surface may indicate that rapid development is advantageous to these more vulnerable stages. The fact that migration may begin noticeably around C I1 or C I11 (e.g. Fig. 36) is suggestive, since this is when temperature begins to affect size (p. 120, Fig. 27). The fact that adult females (which can gain no further increase in clutch size) may, at times, give up migrations when copepodids continue (e.g., May in Fig. 38) or fail t o enter warmer waters at night when older copepodids do so (e.g., p. 167), suggests that at these times a demographic advantage obtains even when an energy bonus (of. previous hypothesis ; possibly useful to full grown migrating females) is not available or useful. Observations

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that migration may be most coherent and vigoroas early in the season (May in Figs 37, 38) may be related to the greater demographic advantages deduced by McLaren (1974) to occur when surface waters are relatively cool. Finally, the observations from Bedford Basin that female copepodids may spend more time in cooler depths than do males (p. 171) suggests that females may be seeking a further advantage (i.e. ultimate clutch size) that males cannot use.

D. Retrospects and prospects Although one of us (McLaren, 1963, 1974) has promoted particular explanations for the adaptive value of vertical migration by Pseudocalanus, our survey of the literature persuades us that no single explanation is likely to apply exclusively. Nevertheless, we stress the profound effect that temperature has on development and growth of Pseudocalanus and also the clear responses of the copepods to seasonal and vertical differences in temperature. We conclude that temperature must play an important role in the consequences of vertical migration, whatever its primary value to the animal. Unfortunately, the observational evidence on vertical migration of Pseudocalanus, and indeed of most zooplankters that have been studied, is deficient. Collections have been made over 24 h periods from populations that were clearly changing with the flow of water masses. Horizontal samples, even when quite closely spaced, may have missed concentrations a t some depths and times. Important variables (sometimes temperature, sometimes feeding status, etc.) have been ignored in most studies. Stages and sexes have been grouped for analysis or illustration. We suggest that a carefully planned seasonal study of vertical distributions of Pseudocalanus would be amply rewarding. This should preferably be done in a stable, landlocked setting at temperate latitudes, using techniques to sample the entire water column in narrow strata (e.g. a pump or comparably continuous collector). At least temperature, food supply and predators should be monitored. A very thorough accounting of all stages should be made-all stages and sexes measured, fat contents and gut contents noted, etc. I n short, we need the sort of laborious “classical” work that seldom attracts the current generation of marine biologists. XIII. PRODUCTION A. General methods By production of Pseudocalanus we mean the total amount of organic matter converted by a population from its food organisms and

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made available for other organisms as food. This is expressed per unit of time under a surface area or in a volume of water. Production occurs, however, whether other organisms use it or not ;a population of Pseudocalanus might, for example, die of salinity shock and decay chemically, yet still have produced organic matter. Thus the concept of production, as currently understood, differs from " yield ", such as might be obtained by man from a fish population. Production by immature stages of Pseudocalanus is obviously a result of growth of somatic tissues. Since adults do not grow in body size (although females may increase their store of oil), all their production must be gonadal. This may be unimportant in males, but clearly production of eggs by females must be included with somatic production by younger stages for a complete estimate of production by Pseudocalanus. Methods for estimating production of copepods are becoming fairly well established. Those that have been applied to Pseudocalanus are of three basic sorts, which are described at length, for example, in Winberg (1971). 1. If life cycles are more or less synchronous (as with annual cycles at high latitudes) it is possible to follow a cohort (a brood or a generation of Pseudocalanus, see p. 135) as its individuals grow in body size and diminish in numbers. The average numbers of individuals present during a time interval (the shorter the better) is multiplied by the average weight of organic matter by which each individual increases during the interval to give the production of somatic matter during the interval. To this must be added the weight of eggs produced during the interval by the non-growing adults. Sometimes " elimination ') (i.e. yield to predators or death and decay) is estimated from the numbers of individuals lost from the population during the time interval and the mean weight of each such individuals. Over the entire life of a cohort, somatic production should be the same as total elimination (except for small errors of integration). However, over a shorter portion of the cohort's life, elimination does not equal production. Furthermore, since increases in mean body size of a cohort can be estimated from sequential samples even when estimates of population decreases are inaccurate, it is generally better to estimate somatic production by multiplying mean number times the increase in weight during an interval. I n general, this " cohort method ') of estimating somatic production is widely accepted, but rarely applicable. Sometimes, as we shall see, it has been applied very crudely to Pseudocalanw, with nevertheless useful results. 2. If life cycles of Pseudocalanus are overlapping and recruitment of young is continuous, so that it is not possible to distinguish cohorts of

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individuals and follow their growth during the season, then the cohort method cannot be used to estimate somatic production. Instead we must use some indirect measure of the rate of growth of each developmental stage in the population, together with rate of egg production, applied with faith or evidence to the natural situation. For Pseudocalanus, growth rates used in production estimates have been based on durations of stages in conditions of excess food in the laboratory. We have seen how durations of stages in these conditions together with weight increments between stages can be used to estimate growth rates as percentages of body weights per time unit (p. 131). Such percentages can be used as production/biomass coefficients (or P/B ratios) which, when multiplied by the biomass (i.e. sum of body weights) of a particular nauplius or copepodid stage, give somatic production per time unit. Rate of egg production can also be expressed as a percentage of weight of a female per time unit (p. 131), thus giving an estimate of the P/B ratio for the biomass of females in nature. Soviet and eastern European investigators, using various groupings of stages and approximations of body sizes and temperature effects (as described in Winberg, 1971), have been particularly vigorous in estimating production of Pseudocalanus and other copepods from P/B ratios based on laboratory estimates of growth rates and egg production. Unfortunately, their detailed assumptions and methods are rarely explicitly stated in their published papers. Some authors have attempted to estimate production of Pseudocalanus using durations of stages or even whole generations to calculate “ turnover times ’ I . Although in principle this approach can give the same results as achieved by estimating growth rates and P/B ratios, i t can lead to conceptual problems, as discussed in Winberg (1971). 3. Production by Pseudocalanus has been estimated by a number of methods, including the so-called “ physiological method ” (Winberg, 1971), that involve use in one guise or another of the “balance equation ” of growth (see p. 132). I n essence, the rates of ingestion, assimilation, respiration and sometimes excretion are estimated experimentally (or often from general metabolic equations) to give estimates of production of organic matter by individuals (whether this appears as growth or eggs) by difference. These estimates can be used directly in P / B ratios. As we have stressed earlier (p. 133), any use of the “ balance equation ” is tenuous at best. However, to the extent that the components of the “ balance equation ’’ can be estimated from experiments on populations in the ‘field, assumptions that laboratory growth rates apply in nature might be avoided. I n the following section we will describe all the attempts known to us

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to estimate the production of Pseudocalunus in nature. We shall pay particular attention to the methods that have been used, with reference to the general approaches that we have described above and to the observations on growth of Pseudocalanus that we have reviewed elsewhere.

B. Production estimates 1. Oguc Lake, Bafin Island

The only attempts to estimate production rates of Pseudoculunw using the cohort method appear to be those of McLaren (1969). He used estimates of dry weights and carbon contents as described in page 125. In Ogac Lake, as we have already seen (p. 138, Fig. 28B), the individual cohorts (broods in this case) could be traced with great assurance, showing decrease in abundance and increase in weights of individuals through the season. Weights of clutches that gave rise t o each brood could also be estimated (p. 128). Although cohorts could almost always be readily separated, there were a few small problems in interpretation. For example, occasional greater mortality of older individuals within a brood gave negative production estimates that were taken as zero, not negative. Although there may be small errors in McLaren’s estimates of production, the cohort method involves no ussumptions about rates of growth, so that his overall estimates are probably quite accurate. Overall production in carbon (including 10-20 mg/m2 during the winter period, that could not be estimated exactly) was about 400, 500 and 350 mg C/m2 in the outer, middle and inner basins of Ogac Lake in 1957. I n 1962, about 430 mg/m2 was produced in the middle basin, between 8 June and 27 August. I n an experiment in 1962 with a fertilized polyethylene column in the middle basin, production was about 710 mgC/m2,or about 1.7 times as high as in the lake. (McLaren, 1969, p. 1509, gives this as 1.9 in error.) Production in 1957 had virtually ceased by September, and the production by overwintered copepodids at the beginning of the seasons only contributed an estimated 1-13% of production in the 1957 and 1962 seasons. Eggs were a large factor (see p. 96), supplying between 26 and 45% of overall production in the various basins and during the two years. Clearly any estimates that ignore such production of eggs by adult females will be in error. The growth of the various summer cohorts made up the remaining production. Because animals became bigger as the season progressed, production was generally sustained through the summer, even though growth was slow and populations greatly reduced by mortality.

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2. BlackSea

Soviet investigators have estimated production rates of a number of Black Sea copepods from body weights and durations of stages established in laboratory studies. Greze et al. (1968)have carried out such calculation for Pseudocalanus in the waters off Sevastopol during at least some of the four seasons for each year from 1960 to 1966. The biomasses of Pseudocalanus tabulated by them varied up to %fold seasonally within years and up to 7-fold between years within the same season. However, their tabulated P/B ratios varied much less: the largest is given as 0.203 for autumn 1960, and the smallest as 0.104 for winter 1965 (from their Table 3, wrongly labelled as " biomasses '' in the English edition). The exact methods of calculation used for these estimates are not described. However, reference is made to standard tables of wet weight of each developmental stage and to the work of Sazhina (1968)which, as we have noted (p. lll), gives acceptable estimates of development times for nauplii and copepodids at 8-10°C. We are not told how egg production might have been incorporated, although estimates of the size and frequency of clutches are also given by Sazhina (1968). The daily P/Bratio were probably calculated as a daily percentage of growth of mean body weights of individuals grouped as nauplii and copepodids in the manner outlined in Winberg (1971).The ratios are said to have been corrected for seasonal differences in temperature, possibly with '' Krogh's normal curve ", commonly used in Soviet work (see Winberg, 1971). The overall mean estimates of daily P/B for each season in Black Sea Pseudocalanus differ little from one another : 0.143 for January to March at 7~8°C;0.173 for 1 April to 20 June a t 12.3"C; 0.159 for 20 June to 1 October at 21.4"C (temperature probably too high for deep living Pseudocalanus at this season-see p. 166); 0.174 €or October to December at 14.1"C. (Greze, 1973, gives seasonal estimates based on the same data that differ slightly for reasons that we cannot determine.) These P/B ratios in turn differ little from the estimates of growth rate in dry weight per day calculated for older Black Sea animals (see p. 130) of l6-18% per day at temperatures of 8 to 15°C. Thus, we believe that the daily P/B ratios in Greze et al. (1968)are in keeping with what we have learned about Pseudocalanus in our review. Conover (1974),assuming carbon to be 5% of wet weight:and a 40 m water-column, rendered the estimates of Greze et al. (1968)for Pseudocalanus as 2.57 mg C/m2/day, or 939 mg C/m2/yr. An estimate for the " neritic zone " of the Black Sea off Sevastopol of 0.32 mg dry wt/mS/ day is given by Greze (1970). Assuming this to be 50% carbon and in

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40 m water column gives rates about twice as high as those estimated by Conover (1974). Conover’s estimate is in turn twice that for Ogac Lake (above). However, Pseudocalanus is one of only two common copepods in Ogac Lake, whereas several other species in the Black Sea contribute to an overall annual production of copepods in the upper waters (using Conover’s conversions for Greze et al., 1968) of some 3,360 mgC/m2/yr,some 4-5 times that of Ogac Lake (about 750 mgC/m2/yr, from McLaren, 1969). Porumb (1972) has estimated production of Pseudocalanus in the upper 50 m of the coastal waters (out to 50 nautical miles offshore) off Romania. Again, she does not detail methods, but refers to those used by Greze et al. (1968) and the references therein. She tabulates mean standing crops, P/B coefficients, the production by months, seasons and the year for the period July 1970 to June 1971. From her annual P/B coefficients it can be inferred that the daily P/B ratio was about 0.089 (32.796/365), rather lower than those estimated by Greze et al. (1968). Assuming with Conover (1974) that carbon is 5% of wet weight, and that Porumb’s estimates of production apply to the upper 50 m, a value of production of 313 mgC/m2/yr can be calculated for Pseudocalanus. This is rather lower than that given above by Greze et al. (1968) for the waters off Sevastopol. Because of the reasonably close resemblance of the P/B ratios used in these Black Sea studies to growth rates of Pseudocalanus that we have calculated for other localities (see above), the production estimates may be reasonable. However, i t must be stressed that they are only acceptable if the Black Sea populations are at times developing at temperature-dependent rates, with no food shortages or “ resting ” phases. This may be plausible for the Black Sea region, but we have seen no published evidence to support the assumptions. I n fact, in the work of Sazhina (1971) we are informed that the period of (abundant?) occurrence of Black Sea P s e u d o c u l u n ~is 180 days from October to March. 3. Sea of J a p a n

Shushkina et al. (1974) have estimated production of Pseudocalanwr C 111-IV from an experiment with radiocarbon tagging of food algae in a mixed population of herbivorous and carnivorous zooplankton in large, unconcentrated samples from the northwestern Sea of Japan. From an elaborate model of the food web €or this group of zooplanktonic species and the distributions of radioactivity among them following the experiment they deduce (by computer solution) a daily P/B ratio of 0.18 1 for C 111-IV Pseudocalanus.

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It is not possible to discover all details of their methods and calculations, but this result compares quite well with estimates based on the assumption that copepods grow at maximal, temperature-dependent rates (see Black Sea and other estimates). This comparison is not made by Shushkina et al. (1974). Rather, they compare their results with those from the " physiological method " as outlined in Winberg (1971), assuming growth efficiencies of 0-4 and 0.3, and deriving estimates of daily P/B ratios of 0.23 and 0.36 respectively. As pointed out earlier, this method is unlikely to be very accurate. Andreeva (197Gb) has evidently estimated production of Pseudocalanus in the northern Sea of Japan using durations of stages in the laboratory (but see p. 111) and body weights. I n her preliminary account, she merely notes that P/B coefficients are similar to those for the Black Sea. North Xea There are two recent, independent estimates of production for Pseudocalanus from the North Sea. Although both are based on the assumption that there is more-or-less continuous development of generations in the areas studied, the approaches and assumptions used are quite different. Evans (1977) has used a statistical means of separating successive generations of adult female Pseudocalanus in samples from Northumberland coastal waters. We have already indicated (p. 147) that this approach may not have distinguished generations during summer when temperatures were high and changing slowly. However, Evans uses this method to indicate the turnover times of adult females during the period between spring 1971 and spring 1972. I n order to estimate production, Evans used weights of copepodids and adults as calculated with Robertson's (1968) weight-length equation. Although this equation is inaccurate outside rather narrow length limits (p. 127)) errors from this source are probably not too serious. Weights of nauplii were estimated by assuming that the same growth increments apply to them as to copepodids, with due consideration of shape differences between nauplii and copepodids. Two estimates of production are made by Evans. The fist simply sums the weights of adult females (means of samples) considered to represent each generation. The weights for each generation are then summed for the entire sampling period. As Evans points out, this estimate of production will be lower than overall production. The second method assumes that the populations are eliminated entirely between each designated generation and a '' more probable production figure is obtained by summing the standing stock of 4.

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juveniles as well as adults ”. Although Evans notes that the possibility of extra generations (the adult females not statistically separated) means that his production estimates would be low, his second method will certainly give underestimates for other reasons as well. Essentially it is an approximate ‘‘ turnover time ” method, in which biomass is divided by the time (generation time) in which it is replaced. It will give a somewhat low approximation of the production to date of a particular generation from time of its origin (with a very low biomass taken by Evans as zero), to the time of sampling (when all biomass is counted as production). Also, it fails to account for production subsequent to sampling and prior to the replacement of the generation. It also does not include egg production. Estimates by Evans using his second method indicate that Pseudocalanus had the highest production of five copepod species off the Northumberland coast, about 47.9 mg dry wt/m3/yr out of 143-4 mg for the five species combined. Production of Pseudocalanus in the 50 m water column sampled by Evans was thus about 1 197 mg C/m2/yr (assuming carbon is 50% of dry wt, see p. 126). This (together with an estimate for all five copepods of 3 585 mg C/m2/yr)is similar to estimates €or the Black Sea, but is probably substantially too low if the number of sources of error are taken into account. Recently Thompson (1976) has estimated the abundance of various stages of Pseudocalanus during a series of sampling periods between 12 December 1967 and 3 June 1968 over a wide area of the southern North Sea. She makes use of her extensive laboratory data (see p. 108, Fig. 25B) to convert numbers in each stage to production of num.bers using the relationship : No./m2 of stage during sampling period = No. produced/m2/day. stage duration in days

____

This, then, is a refined “turnover time ” method in which each stage was treated separately. Her estimates of stage dura,tionsas functions of temperature are based on regressions that give slightly different results from those in Fig. 25B. To convert production of numbers to production of biomass, Thompson used lengths at experimental temperatures and a formula linking wet body weight of copepodids and adults of copepods to total body length from Kamshilov (1951). In this way, she estimated that an average of some 280 >< 108 mg wet weightlday of copepodid and adult Pseudocalanus were produced over the study area in the southern North Sea during the winter-spring sampling period. From her estimate of the area sampled (30640 km2) this is about 1 mg wet wt/m2/day.

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Since only about 5 % of this would be carbon (see p. 183), the implied production rates seem much lower than those for other localities. Although Thompson's actual measurements do not include egg production and probably contain small weight errors, they represent an admirable attempt to combine production estimates based on laboratory growth data with a sampling programme on a sufficiently large scale to be meaningful for a region. 5 . Baltic Sea

Production of Pseudocalanus in the Gdansk Deep area of the Baltic was briefly reported in a symposium abstract by Ciszewski and Witek (1975). By methods that are not detailed, they estimate that annual production of Pseudocabnus amounted to 137 g wet wt/m2/yr, about 5% of which would be carbon (see p. 183). This estimate is higher than any we have from other regions. The V/B ratio reported by them is 10-3, which seems similar t o those from elsewhere, if 0.103 on a daily basis is meant. 6. Norwegian Sea

Pavshtiks and Timokhina (1972) summarizing earlier work by Timokhina, attempt to estimate the production of Pseudocalanus and other major zooplankton species for the entire area of the Norwegian Sea. To do so, they depended on samples taken from the upper 500 m from two east-west and one north-south section during 1959-63 and 1968-69. They use the method of Boysen-Jensen (as described in modern terms in Winberg, 1971), which is essentially a simplified version of the cohort method for populations with an annual life cycle. The population loss during a year is estimated from samples a t the beginning and end of the annual cycle, and this is multiplied by the mean biomass per individual between the beginning and end of the cycle. To this is added the biomass not yet dead at the end of the cycle. Pavshtiks and Timokhina, however, gave no details on the ways in which they evaluated numbers or weights. Estimates for the Norwegian Sea as a whole, are given in a table as millions of tons. For Pseudocalanus, these estimates (wet w t ? ) ranged as low as 1.57 x lo6 tons in 1961 and as high as 5.28 x 106 tons in 1962. Pavshtiks and Timokhina point out that the method of evaluation gives minimal estimates, and also that the catching efficiency of their plankton nets was unknown. Nevertheless, we consider the attempt to be of interest in demonstrating that approximations can be made even from rather unpromising material.

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CHRISTOPHER J. COREETT AND IAN A. MOLAREN

7. White Sea

Fedorov et al. (1975) have estimated production of phytophagous copepods, including Pseudocalanus, in the White Sea from a 24 h experiment on 26-27 June 1970, and a 3-day experiment on 18-21 August 1971. During these time periods they made frequent measurements of biomass of phytoplankton and zooplankton species in the 75 m water column and made less frequent estimates of phytoplankton production rates. They followed the approaches of McAllister (1969) to calculate consumption of phytoplankton by zooplankton, using observed phytoplankton production and changes in biomasses of phytoplankton. The rest of their analysis is based on the " balance equation " approach, with coefficients of assimilation and respiration assumed from the literature, evidently with no account of the (low?) prevailing temperatures. They conclude that production by phytophagous zooplankton (of which Pseudocalanus was 26% of the biomass) was 26 mg C/m2/day. The mean P/B ratio was about 0-17 ; the concordance of their ratio with estimates for Pseudocalanus from the Black Sea is of doubtful significance in view of the fact that some of the assumed rates used by Federov et al. are based on Black Sea studies.

C. Retrospects and prospects Although the estimates of production rate in Ogac Lake by McLaren (1969) are probably the least disputable that we have reviewed, the

environment of Ogac Lake is very special and the opportunity for using the cohort method is not available for most of the geographical range of Pseudocalanus. We believe that we have summarized enough information in this review so that production of Pseudocalanus in many localities (some with published data available for analysis) could be estimated with little or no further information, provided that development rate in the population over the period of interest is not limited by food. We have argued in several places in our review that this assumption is probably valid for a substantial part of the year in many temperate localities, and have demonstrated its validity for Loch Striven, Scotland (p. 145) and less completely for a number of other localities (see Section XI). However, the investigator wishing to validate the assumption for any given locality has been given a number of techniques (egg frequency counts, p. 9 8 ; oil sac sizes, p. 129; and especially intensive sampling and analyses of female sizes and perhaps relative number of specific copepodids, p. 143) that might be used. If it can be assumed or demonstrated that food is not limiting, then the

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following methods (including some approximations and “ short-cuts ”) can be used to estimate the parameters of production. (1) We have shown (p. 11 1) that development rates of the various stages as functions of temperature differ little over various parts of the world, so that Fig. 25A or B might do as approximations. (2) We have shown that volume of a clutch of eggs is about the same function of female length in different regions, so that the potential rate of production of egg matter as a per cent of female body weight is a similar function of temperature everywhere (p. 131). (3) A general weight-length relationship for Pseudocalanus copepodids and adults (p. 127) allows weight estimates t o be made from length, if not made directly. (4)Unfortunately there are no direct estimates available of weights of nauplii. Evans (1977) estimated that “ growth factors ” (length multiples between moults) of 1.17-1.28 (mean 1.22, implying a weight increase of about 1-8 times) apply for Pseudocalanus. Since nauplii contribute very little to overall production, McLaren’s (1969) assumption that they double weight between moults, might do. From weights and stage durations, growth rates of stages can readily be calculated, and applied as P/B ratios to counts or biomasses of animals in samples. It should also be possible to construct purely theoretical models of Pseudocalanus production for various regions, since size-temperature relationships can be added to the above information. Although these are local in application (see, e.g. Fig. 26) they should be readily established from a small number of points from experimental or field data. From estimates of growth rates that we have given for various stages and temperatures, approximations might be made for P/B ratios applying to entire samples. For example, we suggested (p. 130) that older copepodids from the North Sea might grow a t about 17% of body wt/day irrespective of temperature. Younger copepodids and nauplii might be more affected by temperature, but generally contribute little to biomass in nature, so that an overall rate of 20% might do at North Sea temperatures. Production by adult females can be reckoned in terms of egg matter, and is more temperature dependent. Overall, a P/B ratio of 0.20 might suffice for the North Sea for some purposes. None of the attempts to estimate production of Pseudocalanus that we have reviewed take into account all the kinds of information alluded to above. However, there remains a more serious problem for the future of such studies. Only Evans (1977) has attempted (with shortcomings that we have noted) to discriminate generations in the field objectively. Some of the other estimates depend on the assumption that food is in excess, and temperature in control. However, we have also described the way in which overwintering, resting stages begin to accumulate

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in the population in some regions while other members of the population continue to produce a t maximal rates (p. 144). I n our opinion, the most important work remaining to be done on the production biology of Pseudocalanus (and perhaps other copepods as well) is in the establishment of the “ growth status ” of individual stages in samples from nature. Does stored fat in young stages and in adults denote full production potential? Does fat above certain levels in C IV denote ‘‘ resting ”? Are there other morphological criteria of growth status? Of course, determining production of Pseudocalanus should not be viewed as an end in itself. We know very little about the impact this production has in nature, and what we do know is the subject of Section XV.

XIV. PARASITES A. Dinoflagellates 1. Blastodinium hyalinum Chatton, 191 1

(a) Taxonomy Apstein (1911) described and recorded this genus as ‘‘ Parasit 1 ” in a number of copepods including Pseudocalanus. Chatton (191 1) made observations on Apstein’s work and writes “ Dans le MBmoire que j’achhve, en ce moment, sur les Phidiniens parasites, je lui ai rhservh le nom de B. hyalinum, n. sp.”. Chatton (1911) does not give a description of this parasite in that paper although he does say it is closely related to B. contortum Chatton. I n his large monograph Chatton (1920) refers to this parasite as B. contortum hyalinum which differs from B. contortum in lacking torsion and pigmentation ; when removed from the host gut and left in Bea water this variety dies earlier than B. contortum since it is less resistant. Sewell (1951) from extensive study concluded that Chatton (1920) confused more than one species under the name B. contortum hyalinum. Sewell refers to the parasite discussed here as B. hyalinum Chatton, and confines this species to the form found in the North Sea, which was described but not named by Apstein (1911) and to the forms Chatton (1920) recorded from the Mediterranean. We adopt here the view of Sewell and refer to this parasite as B . hyalinum Chatton. I n addition Sewell (1951) examined a number of specimens of Pseudocalanus collected from the North Sea and confirmed that B. hyalinum infected

Pseudocalanw.

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(b) Life history The parasite lies within the alimentary canal of Pseudocalanus and initially consists of a single cell, the trophocyte or trophozoite (Fig. 39A). Transverse or oblique division gives two daughter cells, the anterior (i.e. nearly always at the anterior end of the host) becoming Parasite

/

Gut

Secondary trophozoite

I

0

Tertiary trophozoite

C

Secondary layer of sporocytes

I I

A

First layer of sporocytes

/

F m t loyer o f sporocytes

I

Fro. 39. The dinoflagellateparasit,e Blastodinium hyalinum. A, parasite in the gut of a calanoid copepod. B, monoblastic stage from Pseudoculun~. C, diploblastic stage. D, polyblastic stage. (A after Chatton. 1920; B-D after Sewell, 1961.)

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

a secondary trophocyte and the posterior a gonocyte. The gonocyte forms a large number of sporocytes by repeated division (Fig. 39B). The monoblastic stage in the life cycle occurs when the secondary trophocyte is surrounded by one layer of sporocytes on all sides, apart from a distinct gap which may be present and is called by Chatton (1920) the hilum (Fig. 39D). The secondary trophocyte divides into an anterior tertiary trophocyte and a posterior gonocyte, which in turn undergoes repeated divisions to give a second layer of sporocytes lying within the first layer ; this is the diploblastic stage (Fig. 39C). The polyblastic stage is formed when the trophocyte continues to divide, ultimately into a number of layers of sporocytes (Fig. 39D). I n the genus Blastodinium the trophocyte may not divide into a trophocyte and a gonocyte, but into two trophocytes (i.e. by schizogony), and each of these trophocytes may subsequently undergo division into more trophocytes and gonocytes (Sewell, 1951). The end result is a single primary layer of sporocytes enclosing two separate layers of secondary sporocytes, each enclosing a trophocyte. Usually the release of sporocytes from the host sets the two daughter trophocytes free within the alimentary canal to develop into separate parasites. Schizogony enables more than one parasite to infect a host and hence gives rise to a larger number of sporocytes than would have been possible without schizogony. Chatton (1920) and Sewell (1951) agree that two or more individual parasites in a host have likely arisen by schizogony. Sewell does not state expIicitly that schizogony occurs in B. hyalinum inside Pseudocalanus but does give a table (his p. 329) in which size measurements of parasites are given for four cases of “double infections ” and one “ triple infection ” of B. hyalinurn from the North Sea, from which it may be inferred that B. hyalinum can undergo schizogony in Pseudocalanus. The rupture of the cuticle surrounding the parasite sets the sporocytes free into the alimentary canal of the host. The sporocytes are small immobile cells with two nuclei (Fig. 40A). Sometime after expulsion from the anus and after an unknown number of divisions they form dinospores (Fig. 40B),with 2-4 flagella attached in the region of the equatorial groove. Under unfavourable conditions the dinospores are able to encyst and host infection presumably takes place by ingestion of the dinospores or cysts (Fig. 40C)with the host’s food. (c) Eflect of infection on the host Chatton (1920) found that in host individuals parasitized with Blastodinium the gonad was immature and the genital ducts undeveloped. He also noted that he had never seen a parasitized male.

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Cattley (1948) examined a number of Pseudocalanus from the North Sea, parasitized with 3. hyalinum, and found very distinct changes in the fifth pair of legs. He concluded that in the male the parasite is able to arrest the development of the male characters of the copepodid (externally, the fifth pair of legs), while in the final moult it causes the copepodid to undergo sex reversal and appear externally as a mature but sterile female ; he found spermatophores on three such individuals.

Transverse groove

/

Transverse

flogello

Longitutlinal groove

FIQ.40. The sporocytes (A), dinospore (B),and (C), a cyst of Blastodinium hyaliizurn, a parasite of Pseudocalanus. (After Chatton, 1920.)

Cattley believed that the parasite had no effect on the morphology of female hosts and that the C V moulted into a fully formed but sterile individual. Sewell (1951) examined a large number of individuals from several host species (not Pseudocalanus) infected with 3lastodinium and found both sexes infected with the parasite. In none of the infected males (usually C V, rarely adults) could Sewell detect any change of structure from the normal. I n females, a late infection evidently only partly reduces the development of the ovary and oviduct whereas an early infection has more profound effects. The first two segments of the A.M.B.-15

6

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CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

urosome may not fuse to form the genital segment, as occurs in a normal female (p. 34), but remain, separate as five segments, resembling the male condition. Even with fusion, the genital operculum may not develop, and there may develop a fifth pair of legs similar to that found in the male C V. Sewell (1951), therefore, was of the opinion that intersex individuals in copepods are modified females and not male individuals that have undergone partial or complete sex reversal as proposed by Cattley (1948) for Pseudocalanus. Sewell (1951) makes the further observation that those individuals in which a modified fifth pair of legs is present and which in all other characters appear to be females, but in which no parasite has been found, result from an early infection from which the individual has later recovered.

(d) Occurrence Blastodinium hyalinum is widespread (see ranges in above account) in the most wide ranging study (with Continuous Plankton Recorder). Vane (1952) states that it occurred in 3.8-60% of individual Pseudocalanus (mainly C V and adult females) in samples from the North Sea in 1948-49. It was most common in July-August, especially in the central part of the North Sea, minimal in December-March. 2. Dissodinium pseudocalani Drebes, 1969

(a) Taxonomy Dissodinium pseudocalani is a parasitic dinoflagellate found by Drebes (1969). The reproduction of this species resembles that of D . lunula and so the parasite was provisionally put in this genus. D. pseudocalani has thus far only been observed as an ectoparasite on the eggs of Pseudocalanus. Drebes (1972) subsequently indicated that his Dissodinium pseudocalani Drebes, 1969, is a synonym of Sporodinium pseudocalani Gbnnert, 1936 (see p. 196), and suggested that fwrther nomenclature changes will be made after he has concluded his revision of a few dinoflagellate genera. We accept the possibility that the two species discussed above are synonyms but we prefer here to give an account of the two forms separately, particularly since Sournia, et al. (1975) comment that the two parasitic species seem distinct. (b) f i f e history The life cycle includes a free drifting phase unattached to any host and an ectoparasitic phase on Pseudocalanus eggs. Mature primary cysts (Fig. 41A) usually drift unattached to the

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host. They are spherical or oval, between 150-250 pm, and contain green or orange protoplasm, which is the same colour as the host’s eggs. The protoplasm invaginates in one region (Fig. 41B) and the nucleus divides mitotically to form nuclei on the periphery of the

FIG.41. Diagrammatic representation of the life cycle of Diasodinium pseudocalani, a parasite of Pseudocalanus. A, primary cyst in plankton. B, multinucleate protoplasm, invaginating. C, division into 16 cells. D, development into 16 secondary cysts. E, secondary cysts forming dinospores. F, liberated motile dinospores. G, infeotion of host egg. H, growth of mature trophont. I, the trophont still attached to egg membrane of host. (After Drebes, 1969.)

b

protoplasm, which then divides to form, generally, 16 segments (Fig. 41C),but sometimes 8 or 32. Secondary cysts (Fig. 41D) are formed by the rounding off of the 16 segments formed in the primary cyst; these become oval and about 76 x 47 pm.

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CHRISTOPHER J. CORKETT AND IAN A. MULAREN

Sporulation occurs when the secondary cysts divide to form 16 or 32 flagellated dinospores (Fig. 41E), which are colourless swarmers of the Gyrodinium type (Fig. 41F). Host infection occurs when a dinospore attaches itself to the surface of a host egg (Fig. 41G) and sucks out the contents. The trophont develops as an ectoparasite on the egg, then generally separates and continues its development in a primary cyst in the plankton (Fig, 41H, I). Later developmental stages were occasionally seen with the parasite still attached to the host egg membrane (Fig. 411). (c) Occurrence The parasite appears in the North Sea off Heligoland in the German Bight from late April until early June and is fairly common in the second half of May. The reason for this marked seasonality is not known. 3. Sporodinium pseudocalani Gbnnert, 1936 This parasite was found as a free cyst in the plankton off Heligoland in April 1934. At this time Pseudocalanw was the most numerous zooplanktonic species and one parasitic cyst was found in a clutch of eggs of Pseudocalanus. Gbnnert (1936) was unable to elucidate the complete life cycle, but described the formation of sporoblasts ( = primary cysts) which develop into sporocysts ( = secondary cysts) containing dinoflagellate-type nuclei. As Drebes (1972) observed, Gbnnert (1936) confused the membrane of the primary cyst with the egg wall of the host and therefore concluded that the dinoflagellatewas endoparasitic, although he did not rule out completely the possibility of ectoparasitism . This species is probably synonymous with Dissodinium pseudocalani Drebes, 1969, the life history of which has been described in detail above. 4. Ellobiopsis chattoni Caullery, 1910 Ellobiopsis chattoni is an external parasite of pelagic copepods first described by Caullery (19lo), who provisionally considered it to be a peridinian dinoflagellate, although no developmental stages or dinospores were observed. Apstein (1911, " Parasit 19 ") recorded an external parasite of Pseudocalanus which was identified by Jepps (1937a)as Ellobiopsis. Specimens of Pseudocalanus with Ellobiopsis have been recorded subsequently from the Gulf of St. Lawrence (Pinhey, 1927), Loch Striven (Marshall, 1949), the southern Norwegian Sea (Hansen, 1960) and off Ireland (Fives, 1969). Wing (1975) found a low level of

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infection of Pseudocalanw by Ellobiopsis chattoni in southeastern Alaska. Throughout the year only 1-6 Paeudomlanus were found to be infected each month from many thousands of potential hosts that were examined. Wing found no distinct seasonal trend of infection. The systematic position of this parasite has been disputed. Jepps (1937b) considered that the available evidence suggested a fungus relationship, but a t present the whole group is considered as belonging to the dinoflagellates by Loeblich (1976), and this is the view adopted here. An account of Ellobiopsis chattoni on Calanus finmarchicus is given by Jepps (1937b). It is assumed that spores formed by sporulation are responsible for infection of new hosts, although this process has not been observed.

B. Qregarines Apstein (1911, " Parasit 3 ") recorded gregarines in Pseudocalanus, and Jepps (1937b)observed them in the gut of Calanw. These parasites have not been studied in detail in copepod hosts, but the young parasite (sporozoite) becomes typically intracellular as it grows. It then leaves the host cell and the mature trophozoites adhere externally to the digestive lining. The trophozoites fuse in pairs (syzygy, observed in Calanus by Jepps, 1937b) and ultimately produce young sporozoites.

C. Trematodes Giesbrecht (1882) reported that Pseudocalanus was the most frequent host of a trematode, probably Hemiurw, that was also found free-living in Kiel Harbour. Entry into the host (later copepodids and adults) was between two thoracic segments or between cephalothorax and urosome, using the tapered posterior end for penetration. Other references to trematodes (probably H . appendiczclatus according to Thompson, 1976) in Pseudocalanzcs are Canu (1892), Apstein (1911), Wright (1907) and Marshall et al. (1934). Trematodes (or cestodes) were considered by Fives (1969)to be responsible for the red colouration often found in Pseudocalanus. Thompson (1976) concludes that these parasites do not have any detrimental effect on the copepod, merely acting as an intermediate host, the final hosts being fishes that feed on copepods.

D. Nematodes Apstein (1911, " Parasit 17 ") observed nematodes in Pseudocalanw, and Marshall and Orr (1955) reported them in Calanw as Contracaecum

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sp. The eggs of Contracaecum from the faeces of definitive hosts (birds or sea mammals) develop into larvae that may be eaten by copepods and subsequently by second intermediate hosts (Huizinga, 1966). The genus Thynnascaris parasitizes fish as adults and its larvae occur in fish and invertebrates. Popova and Valter (1965) and Valter (1968) have completed experimental studies on the intermediate hosts of Contracaecum aduncum (= Thynnascaris aduncum according to Norris and Overstreet, 1976) ; eight copepod species were subjected to experimental infection with larvae and Pseudocalanus was the second most infected copepod species (26% became infected). The larvae remained in the body cavity of the copepod where they increased in size (Popova and Valter, 1965).

E. Crustaceans All epicaridean isopods parasitize crustaceans and feed on blood. They undergo a marked metamorphosis in their life cycle (see Kaestner,

PIa. 42. Two mioronisoium larvae of an epicaridean isopod parasite, on Pseudocdanue. (From Sam, 1899.)

1970). The young leave the mother as pelagic epicaridean larvae and survive for a period on stored yolk before attacking a pelagic copepod or other host and transforming into a parasitic microniscium. After several moults on the host, its appendages become reduced and it becomes a free-swimming cryptoniscium stage, which then seeks out a final crustacean host. Sars (1899) gives an account of two microniscia on a female Pseudocalanus a t two different stages of development (Pig. 42), and Marshall (1949) found a microniscium on a female PseudoCaZanus, but nothing is known about final hosts of these parasites.

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.!I Retrospects and prospects

Our knowledge of the parasites of Pseudocalanus (as for marine parasites in general) is fragmentary, and advances will probably have to depend on the efforts of specialists working on the parasitic groups, rather than on the incidental findings of copepodologists. Although some of the parasites are fatal or sexually sterilizing to their hosts, their impact on Pseudocalanus populations is thought to be small (e.g. by Vane, 1952, and Wing, 1975). However, those parasites that use the abundant Pseudocalanus as intermediate hosts could have substantial impacts on less common final hosts. XV. ROLEIN THE FOODWEB We have indicated near the beginning of our review (p. 17) that Pseudocalanus is one of the most widespread and abundant metazoans in the world. Given this status, it is certain to be of substantial importance in the lives of other organisms. Here we review briefly the role of Pseudocalanus as a consumer, as a source of nutrients and as food for others (especially larval fishes), and discuss the first hesitant attempts to include Pseudocalams in descriptive and predictive models of marine food webs.

A. Effect on phytoplankton 1. Peeding on phytoplankton

The feeding of herbivorous copepods clearly removes phytoplankton from the water column, but evidently the impact of Pseudocalanus on its food species has not been fully assessed. Zagorodnyaya (1977) estimates the fraction of edible biomass of phytoplankton in the Black Sea removed by Pseudocalanus during two times of year. To do so she calculated daily rations of copepodids and adults, using the “balance equation” approach (see p. 132). She concludes that these animals removed about 18% of the standing crop daily (compared with 11 yoby Calanzis hegolandicus) in inshore waters in spring, and 8% (compared with 40% by C. helgolandicus) in offshore waters in winter. As these conclusions are local in implication, we feel that the following general account is useful. A population of Pseudocalanus growing or producing eggs a t about 15-20% of its biomass per day (pp. 130,131) might need a minimum of about four times this amount of food (p. 133), which it might obtain from food concentrations as low as 25 pgC/I. (p. 130). If we can imagine

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that such an unconcentrated phytoplankton biomass could, if ungrazed, double its quantity each day, then a population of 3 or 4 adult female Pseudocalanus per litre, each weighing about 10 pgC, could keep the plant population in check. However, we have shown that copepodids can consume up to 140% of their weight per day at phytoplankton concentrations of 200 pg C/1. (p. 67). Since growth rates do not increase with ration above at most 50 pgC/l. (p. 130, despite equivocal analyses from growth efficiencies, p. 133), this seems to us to indicate that " superfluous feeding " by Pseudocalanus could occur at phytoplankton levels substantially below the 390 pgC/l. suggested by Beklemishev (1962) for copepods. Possibly, however, die1 feeding cycles (p. 72) and vertical migration (p. 165) reduce this " wastage " in nature. The faecal pellets produced by Pseudocalanus (whether with superfluous food or not) may carry material below the photic zone. The pellets produced by Pseudocalanus are of the order of 106 to 3 x 106 pm3 (from Corkett, 1966 ; Martens, 1972), and these might sink at up to 100 m/day (Fig. 1 in Smayda, 1969). 2. flupply of nutrients

Although Pseudocalanus removes organic matter from the water column, it also resupplies nutrients to the phytoplankton. We have concluded that rates of excretion of nutrients by Pseudocalanus are strongly dependent on food concentrations (p. 51). Evidently the only estimates of the possible contributions of excretion by Pseudocalanus to phytoplankton requirements come from two studies in Bras d'Or Lake, a landlocked arm of the Atlantic in Nova Scotia. Here Christiansen (1968), using rates of NH3 excretion described elsewhere (p. 44), estimated from population densities that Pseudocalanus supplied about two-thirds of the N excreted by the copepod community, but that this was only about 4.8 mg N/m2/day, which was about 4% of the daily phytoplankton requirements of about 118 mg N/m2/day. I n the same environment, Hargrave and Geen (1968) calculated that the phytoplankton needed about 7.5 mg P/m2/day. From estimates of population densities and excretion rates of P (see p. 49), they suggested that about 15 mg P/m2/day could be supplied by copepods, sometimes predominantly Pseudocalanus. Although they note possible sources of error, they conclude that regeneration of P (unlike that of N, see above) by copepods like Pseudocalanus is important.

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B . Predators I. Fishes There are many records of the occurrence of Pseudocalanus in the diets of fishes, and we cannot refer to them all. Rather, we attempt to outline the possible importance of Pseudocalanus to numerically and commercially important fish species. Where possible we refer to recent, more general studies of diets in which Pseudocalanus figures. We also attempt to give geographical balance in our survey. Although Pseudocalanw occurs in diets of some postlarval fishes, it is much more important as a food for larval fishes. Most marine fishes have very high fecundities that are balanced by high mortalities. Most of this mortality occurs during the larval stage, as was first stressed by Hjort (1914). May (1974) restates ‘ I Hjort’s critical period concept ’’ as a concept that maintains that the strength of a year-class is determined by the availability of planktonic food shortly after the larval yolk has been exhausted ”. May concludes that field and laboratory evidence, although often circumstantial, indicate that starvation is indeed an important cause of larval mortality when yolk has been exhausted, as Hjort hypothesized. I n order to signify the relative commercial importance of each species of fish considered here, we quote the most recent available fishery statistics (P.A.O., 1974) on the nominal catch for 1973.

(a) Salmon Salmon as diadromous fishes breed in fresh water and their young may migrate to the sea at various stages of development. Among them, the pink (= humpback) salmon (Oncorhynchus gorbuscha) goes to sea as plankton-feeding fry. It produced a catch in the northeast Pacific of 34 x lo3 metric tons in 1973. Young pink salmon (36-104 mm) off southern Hokkaido have been shown to contain Pseudocalanus in their stomachs (Okada and Taniguchi, 1971). Parsons and LeBrasseur (1970) showed in the laboratory that young pink salmon 90 mm long fed best off Calanus plumchrw and less well off the smaller Pseudocalanus. I n the Strait of Georgia, however, it appeared that pink salmon less than 30 mm long fed best on smaller copepoda, such as Pseudocalanus (LeBrasseur et at., 1969). The chum (= keta or dog) salmon (0.keta) is an important food fish in Japan and North America, with a catch of 126 x lo3 metric tons in 1973. Okada and Taniguchi (1971) found Pseudocalanus in the guts of juveniles, and LeBrasseur et al. (€969)showed that such small copepods are an important food source.

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(b) Herring The Atlantic herring (Clupea harengus) yields one of the world's great fisheries, with a total catch in the Atlantic of some 1 956 x lo3 metric tons in 1973. Hardy's (1924) classical study in the North Sea showed that Pseudocalanus was numerically 86% of the food of herring between 12 and 42 mm. The importance for herring of food availability after the yolk sac has been absorbed has been investigated by Blaxter (1 963). He related the dimensions of food organisms to the maximum gape of the jaws, which is just sufficient to take an adult Pseudocalanus " end on "when larvae reach 12 mm. He found that Pseudocalanus were not generally taken until the larvae reached 12 mm, and that Calanzls was not taken until the larvae were 30 mm long. He concluded, however, that Calanus were rarely taken by postlarval fish, and then only when the copepods are in younger stages, but that Pseudocalanus is of major importance. Legare and Maclellan (1960) have carried out the most extensive study of herring feeding in relation to zooplankton in the western North Atlantic. They found that Pseudocalanus was the second most abundant copepod in the region of the mouth of the Bay of Fundy. It was third in incidence in stomachs from within Passamoquoddy Bay and, along with Calanus, even more common in stomachs from outside the bay, especially in fish longer than 200 mm. The Pacific herring (Clupea harengus pallmi) produced a catch of some 539 x lo3 metric tons in the North Pacific in 1973. Lowe (1936) investigated food in various sized herring from off southern British Columbia. For fish 9-12 mm long, the most important food items were small eggs and nauplii. Pseudocalanus was present in guts of all larvae longer than 13 mm. (c) Atlantic mackerel . The mackerel (Scomber scombrus) produced some 1 017 x lo3 metric tons in the Atlantic and 11 x lo3 metric tons in the Mediterranean and Black Seas in 1973. Bullen (1908) found that PseudocaZanus was common in stomachs of postlarval mackerel in the English Channel, especially in the month of May. Among " fishermen's signs " was one stating that mackerel are abundant in " yellow water which seldom appears before the last week in April. Bullen showed that phytoplankton was almost absent from " yellow water ", but that copepods were abundant, especially Calanus and Pseudocalanzls. )),

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(d) European pilchard The pilchard or sardine (Sardina pilchardus) makes up several important fisheries. I n 1973, Portugal took 101 x 103 out of a total of 177 x 103 metric tons from the northeastern North Atlantic, probably largely outside the range of Pseudocalanus. There is also an important fishery in the Mediterranean, of some 144 x lo3 metric tons in 1973. Although Lebour (1920) recorded Pseudocalanus among the food items of larval (9-25 mm) pilchard off Plymouth, the fish largely occurs south of the range of Pseudocalanus, except in the Black Sea, where the copepod may be an important food item (I. I. Porumb, 1969). (e) Sprat The sprat (Sprattus sprattus) is an important fishery for the U.S.S.R. and Denmark, which together landed over half of the 507 x lo3 metric tons taken in the northeastern North Atlantic in 1973. There is also a small fishery (9 x lo3 metric tons) in the Mediterranean and Black Seas. Nguyen et al. (1972) found that Pseudocalanus was important seasonally in samples of sprat from the Bornholm Deep in the Baltic, making up about 43% of food items in April and 59% in May. Miller (1969) found that Pseudocalanus also dominated the diet of fish taken in April and May off the Estonian coast, and that most feeding took place a t night. I n the Black Sea, Porumb (1971) found that a reduction of Pseudocalanus populations in April could be traced to its fate as a principal food of sprat, which is a migrant to Romanian coasts at this time of year.

(f) Atlantic cod The cod (Gadus morhua = G . callarim) yields the greatest catch among demersal fishes whose larvae feed on zooplankton. It is important both in the northeastern North AtIantic, where some 1 727 x lo3 metric tons were taken in 1973, and in the northwestern part, where some 808 x lo3 metric tons were caught. Wiborg (1948) investigated the food of larval cod in coastal waters of northern Norway during spring and summer of a number of years between 1930 and 1947. The larvae ranged from 3.1 to 13.0 mm long. When Pseudocalanm was abundant in the plankton in May 1933, copepodids occurred in larvae 4-7-6-5 mm long. Nauplii were found in small larvae, many with a yolk sac, in 1939. Pseudocalanus was not found in larvae taken in 1930 and 1947, evidently because the copepod

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was scarce at the times of sampling. As expected, copepods are not important in the diet of postlarval, " 0-group " cod, which feed almost exclusively on euphausiids, although Pseudocalanus was recorded in some stomachs (Wiborg, 1949). Marak (1960) investigated feeding by larval cod in the Gulf of Maine and over Georges Bank. He found " Pseudocalanus-type " (i.e. including Paracalanus) remains in six larvae 9-16 mm long. (g) Polar cod The small polar cod (Boreogadus saida) forms a significant fishery in the northeastern region of the North Atlantic, some 82 x lo3 metric tons having been taken in 1973. Diet of its larvae has been investigated by Ponomarenko (1967), who found that copepods were important t o larvae between 4.6 and 39 mm long, and that Pseudocalanus occurred in 57% of specimens in the size range 19.1-28.7 mm. Pseudocalanus nauplii have been used as food for polar cod larvae reared in the laboratory (Aronovich et al., 1975). (h) Haddock The haddock (Melanogrammus aeglefinus) offers a huge catch of some 593 x 103 metric tons in the northeastern North Atlantic, compared with only 26 x 103 metric tons in the northwestern part. Marak (1960) found among larvae between 4 and 46 mm long, " Pseudocdanus-type '' copepods (see under cod, above) mainly in larvae between 13 and 23 mm long. Ogilvie (1938) found Pseudocalanus commonly in larvae 3-5-31 mm long from Scottish waters. She considered the largest individuals to be postlarval. Pseudocalanus eggs and younger stages were found in smaller larvae, and adult females were common in those more than 12 mm long. (i) Whiting The whiting (Merlangius merlangus) formed an important fishery of about 207 x lo3 metric tons in the northeastern North Atlantic and a smaller one of about 1 x lo3 metric tons in the Mediterranean and Black Seas during 1973. Lebour (1920) found that Pseudocalanus was a favoured food item of larval whiting up to about 9 mm long during spring and summer off Plymouth. Although the larvae take other copepods, Lebour concluded that they select Pseudocalanus in preference to other like-sized forms, such as Temora.

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(j) European plaice The plaice (Pleuronectes platessa) yielded 164 x lo3 metric tons in the northeastern North Atlantic in 1973. Scott (1922) examined over 600 larvae 13-87 mm long from off the coasts of Wales and the Isle of Man, and found that 22% of them contained Pseudocalanus.

(k) European hake Of the several commercial species of hake, the European hake (Merluccius merluccius) gave an important catch in the northeastern North Atlantic of some 110 x lo3 metric tons in 1973. Lebour (1920) found a Pseudocalunus in a single larval hake (5-5 mm long) in a sample of 12, but felt able to conclude that " it seems likely that young Hake, like the Gadus species, begins by taking Pseudocalanus, Calanus afterwards being frequently taken ". (1) Sandeels

Sandeels (sandlances or sand launces) of several species (Ammodgtes spp.) are important fisheries, especially for Denmark, which landed 283 x lo3of the 307 x 10%metric tons taken in 1973 in the northeastern North Atlantic. Sandeels occur in large shoals in shallow waters and are caught by fine-mesh trawls, so that in this sense they can be classed as demersal fishes. They are important as food for other larger demersal fishes. Lebour (1919b) found Pseudoculunus in the guts of fish 19-21 mm long in a sample of 109 A . tobianus ranging from 3 t o 21 mm long. She also found (Lebour, 1918, 1920) Pseudocalanus frequently in the guts of Hyperoplus lanceolatus ( = A . lanceolatus) 1 P 2 5 mm in length. The Pacific sandlance ( A .personatus) forms an important fishery for Japan, which took 194 x 103 metric tons from the northwestern North Pacific in 1973. LeBrasseur et al. (1969) examined gut contents of larvae (< 30 mm) and young (> 30 mm) of this fish from the Strait of Georgia, British Columbia. At the time of sampling, Pseudocalanus and two larger Calanus spp. were available as food. Fish of the size range 20-40 mm had 48% of their stomachs filled with zooplankton in the 0.5-1.0 mm size range, whereas large (> 40 mm) fish had 85% of their stomachs filled with zooplankton in the 1-1-1.5 mm range. Species of zooplankton were not identified, but it can be inferred from these sizes that Pseudocalanus and young stages of Calanus must have been of prime importance.

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(m) Other denzersal species Pseudocalanus has been identified in the gut contents of a number of other demersal species by Lebour (1918, 1919a, 1919b, 1920). Among these, the ling (Molva molva), common dab (Limanda Eimanda) and turbot (Psetta maxima) are of some commercial importance, producing a combined catch of some 91 x lo3 metric tons in 1973, mostly in the northeastern North Atlantic. 2. Crustaceans

No doubt Pseudocalanus is preyed upon by a variety of planktonic crustacea, but only euphausiids appear to have been implicated explicitly. Ponomareva (1954) concluded that Pseudocalanus was about the right size among copepods of the Sea of Japan as food for Thysanoessa inermis, T . longipes and Euphausia pacifica. I n the aquarium, euphausiids caught Pseudocalanus readily and ate out their soft parts, leaving the exoskeleton. I n nature, parts of Pseudocalanw were regularly found in gut contents of euphausiids. 3. Chaetognaths

Chaetognaths are highly predaceous animals, eating many small animals of suitable size with which they come into contact. Since Pseudocalanus in its various stages is abundant and of suitable size, it is hardly surprising that it is eaten by chaetognaths. The most widespread chaetognath within the range of Pseudocalanus is Sagitta elegans (= Parasagitta elegans), but Sagitta setosa also occurs in the more southern parts of the range of the copepod. Adult female Pseudocalanus were prominent in the guts of 6-19 mm 8.elegans in the North and Celtic Seas in summer and the Irish Sea in winter, according to Rakusa-Suszczewski ( 1969). Pseudocalanus was evidently selected by small chaetognaths in preference to Calanus and Temora, even though Calanus was more abundant than Pseudocalanus in the plankton. Rakusa-Suszczewski (1969) found that most feeding on Pseudocalanus took place at night. Pearre (1973), working with the same samples from Bedford Basin, Nova Scotia, from which vertical distributions of Pseudocalanus had been determined (Fig. 38) found that Pseudocalanw living near the surface in July were largely taken at night. McLaren (1969) gives a detailed account of the ways in which populations ofS. elegans in Ogac Lake, Baffin Island, depend on Pseudocalanus. Adult chaetognaths produced eggs over a protracted period in

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spring, but recruitment of their young was only successful when the seasonal burst of nauplii of Pseudocalanw occurred. The sizes of the subsequent juvenile populations of chaetognaths were determined by the number of adult chaetognaths present at the times of these bursts of nauplii, but not on the number of nauplii present. McLaren also concluded that the overwinter survival of older S. elegans in Ogac Lake was dependent on accessibility of copepods, largely Pseudocalanus, in deeper water. Mironov (1960) studied the feeding of Sagitta setosa in the Black Sea in some detail. He found that Pseudocalanus was not very important aa a food item in Sevastopol Bay in September 1961, but that it constituted about 77% of the daily ration of S. setosa by numbers and 97% by weight in offshore waters in February 1951. RakusaSuszczewski (1969) showed that Pseudocalanus was taken by S. setosa around the British Isles, but that it was never as important in the diet as was Temora. 4.

H ydromedwsae

Lebour (1922) studied feeding of a number of species of hydromedusans and recorded Pseudocalanus in the guts of the following species in samples from nature : Sarsia tubulosa, Rathkea octopunctata, Leukartiara octona (as Turris pileata), Phialidium spp. and Obelia spp. Although Pseudocalanus is eaten by Aglantha digitale in Ogac Lake, Baffin Island, McLaren (1969) found that recruitment of the tiny young of this hydromedusan appeared to be related to availability of phytoplankton, and that its overwinter survival was unrelated to abundance of Pseudocalanw. It seems probable that, in general, hydromedusae are less important than chaetognaths as predators on Pseudocalanus. 5. Ctenophores

The tentaculatan ctenophore Pleurobrachia pileus occurs widely in northern waters and is known to be a predator on Pseudocalanw. Lebour (1923) found Pseudocalanus in a specimen of P. pileus from off Plymouth and Fraser (1970) included the " group Pseudocalanw and Paracalanus " as important to P. pileus in the North Atlantic. Carter (1965) considered that P. pilew had an important impact on Pseudocalanus in Tessiarsuk, a landlocked bay on the coast of Labrador. We infer that P. pilewr feeds on Pseudocalanws in the Black Sea from Petipa et al. (1970). Two studies of predation by P. pileus give more details. Bishop (1968) studied feeding in the laboratory by P. bachei (= P . pilew of

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some authors) from waters of Washington State and found that they fed on adult and copepodid Pseudocalanus a t a rate of 1-7 copepods/ ctenophore/h, at a copepod density of 2511. of each stage. They fed more slowly on nauplii of Pseudocalanus (1.0 nauplii/ctenophore/h), but the significance of this difference cannot be ascertained (SD only given). Anderson (1974) found that 89% of gut contents of P. pileus in St. Margaret’s Bay, Nova Scotia, consisted of C V and adult copepods, including Pseudocalanus. He found that Temora and Centropages were about 10 times more likely than Pseudocalanus to be captured in the sea when the relative concentrations of the copepod species in the sea were taken into account. He attributes the relative invulnerability of Pseudocalanus to its manner of swimming (see p. 53). He also disputes Bishop’s (1968) rates of feeding of P. bachei (see above), suggesting that Bishop’s animals were feeding past satiation levels. However, Anderson does stress the great natural importance of P. pileus on copepods in general. The tentaculatan Bolinopsis microptera was also studied in the laboratory by Bishop (1968) in conditions as described above for P. bachei. He found that B . micropteya fed on Pseudocalanus of all stages at a rate of 0.9 copepods/ctenophore/h, somewhat lower than the rates for P. bachei. Anderson (1 974) found that Bolinopsis infundibulum also fed on Pseudocalanus, but preferred Oithona in particular. We conclude that ctenophores may be significant predators on Pseudocalanus, but probably less so than on some other species of copepods.

C. Significance in the food web A species-by-species account of the species eaten by and the species that eat Pseudocalanus does not altogether fix its significance in the economy of the sea. Cushing (1970) and Steele (1974) are examples of the thrust toward systems analysis of production and the flow of matter or energy throughout the food webs of the sea. That Pseudocalanus is an important secondary producer ‘innorthern seas cannot be doubted. However, an initial note of caution must be made concerning the significance of this production in the food web. Recently Martens (1975) counted the numbers and estimated biomasses of dead Pseudocalanus sedimenting out of the water column and into collectors set in the western Baltic. He found that some 2.8 g C/m2/yr of Pseudocabnus was sedimented in this way. The figure compares with estimates of total production in some localities (see Section XIII). The great bulk of the Pseudocalanus carbon was sedimented in April-

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May, shortly after a numerical maximum in this copepod. It is possible that in regions with highly seasonal life cycles a major fraction of the production of Pseudocalanus can at times end up as detritus, even of “ spent ” adults, unused by predators. Marten’s work cries out for repetition elsewhere. One of the more detailed food chain analyses involving Pseudocalanm is presented by Petipa et al. (1970). They express on diagrams the standing crops and specific transfer rates (rates per unit of biomass) for the major components of the pelagic food web in the Black Sea (it should be noted that the captions in their Figs 1 and 2 are reversed). I n the near-surface waters (the “ epiplankton ”) older stages of Pseudocalanus contribute little, although their young (grouped among nauplii) may be important as consumers of medium-sized phytoplankton. I n deep water (the “ bathyplankton ”) C IV-adults of Pseudocalanus and Calanus are represented as’having a dominant role, forming the largest standing crop among the herbivores and consuming much the greatest amount of the relatively common large phytoplankters at these depths. They conclude that “ at the herbivorous level the most powerful flow of matter and energy is through the migratory copepodites and adult large-sized copepods (Pseudocalanus and Calanus) ”. While it may rest on huge accomplishments in data acquisition and analysis, a detailed assessment of the flow of matter and energy through a food web is nothing more than descriptive science. However, steps are being made toward the study of food webs as a predictive science. The early systems analysis by Cushing (1959) included Pseudocalanus among “ other copepods ”, but made no direct use of its parameters of feeding, growth rates, etc. An example that explicitly contains parameters for Pseudocalanus is by Menshutkin et al. (1974). They developed a mathematical model of the pelagic ecosystem of the Sea of Japan, which includes Pseudocalanus as an important part of the “ mesoplankton ” component of the “ boreal epiplankton ”. They took into account all the variables of growth and advection that influenced the transfer functions of biomass (energy) through the food web, including such variables as currents, temperature, vertical migrations, etc. I n this way they predicted the quantitative distributions of biomass (energy) of various categories throughout the Sea of Japan at various seasons. Although their results have not been fully tested, they do match fragmentary observations of the real patterns in the Sea of Japan. Another example is by Steele and Frost (1977), who construct a simulation model of nutrient-plant-herbivore-carnivore dynamics using parameters for small (= Pseudocalanus) and large ( = Calanus) cope-

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pods. Some of their assumptions match what we have shown: that there is an upper limit to assimilation and growth rates (set at a food level of 80 mg C/m3, which is probably too high, cf. p. 130), and that reproductive rate (as C) is the same as growth rate before maturity (see p. 131). Other assumptions about size-selection of food particles may be premature (see p. 63). Although their model produces some general results that match general observations, it fails in an attempt to simulate the seasonal cycle of Pseudocalanus in Loch Striven, Scotland. Steele and Frost conclude that this failure results largely from misassumptions about hydrographic restoration of nutrients, rather than from errors in the parameters of grazing and growth by copepods.

D. Retrospects and prospects The importance of Pseudocalanus as a consumer, replenisher of nutrients and food base have been amply demonstrated. The genus is of particular significance to fish, and therefore to fisheries, of northern waters. Gulland (1970) notes " that the potential for great expansion of catches are among species lower in the ecological pyramid " ; that is, among fishes that feed on zooplankton. Lishev and Freimane (1970) have found high degrees of correlation between biomasses of Pseudocalanus sampled at different times of year (see critical account of their work, p. 156). They argue that it should thus be possible to predict food supplies for planktivorous fishes. They also suggest that deviations from long-term regressions of population estimates of Pseudocalanus at a given time of year on estimates at an earlier time of year (i.e. deviations from the usual mortality rate) may reflect differences in the amount consumed in the interval by plnnktivorous fishes. This in turn may give a new technique for estimating fish populations from the amount they consume. Altogether these possibilities, which stand apart from the main thrust of food-web studies in the sea, seem very worth exploring. A common aim of systems analyses of production in marine food webs is the substitution by general categories (e.g. trophic levels, biogeographic units, particle sizes) of the actual species involved. We hope that we have said enough in our lengthy review on the biology of Pseudocalanus to indicate that it may not be possible altogether to substitute somewhat abstract categories for the more objective realities of this interesting marine copepod.

XVI. ACENOWLEDGEMENTS We are grateful to Dr R. J. Lincoln and the staff of the Crustacea Section, British Museum (Natural History) for allowing us the use of

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the section library, and also to Miss S. Fullerton and the staff of the ScienceLibrary, DalhousieUniversity, for much bibliographic assistance. The Trustees of the British Museum (Natural History) gave us permission to reproduce Fig. 39B, C and D. The Centre National de la Recherche Scientifique allowed us to reproduce Fig. 39A and Fig. 40. We are grateful to authors of theses who allowed us to make use of often previously unpublished material, and especially to Dr Brenda Thompson, whose thesis we have used so extensively. A large number of scientific colleagues throughout the world have given us advice, information and assistance throughout the preparation of this work. We are especially grateful to Robert Conover, Barry Hargrave, Kenneth Mann, the late Sheina Marshall, Georges Merinfeld, Eric Mills, Serge Poulet and Sharon Smith for reading the typescript of one or more sections and to Georges Merinfeld for bibliographic assistance. Despite their help, inadequacies remain for which we are entirely responsible. Our research has been supported by grants from the National Research Council of Canada to I.A.M.

XVII. REFERENOES Ackefors, H. (1966). Plankton and hydrography of the Landsort Deep. Veroflentlichwngen des Institutes fiir Meeresforschung i n Bremerhaven, Xonderbd., 2 , 381-385. Ackefors, H. (1969a). Ecological zooplankton investigations in the Baltic proper, 1963-1966. Report. Institute of marine Reeearch Lysekil, Seriea Biology, 18, 1-139. Ackefors, H. (1969b). Seasonal and vertical distribution of the zooplankton in the Ask6 area (northern Baltic proper) in relation to hydrographical conditions. Oikos, 20, 480-492. Adler, G. and Jespersen, P. (1920). Variations saisonnihres chez quelques cop& podes planctoniques marins. Meddelelser f r a Kommhsionen for Hawundersegelser, Serie Plankton, 2(1), 1-46. Afrikova, S. G. (1975). Distribution of Pseudocalanus elongatus (Boeck) in the Black Sea. Gidrobiologicheskil zhurnal, 11(5), 92-97. (In Russian.) Afrikova, S. G. (1976). On the die1 vertical migration of Black Sea Pseudocalantia elongatus (Boeck). Biologiya Morya, Kiev, 37, 68-76. (In Russian.) Anderson, E. (1974). Trophic interactions among ctenophores and copepods in St. Margaret’s Bay, Nova Scotia. Ph.D. Thesis, Dalhousie University, Halifax, Nova Scotia. Andreeva, V. V. (1975). Basic features of the biology of some very abundant species of copepods of the Northern part of the Tatar Strait. I n “ Biologicheskie Resursy Morei Dal’nego Vostoka ” (abstracts of an All-Union symposium, Vladivostok, Oct. 1976), pp. 13-16. Tikhookeanskii nauchnoissledovatel’skii institut rybnogo khozyaistva i okeanografii (TINRO), Vladivostok. (In Russian.)

212

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Andreeva, V. V. (1976s). Influence of temperature on the duration of development in Pseudocabnus elongatus (Boeck). I n “ gksperimental’naya Ekologiya Morskikh Bespozvonochnykh ” (symposium held a t the “ Vostok ” Biological Station, 1976), pp. 17-18. Akademiya nauk SSSR, Vladivostok. (In Russian.) h d r e e v a , v. v. (1976b). Production estimates for P8eUdOCdanUS elongatus Boeck in the Northern part of the Japan Sea. I n ‘‘ Vsesoyuznaya konferentsiya molodykh uchenykh ‘Nauchno-tekhnicheskii progress v Wbnoi promjkhlennosti’ ”, pp. 13-14. Ministerstvo rybnogo khozyaistva SSSR, Tsentral’nj% nauchno-issledovatel’skii institut infonnatsii i tekhnikobkonomicheskikh issledovanii, Moscow. (In Russian.) Anraku, M. (1964a). Influence of the Cape Cod Canal on the hydrography and on the copepods in Buzzards Bay and Cape Cod Bay, Massachusetts. I. Hydrography and distribution of copepods. Limnology and Oceanography, 9, 46-60. Anraku, M. (1964b). Influence of the Cape Cod Canal on the hydrography and on the copepods in Buzzards Bay and Cape Cod Bay, Massachusetts. 11.Respiration and feeding. Limnology and Oceanography, 9, 195-206. Anraku, M. (1964~). Some technical problems encountered in quantitative studies of grazing and predation by marine planktonic copepods. Jaz~nzal of the Oceanographical Society of Japan, 20, 221-231. Anraku, M. and Omori, M. (1963). Preliminary survey of the relationship between the feeding habit and the structure of the mouth-parts of marine copepods. Limnology and Oceanography, 8 , 116-126. Apstein, C. (1911). Parasiten von Calanus finmarchicus. Wissenschaftliche Meeresuntersuchunge?a der Kommission fur wissenschaftlichen Untersuchung der deutschen Meere. Abt. Kiel, 13, 205-223. Arashkevich, Y e . G . (1969). The food and feeding of copepods in the northwestern Pacific. Oceanology, 9, 695-709. (English version.) Arndt, E. A. and Stein, H. (1973). Biologische Untersuchungen wlihrend des Internationalen Ostseejahres (International Baltic Year, IBY) 1969/70. 111.Das Zooplankton. Beitrme zur Meereskunde, 32, 33-35. Aronovich, T. M., Doroshev, S. I., Spectorova, L. V. and Makhotin, V. M. (1975). Egg incubation and larval rearing of navaga (Eleginw navaga Pall.), polar cod (Boreogadus saida Lepechin) and arctic flounder (Liopetta glacialis Pall.) in the laboratory. Aquaculture 6 , 233-242. Bainbridge, R. (1952). Underwater observations on the swimming of marine zooplankton. Journal of the Marine Biological Association of the United Kingdom, 31, 107-112. Barnes, H. and Marshall, S. M. (1951). On the variability of replicate plankton samples and some applications of ‘‘ contagious ” series to the statistical distribution of catches over restricted periods. Journal of the Marine Biological Association of the United Kingdom, 30, 233-263. Beklemishev, K. V. (1954a). The discovery of siliceous formations in the epidermis of lower Crustacea. Dokladg Akademii nauk SSSR, 97,543-545. (In Russian.) Beklemishev, K. V. (1954b). Feeding of several common planktonic copepods in far eastern seas. Zoolog~chesk~~ zhurnal, 33, 1210-1230. (In Russian.) Beklemishev, C. W. [K. V.] (1962). Superfluous feeding of marine herbivorous ZOOplankton. Rapport et procb-verbaw des rhunwns. Oonseil permanent international pour l’exploratirm.de la mer, 153, 108-113.

THE BIOLOGY OF PSEUDOCALANUS

213

Bglehrhdek, J. (1935). Temperature and living matter. Protoplaama monographien, 8, 1-277. Bigelow, H. B. (1926). Plankton of the offshore waters of the Gulf of Maine. Bulletin of the United States Bureau of FGheries, 40, 1-509. Bigelow, H. B. (1927). Physical oceanography of the Gulf of Maine. Bulletin of the United States Bureau of Fisheries, 40, 511-1027. Bishop, J. W. (1968). A comparative study of feeding rates of tentaculate ctenophores. Ecology, 49, 9 9 6 9 9 7 . Bjornberg, T. K. S. (1973). The planktonic copepods of the Marchile I Expedition and of the ‘‘ Eltanin ” Cruises 3-6 taken in the SE Pacific. Boletim de Zoologia e Biologia Marinha, N.S., 30, 245-394. Blaxter, J . H. S. (1963). The feeding of herring larvae and their ecology in relation to feeding. Report. California Cooperative Oceanic Fisheries Investigations, 10, 79-88. Boeck, A. (1864). Oversigt over de ved Norges Kyster iagttagne Copepoder henherende ti1 Calanidernes, Cyclopidernes, og Harpactidernes Familier. Forhandlinger, Videnskabsselskabet i Kristiania, 7 , 2 2 6 2 8 2 . Boeck, A. (1872). Nye Slaegter og Arter af Saltvands-Copepoder. Forhandlinger, Videnskabsselskabet i Kristiania, 14, 35-60. Bogorov, B. :G. (1939). Sex ratio in marine copepods. (On the problem of the evaluation of plankton production). Dokladg Akademii nauk SSSR, 23, 706-709. (In Russian.) Bogorov, B. G. (1946). Peculiarities of diurnal vertical migrations of zooplankton in polar seas. Journal of Marine Research, 6 , 25-32. Borradaile, L. A., Potts, F. A., Eastham,.L. E. S. and Saunders, J. T. (1961). “ The Invertebrata The University Press, Cambridge. Bottrell, H. H. (1975). The relationship between temperature and duration of egg development in some epiphytic Cladocera and Copepoda from the River Thames, Reading, with a discussion of temperature functions. Oecologia, 18, 63-84. Bowman, T. E. (1971). The distribution of calanoid copepods off the southeastern United States between Cape Hatteras and southern Florida. Smithsonian Contributions to Zoology, 96, 1-58. Brady, G. S. (1863). Report of dredging operations on the coasts of Northumberland and Durham in July and August 1863. Transactions of Tyneside Naturalists’ Field Club, 6 , 178-194. Brady, G. S. (1865). Report on the pelagic Entomostraca dredged off the coasts of Northumberland and Durham, 1862-64. Natural History Transactions of Northumberland Durham and Newcaatle-on-Tyne, 1, 29-40. Brady, G. S. (1878). “ A Monograph of the Free and Semi-parasitic Copepoda of the British Islands ”, Vol. 1. Ray Society, London. Bradley, B. P. (1975). The anomalous influence of salinity on temperature tolerance of summer and winter populations of the copepod Eurytemora afinia. Biological Bulletin, 148, 2 6 3 4 . Brodskii, K. A. (1948). Free-living Copepoda of the Sea of Japan. Izvestiya Tikhookeanskogo nauchno-issledovatel’skogoinstituta ebnogo khozyabtva d okeanografii, 26, 3-130. (In Russian.) Brodskii, K. A. (1950). “ Calanoida of the Far Eastern Seas and Polar Basin of the USSR (1967 transl. Israel Programme for Scientific Translations, Jerusalem.)

”.

”.

214

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Brodskii, K. A. (1956). The regional distribution of pelagic life in far eastern seas and adjacent waters of the Pacific Ocean as shown by a study of the copepod fauna (Calanoida). T r u d g problemngkh i tematichkikh soveshchanii. Zoologicheskii institut, 6, 22-35. (In Russian.) Bullen, G. E. (1908). Plankton studies in relation to the western mackerel fishery. Journal of the Marine Biological Association of the United Kingdom, 8, 269-302. Butler, E. I., Corner, E. D. S. and Marshall, S. M. (1969). On the nutrition and metabolism of zooplankton. VI. Feeding efficiency of Calanus in terms of nitrogen and phosphorus. Journal of the Marine Biological Association of the United Kingdom, 49, 977-1001. Cairns, A. A. (1967). The zooplankton of Tanquary Fjord, Ellesmere Island, with special reference to the calanoid copepods. Journal of the Fkheries Research Board of Canada, 24, 555-568. Cannon, H. G. (1928). On the feeding mechanism of the copepods Calanus jinmarchicua and Diaptomus gracilis. British Journal of Experimental Biology, 6, 131-144. Canu, E. (1892). Les cophpodes du Boulonnais. Morphologie, embryologie, taxonomie. Travaux d u hboratoire de zoologie maritime de WimereuxAmbleteuse (Pas-de-Calais) 6, 1-292. Also published as separate the&, Danel, Lille. Carlisle, D. B. and Pitman, W. J. (1961). Diapause, neurosecretion, and hormones in Copepoda. Nature, London, 190, 827-829. Carrillo B.-G., E., Miller, C. B. and Wiebe, P. H. (1974). Failure of interbreeding between Atlantic and Paciiic populations of the marine calanoid copepod Acartia clausi Giesbrecht. Limnology and Oceanography, 19, 452-458. Carter, J. C. H. (1965). The ecology of the calanoid copepod Pseudocalanus minutus Krrayer in Tessiarsuk, a ooastal meromictic lake of northern Labrador. Limnology and Oceanography, 10, 345-353. Cattley, J. G. (1948). Sex reversal in copepods. Nature, London, 161, 937. Caullery, M. (1910). Ellobiopsis clhattoni, n.g., n.sp., parasite de Calanus helgolandicus Claus, appartenant probablement aux pdridiniens. Bulletin scientifique de la France et de la Belgique, 44, 201-214. Chatton, E. (1911). Sur divers parasites de copBpodes pdlagiques observes par M . Apstein. Compte rendu hebdomadaire des Sdances de I’Acaddmie des Sciences, 153, 474-476. Chatton, E. (1 920). Les peridiniens parasites. Morphologie, reproduction, Bthologie. Archives de Zoologie expdrimentale et gdndrale, 59, 1-475. Christiansen, F. E. (1968). Nitrogen excretion by some planktonic copepods of Bras d’Or Lake, Nova Scotia. M.Sc. Thesis, Dalhousie University, Halifax, Nova Scotia. Ciszewski, P. and Witek, 2. (1975). Production of copepods Psedocalanzta elongatus and Acartia bi$losa in the Gdansk Deep area. 4th Symposium of Baltic Marine Biologists, Cfdhsk, Poland, 13-18 Oct. 1975, p. 22. Colebrook, J. M. (1964). Continuous plankton records : a principal component analysis of the geographical distribution of zooplankton. Bulletin of Marine Ecology, 6, 78-100. Colebrook, J. M. (1969). Variability in the plankton. Progressive Oceanography, 5, 115-125.

THE BIOLOGY OF PBEUDOCALANUS

215

Colebrook, J. M., John, D. E. and Brown, W. W. (1961). Contribution towards a plankton atlas of the north-eastern Atlantic and the North Sea. Part I1 : Copepoda. Bulletin of Marine Ecology, 5, 90-97. Conover, R. J. (1959). Regional and seasonal variation in the respiratory rate of marine copepods. Limnology and Oceanography, 4, 259-268. Conover, R. J. (1974). Production in marine planktonic communities. Proceedings Pirst International Congress of Ecology, The Hague, 8-14 Sept. 1974, 159-163. Conover, R. J. (In press). Transformation of organic matter. I n “Marine Ecology. Vol. IV, Dynamics ” (0.Kinne, ed.). Wiley, London. Conover, It. J. (1978). Feeding interactions in the pelagic zone. Rapports et prods-verbaw des r6unions. Conseil international pour C’explorationde la mer, 173. Conover, R. J. and Francis, V. (1973). The use of radioactive isotopes to measure the transfer of materials in aquatic food chains. Marine Biology, 18, 272283. Corkett, C. J. (1966). Studies on the biology of the marine planktonic copepods Psez~docalanuselongatus and Temora longicornis under laboratory conditions, t,ogether with morphological studies on the larval stages. Ph.D. Thesis, University of London. Corkett, C. J. (1968). Observations sur les stades larvaires de Pseudocalanus elongatus Boeck et Temora longicornis 0. F. Miiller. Pelagos. Bulletin de l’lmtitut Ockanographipue d’Alger, 8 , 51-67. Corkett, C. J. (1969). Reproduction and oil storage by Pseudocalanus in nature : an approach to copepod production. Unpublished report to Bedford Institute Oceanography, Dartmouth, Nova Scotia. Corkett, C. J. (1970). Techniques for breeding and rearing marine calanoid copepods. Helgolander wissenschuftliche Meeresuntersuchungen, 20, 3 18-324. Corkett, C. J. and McLaren, I. A. (1969). Egg production and oil storage by the copepod Pseudocalanus in the laboratory. Journal of Experimental Marine Biology and Ecology, 3, 90-105. Corkett, C. J. and McLaren, I. A. (1970). Relationships between development rate of eggs and older stages of copepods. Journal of the Marine Biological Association of the United Kingdom, 50, 161-168. Corkett, C. J. and Urry, D. L. (1968). Observations on the keeping of adult female Pseudocalanua elongatus under laboratory conditions. Journal of the Marine Biological Association of the United Kingdom, 48, 97-105. Corkett, C. J. and Zillion, E. J. (1975). Studies on the effect of temperature on the egg laying of three species of calanoid copepods in the laboratory. Bulletin of Plankton Society, Japan, 21, 13-21. Corner, E. D. S. and Cowey, C. B. (1968). Biochemical studies on the production of marine zooplankton. Biological Reviews, 43, 393-426. Corner, E. D. S. and Davies, A. G. (1971). Plankton as a factor in the nitrogen and phosphorus cycles in the sea. Advances in Marine Biology, 9, 101-204. Corner, E. D. S., Cowey, C. B. and Marshall, S. M. (1966). On the nutrition and metabolism of zooplankton. 111. Nitrogen excretion by Calanus. Journal of the Marine Biological Association of the United Kingdom, 45, 429-442. Crawshay, L. R. (1915). Notes on experiments in the keeping of plankton animals under artificial conditions. Journal of the Mwine Biological Association of the United Kingdom, 10,665-676.

216

CHRISTOPHER J. CORRETT AND IAN A. MOLAREN

Curl, H. and McLeod, G. C. (1961). The physiological ecology of a marine diatom, Skeletonema costatum (Grev.) Cleve. Journal of Marine Research, 19, 70-88. Cushing, D. H. (1955). Production and a pelagic fishery. Pishery Investigations. Ministry of Agriculture, Food and Fisheries, 18, 1-104. Cushing, D. IT. (1959). On the nature of production in the sea. F i d e r y Investigations. Ministry of Agriculture, Fisheries and Food, Ser. 2, 22(6), 1-40. Cushing, D. H. (1970). Introduction. I n “ Marine Food Chains ” (J.H. Steele, ed.), pp. 69-73. Oliver and Boyd, Edinburgh. Dngg, M. (1977). Some effects of patchy food environments on copepods. Limnology and Oceanography, 22, 99-107. Davis, C. C. (1949). The pelagic Copepoda of the north-eastern Pacific Ocean. University of Washington Publications in Biology, Seattle, 14, 1-1 17. Davis, C. C. (1976). Overwintering strategies of common planktic copepods in some North Norway fjords and sounds. Astarte 9, 37-42. Deevey, G. B. (1952). Quantity and composition of the zooplankton of Block Island Sound, 1949. Bulletin of the Bingham Oceanographic Collection, Yale University, 13, 120-164. Deevey, G. B. (1956). Oceanography of Long Island Sound, 1952-1954. V. Zooplankton. Bulletin of the Bingham Oceanographic Collection, Yale University, 15, 113-155.

Dewey, G. B. (1960a). The zooplankton of the surface waters of the Delaware Bay region. Bulletin of the Bingham Oceanographic Collection, Yale University, 17, 5-53. Deevey, G. B. (1960b). Relative effects of temperature and food on seasonal variations in length of marine copepods in some eastern American and western European waters. Bulletin of the Ringham Oceanographic Collection, Yale University, 17, 54-84. Deevey, G. B. (1971). The annual cycle in quantity and composition of the zooplttrikton of the Sargasso Sea off Bermuda. 1. The upper 500 m. TAmnology and Oceanography, 16, 219-240. Delalo, E. P. (1964). The die1 feeding rhythm in PseudocaEanw elongatus (Boeck). Trudg Sevastopol’skoi biologicheskoi stantsii. A kademiya nauk SSSR, 15, 94-100. (In Russian.) Digby, P. S. B. (1950). The biology of the small planktonic copepods of Plymouth. Journal of the Marine Biological Association of the United Kingdom, 29, 393-438.

Digby, P. 8. B. (1954). The biology of the marine planktonic copepods of Scoresby Sound, East Greenland. Journal of Animal Ecology, 23, 298-338. Digby, P. S. B. (1967). Pressure sensitivity and its mechanism in the shallow marine environment. Symposia of the Zoological Society of London, 19, 159- 1 88. Dodimearl, A. J., Favorite, F. and Hirano, T. (1963). Salmon of the North Pacific Ocean. Part 2. Review of oceanography of the subarctic Pacific region. Bulletin of the International North Pacific Fisheries Commission, 13, 1-195. Drebes, G. (1969). Dissodinium pseudocalani sp. nov., ein parasitischer Dinoflagellat auf Copepodeneiern. Helgolander wissenschaftliche Meeresuntersuchumgen, 19, 58-67. Drebes, G. (1972). Entwicklung des Ektoparasiten Dissodinium pseudocalani (Dinophyceae). Accompanying pamphlet for scientific film C 1069/1971, Institut fur den wissenschaftlichen Film, CGttingen.

217

THE BIOLOGY OF PSEUDOGALANUS

Dunbar, M. J. (1947). Note on the delimitation of the arctic and subarctic zones. Canadian Field Naturalist, 61, 12-14. Dunbar, M. J. and Harding, G. (1968). Arctic Ocean water masses and plankton -a reappraisal. I n “ Arctic Drifting Stations ” (J.E. Slater, coordinator), pp. 315-326. Arctic Institute of North America. Edinburgh Oceanographic Laboratory ( 1973). Continuous plankton records : 8 plankton atlas of the North Atlantic and the North Sea. Bulletin of Marine Ecology, 7 , 1-174. Edmondson, W. T., Comita, G. W. and Anderson, G. (1962). Reproductive rate of copepods in nature and its relation to phytoplankton population. Ecology, 43, 625-634. Eriksson, S. (1973a). The biology of marine planktonic Copepoda on the west coast of Sweden. Zoon, 1, 37-68. Eriksson, S. (197313). Preliminary zooplankton investigations in the dregrund Archipelago (southwestern Bothnian Sea) during the summer 1970. Zoon, 1, 95-111. Esterley, C. 0. (1916). The feeding habits and food of pelagic copepods and the question of nutrition by organic sitbstances in solution in the water. University of Calijornia Publications i n Zoology, 16, 171-184. Evans, F. (1973). The permanent zooplankton of Northumberland coastal waters. Proceedings of University of Newcastle upon Tyne Philosophical Society, 2 , 25-68. Evans, F. (1977). Seasonal density and production estimates of the commoner planktonic copepods of Northumberland coastal waters. Estuarine and Coastul Marine Science, 5, 223-241. F A 0 (1974). Yearbook of Fishery Statistics. Vol. 36. Catches and Landings, 1973. U.N. Food and Agriculture Organisation. Faber, D. J. (1966). Free-swimming copepod nauplii of Narragansett Bay with a key to their identification. Journal of the Fisheries Research Board of Canada, 23, 180-205. Falconer, D. S. (1960). ‘‘ Introduction to Quantitative Genetics Ronald Press, New York. Farran, G. P. and Vervoort, W. (1951). Copepoda, sub-order: Calanoida; family: Pseudocalanidae. Fiches d’identi,fication d u zooplankton, Conseil permanent international pour Z’Exploratwn de la Mer, No. 37. Fedorov, V. D., Zhitina, L. 5. and Polyakova, T. V. (1975). Daily production of mass species of White Sea zooplankton. Izvestiya Akademii Nauk SSSR, 8er. biol., 5, 719-726. (In Russian.) Fish, C. J. (1925). Seasonal distribution of the plankton of the Woods Hole region. Bulletin of the United States Bureau of Fisheries, 41, 91-179. Fish, C. J. (1936). The biology of Pseudocalanus minutus in tho Gulf of Maine and the Bay of Fundy. Biological Bulletin, 70, 193-216. Fives, J. M. (1969). Investigations of the plankton of the west coast of Ireland. 2. Planktonic surveys during the years 1958-63. Proceedings of Royal Irish Academy, 67, 233-259. Fleminger, A. (1967). Distributional atlas of calanoid copepods in the California Current region, Part 11. Culifornk Cooperative Oceanic Fisheries Inuestigations Atlas 7 , 1-213.

”.

218

CFIRISTOPHEE J. CORKETT AND IAN A. MOLAREN

Fontaine, M. (1955). The planktonic copepods (Calanoida, Cyclopoida, Monstrilloida) of Ungava Bay, with special reference to the biology of Pseudocalanw minutus and Calanus finmarchicus. Journal of the Pishm%es Research. Bomd of Canada, 12, 858-898. Fraser, J. H. (1970). The ecology of the ctenophore Pleurobrachia pileus in Scottish waters. Journal du Conseil. Conseil permanent international pour l’exploration de la mer, Copenhague, 33, 149-168. Frost, B. W. (1974). Feeding processes a t lower trophic levels in pelagic food communities. In “ The Biology of the Pacific Ocean ” (C. B. Miller, ed.), pp. 59-77. Oregon State Univ. Friichtl, F. (1920). Planktoncopepoden aus der nordlichen Adria. Sitzun.gsberichte der Alcademie der Wwseiwchaften Wien, Math.-naturw&8. Kl., Abt. 1, 129, 463-509. Furuhashi, K. (1961). On the possible segregation found in the copepod fauna in the deep waters off south-eastern coast of Japan, Publications of the Set0 Marine Biological Laboratory, 9, 1-13. Furuhashi, K. (1966). Studies on the vertical distribution of copepods in the Oyashio region east of Japan and in the Kuroshio region south o f Japan. Publications of the Set0 Marine Biological Laboratory, 14, 295-322. Gaudy, R. (1962). Biologie des copepodes pelagiques du golfe de Marseille. Recueil des Travaux de la Station marine d’Endoume, Pacult6 dea Sciences de Marseille, 42 (Bull. 47), 93-184. Gauld, D. T. (1951). The grazing rate of planktonic copepods. Journal of the Marine Biological Association of the United Kingdom, 29, 695-706. , Gauld, D. T. (1964). Feeding in planktonic copepods. In “ Grazing in Terrestrial and Marine Environments ” (D. J. Crisp, ed.), pp. 239-245. Blackwell, Oxford. Gauld, D. T. (1966). The swimming and feeding of planktonic copepods. I n “ Some Contemporary Studies in Marine Science ” (H. Barnes, ed.), pp. 313334. Allen and Unwin, London. Gem, G. H. and Hargrave, B. T. (1966). Primary and secondary production in Bras d’Or Lake, Nova Scotia, Canada. Verhandlungen Internationaleiz Vereinigung f u r Theoretische und Angewandte Limnologie, 16, 333-340. Gibson, V. R. and Grice, G. D. (1977). Response of macro-zooplankton POpUlEtions to copper: Controlled Ecosystem pollution experiment. Bulletin of Marine Science, 27, 85-91. Giesbrecht, W. (1881). Vorliiufige Mittheilung aus einer Arbeit uber die freilebenden Copepoden des Kieler Hafens. Zoologischer Anzeiger, 4, 254-268. Giesbrecht, W. (1882). Die freilebenden Copepoden der Kieler Foehrde. Bericht Kommision zur wissenschaftlichen Untersuchung des deutschert Meere, Kiel, 4, 85-168. Glover, R. S., Robinson, G. A. and Colebrook, J. M. (1974). Marine biological surveillance. Environment and Change, 2, 395-402. Gonnert, R. (1936). Sporodinium pseudocalani n.g., n. sp., ein Parasit aus Copepodeneiern. Zei&chr$t f u r Parasitenkunde, 9, 140-143. Grainger, E. H. (1959). The annual oceanographic cycle at Igloolik in the Canadian arctic. 1. The zooplankton and physical and chemical observations. Journal of the Fisheries Research Board of Canada, 16, 453-501. Grainger, E. H. (1965). Zooplankton from the Arctic Ocean and adjacent Canadian waters. Journal of the Fisheries Reseamh Board of C a d a , 22, 643-664.

THE BIOLOGY OF PSEUDOCALANUS

219

Greze, V. N. (1970). The biomass and production of different trophic levels in the pelagic communities of south seas. I n " Marine Food Chains " (J. H. Steele, ed.), pp. 458-467. Oliver and Boyd, Edinburgh. Greze, V. N. (1973). Secondary production in southern seas. Oceanology, 13, 80-85. (English edition.) Greze, V. N., Baldina, E. P. and Bileva, 0. K. (1968). Production of planktonic copepods in the neritic zone of the Black Sea. Oceanology, 8 , 839-843. (English edition.) Grice, G. D. (1961). Calanoid copepods from equatorial waters of the Pacific Ocean. Fkhery Bulletin, Fish and Wildlife Service, United States Department of the Interior, 61, 171-246. Grice, G. D. (1962). Copepods collected by the nuclear submarine Seadragon on a cruise to and from the North Pole, with remarks on their geographic distribution. Journal of Marine Research, 20, 97-109. Grice, G. D. and Hulsemann, K. (1965). Abundance, vertical distribution, and taxonomy of calanoid copepods a t selected stations in the northeast Atlantic. Journal of Zoology, 146, 213-262. Grice, G. D., Wiebe, P. H. and Hoagland, E. (1973). Acid-iron wmtc as a factor affecting the distribution and abundance of zooplankton in the New York Bight. I. Laboratory studies on the effects of acid waste on copepods. Estuarine and Coastal Marine Science, 1, 45-50. Griffiths, A. M. and Frost, B. W. (1976). Chemical communication in the marine planktonic copepods Calanus paci$cus and Pseudocalanua sp. Cruatacmna, 30, 1-8. Gulland, J. A. (1970). Food chain studies and some problems in world fisheries. I n " Marine Food Chains " (J. H. Steele, ed.), pp. 296-315. Oliver and Boyd, Edinburgh. Gurney, R. (1931). " British Fresh-water Copepoda ", Vol. I. Ray Society, London. Gurney, R. (1942). " Larvae of Decapod Crustacea ". Ray Society, London. Hansen, K. V. (1951). On the diurnal migration of zooplankton in relation to the discontinuity layer. Journal du conseil. Conseil permanent international pour I'Exploration de la Mer, Copenhague, 17, 231-241. Hansen, K. V. (1960). Investigations on the quantitative and qualitative distribution of zooplankton in the southern part of the Norwegian Sea. Meddelelser fra Danmrks Fiskeri- og Havundersegelser, N.S., 2(23), 1-63. Harding, G. C. H. (1966). Zooplankton distribution in the Arctic Ocean with notes on life cycles. M.Sc. Thesis, MoGill University, Montreal. Rarding, G. C. H. (1972). Ecological observations on North Atlantic deep sea copepods. Ph.D. Thesis, Dalhousie University, Halifax, Nova Scotia. Hardy, A. C. (1924). The herring in relation to its animate environment. Part 1. Fishery Investigations. Ministry of Agriculture, Food and Fisheries, Ser. 2 , 7(3), 1-53. Hardy, A. C. (1956). " The Open Sea. Its Natural History. Part 1, The World of Plankton ". Collins, London. Hargrave, B. T. (1966). Some aspects of phosphorus excretion by planktonic and benthic crustaceans of Bras d'Or Lake, Nova Scotia. M.Sc. Thesis, Dalhousie University, Halifax, Nova Sootia. Hargrave, B. T. and Geen, G. H. (1968). Phosphorus excretion by zooplankton. Limwlogy and Oceanography, 13, 332-342.

220

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Hargrave, B. T. and Geen, G. H. (1970). Effects of copepod grazing on two natural phytoplankton populations. Journal of the Fisheries R M ~ T C A Board of Canada, 27, 1395-1403. Harris, R. P. and Paffenhofer, G.-A. (1976). The effect of food concentration on cumulative ingestion and growth efficiency of two sniall marine planktonic copepods. Journal of the Marine Biological Association of the United Kingdom, 56, 875-888. Hart, R. C. and McLaren, I. A. (1978). Temperature acclimation and other influences on embryonic development in the copepod Pseudocalanus. Marine Biology, 45, 23-30. Heinrich, A. K. (1962). The life histories of plankton animals and seasonal cycles of plankton communities in the oceans. Journal d u comeil. Conaeil permanent international pour I’Exploration de la Mer, Copenhague, 27, 15-24. Hessle, C. and Vallin, S. (1934). Undersokningar over plankton och dess viixlingar i Ostersjon under Bren 1925-1927. Svenska hydrografisk-bwlogislca kommissionens skrijter. Biologi. ( N . S . ) 1(5), 1-132. Hirschfeld, H. S. (1974). Some observations on the breeding of Pseudoccllanwr in tho laboratory and comparisons with other calanoid species. B.Sc. Hons. Thesis, Dalhousie University, Halifax, Nova Scotia. Hjort, J. (1914). Fluctuations in the great fisheries of northern Europe viewed in the light of biological research. Rapport et Procka- Verbaux des Rdunwna. Conseil permanent international pour 1’Exploration de la Mer, 20, 1-228. Huizinga, H. W. (1966). Studies of the life cycle and development of Contracaecum apiculigerum (Rudolphi, 1809) (Ascaroidea : Heterocheilidae) from marine piscivorous birds. Journal of the Elisha Mitchell Scienti,fic Society, 82, 181-195.

Hum, J. and Scotto di Carlo, B. (1968). Comparazione tra lo zooplancton del Golfo di Napoli e dell’Adriatico meridionale presso Dubrovnik. I. Copepoda. Pubblicazioni della Stazione zoologica di Napoli, 36, 21-102. Hum, J. and Scotto di Carlo, B. (1969). Copepodi pelagici dell’Adriatico settentrionale nel periodo gennaio-dicembre 1965. Pubblicazioni della Stazione zoologica d i Napoli, 37, Suppl. 2, 173-195. Ikeda, T. (1970). Relationship between respiration rate and body size in marine plankton animals as a function of the temperature of habitat. Bulletin of the Faculty of Pisheries, Hokkaido University, 21, 91-112. Ikeda, T. (1974). Nutritional ecology of marine zooplankton. Memoirs of the Faculty of Fisheries, Hokkaido Univer&y, 22, 1-97. Itoh, K. (1970). A consideration on feeding habits of planktonic copepods in relation to the structure of their oral parts. Bulletin of Plankton Society of Japan, 17, 1-10. (In Japanese, English summary.) Jeffries, H. P. (1962). Copepod indicator species in estuaries. Ecology, 43, 730-733.

Jeffries, H. P. and Alzara, L. (1970). Dominance-diversity relationships of the free amino acids in coastal zooplankton. Comparative Biochemistry and Phy~iology,37, 215-223. Jepps, M. W. (19378). Note on Apstein’s parasites and some very early larval Platyhelminthes. Parmitology, 29, 554-558. Jepps, M. W. (193713). On the protozoan parasites of Calanue finmarchicus in the Clyde Sea area. Quarterly Journal of Microscopical Science, 79, 589-668. Jespersen, P. (1939). Investigations on the copepod fauna in East Greenland waters. Meddelelser om Grenland, 119(9), 1-106.

THE BIOLOGY OF PA'EUDOCALANUS

221

Kaestner, A. (1970). " Invertebrate Zoology ", Vol. 111. Interscience, New York. Kamshilov, M. M. (1951). Determination of the weight of Calanus$nmarchicus Gunner, on the basis of body length measurements. Dokladfj Akademii m u k SSSR, 76, 945-948. (In Russian.) Kamshilov, M. M. (1961). Data on the biology of Pseudocalanus elongatus Boeck in the Barents and White Seas. I n " Hydrological and Biological Features of the Inshore Waters of Murman" (M. M. Kamshilov, ed.), pp. 109-126. Murmanskoe Knizhnoe Izdatel'stvo, Murmansk. (In Russian.) Katona, S. K. (1973). Evidence for sex pheromones in planktonic copepods. Limnology and Oceanography, 18, 5 7 4 4 8 3 . Katona, S. K. and Moodie, C. F. (1969). Breeding of Pseudocalanus elongatus in the laboratory. Journal of the Marine Biological Association of the United Kingdom, 49, 743-747. Kimor, B. and Wood, E. J. F. (1975). A plankton study in the eastern Mediterranean Sea. Marine Biology, 29, 321-333. Klyashtorin, L. B. and Yarzhombek, A. A. (1973). Energy consumption in active movements of planktonic organisms. Oceanology, 13, 575-580. (English edition.) Kolosova, Ye. G. (1975). Effect of temperature on the distribution of mass species of zooplankton in the White Sea. Oceanology, 15, 92-95. (English edition, published in 1976.) Kovalev, A. V. (1967). Size composition of planktonic copepods at various depths in the Black Sea. Gidrobiologicheskiizhurnal, 3(3), 74-77. (InRussian.) Kovalev, A. V. (1968). On the size of the pelagic copepods in the Black Sea, Mediterranean Sea, and Adriatic Sea in relation to habitat conditions. I n " Biological Investigations of the Black Sea and its Biological Resources " (V. A. Vodyanitskii, ed.), pp. 63-66. Nauka, Moscow. (In Russian.) Kraefft, F. (1910). mer das Plankton in Ost- und Nordsee und den Verbindungsgebieten mit Besonderer Beriicksichtigung der Copepoden. Wissenschaftliche Meeresunterwchungen der Iiornmksion zur Wissenschaft,?ichenUnterauchung der deutschen Meere, Abt. Kiel, 11, 29-107. Kraneis, W. and Martens, P. (1975). Ecological studies on the plankton in the Kiel Bight 11. Zooplankton. 1MerentutkimuslaitoksenJulkaisu, 239, 187-194. Kreyer, H. (1842-45). Plate 41, fig. 4. I n " Voyages de la Commission Scientifique du Nord en Scandinavie, en Laponie, au Spitzberg et aux FBroe pendant les annBes 1838-40, sur la corvette La Recherche commandbe par M. Fabvre. Zoologie, Atlas " (P. Gaimard, ed.). Paris. Kreyer, H. ( 1848). Karcinologiske Bidrag. Naturhiatorisk Tidsskrqt, Ser. 2, 2, 527-609.

Krylov, V. V. (1968). Relation between wet formalin weight of copepods and copepod body length. Oceanology, 8, 723-727. (English edition.) Kuznetsov, V. V. (1973). Copepod fauna of the brackish water zone of the Laptev Sea. Nauchnfje dokladg vfjsshet ahkolfj, Biologicheskie nauki, 16(5), 22-24. (In Russian.) Lacroix, G. and Filteau, G. (1971). Les fluctuations quantitatives du zooplankton de la Baie-des-Chaleurs (Golfe du Saint-Laurent). 111. Fluctuations des copBpodes autres que Calanus. Natu~alisteCanadien, 98, 775-8 13. Laurence, G. C. (1976). Caloric values of some North Atlantic calanoid copepods. Fishery Bulletin, U.S. National Marine Fisheries Service, 74, 218-220.

222

CHRISTOPHER J. CORKETT AND JAN A. MCLAREN

Lawson, T. J. and Grice, G. D. (1977). Zooplankton sampling variability: controlled ecosystem pollution experiment. Bulletin of Marine Science, 27, 80-84. Lebour, M. V. (1918). The food of post-larval fish. Joztrnal of the Marine Biological Association of the United liingdom, 11, 433-469. Lebour, M. V. (1919a). Feeding habits of some young fish. Journal of the Marine Biological Association of the United Kingdom, 12, 9-21. Lebour, M. V. (1919b). The food of post-larval fish. I1 (1918). Journal of the Marine Biological Association of the United Kingdom, 12, 22-47. Lebour, M. V. (1920). The food of young fish. I11 (1919). Journal of the Marine Biological Association of the United Kingdom, 12, 261-324. Lebour, M. V. (1922). The food of plankton organisms. Journal of the Marine Biological Association of the United Kingdom, 12, 644-677. Lebour, M. V. (1923). The food of plankton organisms. 11. Journal of the Marine Biological Association of the United Kingdom, 13, 70-92. LeBrasseur, R. J., Barraclough, W. E., Kennedy, 0. D. and Parsons, T. R. (1969). Production studies in the Strait of Georgia. Part 111. Observations on the food of larval and juvenile fish in the Eraser River plume, Eebruary to May, 1967. Journal of Experimental Marine Biology and Ecology, 3, 51-61. Lee, J. W. and Williamson, D. I. (1976). The vertical distribution and diurnal migration of copepods a t different stations in the Irish Sea. Journal of the Oceanographical Society of Japan, 31, 199-21 1. Lee, R. F., Takahashi, M., Beers, J. R., Thomas, W. H., Seibert, D. L. R., Koeller, P. and Green, D. R. (1977). Controlled ecosystems: their use in the study of the effects of petroleum hydrocarbons on plankton. J n “ Physiological Responses of Marine Biota to Pollutants ” (F. J. Vernberg, A. Calabrese, F. P. Thurberg and W. B. Vernberg, eds.), pp. 323-342. Academic Press, New York, London. Legar6, J. E. H. and Maclellan, D. C. (1960). A qualitative and quantitative study of the plankton of the Quoddy region in 1957 and 1958 with special reference to the food of the herring. Journal of the Fisheries Research Board of Canada, 17, 409-448. Lishev, M. N. and Freimane, S. 0. (1970). The use of correlation analysis for studying the life cycle of Pseudocalanus elongatus Boek and Limnocalanus grimaldii Guerne. Trudg Bctltitskogo nauchno-issledouatel’skogo instituta morskogo rgbnogo khozyattua i okeanogra$i, 4, 68-78. (In Russian.) Lock, A. R. (1968). A laboratory and field study of the role of temperature in vertical migrations of the copepod Pseudocalanus minutus. M.Sc. Thesis, Dalhousie University, Halifax, Nova Scotia. Lock, A. R. and McLaren, I. A. (1970). The effect of varying and constant temperatures on size of a marine copepod. Limnology and Oceanography, 15, 638-640. Loeblich, A. R., I11 (1976). Dinoflagellate evolution : speculation and evidence. Journal of Protozoology, 23, 13-28. Lowe, C. W. (1936). Observations on some Pacific diatoms as the food of copepods and fishes. Journal of the Biological. Board of Canada, 3, 12-19. Lowndes, A. G . (1935). The swimming and feeding of certain calanoid copepods. Proceedings of the Zoological Society of London for 1935, 687-716.

THE BIOLOGY OB PSEUDOCALANUS

223

Mackas, D. and Bohrer, R. (1976). Fluorescence analysis of copepod gut contents and an investigation of die1 feeding patterns. Journal of Experimental Marine Biology and Ecology, 25, 77-85. Malikowski, W. and Ciszewski, P. (1962). The vertical migration of the Baltic zooplankton in a 24-hour cycle in the various seasons of the year. Rapport et ProcBs-verbaux des Riuniona. Conseil permanent international pour 1’Exploration de la Mer, 153, 220-221. Marak, R. R. (1960). Food habits of larval cod, haddock, and coalfish in the Gulf of Maine and Georges Bank area. Journal du Conseil. Conseil permanent international pour 1’Exploration de la Mer, 25, 147-167. Marshall, S. M. (1949). On the biology of the small copepods in Loch Striven. Journal of the Marine Biological Association of the United Kingdom, 28, 4 6-1 22. Marshall, S. M. (1963). ;In Scottish Marine Biological Association Annual Report for 1963, p. 19. Marshall, S . M. (1973). Respiration and feeding in copepods. Advances in Marine Biology, 11, 57-120. Marshall, S . M. and Orr, A. P. (1 652). O n the biology o f Calanw finmarchicus. VII : Factors affecting egg production. Journal of the Marine Biological Association of the United Kingdom, 30, 527-547. Marshall, S. M. and Om, A. P. (1954). Hatching in Calanzcsfinmarchicus and some other copepods. Journal of the Marine Biological Association of the United Kingdom, 33, 393-401. Marshall, S. M. and Orr, A. P. (1955). “ The Biology of a Marine Copepod Calanus finmarchicus (Gunnerus) Oliver and Boyd, Edinburgh. Marshall, S. M. and Orr, A. P. (1966). Respiration and feeding in some small copepods. Journal of the Marine Biological Association of the United Kingdom, 46, 513-530. Marehall, S. M., Nicholls, A. G . and Orr, A. P. (1934). On the biology of Calanus ]%&marchicus.V. Seasonal distribution, size, weight, and chemical composition in Loch Striven in 1933, and their relation to the phytoplankton. Journal of the Marine Biological Association of the United Kingdom, 19, 793-827. Martens, P. (1972). Zur Charakterisierungder Kotballen von vier dominierenden Copepodernarten der Kieler Bucht. Diplomarbeit Thesis, ChristianAlbrechts University, Kiel. Martens, P. (1975). uber die Qualitfit und Quantitlit der Sekundar- und Tertiarproduzenten in einem marinen Flachwasserokosystemder westlichen Ostsee. Doctoral Thesis, Christian-AlbrechtsUniversity, Kiel. May, R. C. (1974). Larval mortality in marine fishes and the critical period concept. I n “ The Early Life History of Fish ” (J. H. S. Blaxter, ed.), pp. 3-19. Springer-Verlag, New York. Mayr, E. (1963). “ Animal Species and Evolution Harvard University Press, Cambridge, Massachusetts. McAllister, C. D. (1969). Aspects of estimating zooplankton production from phytoplankton production. Journal of the Fakheries Research Board of Canada, 26, 199-220. McLaren, I. A. (1963). Effects of temperature on growth of zooplankton and the adaptive value o f vertical migration. Journal of the Fisheries Research Board of Canada, 20, 685-727.

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”.

224

CIIRISTOPHER J. CORKETT AND IAN A. MCLAREN

McLaren, I. A. (1965). Some relationships between temperature and egg size, body size, development rate and fecundity, of the copepod Pseudocalanwr. Limnology and Oceanography, 10, 528-538. McLaren, I. A. (1966). Predicting development rate of copepod eggs. Biological Bulletin, 131, 457-469. McLaren, I. A. (1969). Population and production ecology of zooplankton in Ogac Lake, a landlocked fiord on Baffin Island. Journal of the Fisheries Research Board of Canada, 26, 1485-1559. McLaren, I. A. (1974). Demographic strategy of vertical migration by a marine copepod. American Naturalist, 108, 91-102. McLaren, I. A. (1976a). Inheritance of demographic and production parameters in the marine copepod Eurytemora herdmani. Biological Bulletin, 151, 200-213.

McLaren, I. A. (1976b). Ecological and genetical strategies in Pseudocalanw, production. International Council for the Exploration of the Sea, Plankton Committee, document C.M. 1976/L : 5, 1-8. McLaren, I. A., Woods, S. M. and Shea, J. R. (1966). Polyteny : a source of cryptic speciation among copepods. Science, New York, 153, 1641-1642. McLaren, I. A., Walker, D. A. and Corkett, C. J. (1968). Effects of salinity on mortality and development rate of eggs of the copepod Pseudocalanw minutus. Canadian Journal of Zoology, 46, 1267-1269. McLaren, I. A., Corkett, C. J. and Zillioux, E. J. (1969). Temperature adaptations of copepod eggs from the arctic to the tropics. Biological Bulletin, 137, 486-493.

Mednilrov, B. M. (1961). On the sex ratio in deep water Calanoida. Crz&meana, 3, 105-109.

Menshutkin, V. V., Vinogradov, M. Ye. and Shushkina, E. A. (1974).Mathematical model of the pelagic ecosystem in the Sea of Japan. Oceanology, 14, 717723. (English edition.) Meshcheryakova, I. M. (1964). The quantitative distribution of plankton in the southeastern Bering Sea in the summers of 1958 and 1959. I n “ Soviet Fisheries Investigations in the Northeast Pacific ” (P. A. Moiseev, ed.), Part 11, pp. 141-150 (Trudg Vsesoyuznogo nauchno-wsledovatel’akogo tutu rgbnogo khozyaistva i okeanografii, 49, and Izvestiya Tilchookeanskogo nauchno-iaaledovatel‘skogo instituta rgbnogo kiiozyaistva i okeanograji, 51.) (In Russian.) Miller, A. (1969). Circadian rhythm of feeding of sprat (Clupea sprattus balticus) from the Baltic Sea. Izvestiya Akademii nauk &stonskoi S S R , Ser. biol., 18(1), 16-28. (In Estonian.) Minoda, T. (1967). Seasonal distribution of Copepoda in the Arctic Ocean from June to December, 1964. Record of Oceanographic Works i n Japan, 9, 161-168.

Minoda, T. (1971). Pelagic Copepoda in the Bering Sea and the north-western North Pacific with special reference to their vertical distribution. Memoir8 of the Faculty of Fisheriea, Hokkaido University, 18, 1-74. Minoda, T. and Osawa, K. (1967). Plankton in the sonic scattering layer in the Okhotsk Sea, summer 1963. Bulletin of the Faculty of Fiaheries, Hokkaido University, 18, 9-19. (In Japanese.) Mironov, G. N. (1960). The feeding of plankton predators. 11. The feeding of sagittae. Trudg Sevaatopol’skoi biologicheskoi stantsii. Akademiya nauk SSSR, 13, 78-88. (In Russian.)

225

THE BIOLOGY OF PSEUDOCALANUS

Mori, T. (1937). “ The Pelagic Copepoda from the Neighbouring Waters of Japan ”, published by the author (reprinted 1964, the Soyo Co., Tokyo). Morioka, Y. (1973). The occurrence of cold-water calanoid copepods in the coastal waters of the San’in District, Japan Sea. Bulletin of the Japan Sea Regional Fwheries Research Laboratory, 24, 77-84. Motoda, S. and Minoda, T. (1974). Plankton of the Bering Sea. I n “ Oceanography of the Bering Sea ” (D. W. Hood and E. J. Kelley, eds),pp. 207-241. Institute of Marine Science, University of Alaska, Fairbanks. Muhammad, M. (1965). Succinic acid dehydrogenase activity and oxygen uptake in zooplankton of Bras d’Or Lake, Cape Breton Island, N.S. B.Sc. Hons. Thesis, Dalhousie University, Halifax, Nova Scotia. Mullin, M. M., Sloan, R. R. and Eppley, R. W. (1966). Relationship between carbon content, cell volume, and area in phytoplankton. Limnology and Oceanography, 11, 307-311. Nakai, Z. (1955). The chemical composition, volume, weight and size of the important marine plankton. Tokai regional fisheries research Laboratory, special Publication, 5, 12-24. Nguyen,V. K., Drzycimski, I. and Chojnacki, J. (1972). Feeding and food composition of sprat from Bornholm Depth. Acta ichthyologica et piscatoria, 2, 55-66. Nilritine, B.(1929). Les migrations verticales saisonnieres des organismes planktoniques dans la mer Noire. Bulletin de 1’Institut Oc&anographique,Monaco, 26, NO. 540, 1-24. Nival, P., Malara, G., Charm, R., Palazzoli, I. and Nival, S. (1974). &ude de la respiration et de l’excr6tion de quelques cop6podes planctoniques (Crustacea) dans la zone de remont6e d’eau profonde des c6tes marocaines. Journal of Experimental Marine Biology and Ecology, 15, 231-260. Norris, D. E. and Overstreet, R. M. (1976). The public health implications of larv a1 Thynnascaris nematodes from shellfish. Journal of &!ilk and Food Technology, 39, 47-54. Oberg, M. (1906). Die Metamorphose der Plankton-Copepoden der Kieler Bucht. Wissenschaftliche Meeresuntersuchungen der Kommission zur wissenschaf tlichen Utatersuchung der deutschen Meere, Abt. Kiel, 9, 37-103. Ogilvie, H. S. (1938). The food of post-larval haddock with reference to the annual fluctuations in the haddock broods. Rapport et procbs-verbaux des rbunions. Conseil permanent international pour I’Exploration de la Mer. 107, 57-66. Ogilvie, H. S. (1953). Copepod nauplii (1). Fiches d’identiJication d u zooplancton. Conseil permanent international pour 1’Exploration de la Mer, No. 50. Okada, S. and Taniguchi, A. (1971). Size relationships between salmon juveniles in shore waters and their prey animals. Bulletin oj the Faculty of Fisheries, Hokkaido University, 22, 30-36. Omori, M. (1965). The distribution of zooplankton in the Bering Sea and northern North Pacific, as obRerved by high-speed sampling of the surface waters, with special reference to the copepods. Journal of the Oceanographic Society of Japan, 21, 18-27. IZlstvedt, 0.-J.(1955). Zooplankton investigations from Weather Ship M in the Norwegian Sea, 1948-49. Hvalrddets Skrijter, 40, 1-93. Owre, H. B. and Foyo, M. (1967). “ Copepods of the Florida Current Fauna Caribaea No. 1. Institute of Marine Science, University of Miami.

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10

226

CHRISTOPHER J. CORKETT AND IAN A. MCLAREN

Paffenhofer, G.-A. and Harris, R. P. (1976). Feeding, growth and reproduction of the marine planktonic copepod Psewlocalanus elongatus Boeck. Journal of the Marine Biological Association of the United Kingdom, 56, 327-344. Parsons, T. R. and LeBrasseur, R. J. (1970). The availability of food to different trophic levels in the marine food chain. I n “ Marine Food Chains ” (J. H. Steele, ed.), pp. 325-343. Oliver and Boyd, Edinburgh. Parsons, T. R., LeBrasseur, R. J. and Fulton, J. D. (1967). Some observations on the dependence of zooplankton grazing on the cell size and concentration of phytoplankton blooms. Journal of the Oceanographic Society of Japan, 23, 10-17. Parsons, T. R., LeBrasseur, R. J., Fulton, J. D. and Kennedy, 0. D. (1969). Production studies in the Strait of Georgia. Part 2. Secondary production under the Fraser River plume, February to May 1967. Journal of Experimental Marine Biology and Ecology, 3, 39-50. Pavlova, E. V. (1975). On certain factors influencing rate of oxygen consumption of marine planktonic animals. Biologiya Morya, Kiev, 33, 73-78. (In Russian. ) Pavlovskaya, T. V. and Pechen’-Finenko, G. A. (1975). Comparison of the relative role of living and non-living organic matter in the nutrition of Pseudocalanus elongatus (Boeck). Biologiya Morya, Kiev, 34, 66-70. (In Russian.) Pavshtiks, E. A. and Timokhina, A. F. (1972). History of investigations on plankton in the Norwegian Sea and the main results of Soviet investigations. Proceedings of the Royal Society of Edinburgh, B, 73, 267-278. Pearre, S. (1973). Vertical migration and feeding in Sagitta elegam V e r d . Ecology, 54, 300-314. Pearse, A. S . (1936). Estuarine animals at Beaufort, North Carolina. Journul of the Elisha Mitchell Scientijk Society, 52, 174-222. Peterson, W. T. and Miller, C. B. (1975). Year-to-year variations in the planktology of the Oregon upwelling zone. Fishery Bulletin. Fish and Wildlye Service. United States Department of the Interior, 73, 642-653. Petipa, T. S. (1975). Origin and classification of certain ecological types of feeding of Calanoid Copepoda. Biologiya Morya, Kiev, 33, 27-49. (In Russian.) Petipa, T . S., Sazhina, L. I. and Delalo, E. P. (1963). Vertical distribution of Zooplankton in the Black Sea. Trudfj Sevastopol’skoi biologicheskol stanti% Akademiya nauk SSSR, 16, 119-137. (In Russian.) Petipa, T. S., Pavlova, E. V. and Mironov, G. N. (1970). The food web structure, utilization and transport of energy by trophic levels in the planktonic communities. I n “ Marine Food Chains ” (J.H. Steele, ed.), pp. 142-167. Oliver and Boyd, Edinburgh. Pinhey, K. F. (1927). Entomostraca of the Belle Isle Strait Expedition 1923 with notes on other planktonic species. Part I. Contributions to C a n a d h Biology and Fisheries, 3, 179-234. Polyakova, T. V. and Perueva, F. G. (1976). Oxygen consumption by some planktonic organisms from the White Sea. Zhurnal obshchel biologii, 31, 450-458. (In Russian.) Ponomarenko, V. P. (1967). Feeding of the larvae and fry of the arctic cod (Boreogadus saida Lepechin) in the Barents and Kara Seas. Materia@ rfjbokhozyahtvenngkh issledovanii severnogo bamelna, 10, 20-27. (English transl. Bureau of Sport Fisheries and Wildlife, Narragansett, Rhode Island.)

THE BIOLOGY OF PSEUDOCALANUS

227

Ponomareva, L. A. (1954). On the feeding by euphausids of the Sea of Japan on copepods. Dodladg Akademii nauk SSSR, 98,153-154. (In Russian.) Popova, T. I. and Valter, E. D. (1965). On the elucidation of the life cycle of the fish nematode Contracaecum aduncum (Rudolphi, 1802) Baylis, 1920 (Ascaridata). I n “ Material9 nauchnoi konferentsii Vsesoyuznogo obshchestva gel’mintologov, 1965 ” (V. G. Gagarin, ed.), Part 1, pp. 175-178. (English transl. Fisheries Research Board of Canada, no. 1797.) Porumb, F. I. (1971). Sur la biologie des copbpodes pblagiques des eaux roumaines de la mer Noire. Cercetciri marine, 1, 129-147. Porumb, F. I. (1972). Contributions B la connaissance de la dynamique des populations et B la production de cop6podes dans les eaux roumaines de la mer Noire. Cercetiiri marine, 4, 57-94. Porumb, I?. I. (1973). Recherches sur le zooplankton au-dessus des fonds rocheux du littoral roumain de la mer Noire (aspect printanier). Rapport et procksverbaux des rdunions. Commission internationale pour l’exploratwn scientifique de la Mer Mdditerrande, 21, 533-535. Porumb, I. I. (1969). Contributions h1’6tude de la biologie de Sardina pilchardus sardina. La nourriture devant le littoral roumain de la mer Noire. Lucrcirile statiunii de cerceta’ri marine “ Prof. Ioan Borcea ”, Agigea, 3, 101-112. Poulet, S. A. (1973). Grazing of Pseudocalanwr minutus on naturally occurring particulate matter. Linznology and Oceanography, 18, 564-573. Poulet, S. A. (1974). Seasonal grazing of Pseudocalanus minutus on particles. Marine Biology, 25, 109-123. l’oulet, S. A. (1976). Feeding of Psezldocalanus minutus on living and non-living particles. Marine Biology, 34, 117-125. Poulet, S . A. (1977). Grazing of marine copepod developmental stages on naturally occurring particles. Journal of the Fisheries Research Board of Canada, 34, 2381-2387. Poulet, S . A. and Chanut, J. P. (1975). Non-selective feeding of Pseudocalanus minutus. Journal of the Fisheries Research Board of Canada, 32, 706-713. Prbfontaine, G. and Brunel, P. (1962). Liste d’invert6br6s marins recueillis dans l’estuaire du Saint-Laurent de 1929 B 1934. Naturaliste canadien, 89, 237263. Raymont, J. E. G. (1959). The respiration of some planktonic copepods. 111. The oxygen requirements of some American species. fimnology and Oceanography, 4 , 479-491. Rakusa-Suszczewski, S. (1969). The food and feeding habits of Chaetognatha in the seas around the British Isles. Polskie archiwum hydrobiologii, 16, 213-232. Reeve, M. R., Gamble, J. C. and Walter, M. A. (1977). Experimental observations on the effects of copper on copepods and other zooplankton :Controlled Ecosystem Pollution Experiment. Bulletin of Marine Science, 27, 92-104. Robertson, A. (1968). The Continuous Plankton Recorder : a method for studying the biomass of calanoid copepods. Bulletin of Marine Ecology, 6 , 185-223. Robertson, S. B. and Frost, B. W. (1977). Feeding by an omnivorous planktonic copepod Aetideus divergene Bradford. Journal of experimental marine Biology and Ecology, 29, 231-244. Roe, H. S. J. (1972). The vertical distributions and diurnal migrations of calanoid copepods collected on the SOND Cruise, 1965. I. The total population and general discussion. Journal of the Marine Biological Aasociation of the United Kingdona, 52, 277-314.

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IZoquette, M. de la (1842). Notice annuelle des progrds des sciences g6ographiques et des travaux de la Soci6t6 de geographie pendant l'ann6e 1842. Bulletin de laSocidtd de Gkographie. Paris [2] 18, 397-621. Russell, F. S. (1934). The study of copepods as a factor in oceanic economy. Proceedings of the fifth. Pacific Science Congress, 1933, 3, 2023-2034. Russell, F. S., Southward, A. J., Boalch, G. T. and Butler, E. I. (1971). Changes in biological conditions in the English Channel off Plymouth during the last half-century. Nature, London, 234, 468-470. Ruud, J. T. (1929). On the biology of copepods off Mare, 1925-27. Rapport et Procbs- Verbaux des Reunions. Conseil permanent international pour I'Exploration de la Mer, 56, 1-84. Sars, G. 0. (1899). " An Account of the Crustacea of Norway Vol. 11. Bergen Museum, Bergen. Sars, G. 0. (1900). Part V, Crustacea. Scientific Results. The Norwegian North Polar Expedition 1893-1896 (F. Nansen, ed.), Vol. I , 1-137. Sars, G. 0. (1903). " An Account of the Crustacea of Norway Vol. IV. Bergen Museum, Bergen. Savage, R. E. (1926). The plankton of a herring ground. Fishery Investigations. Ministry of Agriculture, Food and Fisheries, Ser. 2 , 9( l ) , 1-35. Savage, R. E. (1931). The relation between the feeding of herring off the east coast of England and the plankton of the surrounding waters. Fishery Investigations. Ministry of Agriculture, Food and Fisheries, Ser. 2, 12(3), 1-88. Sazhina, L. I. (1968). On individual fecundity and duration of development of certain mass pelagic Copepoda of the Black Sea. Bidrobiologicheskit zhurnal, 4(3), 69-72. (In Russian.) Sazhina, L. I. (1971). Fecundity of mass pelagic Copepoda in the Black Sea. Zoologicheskit zhurnal, 50, 586-588. (In Russian.) Sazhina, L. I. (1974). Rate of reproduction of pelagic Copepoda of the Black and Mediterranean Seas. I n " Biological Production of Southern Seas " (T. G. Kondratskaya, ed.), pp. 175-182. Naukova Dumka, Kiev. (In Russian.) Schnack, S. (1975). Untersuchungen zur Nahrungsbiologie der Copepoden (Crustacea) in der Kieler Bucht. Doctoral Thesis, Christian-Albrechts University, Kiel. Scott, A. (1922). On the food of young plaice (Pleuronectes platessa). Journal of the Marine Biological Association of the United Kingdom, 12, 678-687. Sewell, R. B. S. (1948). The free-swimming planktonic Copepoda. Geographical distribution. John Murray Expedition. Science Reports, B.M. ( N . H . ) , 8, 317-592. Sewell, R. B. S. (1951). The epibionts and parasites of the planktonic Copepoda of the Arabian Sea. John Murray Expedition. Science Reports, B . M . ( N . H . ) , 9, 255-394. Sherborn, C. D. and Woodward, B. B. (1901). Dates of publication of the zoological and botanical portions of some French voyages. Part 11. Annuls and Magazine of Natural History, Ser. 7, 8, 491-494. Sherman, K. and Honey, K. A. (1970). Vertical movements of zooplankton during a solar eclipse. Nature, London, 227, 1156-1158. Sherman, K. and Honey, K. A. (1971). Size selectivity of the Gulf I11 and Bongo zooplankton samplers. Research Bulletin of the International Cornmission for the Northwest Atlantic Fisheries, 8, 45-48.

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Shushkina, E. A., Kislyakov, Yu. Ya. and Pasternak, A. F. (1974). Estimating the productivity of marine zooplankton by combining the radiocarbon method with mathematical modelling. Oceanology, 14, 259-265. (English edition.) Skud, B. E. (1968). Responses of marine organisms during the solar eclipse of July 1963. Pishery Bulletin. Fish and Wildlije Service. United States Department of the Interior, 66, 259-271. Smayda, T. J. (1969). Some measurements of the sinking rate of fecal pellets. Limnology and Oceanography, 14, 621-625. Smith, L. R., Miller, C. B. and Holton, R. L. (1976). Small-scale horizontal distribution of coastal copepods. Journal of Experimental Marine Biology and Ecology, 23, 241-253. Smith, S. L. (1975). The role of zooplankton in the nitrogen dynamics of marine systems. Ph.D. Thesis, Duke University, Chapel Hill, North Carolina. Snodgrass, R. E. (1956). Crustacean metamorphosis. Smithsonian naiscellaneow Collections, 131(10), 1-78. Sournia, A., Cachon, J. and Cachon, M. (1976). Catalogue des especes et taxons infraspecifiques de dinoflagellbs marins actuels publies depuis la revision de J. Schiller. 11-DinoflagellBs parasites ou symbiotiques. Archiv fiir Protistenkunde, 117, 1-19. Harvard UniSteele, J. H. (1974). “The Structure of Marine Ecosystems versity Press, Cambridge, Massachusetts. Steele, J. H. and Frost, B. W. (1977). The structure of plankton communities. Philosophical Transactions of the Royal Society of London. B . Biological Sciences, 280, 485-534. Sullivan, B. K., Miller, C. B., Peterson, W. T. and Soeldner, A. H. (1975). A scanning electron microscope study of the mandibular morphology of boreal copepods. Marine Biology, 30, 175-182. Svetlichnyi, L. S., Zagoradnyaya, Yu.A. and Stepanov, V. N. (1977). Bioenergetics of migrations of Pseudocalanus elongatua (Boeck, Biologiya Morya, Vladivostolc, No. 6, 41-49. (In Russian.) Thompson, B. M. (1976). The biology of Pseudocalanus elongatus (Boeck). Ph.D. Thesis, University of East Anglia, Norwich. Tregouboff, G. and Rose, M. (1957). “ Manuel de Planctonologie m6diterraneenne I, 11. Centre National de la Recherche Scientifique, Paris. Urry, D. L. (1964). Studies on the food, feeding and survival of Pseudocalanus elongatus Boeck under laboratory conditions, with observations on other genera of Copepoda. Ph.D. Thesis, University of London. Urry, D. L. (1965). Observations on the relationship between the food and survival of Pseudocalanus elongatus in the laboratory. Journal of the Marine Biological Association of the United Kingdom, 45, 49-68. Ussing, H. H. (1938). The biology of some important plankton animals in the fjords of East Greenland. Meddelelser om Grenland, 100(7), 1-108. Valter, E. D. (1968). On the hosts of Contracaecum aduncum (experimental infection of animals with larvae of the parasite). I n “ Sed’maya sessiya Uchenogo soveta PO probleme ‘ Biologicheskie resursg Belogo morya i vnutrennikh vodoemov Karelii ’, Mart,1968 goda. Tezisy dokladov. Petrozavodsk.” (English transl. Pisheries Research Board of Canada, no. 2031 .) Vane, F. R. (1952). The distribution of Blastodinium hyalinum in the North Sea. Challenger Society, 3(4), 23-24.

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Vervoort, W. (1965). Notes on the biogeography and ecology of free-living, marine Copepoda. Monographiae biologicae, 15, 381-400. Vinogradov, M. E. and Voronina, N. M. (1963). Distribution of plankton in the waters of the Equatorial Current of the Pacific Ocean, I. Distribution of biomass of plankton and horizontal occurrence of certain species. Trudg Instituta okeanologii. Akademiya nauk SSSR, 71, 22-59. (In Russian.) Vives, F. (1967). Los cop6podos plancthicos del mar Tirreno en septiembre y octubre de 1963. Investigacidn pesquera, 31, 539-583. Vives, F. (1970). Distribuci6n y migracih vertical de 10s cop6podos plancthicos (Calanoida) del SO. de Portugal. Investigacion pesquera, 34, 529-564. Vutieti6, T. (1957). Zooplankton investigations in the sea water lakes “ Malo Jezero ” and “ Veliko Jezero ” on the island of Mljet (1952-1953). Acta Adriatica, 6(4), 1-51. VuEetib, T. (1961). Vertical distribution of zooplankton in the bay Veliko Jezero on the island of Mljet. Acta Adriatica, 6(9), 1-20. Wiborg, K. F. (1940). The production of zooplankton in the Oslo Fjord in 1933-34, with special reference to the copepods. Hvalrcidets Skrqter, 21, 1-87.

Wiborg, K. F. (1944). The production of zooplankton in a landlocked fjordthe NordQsvatn near Bergen in 1941-42. Fiskeridirektoratets skrifter. Serie Havunders0kelser, 7(7), 1-85. Wiborg, K. F. (1948). Investigation on cod larvae in the coastal waters of northern Norway. Fiskeridirektoratets skrvter. Serie Havundersekelser, 9(3), 1-27.

Wiborg, K. F. (1949). The food of cod (Gadus cullarias L.) of the 0-11-group from deep water in some fjords of northern Norway. Fiskeridirektoratets skrifter. Serie Havundersekelser, 9(8), 1-27. Wiborg, K. F. (1954). Investigations on zooplankton in coastal and offshore waters of western and northwestern Norway. Fiskeridirektoratets skrvter. Serie Havundersekelser, 11(l ) , 1-246. Wilson, C. B. (1942). The copepods of the plankton gathered during the last cruise of the Carnegie. Publications, Carnegie Institution of Washington, No. 536. Wilson, C. B. (1950). Copepods gathered by the United States Fisheries Steamer “ Albatross ” from 1887-1909, chiefly in the Pacific Ocean. Bulletin. United States National Museum, 14(4) No. 100, 141-441. Winberg, G. G. (ed.) (1971). “ Methods for the Estimation of Production of Aquatic Animals ”. Academic Press, London, New York. Wing, B. L. (1975). New records of Ellobiopsidae (Protista (incertae sedis))from the North Pacific with a description of Thalassomyces albatrossi n.sp., a parasite of the mysid Stylomysis major. Fishery Bulletin, U.S. National Marine Fisheries Service, 73, 169-185. With, C. (1915). Copepoda I. Calanoida Amphasoandria. Danish Ingo2f Expedition, 3(4), 1-260. Woods, S. M. (1969). Polyteny and size variation in the copepod Pseudocalanus from two semi-landlocked fiords on Baffin Island. Journal of the Fisheries Research Board of Canada, 26, 643-556. Wright, R. ‘R. (1907). The plankton of eastern Nova Scotia waters. An account of floating organisms upon which young food-fishes mainly subsist. Contributions to Canadian Biology, 1902-1905, 1-19.

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Zagorodnyaya, Yu. A. (1974). Nutrition and migration o f Black Sea Pseudocalanus elongatus (Boeck) in the winter period. Gidrobiologicheskii zhurnal, 10(6), 49-56. (In Russian.) Zagorodnyaya, Yu. A. (1975). Vertical migration and daily ration of the copepod Pseudocalanus elongatus (Boeck) in the Black Sea. Biologiya Morya, Kiev, 33, 11-17. (In Russian.) Zagorodnyaya, Yu. A. (1977). Estimation of the value of the die1 phytoplankton grazing by the copepod Psezdocalanus elongatus (Boeck) on the basis of a physiological calculation of its ration. Biologiya Morya, Kiev, 42, 95-1 00. (In Russian.) Zagorodnyaya, Yu. A. and Svetlichnyi, L. S. (1976). Die1 dynamics of the specific gravity and vertical distribution of Pseudocalanus elongatus (Boeck). Biologiya Morya, Kiev, 39, 39-42. (In Russian.) Zelikman, €?.A. (1961). Mass occurrence of Pseudocalanus elongatus Boeck (Copepoda) in the coastal area of eastern Murman in 1956 and its causes. In " Hydrological and Biological Features of the Shore Waters of Murman ", pp. 127-135. AkademiyaNauk SSSR, Kol'skii Filial S. M. Kirova, Murmansk. (In Russian.) Zelikman, €?.A. (1966). Notes on composition and distribution of zooplankton in the southeast part of the Barents Sea in August-October, 1959. I n '' Composition and Distribution of Plankton and Benthos in tho Southern Part of the Barents Sea ", T r u d g Murmanskogo morskogo biologicheskogo instituta, 11(15), 34-44. (In Russian.) Zelickman, E. A. and Golovkin, A. N. (1972). Composition, structure and productivity of neritic plankton communities near the bird colonies of the northern shores of Novayrt Zemlya. Marine Biology, 17, 265-274.

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The study of marine biology may be said to have started as long ago as when Aristotle described many of the animals which live in the sea and wrote in detail on such matters as the migrations of the tunny. These marine animals always held a fascination for the early naturalists because of their great diversity of form. But it was only when enough was known to encourage attempts at classification that their study became truly scientific, developing through Linnaeus to Cuvier and finally to Darwin. It is noteworthy that both Huxley and Darwin went on voyages on which the investigation of marine life occupied much of their time. Huxley, for instance, on the voyage of the “ Rattlesnake ” disclosed the true affinities of the coelenterates, or Cnidaria as they are now known, with their two cell layers, thus amending Cuvier’s classification which combined jellyfish with echinoderms as radiate animals. Darwin produced a classic monograph on barnacles, published by the Ray Society. It was not so very long after the publication of Darwin’s “ Origin of Species ” that an international marine biological laboratory was founded in 1874 by Anton Dohrn a t Naples. This was to be followed during the next decade by similar laboratories in the United Kingdom, France and the United States, so that by the close of the century

* Being the third John Harris Memorial Lecture, delivered a t Bristol University on 4 M a y 1976. (Reprinted from Biology and Human Affaira, Vol. 42, 1976). 233

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there were twenty-one European marine laboratories, either independently established or under university management. The setting up of such laboratories had three main objects. First, the purely academic study of the structure and habits of the many animals that live in the sea, whose diversity and number of species apart from the insects far exceeds those on land. Second, the investigation of marine animals for the furtherance of physiological knowledge, for different tissues and organs are to be found in their simplest form in some of these animals. And, third, and slightly later, for the essentially practical study of the habits and life histories of fish, because fears were beginning to be expressed that the sudden increase in trawling due to the introduction of steam might damage the commercial fisheries. From these small beginnings there has grown an immense organization, found necessary as a result of the realization that many aspects of marine biology have a bearing on the practical affairs of life. Marine biological and fishery research laboratories around the world can now be numbered in their hundreds, while the world fleet of research vessels of all kinds now exceeds a thousand. I propose to mention some of the more important discoveries and fields of research in marine biology that have proved useful, and indeed necessary, for mankind.

I. FOOD FROM THE SEA I suppose that the most obvious field is that of food from the sea, which is now estimated to yield something of the order of nearly a hundred million tons of fish a year. I n Great Britain as long ago as the fourteenth century some were calling for fishing restrictions, and in the sixteenth and seventeenth centuries objections were continually being raised against foreigners fishing in so-called " British waters ". I n succeeding years numerous regulations were brought into force restricting methods of fishing in different areas. Commissions were set up in 1854, 1865 and 1878 to study the availability of fish and no trustworthy evidence could be found that there was any overfishing. The seas were thought to be inexhaustible. As a result an Act was passed in 1868 ruling " that all Acts of Parliament which profess to regulate or restrict the mode of fishing pursued in the open sea be repealed, and that unrestricted fishing be permitted hereafter ". Before the end of the century further experimental closing of certain areas off the coasts of Scotland in which scientific observations were made still led to the conclusion that no serious damage could be done by commercial fishing.

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One of the main contentions by the fisherman was that beamtrawling by steam vessels was destroying the spawning grounds of commercially important fish. Many fishermen thought that marine fish laid their eggs on the sea floor or attached to weeds or rocks. But as early as 1864 the Norwegian naturalist G . 0. Sars had shown that the eggs of cod and gurnards were in fact planktonic, being spawned in the upper water layers well above the sea floor. So one of the first practical investigations at the marine laboratories was to study in detail the early life histories of fish in the sea, and to describe their eggs and young stages so that their distribution could be plotted. Despite the earlier predictions that the supply of fish was inexhaustible, the increasing numbers of fishing vessels and advances in the efficiency of fishing gear provided evidence that there were indeed dangers of overfishing. So, in 1902, an International Council for the Exploration of the Sea was set up, supported by the Governments of a number of European countries, with the object of obtaining by cooperative research sufficient knowledge to enable stocks of fish in the sea to be rationally exploited. For this a thorough knowledge of the biology of marine organisms and of the chemistry and physics of sea water was required, for it is not possible to divorce the fish from their ecological environment. The example of this International Council has now been followed in most regions of the world, and similar organizations have been set up for different areas of the oceans, as well as special Commissions concerned with individual species of fish such as halibut, tuna and salmon. Besides fish, other organisms are much sought after from the sea; seaweeds and squid are eaten in quantities by eastern nations, and shrimps and prawns now form a significant part of the food from the sea. These need also to be guarded. These International organizations have enabled cooperative research to be undertaken over wide areas. Up to the last war this was mainly done to obtain knowledge on the bioIogy of marine fish and their ecology, but since the war, research has been aimed especially at problems of the estimation of fish stocks and their rates of recruitment. While the scientific information thus made available in the North Eastern Atlantic has been sufficient to enable some advice to be given on management of the fisheries, on the administrative level this has not been fully made use of and some species are becoming seriously overfished. This situation should change when legislation on national limits is enacted and countries bordering the sea will have sovereignty over their own areas. This should enable much greater management of local stocks by those countries determined to do so, but it may restrict

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the operations of rescarcli vessels because, already, permission is now needed to visit waters controlled by other countries. The establishment of quota systems for different countries has already started. Much monitoring of biological conditions will thus be required in the future. Indeed the whole problem of stock management still requires a great deal of investigation to find the effects of fishing on the recruitment of each species and the relationship between one species and other competing species and with the ecological environment.

11. FISHFARMING But the supply of food from the sea, even if well managed, is not inexhaustible. It can be supplemented to some extent by the farming of fish and shellfish in confined areas. While the cultivation of freshwater fish has long been practised, as has that of shellfish such as oysters and mussels, the cultivation of marine fish was found to be difficult and awaited the necessary discovery. The difficulty was that most sea fish hatch at a very small size, usually about 3 mm long, and have extremely small mouths. When they have absorbed their yolk they are still only able to capture and eat tiny organisms. It was found just before the last war that a food supply of animals of the right size was available in salt pans in which the brine shrimp Artemia lays its hardy eggs. These eggs can be transported in sufficient quantities to enable their newly hatched nauplius larvae to be used as food. The earlier work on rearing in Great Britain was concentrated on the plaice, for which it was soon possible to obtain survival values of 70% or more. This made the suppIy of baby fish ample for rearing purposes seeing that one female plaice might produce 50 000 eggs. But the cost of rearing plaice to marketable size is still somewhat higher than that of sea caught fish, and the main experimental emphasis is at present on luxury species such as the sole, turbot and halibut. There is no doubt that mariculture will play an increasing part in the future supply of food, but it should be remembered that fish require protein for their nourishment and are thus likely to remain a luxury food. This is not so with shellfish which feed mostly on planktonic plants and can be cultivated on a large scale in unpolluted estuaries. The rearing of shellfish such as oysters and mussels was much advanced by a systematic, study and culture of unicellular flagellates: on which their larval stages can be fed.

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The cultivation of different kinds of shrimps is also now receiving much attention, for shrimps form quite a large portion of our food supply from the sea. Seaweeds are cultivated by the Japanese as food, especially species of the red seaweed Porphyra, or laver. The discovery in 1949 by the British algologist Kathleen M. Drew that there was a stage in the life history of this weed which bored into mollusc shells (the so-called Conchocelis stage) was seized upon by the Japanese, who now culture this stage so that they have a continuous supply of spores for the next stage of development. As a result, the amount of weed cultivated in Japan has grown enormously. Indeed, so beneficial was this discovery to the industry that fishermen at Uzuchi near Tokyo erected in 1963 a polished grey granite memorial on which was inset in bronze a portrait of Dr Drew, their “Mother of the Sea”. Each year they meet in commemoration of her.

111.POISONOUS AND VENOMOUS PLANTS AND ANIMALS Every kind of marine fish is not necessarily good to eat. Many can be very harmful. From time immemorial people have been warned against poisonous species. It was.an ancient Jewish rule that fish with no fins or scales should not be eaten. The puffer fish was recognized as poisonous in the times of the Pharaohs, and the Japanese have for long had regulations about their preparation as food. I n World War I1 American troops were warned never to eat fish which blew themselves up like balloons. It was in fact during that war that attention was really drawn to the dangers of poisonous food from the sea, when so many troops were dispersed over so wide an area of the oceans. More attention was then paid also to the occurrence of venomous animals. I n 1942 the Japanese produced a survival manual for the fighting forces with beautiful illustrations, and in 1943 a book was published in Australia on poisonous and harmful marine fishes. These have been superceded by the great three volume work by Dr Bruce Halstead published in 1965-70. Much research is now directed to the extraction and identification of poisons from fish, which may be dangerous to eat throughout the year or only at certain seasons. The poisons from organisms upon which shellfish may feed is also a special field for research. An outbreak of paralytic shellfish poisoning was recorded in 1793, but it was the much later outbreak which occurred near San Francisco in 1927 which resulted in the instigation of long-term investigations. These showed, ten years later, that the unicellular flagellate

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Goniaulax was the causative organism. The excessive blooming of this flagellate may give rise at times in the sea to " red tides ", whose resulting depletion of oxygen produces mass mortality of fish. Apart from this kind of poisoning, shellfish may of course also be harmful if taken from sewage polluted waters. I cannot leave the subject of dangerous marine animals without mention of the jellyfish Chironex which lives in waters along the Queensland coast of Australia and has been the cause of many bathing fatalities. A person badly stung by this jellyfish may be dead within two minutes. It is in fact the most venomous animal at present known. Its presence is thus a danger comparable with that of sharks in areas frequented by tourists.

IV. UNDERWATER STRUCTURES One of the early practical aspects of marine biology was the attempt to combat damage to the hulls of wooden ships by the boring shipworm Teredo. The Phoenicians and Trojans are known to have sheathed their ships with lead, and Pliny and Ovid wrote about shipworm and the use of lead. I n more modern times the rise of naval fleets and the building of harbours and dockyards increased the need for the scientific study of the deterioration of structures in sea water. Apart from boring animals such as Teredo and a few crustaceans, marine growths in general can also cause trouble. Corrosive pitting may arise in steel hulls under decaying masses of sedentary plants and animals. But even greater harm is caused by the growth of fouling organisms beneath the water line which increases the drag on ships, thus slowing them down and leading to great financial loss. One of the most general methods for the prevention of this growth is by the use of antifouling paints, whose principle is the gradual leaching out of toxic substances such as copper and mercury. It is easy to realize how much fundamental knowledge on the life histories, feeding habits and rates of growth of boring and fouling organisms must be acquired in such research. I remember occasions when we were asked how long ago enemy mines recovered from the sea had been laid. From the size of the marine organisms on them and a knowledge of their rate of growth it. was possible to supply the answer.

V. SHE-DESIGN While the study of the shapes and flight of birds has undoubtedly influenced the design of aircraft, observations on fish have probably played little part in the design of ships. Fish swim totally submerged,

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whereas, except for submarines, ships move on the surface of the sea. No doubt the very idea of a streamline shape must have originated from looking at fish, and this was certainly in the mind of Sir George Cayley, a founder of modern hydrodynamics, and it is on record that fish have been towed in experimental tanks by ship designers. There is of course another factor; fish propel themselves through the water by bending the body. I seem to remember a suggestion that one of the universities might experiment with a flexible boat in which the oarsmen rowed with a metachronic rhythm. Be that as it may, there is still very much interest in finding out how fish get their swimming efficiency, whether by using mucous secretion to reduce friction, by having porous skins, or by changes in the shape of the body. Certain skin movements are indeed to be seen in swimming dolphins. The bulbous swelling now given to some ships beneath the water line to reduce drag is reminiscent of the shape of fish and whales.

VI. ECHO-SOUNDING AND NOISE The introduction of echo-sounding after the first war added a new tool for marine biological research. It was noticed that some of the records showed not only a tracing of the bottom of the sea, but also, a false bottom in mid-water. It was soon realized that these must be caused by shoals of fish, and the use of echo-sounders for locating fish is now general throughout the fishing industry. But it was also found that in the open ocean such traces appeared at different depths and that they moved up and down through the twenty-four hours. These traces have proved to be caused by small oceanic fish and by some plankton animals which make nightly vertical migrations towards the surface. The method has been used to demonstrate that plankton animals move upwards during an eclipse of the sun, the vertical migration being stimulated by decreasing light intensity. Underwater hydrophones are used in war for detecting the presence of enemy vessels by the sounds they make. During the last war listeners were often confused by background crackling noises which rendered sonic listening valueless. These noises were found to be made by a small shrimp AZphaeus, known as the snapping or pistol shrimp because of the clicking noise it can make with its claw. As with birds, these sounds made by the shrimps are now found t o be related to their territorial behaviour. The important thing is that this discovery stimulated almost a new branch of marine biology to study under-

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water noises, which are now known to be made by many marine animals. The playing of records of the mating calls of such fish as the cod may have practical results.

VII. PHYSIOLO~ICAL AND MEDICALASPECTS I n my introduction I referred to the use of marine animals and plants for physiological research. I have also mentioned the puffer fish whose flesh is poisonous to eat at certain times of the year. The poison extracted from this fish has proved of use in research, and this leads me on to the subject of physiological and medical aspects of marine biology. It was early realized that animals would prove useful for physiological research. T. H. Huxley, on the occasion of the founding of the Marine Biological Association of the United Kingdom, remarked especially on this. It is perhaps significant that Huxley, as first President of the Association, has been followed in recent years by A. V. Hill and A. L. Hodgkin. Many marine animals are remarkable in having some organ or tissue which is specially suitable for physiological study. It is probably not generally realized for instance that the first direct evidence that insulin comes from the islet tissue of the pancreas was provided by the angler fish Lophius. This fish has islets up to one centimetre in diameter and it was in this easily isolated tissue that McLeod showed the presence of insulin in 1922. The fact that this special tissue was thus available in an exaggerated size in Lophius supports the hope that in the animal kingdom there will be found one specially suited for each physiological problem. A detailed knowledge of the structure and biology of marine animals is thus worth while. I n this respect none has proved more useful than the squid Loligo, in which J. Z. Young demonstrated the presence of giant nerve fibres. These radiate from two ganglia to different areas of the mantle to enable a simultaneous contraction resulting in the violent backward movement by jet propulsion. The largest of these long fibres is one millimetre in diameter allowing the insertion of an electrode or the rolling out of the axoplasm and refilling with fluids of known composition. I n conjunction with research on the nerve fibre of the squid two other marine animals have proved of great value. A common jellyfish Aequorea has the habit of luminescing. It was found that the luminescent reaction is triggered off by the presence of calcium, even in

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molecular quantities. The substance has been extracted and suitably named Aequorin. It is now in general use in research on the squid axon, for it immediately lights up on the passage of calcium through the axon membrane. The puffer fish Tetraodon produces a poison which on extraction is suitably named Tetrodotoxin. This substance blocks the membranes of nerve and muscle fibres so that they are no longer highly permeable to sodium ions. It is 100 000 times more active than local anaesthetics such as procaine and cocaine and has been used as an analgesic. A somewhat similar poison, Saxitoxin, extracted from flagellates such as Goniaulax, has comparable characteristics. I n the study of nerve physiology the sea hare Aplysia is complementary in value to the squid, for it has a giant nerve cell suitable for similar experiments to those on the squid axon. Marine animals have been much used for research on muscle physiology and here again the common barnacle Balanus has proved most useful for it has a specially large muscle fibre. . It was in the first decade of this century that C. R. Richet and P. Portier, working with extracts from the tentacle of the stinging Portuguese Man of War, Physalia, demonstrated the phenomenon of maphyllaxis or allergy, whereby the subject becomes hypersensitive as successive doses of the extract are administered. A few other examples of the special use of marine animals may be given. The horse-shoe crab Limulus is used for its eye and the skate for its labyrinth. The hagfish Myxine, which lacks a thymus, can be useful in immunological studies ; the sand-dollar Echinorachis has been used for studying anti-cancer drugs; and the spiny dogfish Squalus acanthias for investigating the transport of compounds across the blood-brain barrier. And one should not omit the egg of the sea urchin for the part it has played in developmental biology. And now one hears that the spines of this sea urchin may possibly be used as templates for making artificial blood vessels for coronary surgery. The realization that marine organisms may contain unusual substances has led in recent years to a widespread search throughout the plant and animal Kingdoms. The variety of animal life is so much greater in the sea than on land that it is likely that eventually many useful substances will be found. Many organisms have already been screened, especially for the presence of antibiotics and growth-inhibiting substances that might prove useful against cancer. Many have indeed been found to have cell-growth inhibiting characters, such as the extract from the liver of the clam Mercenaria, but apparently only one has so far proved to be A.M.B.-lB

11

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of practical value. This is an extract from a Carribean sponge Cytotehya crypa. This substance has now been synthesized and is used in the treatment of acute leucaemia. But the search goes on, and since there are thousands of different marine organisms it is necessary that the study of systematics should be encouraged, for the specimens must be correctly identified. Among substances obtained from marine plants alginic acid from brown seaweeds has many uses. It is much used in the food and cosmetic industries. Artificial fibres can be made from it and during the war it was in demand for making camouflage material. It has pharmaceutical and medicinal uses. Gauze impregnated with calcium alginate will prevent bleeding. It is used in material for making dental impressions. It has been shown to have a high specificity for binding strontium, and can thus be used in the inhibition of absorption of radioactive strontium by the intestine. The world supply of seaweed for alginate was 12 800 metric tons in 1970. I n the United States its proportional use in 1966 was as follows : 40% for laboratory, pharmaceutical and dental uses; 30% for bakery and confectionery industries ; and 10% for meat packing. Carageen from a red seaweed was used as a laxative in Roman times. An antibiotic which was extracted from a marine fungus obtained from a sewage outlet off the coast of Sardinia was found to be active against some bacteria which are resistant to penicillin and is now widely used under the name " Keflin ". VIII. PESTICIDE The search for possible useful substances has produced some of other than medicinal value. For instance, fishermen in Japan, using a polychaete worm Lumbrinereis brevicirra as bait, noticed that flies and ants died when they settled on the worms. Isolation of the so-called " nereistoxin ", which was found to have a ganglionic blocking action, led to the production of a new insecticide. This has been marketed as " Padan " since 1967 and is used in the control of the larvae of rice stem borers and other insects. I X . GEOLOGY AND METEOROLOGY The study of fossil marine organisms has always been a prominent feature in geological science. It is, however, only rather recently that there have been major advances in micropalaeontology made possible by investigations on the life histories of marine nanoplankton

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organisms in culture, and of their systematics which have been so advanced by the use of the electron microscope. A knowledge of the present distribution in the oceans of foraminifera has been useful in the study of cretaceous climatic zones. Increased knowledge of the life histories of coccolithophores should prove valuable, for these organisms form stratigraphical markers and are associated with oil shales. Microfossils, whose identity was previously unknown, have been shown by culturing to be the resting spores of dinoflagellates. Their distribution on the sea floor is at present little known, but it is thought that when this has been worked out these cysts will prove useful for studying palaeoclimatic changes and in stratigraphy. Only quite recently the elucidation of the life history of planktonic algae of the genus Pterosperrna, previously thought to be dinoflagellates, has proved of interest, for these organisms whose identity was unknown are used by geologists in stratigraphic studies of oil bearing rocks. There is increasing interest by geologists in the systematics of diatoms, while a study of the habits of living animals is being more used to understand the ecological conditions in past ages. The daily deposition of calcium by hermatypic corals is even being examined in the hope that it will throw light on the length of the day millions of years ago. Short term changes in weather and climate also have a notable effect on marine organisms, whether by altering the sea temperature on which their distribution depends, or on a greater scale such as the cooling or warming of the a.rctic which affects the pattern of oceanic circulation, a phenomenon that we have experienced over the last half century.

X. POLLUTION Until after the last war i t was probably a general opinion that the oceans are so vast that they can afford sufficient dilution and dispersion to allow industrial wastes to be poured into them without causing harm. But ever since World War I, when oil was beginning to replace coal in steam ships, damage to amenities on bathing beaches and the oiling and death of sea birds has become increasingly noticeable. The risks from oil pollution were highlighted by the " Torrey Canyon " disaster in 1967 when some 177 000 tons of crude oil were released to come ashore along the coasts of Cornwall and Brittany. This is the most documented of any such disaster, and this was made possible by

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the existence in the vicinity of an independent marine research laboratory at Plymouth, whose staff was composed of research workers in widely different scientific disciplines. Observations and experiments could thus immediately be made by chemists, physiologists, zoologists and botanists, all with the necessary ecological background and outlook. This is a very strong argument for the retention of such independent establishments in which the staff are not committed to some previously designed aim and whose research is in no way planned. On this occasion it was in fact the detergent oil dispersant which did the most damage to shore life. Now less toxic substances are available, and most Governments have organizations with stocks of material ready to deal with such emergencies. Little damage to marine life is likely to occur in the open ocean from oil which is biodegradable, but this is not so in enclosed seas. I n the Black Sea and in the Caspian, oil has caused damage by reducing the reproduction of algae which form the base of the food chain. It was perhaps in the year 1967 also when it was first fully realized how widespread in the ocean might be pollution from other substances. It was in that year that the occurrence of the pesticide DDT was recorded from penguins in the Antarctic. Since then there have been serious occurrences such as the death of 52 persons from mercury poisoning after eating fish in Japan, and similar pollution has occurred in the Baltic. Most enclosed seas have suffered a gradual build up of pollution and this is even noticeable in so large a sea as the Mediterranean. It should be realized that substances taken up by the smaller organisms get passed up the food chain to be concentrated in the larger animals and plants eaten by man. Damage can be caused to marine life, especially in estuaries, by excessive rise of water temperature created by heated effluents from power stations and other industrial plants requiring water for cooling purposes. This is, however, only a serious problem in tropical area.s, for in those regions marine organisms are often living very near their upper lethal limits of temperature. I n temperate regions damage is rarely found, because the animals and plants are adapted in their normal environment to a wide range of temperatures between winter and summer. Slight warming of the water in winter may in fact be beneficial to some shellfish in very cold winters. It is evident that a major effort in marine biology must in the future be concerned with the monitoring of the environment for pollution and with studying the effects of different substances on marine life before they are allowed to drain into the sea.

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XI. CONSERVATION The part played by marine biological research in the conservation of food supplies has already been touched on. But there is now need to have conservation in littoral and sublittoral regions for other reasons. Pollution, dumping and dredging may upset the natural ecology if not carefully controlled. Tourism may cause damage by over population of beaches, and the recent increase in subaqua diving and the growing demands for marine specimens for teaching purposes may cause some animals to be a t risk. One of the first instances of the effects of subaqua diving was that of the fishery for ormers, Haliotis, in the Channel Islands. These shellfish, which are much enjoyed as food, occur intertidally where they may be collected by hand. With the increase in tourism fears arose that the stocks might be depleted by the many visitors who collected these molluscs on the shore. Diving investigations showed a t first, however, that there was a large enough sublittoral population of ormers to keep the stocks going. But these were soon discovered by increasing numbers of visitors with subaqua equipment and the species is thus a t risk. The British Subaqua Club, which probably has something like 20 000 members, draws up Codes of Practice advising members not to cause unnecessary damage to certain species by over-collecting and spear fishing. Apart from the ormer, the gorgonid coral Eunicella is a favourite species for hand collecting, as is the wrasse for spear fishing. There are, however, many independent divers who collect specimens for profit. Sea urchins are special favourites and also starfish, whose dried skeletons are to be seen on sale in many craft and souvenir shops. While much of this type of collecting is very local and not likely to damage on a large scale, it is probably quite otherwise in the tropics where exot,ic shells and corals are collected in quantities for sale all over the world. Before the last war the few marine laboratories in the British Isles collected specimens for sale to schools and universities for teaching purposes and ran courses in marine biology. Thus there were known collecting areas which became examples of different types of habitat for teaching ecology. Since the war the teaching of biology has so increased that the pressure on some marine plants and animals has grown significantly. It is natural that the most popular grounds to be visited are those whose ecology has been fully described in the literature. There is thus a growing danger that certain animals typical of different habitats may be over-collected and the grounds themselves

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disturbed, not only by students but also by the crowds of tourists. I remember that even early in this century the old fisherman collector a t the Plymouth laboratory, when he found a rare animal, used to plant it out in a special place which he kept secret. When one was asked for he could thus miraculously supply it. Some animals may be wanted for special investigations or demonstrations. For instance, after the first war the spider crab Maia became in great demand for the supply of its blood for demonstrating the chemistry of haemocyanin. It is still in demand, but fortunately it is a common and abundant amimal. A less common animal might be required and its collection would need to be watched. It may in fact prove necessary one day to rear some animals for teaching, just as many unicellular plants are now cultured for feeding and rearing experiments. Thus, it can be seen that there is a need for the designation of some localities as marine conservation areas, both above and below low water mark, and for the limitation of collection of certain animals. I n the British Isles the only area so controlled is Lundy Island, but the whole question is under active consideration. I n other parts of the world there are now many so-called " marine parks ". These may be set aside for recreational enjoyment, but will have restrictions on fishing and collecting. There are many such areas on the coast of Queensland in the Great Barrier Reef area, in America, and in the Pacific. Typical is the Eilat Coral Nature Reserve in the northern end of the Gulf of Aqaba, where a coastal strip 1 2 0 0 metres long is fully conserved. There must be close on 100 such areas now regulated or under consideration. Before any area can be selected a detailed knowledge of its ecology is first required. As long ago as 1938 the coral reef surrounding Green Island off the coast of Queensland was designated a marine reserve, and except for recreational fishing full protection was given to marine organisms. At the last International Conference on Marine Parks and Reserves held in May 1975 at Tokyo, the Government of the Cook Islands offered the atoll of Manuae as a World Marine Park. This was one of the first islands in the South Pacific to be discovered by Captain Cook, who named the two small islands the Hervey Islands in 1773. To recapitulate, the services of marine biologists will be required for keeping watch on the ecology of the following types of reserves. 1. 2.

Scientific Reserves, for the study of marine and estuarine ecology. Conservation Reserves, to maintain unspoilt areas of different ecological types, or areas where rare species occur.

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Educational Reserves, to allow access for teaching purposes to different types of grounds. 4. Recreational Reserves, for the enjoyment of tourists in areas of special scenic beauty where underwater conditions are of special interest and can be viewed in underwater glass-walled buildings or by subaqua diving. 3.

XII. MANAND

THE

MARINE ENVIRONMENT

As well as changing the ecology by overfishing or pollution, man can alter the environment deliberately to his advantage. For instance, in the Caspian Sea the Russians noticed that there was a large area of bottom composed of sandy silt with much detritus, rich in organic matter, in which the fauna was very sparse. They accordingly introduced the polychaete worm Nereis diversicolor from the Sea of Azov in 1939 and 1940. The worm, which is a detritus feeder, thrived and by 1956 its biomass was estimated as one million tons with an annual production two or three times greater. This formed a greatly increased food supply for fish, including the sturgeon. Of course, a number of animals have been introduced in different parts of the world for intensive culture, but there is always a danger that an undesirable pest may be brought with them, such as the slipper limpet which competes with the oyster for food. Biological balance can thus be upset. I n this respect the introduction of species requires very careful consideration. The possibility of introducing the giant kelp into European waters as a source of alginic acid is at present under consideration, and this should be resisted. The Japanese brown seaweed Sargasswm has appeared off the Isle of Wight. It is spreading and will probably prove impossible to control. Other major developments by man may have unforeseen results. There used to be a major fishery for sardines, which appeared off the coast of Egypt at the time of the Nile Flood each year. But with the building of the Assouan Dam the Nile has ceased to pour its seasonal supply of nutrient rich water into the sea and the sardines no longer come to feed there. When the Suez Canal was built, the presence of the Bitter Lakes along its course formed an effective barrier to the migration of animals from the Red Sea to the Mediterranean. But over the years the continual stirring of the water in these lakes by passing ships has brought the bottom salt deposits into solution and the gradual dilution has so reduced the salinity of the water that it no longer forms a barrier. As a result, a number of different animals from the Red Sea are now finding

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their way into the Eastern Mediterranean. If poor quality fish were t o replace good quality fish such an interchange would not be beneficial. It is for this reason that it is necessary to know the fauna and flora of different regions and gain a full understanding of their ecology before some major undertaking can be started. It has, for instance, been suggested that the Panama Canal be replaced by a direct excavated channel, thus putting the Pacific fauna into complete connection with that of the Atlantic. The two faunas are very different and the effect of their mixing would be very difficult to forecast. I n this brief survey I have tried to show how a knowledge of marine biology bears on human affairs. It is absolutely essential for the management of our food resources, and is also necessary so that we may guard against excessive polhtion of the sea, and maintain and conserve some species and their environment. It has practical uses, such as in the prevention of structural deterioration and the growth of fouling organisms. A knowledge of the structure and biology of marine animals and plants is of undoubted value for physiological research and will no doubt play an increasing part in the development of medical science. It is necessary so that the effects of proposed major building works by man may if possible be foreseen. Above all perhaps it is necessary for its own sake, for the great variety of living forms to be found in the sea makes it essential in the teaching of biology. The seas cover nearly three-quarters of the earth’s surface and are nearly two miles in average depth. They thus afford infinitely more living space than can be found on land and will no doubt for many years to come yield for our interest many new and fascinating forms of life, if we will continue to search for them.

Adv. mar. Biol., Vol. 15, 1978, pp. 249-287.

NUTRITIONAL ECOLOGY OF CTENOPHOR€SA REVIEW OF RECENT RESEARCH M. R. REEVEAND M. A. WALTER University of Miami, School of Marine and Atmospheric Science, Miami, Florida, U . S . A .

.. ,. ,. .. .. .. I. Introduction . . .. .. .. 11. Feeding Mechanisms and Behavior . . A. Feeding Mechanism and Behavior in Mnemiopsia . . B. Comparison of Feeding Behavior in Other Tentaculata C. Food of Tentnculata . . .. .. .. .. D. Food and Feeding Behavior of Nuda .. .. 111. Ctenophore Predators . . .. .. .. .. .. .. .. .. .. .. IV. Chemical Composition . . .. .. .. . . . .. .. V. Ingestion Rates .. .. .. .. .. .. .. VI. Digestion .. .. .. .. .. VII. Respiration and Excretion .. .. .. .. . . .. .. VIIT. Growth Rate .. .. .. .. . . .. .. IX. Fecundity .. .. .. .. .. .. X . Growth Efficiency .. .. XI. Seasonal Variations in Ctenophore Populations .. . . .. .. .. .. .. XII. Conclusion XIII. References .. .. ..

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249 251 261 259 263 265 267 269 269 271 272 273 215 276 277 282 283

I. INTRODUCTION There has been a tremendous increase in quantitative studies on the ecological role of planktonic ctenophores in the marine environment during the last decade. Most earlier studies were restricted to observations on ctenophore blooms and their correlation with the rapid decline of the standing stock of the rest of the zooplankton community (Bigelow, 1915; Nelson, 1925; Bigelow and Leslie, 1930; Russell, 1931, 1935; Barlow, 1955; Praser, 1962, 1970; Cronin et al., 1962; Hopkins, 1966; and others). The few earlier observations of living ctenophores were concerned with feeding mechanisms and food types (e.g. Lebour, 1923; Nelson, 1925; Main, 1928; Weill, 1935; Hyman, 1940 ; Nagabhushanam, 1959 ; Kamshilov, 1960a, b) which are reviewed in detail by Fraser (1970). The phylum Ctenophora contains two classes and five orders (Bayer and Owre, 1968). 249

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I. Class Tentaculata. All with tentacles, at least as larvae. A. Order Cydippida. Well-developed tentacles throughout life. B. Order Lobata. Tentacles reduced in adults, with development of large oral feeding lobes, moderately compressed. C. Order Cestida. Body extremely compressed and laterally expanded, tentacles reduced in size. D. Order Platyctenea. Compressed, oral creeping surface, usually two well-developed tentacles, sessile. 11. Class Nuda. All lacking tentacles, even a.s larvae. A. Order Beroida. Carnivores on other ctenophores and other soft-bodied zooplankton. Nearly all detailed studies on distribution, seasonal population changes and ecology refer t o the genera Pleurobrachia, Bolinopsis, Mnemiopsis and Beroe, which are common to inshore waters. Within the class Tentaculata, Pleurobrachia belongs to the order Cydippida, whose members feed using tentacles exclusively. With an adult size in the range 10-20 mm, it is generally smaller than Bolinopsis and Mnemiopsis (order Lobata), which outgrow the tentaculate phase and develop oral lobes and other projections for feeding as adults, in the range 50-100 mm and larger. Beroe on the other hand, in the class Nuda, never develops tentacles or lobes, attains an intermediate size, and preys almost exclusively on other planktonic ctenophores. The phylum is not restricted to inshore waters, however, and members of both the Lobata and Cydippida have been reported in oceanic environments, as well as species of the order Cestida, which are long and ribbon-like, being compressed laterally in the sagittal plane. A fourth order of Tentaculata-Platyctenea are flattened in the oralaboral axis and are partially or entirely non-planktonic as adults. Beyond occasional records of occurrence, very few reports exist which provide information on the ecology of either the oceanic or benthic ctenophores. It may be premature, however, to assume that ctenophores in these environments are rare, or of little significance in terms of their predatory impact. There may be other reasons for their obscurity. Most ctenophores do not preserve in formalin solutions. Exceptions to this general rule are Pleurobrachia and Beroe. Many ctenophores are 80 fragile that anything but the gentlest of collection methods quickly fragments them or renders them as unrecognizable gelatinous masses. Mayer (1912) and Harbison et al. (1978) reported that Leucothea is destroyed by even a slight current of water. They questioned the worth of traditional collection methods for such animals. The latter authors have made numerous observations by SCUBA

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across the Atlantic and reported that ctenophores occur more widely and nearly as abundantly in the open ocean as they do close to shore and suggested that they occupy an important place in the community structure of the open ocean. This review summarizes information on their nutritional biology (from observations on living animals mostly in the laboratory) and discusses attempts to utilize laboratory data to interpret environmental fluctuations.

11.FEEDING MECHANISMS AND BEHAVIOR The feeding behavior of Pleurobrachiu, Bolinopsis and Beroe were well documented by the Institute for Scientific Film, Gottingen, West Germany, under the supervision of Wulf Greve who also described the contents of the films (Greve, 1975a, b). These films may be rented or purchased. We have maintained Mnemiopsis rnccradyi through multiple generations in the laboratory for several years and made detailed observations of its behavior. We have also made observations on living ctenophores in the laboratory representative of all the other orders. We also draw heavily on the report of Harbison et al. (1978) to provide information on oceanic forms. A. Feeding mechanism and behavior in Mnemiopsis The newly hatched larvae of M . mccradyi Mayer are equipped with the typical cydippid tentacles and sheaths and eight equally spaced meridional rows of plates composed of transverse bands of long fused cilia known as comb rows. This is the tentaculate stage (Fig. 1A). As the larva grows, it loses the tentacular sheath and the tentacles move orally to lie alongside the mouth in a very reduced form (Fig. 2F). The compression of the oval body of the larva can be seen in 5-6 mm animals (Fig. lB), which results in 4 of the 8 comb rows being longer. The 4 shorter rows form processes known as auricles, with a ciliated edge above the mouth, two on each side, and begin to appear in animals of 8-9 mm in length (Pig. 2F). The longer ones form rounded muscular lobes on either side of the mouth, and by the time the animal attains 1 6 1 5 mm the transition is complete (Fig. 2A-D). Both the larva and adult feed on actively swimming prey such as copepods. During the transition period both tentaculate and lobate methods of feeding may be employed, in proportion to the relative size of tentacles and lobes (Fig. 1B-D). Mnemiopsis larvae can entirely retract the two tentacles into their

FIQ.1. Mnemiopsis. A. A 6 mm larva sets its tentacles for fishing. B-D. Prey capture sequcnce in an 8 mm larva showing the beginning of lobe development. B. Several copepods are entangled in the tentacle which is rapidly contracted. C. The larva rotat,es to position the prey-bearing tentacle so that further contraction (D) will draw the copepod t o the mouth. (p prey, t tentacle).

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tentacular sheaths on either side of the body, between the oral and aboral poles. In order t o " set " their tentacles for food collection animals swim vigorously, oral pole forward, often in a curved or helical pathway, relaxing and trailing their tentacles behind them, while the lateral branches also relax and expand up to two bodywidths from the main tentacle. Figure 3A-D and E-F show two sequences of animals (photographed in aquaria) setting tentacles. As the tentacles reach an extension over 10 times the body diameter, the ctenophore body comes to rest as if restrained by the drag force of total tentaculate area. The tentacles can be some 25 times longer than the body diameter of the animal. The animals drift in this manner, often until disturbed by contact, such as with another animal or air-bubble (in the experimental container) or food organism such as a copepod. I n the absence of contact, animals will contract and then reset their tentacles, sometimes within a few seconds of the first " trial ". However, if prey is captured, it becomes entwined in the tentacular branches which are lined with sticky colloblast cells. It is generally believed (see Hyman, 1940; Fraser, 1970) that prey contact stimulates the release of colloblast filaments from the colloblasts borne on the tentacles, in a manner similar to the nematocysts of Cnidaria, but sticky rather than poisonous. These serve the purpose of restraining the prey as the main tentacle and branches are rapidly contracted to bring the food adjacent to the mouth. Weill (1935), Hyman (1940) and Ralph and Kaberry (1950) believed a poison was involved, but our observations indicate that after capture a copepod continues to struggle and occasionally frees itself and swims away. The ctenophore manipulates its tentacle into the mouth, deposits the prey, and withdraws its tentacle (Fig. 1B-D). Food is positioned at the mouth of the animal by quickly retracting its prey-carrying tentacle and by performing a 180" rotation which moves the foodbearing tentacle adjacent to the mouth (Fig. lC, D). The animal immediately resets its tentacle. Examining the prey after ingestion, small sections of the tentacle can be seen wrapped around it (Walter, 1976). The tentacles must presumably be continuously growing from the base to compensate for this loss. Lobate ctenophores searching for food swim mouth forward (Fig. 2C) with their lobes spread open like wings, upon the surface of which prospective food organisms impinge. The prey appear to become enmeshed on the mucus-lined inner surface of lobes which also contain colloblast cells. The currents set up by the auricular cilia move the prey towards the small tentacles (which further immobilize it), and dong the pharyngeal grooves into the mouth (Fig. SC, F). Newly

FIG.2.

(For legend see facing page)

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FIG.2. Mnemiopsis adult (total length 70 mm). A-B. Tentacular and sagittal plane vicws of animal. C. Oral view of animal swimming with lobes extended. D. Animal with lobes folded to enclose a Chamber. E. A bolus of captured copepods partially ingested. F. Close-up oral view. (p prey, t tentacle, 1 lobe, a auricle, m mouth, pg prey copepod in pharyngeal groove, c comb row).

ingested copepods can be seen to be still active as they move up the pharynx into the stomach area (Fig. 2F). Digestion time varies from 1-3 h from larva to adult a t 21°C. Larvae of M . mccradyi exhibit feeding behavior patterns in small containers which are dependent upon their level of starvation. When starved for 24 h the larvae extend their tentacles fully, frequently changing position and re-setting as described above. If no food is encountered, these search periods are followed by periods of inactivity. When food becomes abundant tentacle length is reduced to 1-2 x their body diameter (Fig. 3A), and animals make frequent contact with food as they swim actively until their guts are full. At this point, active swimming and feeding cease and their tentacles become extended to 2-3 x their body diameter as they drift. This behavior of the larvae is comparable to that described by Rowe (1971) for Pleurobrachia. I n a tank they remain active, although not necessarily feeding, if there is food present. However, in a tank with no food they tend to clump together at the surface. Lobate adults appear bell-shaped under starved conditions

FIQ.3.

(For legend see facing page)

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FIG.3. Mnemiopis larvae (8 mm). A-D and E, F. Two sequences showing setting of tentacles, the first in a curved and the second in a helical pathway.

(Fig. 2A-C). They often swim vertically up and down in an aquarium with their lobes extended a t right angles t o their direction of travel. The lobe width in this condition can be up t o 116% of the animal length. Upon the addition of food to the aquarium the lobe width decreases to about 79% of the body length (Fig. 2B) while the animal feeds. When a number of copepods are in the oral area, the ctenophore will fold its muscular lobes inward (Fig. 2D), which prevents the copepods from escaping and thereby ensures their ingestion. As with the larvae, adults alternate periods of inactivity a t either the surface or bottom of the aquarium, with active searching under prolonged conditions of starvation. Introduction of food invariably stimulates activity. I n high food concentrations (see Walter, 1976) after having been starved for 24 h, lobate ctenophores begin to exhibit what appears to be superfluous feeding. Once their guts are full they still continue to " feed ') by entangling prey in mucus, which results in a bolus a t the oral region (Fig. 2E). Quite frequently they will either " spit out " this enmeshed ball of copepods or completely evacuate their guts, and continue to feed. At the same time mucus strands are released into the water column in which other copepods become entangled. At times mucus strands containing 2-3 copepods can also be seen trailing from the oral region of the ctenophore, which eventually

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break away. This behavior pattern can continue for several hours until the concentration of food is reduced to a point where all the copepods that are captured can be ingested. By this time, the bottom of the experimental container is covered with a layer of partially digested, and undigested but dying copepods. Examination of the guts of these animals after one hour of exposure to low and high concentrations of food indicates a definite difference in the amount of digestion that has taken place. In high concentrations the food organisms are still whole and recognizable, whereas in low concentrations the copepods are partially digested and, in some cases, indistinguishable. Visual observations were made of lobate Mnerniopsis passing under the RSMAS laboratory dock on several occasions, and SCUBA observations were made off Bimini in the Bahamas on two patches of ctenophores of the genus Bolinopsis. I n most cases the animals were oriented against the current as in the laboratory, which potentially brought food to impinge on their extended lobes. An adult Mnerniopsis which had been starved in the laboratory for three days was also set free into the sea and observed for an hour in the natural environment. It exhibited the same behavior seen in the laboratory, i.e. swimming vertically up and down in the water column. On seven separate occasions when patches of ctenophores were observed in the water off the laboratory dock, lobate animals were individually withdrawn and isolated in copepod-free water for immediate microscopical examination of their gut contents. Additionally, quantitative 200 pm mesh net tows were made to obtain an estimate of the density of copepods in the vicinity o f the ctenophores a t the time of capture (Walter, 1976). At no time was the environmental food concentration in excess of 1 0 0 0 copepods/m3. Animals appeared to be feeding actively down to a t least 100/m3. The number of food organisms in the guts of some 200 ctenophores was also recorded as a function of animal length (Walter, 1976). These data confirmed laboratory observations (Reeve et al., 1978) that large lobate ctenophores can hold up to 100 food items in their gut a t any one time, but are contrasted to laboratory observations in that they accomplish this a t much lower food concentrations. At 1 000/m3 in the laboratory, ctenophores are rarely seen with food in their guts. Reeve, et al., (1978) suggested that this discrepancy may be due to the wild animals encountering small patches of food at much higher concentrations. These observations, measurements and photographs on both the larvae and adults under different conditions of food availability show that the mechanism of feeding can be adjusted to the degree of starvation of the ctenophore and the concentration of the food. The

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presence of such a wide range of control of the food collecting mechanism is surprising in light of the quantitative observations reported in Section V on the relatively automatic nature of the ingestion process. Because the food collection process is continuous in lobate ctenophores, i.e. not interrupted by the act of ingestion, there appears to be no practical upper limit to food density at which collection rate becomes independent of food density (i.e. no critical food concentration). I n tentaculate ctenophores, food collection is discontinuous because the tentacle must be withdrawn from fishing t o permit ingestion. As food concentration increases, the proportion of non-fishing time must also increase and Reeve et al., (1978) showed that at very high food densities their ingestion rate reaches a maximum and becomes independent of further food density increase. B. Comparison of feeding behavior in other !l'entacuZata Our observations on the feeding behavior of Pleurobrachia bachei Agassiz from British Columbian waters also cultured in our laboratory differ in some details from that described for Mnemiopsis larvae. I n laboratory aquaria, similarly-sized members of the two genera can be distinguished in their behavior because Pleurobrachia (Fig. 4A) is a more active swimmer trailing short tentacles, and rarely drifts with tentacles a t their maximum extension. I n this way it is more of it search than an ambush predator, when compared to larval Mnemiopsis. Greve (1975a, b) reported that an animal can contract one tentacle which has caught a prey organism while permitting the other to remain extended and fishing. As the tentacle contracts and prey approaches the mouth, Pleurobrachia begins to rotate in a plane which draws the tentacle across the mouth region. The portion of the tentacle bearing the prey gains entry to the mouth (Greve, personal communication) or is draped over the mouth and the prey is " wiped into ') the mouth (Rowe, 1971). I n Pleurobrachia (Greve, 1975a, b ; Rowe, 1971 ; Walter, unpublished) the animal may complete several rotations, but Mnemiopsis larvae (Walter, 1976) rarely rotate more than 180" to accomplish the same end. The portion of their tentacle attached to the copepod breaks off in the mouth and is digested. Very little is known in detail about the feeding of other members of the Cydippida. Representatives of such genera as Hormiphora, Tinerfe and Callianira have been observed by ourselves and Harbison et al. (1978). Unlike the simple filamentous threads of Pleurobrachia and Mnemiopsis, the tentacles of Hormiphora bear more than one kind of side branch (Mayer, 1912) which do not extend very far from the

Fro. 4. (For legend see facing page)

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FIG.4. A. PEeurobrachia. B. Beroe, showing mouth, 45 mm. C and D. VaEticula 15 mm. (body of animal not visible on substrate in C). E. Bolinopsis, 40 mm. F. Ocyropsis, 30 mm, swimming by muscular flapping of lobes. All photographs are of living animals, and in aquaria (except D, which was attached to glass in a dish).

body of the animal, and give it the appearance of already having captured small prey organisms. H . plumosa Agassiz carries large batteries of colloblasts on its tentacles (Harbison et al., 1978) similar to the batteries Qf nematocysts carried by some siphonophores, which might indicate an adaptation to the capture of larger prey such as small fish. Callianira is not readily recognizable as a cydippid in life because its pattern of movement through the water is very uncharacteristic. Instead of the sedate gliding motion of most ctenophores, this animal moves vigorously and rapidly through the water, making observations on its feeding very difficult. Amongst the lobate adult forms, Bolinopsis (Fig. 4E), which we have observed often in the laboratory, moves and feeds in a manner similar to Mnemiopsis. It is more susceptible to damage than Mnemiopsis and frequently suffers lobe atrophy in laboratory aquaria. It is also similar to Mnemiopsis in occurring in dense swarms in inshore waters. Harbison et al. (1978) described the feeding behavior of the lobate genera Eurhamphaea, Leucothea and Ocyropsis. The first two are progressive elaborations of the lobate feeding mechanism. In Euramphaea

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the wide-spread mucus-covered lobes enmesh copepods impinging on their inner surface, and contract in the region of contact moving the prey nearer the labial ridge, along which it is moved by tentacles and cilia to the mouth. The auricles move back and forth and may trap food themselves, or push a copepod onto the lobe surface. While feeding, Euramphnea is either motionless or moves slowly, lobes forward (as does Mnemiopsis). Harbison et al. (1978) reported that Leucothea multicornis Quoy & Gaimard, reaching 20 cm in diameter, has developed lobes larger than other members of the order. The huge diaphanous lobes are very delicate and the animal remains motionless while feeding, apparently relying on the constantly moving auricles to keep the lobes spread out by maintaining gentle water currents. I n addition to the auricles and mucus lined lobes, this ctenophore also has two very long tentacles, which extend beyond the confines of the lobe area. L. ochracea Mayer was reported to have numerous simple side branches on these tentacles (Mayer, 1912), which suggested to Harbison et al. (1978) the possibility that this genus also retained the typical cydippid tentaculate fishing ability. Ocyropsis (Fig. 4F) cannot be confused with any other lobate ctenophore when alive, because it is unique in being able to propel itself vigorously through the water in an aboral direction by a series of bursts of activity, involving muscular contraction of the oral lobes in unison, to give the impression of flapping wings (see descriptions of Mayer, 1912, and Harbison et al., 1978). It is also capable of the normal oral or aboral gliding movements produced by the comb rows. Both reports noted a lack of tentacles in the adults, and Harbison et al. (1978) stated that no mucus was evident on the lobes. They described a feeding mechanism quite different from other lobate adults, which we have also observed in the Iaboratory. The mouth of Ocyropsis is muscular and quite prehensile and " when a prey organism touches the inner lobe surfaces the lobe edge quickly curls over it. The entire lobe contracts somewhat toward the mouth, which simultaneously reaches over and snatches the prey out of the lobe. The feeding action takes a second or less '' (Harbison et al., 1978). They reported also that it could capture larger zooplankton such as small fish and euphausids. I n the mobility and extensibility of its mouth we are reminded of Beroe, although even Pleurobrachia has some ability to manipulate food with its lips (see above). The order Cestida, the ribbon ctenophores, are not uncommon in tropical waters and well-known for their serpentine swimming motion. Harbison et al. (1978) reported, however, that in nature they rarely

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move in this manner, except when disturbed, Instead, the animal hangs motionless in the water, or swims slowly forward with the oral edge leading (as do most lobate ctenophores), its oral tentacles extended up t o the width of its body. Copepods contacting the tentacles are transferred to the oral groove by contraction of the tentacle, and moved along the groove to the mouth. We have had the opportunity to observe the feeding behavior of the Platyctenea as represented by Vallicula (Fig. 4C, D), a relatively small creeping form, which has lost its comb rows and attaches to macroalgae such as Halimeda. Valliczcla is easily overlooked, being inconspicuous, flattened and colorless. It is usually recognized first when an aquarium tank containing live algae in the laboratory is seen to have long delicate single side-branched tentacles (Fig. 4C) stretching and undulating in the circulation throughout the water column. On closer inspection these tentacles emanate not from obvious planktonic cydippid-type larvae, but small slime-like blobs attached to the surface of the algae. The delicacy and usual extension of the tentacles are reminiscent of lobate larvae such as Mnemiopsis, and contact with food elicits the usual ctenophore response of rapid tentacle contraction. I n good food conditions, they grow quickly and multiply by " a kind of fission which recalls pedal laceration in anemones " (Bayer and Owre, 1968). I n an aquarium, a t least, within a few days, a mass of delicate tentacles fill the available water column. Although they do not favor attachment to the bottom sediment, aquarium side-walls provide an acceptable site for them.

C. Food of Tentaculata The Cydippida and Lobata have almost entirely been cansidered to be exclusively carnivores but suggestions have occasionally been made (e.g. Nelson, 1925; Miller, 1970; Miller and Williams, 1972) that they must also rely on phytoplankton or detritus a t times. Baker and Reeve (1974) established experimentally that Mnemiopsis mccradyi starved when exposed to phytoplankton or detrital suspensions only. Hirota (1974) noted that on the rare occasions when phytoplankton was seen in the guts of preserved Pleurobrachia, it could have been originally in the gut of ingested copepods. Harbison et al. (1978) reported that Eurhamphaea vexilligera Gegenbaur could trap fine particles, such as carmine, and ingest them, but they did not know if this had any significance. Baker and Reeve (1974) noted that in a densely colored phytoplankton culture, some coloration of the gut of Mnemiopsis was observable, due no doubt t o adhesion of some cells

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to the mucus on the lobes and its subsequent ingestion. Survival of the ctenophores compared to animals in filtered water was not extended. To a greater or lesser extent, probably most zooplankton animals are susceptible to predation by Cydippida and Lobata, depending on the relative size of the organisms involved, and other considerations such as swimming ability of the prey. Fraser (1970), on the basis of gut analyses of preserved specimens, confirmed Lebour’s (1922, 1923) observations that they were miscellaneous carnivores. I n virtually any plankton sample, crustaceans, particularly copepods, predominate and this alone assures that copepods are their main source of food (Fraser, 1970; Hirota, 1974). Bishop (1968)reported that Bolinopsis and Pleurobrachia select small (1 mm) Pseudocalanufl in preference to Epilabidocera (3 mm), but both Lebour (1922)and Kamshilov (1959) considered the large Calanus to be the major food source of Bolinopsis. Walter (1976)has seen adult Calanw actively struggle free from the tentacles of Pleurobrachia. The experimental conditions (density of ctenophores and prey, size of container and condition of organisms) probably greatly affects the outcome of such experiments. Kremer (in press) found that in natural mixtures containing mostly cyclopoid copepods and veliger larvae, feeding rate of Mnemiqsis leidyi Agassiz was much reduced compared to when feeding on calanoid copepods and cladocerans. She cited prey behavior and palatability as possible reasons, but the small size of the former group may also be significant. Whenever food, which was offered to tentaculate ctenophores in aquaria, contained significant percentages of barnacle nauplii, we always observed that by the next day only barnacle nauplii remained. We subsequently observed encounters between these nauplii and the outstretched tentacles. When copepods of similar size make contact, they are immediately enmeshed in adjacent sidebranch tentacle strands, and rapid contraction towards the mouth occurs. Swimming barnacle nauplii do not disturb the fishing tentacles and the nauplii do not adhere to them. Preliminary experiments suggest that the ctenophores do not actively avoid capturing barnacle nauplii. Extracts of fresh nauplii in the surrounding water do not prevent the normal capture of copepods, nor does the extract of copepods stimulate capture of nauplii. The passive food capture mechanism of the lobate Mnemiopsis does not discriminate between copepods and barnacle nauplii , which both appear to be trapped on the mucus-lined lobes. Quantitative feeding experiments over four days confirmed that tentaculate Pleurobrachia and Mnemiopsis larvae lost weight when only barnacle nauplii

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were available. It is possible that the slow sculling action of the swimming barnacle nauplius does not stimulate the tentacle in the way that the copepod does, with its higher frequency vibrations. Other workers have suggested that swimming behavior affects chances of catchability by tentaculate ctenophores. Anderson ( 1974) made a study of the vertical distribution of zooplankton in St Margaret’s Bay, Nova Scotia. He attributed the small proportion of the copepod Pseudocalanus in the guts of captured P ~ ~ u r o ~ r a c ht oi athe fact that the population center of the copepod was deeper, and to its tendency t o a hop and sink swimming pattern which caused it to retrace its path on sinking. Temora, on the other hand, lives closer to the surface and swims more actively in a horizontal direction. Its chance of encountering tentacles is greatly increased and was reflected, according to Anderson, in its capture rate relative to its abundance. Oithona moves relatively little and was caught less frequently than any of the other common copepods. The same copepod, however, formed the largest fraction of the diet of the lobate Bolinopsis, possibly because passivity is no defense against the feeding mechanism of this ctenophore, and activity could be a potential survival factor. Since both types of ctenophore coexist, however, the swimming behavior of Oithona may have developed more through its own predatory requirements than any defense mechanism. According to Hirota (1974) potential prey of Pleurohrachia can be divided into three categories, those which are too strong and break away from the tentacles, those which struggle but get more entangled, and those which are passive ‘‘ often being dislodged from the tentacle hold, or, like some decapod larvae, are too spiny to be readily ingested and not eaten ”. Ctenophores have long been suspected as having serious effects on fisheries, not only through competing for the same food source, but by providing an alternative but less nutritious food which delayed summer fattening of herring (Manteufel, 1941) and perhaps (Scott, 1913) by retarding the sexual maturity of mackerel, and possibly by destroying fish eggs. Fraser (1970) discounted this latter possibility in Scottish waters on the basis of gut contents over four years, but cited the earlier workers’ positive beliefs regarding predation on the larvae of oysters and other shellfish.

D. Food and feeding behavior of Nuda The feeding behavior of Beroe (Fig. 4B), which preys on other lobate and tentaculate ctenophores, has been studied by Kamshilov

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(1959, 1960a, b), Horridge (1965), Greve (1970), Anderson (1974) and Swanberg (1974). Greve (1970) found prey specificity in that Beroe cucumis Fabricius fed successfully only on Bolinopsis infundibulum (Martens),while B. gracilis fed entirely on Pleurobrachia pileus Fabricius. Although Anderson (1974) reported that in aquaria B. cucumis would feed on either Pleurobrachia or Bolinopsis, Greve et al. (1976) believed this to be a rare occurrence and noted that although B. cucumis was found in the North Sea in the presence of Bolinopsis, i t was not abundant. They speculated that the Bolinopsis population did not support population growth of the beroid. They cautioned care in the interpretation of earlier literature regarding prey preferences of these two species because Bolinopsis do not preserve in formalin, and young of the species of Beroe are particularly difficult to separate. I n our limited experience of temperate-water Beroe cucumis we have been unable t o induce them to feed on Pleurobrachia. Swanberg (1974), on the other hand, stated that he observed B. cucumis and B. ovata Bosc to feed on any ctenophore with which they came in contact. These did not include Pleurobrachia spp. which were absent from his locality but did include such aberrant forms as Cestum veneris Lesueur (Venus’ girdle). Authors generally agree that beroids feed exclusively on other ctenophores. Greve (1971) and Swanberg (1974) reported instances of “accidental” cannibalism in which the smaller ingested animal was egested alive. Hernandez-Nicaise (personal communication), however, has fed Beroe on salps. Greve et al. (1976) suggested that warmer water beroids became progressively less specialized in their feeding in response to the requirements of “ optimal foraging ” in the two systems, and speculated on pathways of speciation of the class in relation to its food. A great deal of experimental work on food preferences, survival and growth rates are required before the somewhat clouded picture of feeding habits of Nuda can be clarified. Beroids feed on prey which range greatly in relative size. I n the case of prey larger than themselves, they appear to attach themselves and suck the prey tissues into their mouth (see Greve, 1971). According to Swanberg (1974), however, some 3 000 macrocilia, arranged in hexagonal arrays and forming part of a ciliary band around the inside of the entire mouth, beat inwards forcing prey tissue into the pharynx. When filled to capacity the enveloping lips contract, and the sturdy cilia cut through the tissue ‘‘ in the manner of moving teeth ”. Horridge (1965) described the fine structure and Swanberg ( 1974) illustrated his account by scanning electron micrographs. A young Beroe, introduced accidentally with a live sample of zoo-

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plankton as food for a population of Mnemiopsis, can easily go unnoticed. On more than one occasion we have discovered a large Beroe and no Mnemiopsis on revisiting the aquarium several days later. We have also observed large beroids t o quickly ingest smaller Bolinopsis in a manner graphically described by Horridge. The animal swims slowly oral end forward, and when its lips contact the smaller prey organism it “ opens its mouth (Fig. 4B)and gives a great gulp. The gulp is caused by the contraction of radially arranged muscle fibers throughout the whole animal. The mesogloea stiffens and the animal expands in size, drawing water rapidly into the mouth ” and the prey with it. Swanberg (1974) showed that Beroe swims more actively in the presence of prey organisms and suggested chemical recognition of food. Kamshilov (1960a) noted that swimming activity decreased with progressive starvation, which we have also seen for Mnemiopsis and Pleurobrachia. I n a laboratory aquarium the latter organisms tend t o clump together a t the water surface after 2-3 days of starvation, but separate and become active immediately when fresh food is provided.

111. CTENOPHOREPREDATORS Besides Beroe, the population dynamics of which have been extensively investigated in relation t o populations of other ctenophores (Greve, 1971; Hirota, 1973, 1974) there appear t o be no usual predators of large blooms of ctenophores, although this assumption might be attributable to difficulties in recognizing their remains in the gut contents of potential predators. Our unpublished observations of Biscayne Bay, Florida, populations of Mnemiopsis mccradyi and Saanich Inlet, British Columbia, Pleurobrachia and Bolinopsis where beroids are rare, suggest no major predator. Fraser (1970) reviewed several earlier observations showing that they were occasionally seen in the diet of some fish and the medusa Chrysaora. This scyphomedusan was reported t o be an important predator by Miller (1974) in the Pamlico River Estuary, North Carolina. Greve (1 972) reported the results of numerous encounters between Pleurobrachia and potential predators. These included the annelid worm Tomopteris, fish larvae, a hermit crab, a crab and a shrimp. On being contacted by medusae, Pleurobrachia discharged mucus which decreased the numbers of successful catches making them more easily detached. Greve (1972) also described experiments which suggested that

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newly-hatched Pleurobrachia were susceptible to having their tentacles tangled in the appendages of copepods which could result in the " destruction " of the ctenophore, or a t least, damage to its tentacles. He considered (personal communication) that large copepods such as Calanus could exert a drastic effect on larval ctenophore populations in this way. The work of Greve was done in small dishes where conditions were far from natural. Our observations in 30 1. aquaria, in which copepod nauplii were provided as food, suggest that mortality of the newly-hatched larvae of Mnemiopsis is very high a t densities of Acartia of 100/1., but a t lO/l., there is no effect. The lower density is rarely exceeded for total zooplankton in a 200 pm mesh net in Biscayne Bay from whence the ctenophore and copepods are derived. Colder waters are more likely to have higher densities a t peak production periods. The situation is made more complicated by the fact that a t intermediate copepod densities some mortality was observed, but those ctenophores which survived through the first 3-4 days became large enough to make use of the copepods and grow much more rapidly than the control animals. The analysis of this type of dynamic interaction of prey and predator requires further study. Fish have been reported a t various times as predators of Ctenophores, the most recent and quantitative account being provided by Oviatt and Kremer (1977). These authors reviewed a variety of reports concerning predation by the ocean sun fish, bluefin tuna, cod, sardine and flying fish. They reported that in Narragansett Bay (Rhode Island), the butterfish (Peprilus) could consume enough Mnemiopsis to support the metabolism of the fish, and account for observed population declines of the ctenophore a t certain times of the year. Hirota (1 974) reported on the infestation of a Pleurobrachict bachei population off California by the endoparasitic larval stages of the amphipod Hyperoche. Harbison et al. (1977) and Harbison et al. (1978) discussed predatory and parasitic associations of hyperiid amphipods with a variety of gelatinous zooplankton, including ctenophores. Some of these could be considered parasitic when small and predatory when large. Flores and Brusca (1975) did not see any permanent damage by two species of Hyperoche on Pleurobrachia, even though amphipods were sometimes present in 100% of the population. Harbison et al. (1978) noted that Oxycephalus can reduce ctenophores and salps to fragments within minutes. A female with young, however, does not feed, but releases the young from her marsupium onto the host, where they commence to feed. These authors have also seen predatory attacks on ctenophores by the heteropods Pterotrachea and Cardiapoda, and the hydromedusa Aequorea.

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COMPOSITION IV. CHEMICAL Ctenophores have long been recognized t o be relatively poor for their size as a food source for other animals on account of their watery body tissue. The percentage of dry material t o live (wet) weight ranged between 3 - & 5 * 0 ~(Cooper, 0 1939 ; Curl, 1962 ; Reeve and Baker, 1975 ; Kremer, 1976a). Most of this dried material is, necessarily, inorganic (ash), the ash-free dry weight (by ignition) being some 20-37% of the dry weight according t o these authors and Hirota (1972). Ash-free and organic dry weight are usually taken to be synonymous terms and the organic carbon content of living tissue comprises about half of this. Curl (1962) gave a value of 21% for Mnemiopsis. Baker (1973) and Reeve and Baker (1975) reported, however, that carbon content in Mnemiopsis mccradyi was only 9% of its ash-free dry weight, which was similar to a value for Pleurobrachia bachei determined by Mullin and Evans (1974) and Kremer (1976a) for Mnemiopsis leidyi. Reeve and Baker (1975) and Mullin and Evans (1974) discussed possible reasons for these discrepancies, and the former agreed with the suggestion of the latter authors that the determination of dry weight was probably a t fault because a substantial fraction of '' bound " water was not driven off a t the temperatures used. This, in turn, led to an overestimation of ash-free dry weight, which was only 12% of that calculated. The percentage of organic carbon of the total " dry )' weight ranges between about 2 4 % for Pleurobrachia and Mnemiopsis (Reeve and Baker, 1975; Mullin and Evans, 1974; Kremer, 1976a; Reeve et al., 1978). Various of these authors determined the nitrogcn and phosphorus contents t o be 0.2-1-3~oand 0-03-0.23~0of the dry weight respectively. Most of these data are summarized in a, table by Krerner (1976a). RATES V. INGESTION Over the past few years feeding rates of cydippid and lobate ctenophores have been measured in the laboratory in order to estimate their effects as predators in the environment. Perhaps the most significant aspect of their feeding behavior is that over an extremely wide range of prey concentration their ingestion rate is proportional t o food concentration. This was first demonstrated by Bishop (1967) for Mnemiopsis leidyi over a copepod food concentration range of 1053600/1., and subsequently by Miller (1970) and Kremer (in press) for the same species over ranges of 4-44 and 1-100 food organisms/l. respectively. Reeve et al. (1978) found this relationship t o hold

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over the entire tested range of 1-3 000 copepods/l. for two sizes of adults of Mnemiopsis mccradyi. For three sizes of larval animals of this species and for Pleurobrachia bachei adults (i.e. both tentaculate) on the other hand, ingestion rate appeared t o level off above 2OOjl. becoming independent of food concentration. Rowe (1971) also showed levelling off of ingestion rate for P. bachei above 400/1., although, unlike all the other studies, his animals were offered nauplii rather than adult copepods. At the highest laboratory food concentrations, the term (‘ingestion” is a misnomer for lobate forms which, as explained in an earlier section, reject mucus-entangled boli of copepods as they collect them a t rates faster than can be ingested and digested. The ecological consequences are similar, however, in as much as the food organisms are destroyed ”. Unlike many other zooplankton herbivores and carnivores, therefore, ctenophores do not appear to encounter in nature a critical ” food concentration a t which their ingestion rate becomes constant and independent of further food concentration increases. Their feeding behavior is not, however, completely automatic. Walter (1976) showed that starved Mnemiopsis ingested copepods at a higher initial rate and levelled off after a few hours t o a rate characteristic of animals which had not been starved. I n animals which had been starved from 1 to 5 days, higher ingestion rates were maintained for longer periods as a function of length of prior starvation. Also, as noted above, feeding activity in terms of tentacle and lobe extension, as well as swimming activity, are also correlated with feeding history. The relationship between food concentration and ingestion rate implies that the daily ration of ctenophores (i.e. the daily amount of organic carbon they ingest as a ratio of the body organic carbon weight) can become very large. Hirota (1972) indicated that this ratio rarely exceeded 40%, but his assumption that Pleurobrachia had a carbon content of 50% of its dry weight was in error (see above). Corrected for 2% carbon, this value would become 1000%. Kremer (in press) calculated that at a food concentration of 100 copepods/l., the ratio would be 115%. Walter (1976) computed the ratio for Mnemiopsis on the same food source as ranging from 10% to nearly 10 000% over a food range between 10 and 3 000 copepods/l., with values comparable to those of Kremer at about lOO/l. She computed a ration of 67 yo based on maximum estimates of zooplankton biomass from the natural environment. Several workers expressed the feeding activity of ctenophores in terms of volume of water swept clear ” per unit of time following ((

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numerical methods developed for copepods feeding herbivorously (e.g. Gauld, 1951). Walter (1976) reported that clearance rates of Mnemiopsis mccradyi between 5 and 70 mm total length (2-1 000 mg dry weight) ranged from 1 t o 74 l./animal/day a t 26°C. These values are some 2 times greater than those obtained by Kremer (in press) for M . leidyi between 20 and 25°C. Both workers found that weight specific clearance rates fell with size increase. Miller (1970) whose clearance rates for the latter species were intermediate, saw no such weight specific decrease, although his data were very variable and may, as Kremer suggested, simply have obscured such a n expected trend. VI. DIGESTION There are very few observations on the process of digestion in ctenophores. Kamshilov (1960b) observed the process in Beroe, which required three hours or more to completely digest Bolinopsis. He also noted that in situations where there was food in the gut of the ingested Bolinopsis, such as a copepod or fish larva, it was subsequently egested more or less intact. On the other hand, in one case in which the copepod had already been digested by the Bolinopsis to the point where it ruptured, Beroe completed the process, only egesting the chitinous exoskeleton. It appeared that in this case the Beroe was able to complete the process once the non-ctenophore food had been partially digested. He observed digested material to pass out of both the mouth and anal pores. Swanberg (1 974) reported a digestion time for Beroe feeding on Bolinopsis of 4.5 h (average). Anderson (1974) measured a digestion time of 2.4 h for Pleurobrachia feeding on copepods. Walter (1976) and Reeve et al. (1978) made quantitative observations on the digestive efficiency of Mnemiopsis feeding on adult copepods (Acartia). Ctenophores were allowed to feed for one hour a t various food concentrations, then removed t o clean water, and all material subsequently egested was collected for analysis. Up to a concentration of 100 copepods/l., digestion efficiency was about 74% on a copepod dry weight basis, which is comparable t o the digestive efficiency of chaetognaths and herbivorous zooplankton (see review in Cosper and Reeve, 1975). Beyond this concentration up to 1 000/1. values became much more erratic, yielding negative figures in some cases. This was because animals egested food boli without digestion which also included ctenophore-produced mucus. Average digestive efficiency at the highest food concentration was 20%,

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although values ranged widely. Unless food was rapidly egested, gut residence time beyond the initial hour during which feeding was permitted was fairly constant and not dependent on food quantity, and ranged between 29 to 34 hours. Fecal material always appeared to be evacuated through the mouth. The fecal material of ctenophores does not form a distinct pellet as does that of many other zooplankton (Reeve, Cosper and Walter, 1975), but is mostly loosely bound by mucus, and sinks to the bottom of the experimental container.

VII. RESPIRATION AND EXCRETION Several workers have made estimates of oxygen consumption and nitrogen excretion rates in ctenophores, usually in an effort to use these data to interpret the ecological interrelationships of ctenophores to the other organisms in the water column. Oxygen consumption measurements for Pleurobrachia were made by Lazareva (196l), Rajagopal (1963), Hirota (1972), Biggs (1977) and Reeve et al. (1978), and for Mnemiopsis by Williams and Baptist (1966), Miller (1970), Miller and Williams (1972), Baker (1973) andKremer (1978). Respiration increased with temperature in Mnemiopsis over a range of 4-29°C with a Qlo ranging from 7.1-1.1 (Miller, 1970), 21-31°C with a mean Qlo of 1.9 (Baker, 1973) and 10-26°C with a Ql0 up to 3.7 (Kremer, 1978). Williams and Baptist (1966) cited a value of 2.3. Kremer (1978) noted that her animals were probably more sensitive to temperature change at higher temperatures compared to those of Miller (1970), or Baker (1973), because her animals were at the extreme upper limit of their range. Baker (1973) noted that at comparable temperatures, the data of Miller (1970) were twice as high as those she obtained (on a pgO,/mg ash-free dry weight basis), which she suggested could have been attributed to the very low salinities employed by Miller (1970), which are known to result in higher energy expenditure in some marine animals (Prosser and Brown, 1961). Both Miller (1970) and Baker (1973) found that weight specific oxygen consumption decreased with increasing ctenophore size, but Williams and Baptist (1966), Hirota (1972) and Kremer (1978) reported that the reverse was the case. Kremer (1975a) noted that the latter situation was unusual and suggested it might reflect the “extremely simple nature” of ctenophores. Reeve, et al., (1978) confirmed the results of Hirota (1972) using the same species (Pleurobrachia bachei) and temperature (13°C). Information on excretion rates is minimal. Apart from isolated values (e.g. Jawed, 1973) measurements were made on a range of sizes

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of animals by Kremer (1975b, 1978) for Mnemiopsis and Reeve et al., (1978) for Pleurobrachia. Kremer measured ammonia, total dissolved nitrogen, urea, nitrite, nitrate, phosphorus and dissolved organic carbon. Urea, nitrite and nitrate did not constitute a signifi: cant proportion of excreted material but dissolved organic nitrogen was 46% of the total, the balance of which was in the form of ammonia. Kremer (1976a)reviewed previous literature which indicated that large percentages of excreted dissolved organic nitrogen are relatively rare for zooplankton. Reeve, et al., (1978) found 0: N ratios on the basis of ammonia nitrogen of about 15 for Pleurobrachia bachei, which was the same as Kremer reported for ivnemiopsis leidyi. They did not measure total organic nitrogen. Kremer (1976s) reported that ammonia excretion was independent of size on a weight specific basis. I n experiments with Pleurobrachia at low food concentrations they found that 45% of the nitrogen ingested was excreted. This fraction dropped to 16% at high food concentrations because a greater proportion of the food was unassimilated, but presumably quickly mineralized from fecal material.

VIII. GROWTHRATE Comparison of length measurements between workers is complicated because for Pleurobrachia (and larval lobate forms) body diameter may be measured in either the equatorial (Hirota, 1972) or polar plane (Reeve and Walter, 1976). For lobate adults, length may include the lobes (Baker and Reeve, 1974; Walter, 1976) or not (Greve, 1970). Comparison measurements were made for Pleurobrachia bachei (Walter, unpublished). The equatorial diameter was 80% of the polar diameter in adults. There are relatively few measurements of growth rate for ctenophores in the laboratory. Of the 4 genera commonly maintained in the laboratory, Bolinopsis is particularly sensitive to physical damage, its lobes frequently atrophying and growing back again. Our (unpublished) data for Bolinopsis growth rate is very similar to that of Greve (1970). Over a period of about 3 days his fastest and slowest growing animal increased in length from 4 mm to 40 and 8 mm respectively a t 16°C. I n our case, starting with a newly-hatched population, animal sizes ranged between 6-25 mm after 36 days at 13°C. Such wide variation is almost certainly an artifact of culture condition. Growth rate, when expressed as the logarithm of biomass increase (e.g. volume, dry weight, organic carbon) as a, function of time, can frequently be represented by a straight line over some or most of the A.M.B.-15

12

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life history of invertebrates (Winberg, 1971). Growth, in this case, takes place at a constant relative rate and is exponentialIy related to time. This coefficient of exponential growth (k) can be used as a basis of comparison between different animals. Hirota (1972) presented growth rate data for Pleurobrachia for which he subsequently (1974) computed growth coefficients. At 15"C, animals reached a maximum size of 14 mm diameter (population mean) after 80 days. At 20°C they reached a diameter of 6 mm in 35-40 days, which was some 1&15 days faster than a t 15°C. His animals grew slowly at first up to 2 mm (40 days), then rapidly up to 6.5 mm and then more slowly again. Growth coefficient ranges over these three periods were 0.12-0.17, 0.21-0.47 and 0-OPO-17 respectively. Hirota (1972) also obtained comparable growth rates in large (70 m3) tanks a t lower food concentrations. Reeve and Walter (1976), working with the same species, obtained much higher mean growth rates, although the same 3 phases were evident. The first phase (up to 2.5 mm) required only 5 days following which, up to the 20th day (6.0 mm) animals increased their biomass about 50% per day (k = 0.47), then grew more slowly up to about the 40th day (k = 0.09) after which they levelled off at about 10 mm. Growth rates were somewhat faster at 20"C, the maximum coefficient being 0-76 and final size being reached in 37 days. The final size of our laboratory populations of Pleurobrachia was considerably smaller than that reported by Hirota (1972) or Greve (1972). Greve, whose species was P. pileus, kindly identified our species as P. bachei, which was the same as that referred to by Hirota. Reeve and Baker (1975) reported on laboratory growth rates of Mnemiopsis mccradyi a t 21, 26 and 31°C. The intermediate temperature produced faster growth rates, and growth was divided into three phases of decreasing exponential growth rate from hatching. At 26°C Mnemiopsis reached 30 mm length in about 10 days (coefficient 0.78) by which time they had become completely lobate. Growth slowed over the next 10 days (k = 0-23, up to 40 mm) and again from 20 to 40 days (k = 0.07, up to 68 mm). The duration of these experiments did not permit animals to reach a final size, but we have had animals reach a total length of over 90 mm in the laboratory. It is worth noting that our highest growth rates recorded for young animals involved a daily doubling of biomass. They approach the extraordinary growth rates reported by Heron (1972) for population increase of the salp Thalia dernocratica, which he stated at that time to be " considerably higher than any other multicellular animals yet measured ".

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Miller (1970) attempted some growth rate studies with Mnenziopsis teidyi, although he considered conditions not very satisfactory because at 20°C his animals ceased growing above an average volume of 11 ml (about 40 mm long). He used a range of temperatures up to 20"C, at which fastest growth rates were observed (k = 0.25). I n order to compare other growth rate data in the literature which was expressed only in terms of length, we have estimated rough growth coefficients using the weightldiameter ratio of Hirota (1972). The assumption is made that Beroe and Bolinopsis (without lobes) are approximately spherical. Greve (1970) reported the maximum growth rate for Bolinopsis was 10-40 mm in 20 days at 16°C (k = 0.2). He reported similar growth rates for PEeurobrachia (1970, 1972) and a somewhat faster rate for Beroe (1970) which increased from 5-15 mm in 8 days (k = 0-4). Kamshilov (1960a) made several measurements of length increase in Beroe in which k ranged from 0.02-0-04 (e.g. 15-30 mm in 28 days). Several authors have reported the ability of ctenophores to undergo periods of food shortage or absence, by utilizing their own body tissue for their metabolic requirements and so gradually shrink in size (e.g. Kamshilov, 1960b;Greve, 1972; Kremer, 1976b; Walter, 1976). Kremer (197623) found that Mnemiopsis lost weight at a copepod concentration of 1011. and gained weight a t 100/1. Reeve, Walter and Ikeda (1978) computed growth coefficients for Mnemiopsis over a food concentration range from 0 to 350/1. and showed that growth decreased to zero at l/l., below which animals lost weight.

IX. FECUNDITY Pianka (1974) summarized information on the considerable regenerative powers of ctenophores. This is not known to be used as a means of reproduction in planktonic forms, but Greve (1970) noted feeding patterns in both the young and adult beroids where the predator would not consume completely the prey ctenophore, but detach itself, which would permit the prey animal to regenerate its missing parts. Coonfield (1938), for instance, showed that Mnemiopsis would completely regenerate new individuals when cut into quarters, and often much smaller fractions also regenerated. It was noted earlier that a form of asexual budding was common amongst some of the Platyctenea. All planktonic ctenophores are simultaneous hermaphrodites and capable of self fertilization, and thus viable offspring can be produced from a single adult, as we and others (e.g. Pianka, 1974;

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Hirota, 1973) have regularly observed. Planktonic ctenophores can produce offspring long before they reach their upper limit of size. Pianka (1974) reviewed cases of paedogenesis (sexual maturity of larvae and juveniles) and dissogony (sexual maturity of larvae followed by regression of gonads and subsequent rematuring of adults). Hirota (1972) documented an individual (Pleurobrachia bachei)which produced a total of 76 offspring as a larva between the 25th and 47th day and resumed egg-laying on the 69th day reaching a maximum of almost 1000 eggslday. Reeve et al., (1978) saw egg production begin at about 4-6 mm, which although a much shorter period from hatching than Hirota recorded, was at a later developmental stage. No indications of dissogony were apparent. Pianka (1974) and Greve (1970) also noted a similar size for the initiation of spawning in Pleurobrachia with no earlier larval maturity. Although dissogony had been mentioned for Bolinopsis in early reports, Pianka (4.v.) did not see spawning until animals had become lobate a t 10 mm, and it has not been reported for Mnemiopsis. Kremer (1975a) and Baker and Reeve (1974) investigated fecundity of Mnemiopsis by collecting wild individuals of various ages and leaving them in the laboratory overnight, after which any eggs and larvae produced were counted. I n both cases egg production was a function of size, the largest animals producing up to 9 990 (Baker and Reeve) and 14 000 eggs (Kremer). Baker and Reeve (1974) followed six individuals from newly-hatched larvae for 23 days. They started to produce eggs 13 days after hatching a t 26 mm total length and the maximum total production over the period was over 12 000 eggs each. There is a wide range of numbers of eggs produced for any given size of ctenophore and food supply is no doubt a very important controlling factor. Kremer (1975a)and ourselves have found differences in fecundity in wild animals brought into the laboratory at different times of the year which she attributed to differences in food supply. The provision of food is well known to be one of the ways to stimulate egg production of ctenophores in the laboratory (Greve, 1970 ; Baker, 1973). Reeve et al., (1978) fed young Pleurobrachia at two food concentrations (10 and 100 copepods/l.) over 6 days. They grew from 5-3 mm (polar diameter) to 7.3 mm and 9.6 mm respectively, and produced a total of 7 and 186 eggs/animal. The dry weight of each egg was 0.35 pg (0.02 pgC).

X. GROWTH EFFICIENCY Hirota (1972) estimated the growth efficiency of Pleurobrachia bachei as 60%. When we first started making similar computations

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from experimental data we arrived at values well in excess of loo%, on a dry weight basis. This stimulated a, search into problems of measuring the biomass (organic tissue weight) of ctenophores (Reeve and Baker, 1975), and the conclusion that organic carbon was only about 2% of the dry weight, compared to 40-45% in the food organisms (copepods). On a carbon basis growth efficiencies varied from 7% a t the lowest food concentration ( 3 copepods/l.) (Reeve et al., 1978) down to 2% a t the highest (300/1.). I n similar measurements made on Pleurobrachia at two food concentrations (10 and 100 copepods/l.) efficiency was 11 and 3% respectively. The 60% value of Hirota (1972) was recomputed to be about 9%. These values are very low compared to values obtained for other zooplankton (reviewed by Reeve, 1970). Even a t food concentrations where digestive efficiency is known to be consistently high (see above), growth efficiency is low, although animals grow as fast as zooplankton which have high growth efficiencies. An explanation of this apparent paradox appears to be in the relatively high energy requirements of Pleurobrachia. Reeve et al. (1978) calculated that these ctenophores required about 63% of their food intake to satisfy their metabolic activities, at food concentrations where digestive efficiency is high. The authors pointed out that the organic carbon content of a ctenophore whose dry weight is the same as that of a copepod would only be 5% of that of the copepod, i.e. the amount of living material generating the energy to move the body bulk of the ctenophore in its environment is, similarly, a tiny proportion compared to the copepod. This great disparity in the ratios suggests why a greater proportion of the ingested food must be directed to energy demands. XI. SEASONAL VARIATIONS IN CTENOPHORE POPULATIONS Fraser (1970), after many years of experience of the seasonal variations of ctenophores and other zooplankton in temperate waters, agreed with the remarks of Kramp (1913) that it is difficult to make reliable predictions of the seasonal appearance of any species, and often their appearance in great numbers seems fortuitous, with no obvious correlation with temperature, season or depth. They can occur throughout the entire year in great numbers (McIntosh, 1926) off the east coast of Scotland, reoccur in large numbers with regularity throughout P 5 months of the summer in Saanich Inlet, British Columbia (personal observations), appear once or twice at particular times of the year, especially later summer and autumn (Fraser, 1970), or sometimes pass through an entire year without appearing in bloom

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proportions (Kramp, 1913, for the English Channel ; our observations in south Florida inshore waters). Many references to ctenophore abundance quote numbers only, often not related to water volume collected, using a variety of different collection methods and meshes and related to differently-sized species. Even biomass, therefore, cannot be compared very closely. Production rates have been determined rarely. Reeve and Baker (1975) estimated the production of Mnemiopsis mccradyi in Biscayne Bay, Florida to be 61-123 mgC/m3 annually, with a daily production : biomass ratio of 0.12. The Biscayne Bay water column is very shallow (3 m) so this translates to a value of 369 mgC/m2/year. Hirota (1974) computed the production of Pleurobrachia bachei off California in terms of ash-free dry weight. When converted to carbon using a factor of 8.7% carbon in ash-free dry weight (Reeve and Baker, 1975) annual production was 472 mgC/m2/year with a production: biomass ratio of 0.02. Water column depth off southern California was in excess of 40 m, however, so that on a m3 basis the warmer-water Biscayne Bay ctenophore population had a much higher production rate, as might be inferred by their much faster growth rates noted in an earlier section. Kremer (1976b) developed a computer production model (discussed below). I n their experiments, Reeve and Baker (1975) maintained an abundant supply of food, which they noted was no less artificial than trying to maintain a uniform environmental average level of food, because it is highly likely that food is distributed in patches, and animals may be exposed to concentrations varying by several orders of magnitude over 24 hours. The bulk of production took place over a short period of the year in both studies, and it could be argued that ctenophores were not likely to be very food limited over this period of rapid population growth rate. Most recent studies which have related population changes of ctenophores to observations made in the laboratory have been designed to estimate the predatory effect of ctenophores on the rest of the plankton. Reeve and Baker (1975) and Hirota (1974) did this by relating ctenophore production to estimates of herbivore zooplankton production. For Biscayne Bay, Reeve and Baker estimated the ctenophore production as about 20% of the copepod production, and the production of another major carnivore, the chaetognath Sagitta hispida Conant, as 12% of the copepod production. Assuming a 32% conversion efficiency from copepod to carnivore biomass, two predators would account for all the copepod production of the bay. Hirota (1974) computed a transfer efficiency of 11% between various levels of the food chain including zooplankton to Pleurobrachia.

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Williams and Baptist (1966) reported (in abstract) that on the basis of respiration measurements of Mnemiopsis leidyi from estuaries around Beaufort, North Carolina, a large animal required all the copepods in 4-100/1. of water. Bishop (1967) directly measured feeding rates of the same species and related them to copepod abundance in the Patuxent River, Maryland. He estimated that 52% of the mortality of the dominant copepod, Acartia tonsa Dana, could be accounted for by predation. Burrell (1968) and Burrell and Van Engel (1976) also performed feeding experiments and estimated that M . Zeidyi was the major predator on zooplankton in the York River estuary, Virginia, accounting for 73% of the total predation exerted by a group of predators including medusae, chaetognaths and fish. He also noted that when Beroe made an appearance, Mnemiopsis populations were eliminated wherever it occurred. Populations of Pleurobrachia and Bolinopsis also decreased dramatically, according to the data of Anderson (1974), when Beroe increased in numbers in St. Margaret's Bay. As already noted, the first two species did not appear to be in direct competition because their gut contents showed that they relied on different components of the copepod population. Between them, they removed 40% of the total accounted for by the large Sagitta population. Miller (1970) used his estimates of ctenophore water clearance rate in the laboratory to compute total water clearance rates for the ctenophore population of the Pamlico River, North Carolina. When ctenophores were at their peak biomass they removed up to 48% of the copepod biomass, i.e. their daily total population clearance rate was 480 l./m3. Most of the season, ctenophore populations were much lower, yielding computed summer and winter mean clearance rates of 5.4 and 0.7% of the copepod population respectively. Since Miller also measured respiration and growth rates, he computed total population respiration and growth requirements in energy units and suggested that these requirements were in excess of food intake, if only copepods were considered as food. Averaged over the year, he estimated that ctenophores could obtain only 23% of their energy requirements from zooplankton. This led him to the suggestion that they must also, of necessity, have to make use of phytoplankton and detrital material present in the water at the same time. Miller and Williams (1972) reinforced this conclusion by taking ctenophore, zooplankton and phytoplankton population data from the Patuxent estuary from Herman et al. (1968) and respiration data from Williams and Baptist (1966), from which they concluded that during most of the season there was not enough zooplankton to satisfy even

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the respiratory needs of both the ctenophore and jellyfish population present. Heinle (1974) pointed out a substantial error in this treatment because ctenophore biomass had been incorrectly reported by Herman et al. to be some three orders of magnitude greater than really occurred. This meant that in some cases there would be enough zooplankton to account for the energetic needs of ctenophores, although he suggested that ctenophores could probably not rely entirely on other zooplankton for their food. Kremer (in press) also made calculations based on laboratory feeding rates and environmental biomass, and reached a similar conclusion as Miller (1970) regarding the fraction of the copepod standing stock removed. A t the time of their greatest abundance, ctenophores were estimated to achieve a maximum of 30% removal rate in a day. Kremer went on to calculate more precisely the effects of the ctenophores on the rest of the zooplankton by estimating the production capacity of the copepods rather than simply their standing stock. She estimated that ctenophores might be responsible for between 20 and 50% of the entire copepod mortality over the summer, but urged caution in generalizing from such computations. A major problem was that often the sequence of copepod decline did not correspond to maximum ctenophore predation pressure, suggesting that there could be other major unknown forces at work affecting the copepod populations independently of ctenophore predation. Ctenophores must recycle a significant fraction of their ingested nitrogen either though excretion of dissolved metabolic waste products, or, when feeding at very high food concentrations, by releasing either partly digested or dying copepods which subsequently become remineralized by bacterial action. Kremer (197510) calculated that at their population peak, they were responsible for the turnover of as much nitrogen as the rest of the zooplankton population. We have also performed similar calculations (Reeve et al., 1978) to indicate tha.t at the time of peak ctenophore standing stock in Biscayne Bay (from Baker, 1973) they could consume some 10% of the standing stock of copepods per day, although this fraction would be much less during most of the year. Nevertheless, taking the average population production rate estimated by Reeve and Baker (1975) it can be estimated that within seven days copepods would be reduced to only 25% of their original biomass assuming no copepod production over the period. Approximately the same figure would result assuming a high (0.3) daily production to biomass ratio for both populations. The characteristically rapid population increase of ctenophores is so striking that it is tempting to attribute it to migration of already

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developed populations from elsewhere. Kremer (1976b) used a simple computer model to show that in Narragansett Bay, the fecundity estimates derived in the laboratory could explain the rapid increase from a winter minimum of about 1 animal/lO 000 m3 over 5 orders of magnitude. Reeve et al. (1978) took the data of Baker and Reeve (1974) and Reeve and Baker (1975) for Biscayne Bay using a model similar to that described by Kremer . These computations suggested that the observed ctenophore population biomass increase over 45 days of the major seasonal ctenophore bloom in Biscayne Bay (2.5 orders of magnitude range) would require a production/biomass ratio of less than 0.2 at fecundities observed in the laboratory. Rapid growth rates and high fecundity in ctenophores mean that it is rarely feasible to follow cohorts in the environment and estimate patterns of growth and mortality directly. Por this reason estimates of the kind outlined above, which make use of laboratory data to interpret natural populations, are essential. There have been some attempts, however, to follow populations in situations where the same population is sampled successively, because it is confined in some way. Mullin and Evans (1974) established a phytoplankton/copepod/Plezcrobrachia food chain in a tank of approximately 70 m3 volume. The daily production/biomass ratio of the predators was 0.16. The production of carnivores was 3% of the primary production, which indicated a food chain efficiency greater than 10% between each trophic level. Reeve et aZ. (1976) reported on the populations of zooplankton in transparent closed plastic columns of 68 m3 volume floating at the sea surface to some of which had been added copper. Copepod populations declined throughout the experiment due to the presence of ctenophores aa well as copper, and estimates of the fraction of the observed mortality due to predation were made by computing ctenophore food requirements from their respiratory demands. I n a subsequent experiment, in which amounts of nutrients were varied in four containers (Parsons et al., 1977), ctenophore production was almost doubled in the container receiving most enrichment, compared with that receiving no enrichment, although the percentage of phytoplankton production converted to ctenophore production was much less (1.9 compared to 4.8%). Reeve and Walter (1976) illustrated how such containers could bo biologically manipulated. They described the withdrawal of most of the larger Pleurobrachia by a selective sampling technique which did not disturb the copepod population. Within a week copepods had more than doubled in this container. This provided a clear demonstration of the predatory significance of ctenophore populations at high

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density. The same containers also demonstrated the rapid ability for population increase of ctenophores initially a t very low density. The sudden appearance of a ctenophore bloom, therefore, and its subsequent effect on copepod populations, can be predicted from the observations of populations in the natural environment, estimated by application of laboratory data, and actually observed in a captured population. This does not necessarily imply that the population dynamics of ctenophores can be so readily explained under all circumstances. Kremer (in press) noted that there were often variations in natural populations of ctenophores and copepods which could not be effectively explained by trophic interactions, and this is certainly the case in the example cited by Reeve and Walter (1976). It is unclear, for instance, what caused the progressive mortality of ctenophores in containers in which there were no known predators, or why populations of adult copepods died off very rapidly in the container from which most of the ctenophores had been removed. Reeve and Baker (1975) suggested that in comparing two dissimilar plankton carnivores-ctenophores and chaetognaths-which depended on the same food source in south Florida inshore waters, the conclusion could be drawn that ctenophores needed a higher food density to enable population growth to occur. This could be deduced from the general correlation of absence of ctenophores in regions of low zooplankton biomass where chaetognaths continued t o occur, and their relatively infrequent appearance, compared to chaetognaths, which was associated in a general way with seasons of the highest copepod biomass. Laboratory data indicating their requirements of very high food concentrations for maximum growth rates and their continued capacity to increase ingestion at these high food concentrations also confirm their dependence on food abundance for rapid population increase. Their potential for self-fertilization, extremely high fecundity and rapid growth potential explain how, given good food abundance, their population can ‘(explode ”. They appear to be environmental specialists which can overwhelm the biomass of any competitor under conditions of peak food supply.

XII. CONULUSION Fraser (1962), Kremer (1976a), Greve (personal communication) and ourselves have pointed out that ctenophores should not be considered merely wasteful ‘(dead ends ’’ in the food chain. They may act sometimes to balance the ecosystem by restraining an overabundance

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of copepods from virtually eliminating all phytoplankton from the water column at a time when other more “ useful ” predators are failing to do this. The phytoplankton would also receive a positive stimulation to renewed growth by virtue of the high percentage of ingested nitrogen being returned in a dissolved form back to the water column. It is possible that the result might otherwise be the production of dead copepod biomass which accumulated in the sediments and was lost (at least on a, short time scale) to the water column. XIII. REFERENCES Anderson, E. (1974). Trophic interactions among ctenophores and copepods in St Margaret’s Bay, Nova Scotia. Ph.D. Dissertation, Dalhousie University. Baker, L. D. (1973). Ecology of the ctenophore Mnemiopsis mccradyi Mayer, in Biscayne Bay, Florida. Rosenstiol School of Marine and Atmospheric Science, University of Miami, Technical Report UM-RSMAS-73016. Baker, L. D. and Reeve, M. R. (1974). Laboratory culture of the lobate ctenophore Mnemiopsis mccradyi with notes on feeding and fecundity. Marine Biology, 26, 57-62. Barlow, J. P. (1955). Physical and biological processes determining the distribution of zooplankton in a tidal estuary. Biological Bulletin (Woods Hole), 109, 21 1-225. Bayer, F. M. and Owre, H. B. (1968). “ The Free-Living Lower Invertebrates”. MacrniHan, New York. Bigelow, H. B. (1915). Exploration of the coast water between Nova Scotia and Chesapeake Bay, July and August, 1913, by the U.S. Fisheries Schooner, Grampus. Oceanography and plankton. Bulletin of the Museum of Comparative Zoology at Harvard College, 59, 149-359. Bigelow, H. B. and Leslie, M. (1930). Reconnaissance of the waters and plankton of Monterey Bay, July 1928. Bulletin of the Museum of Comparative Zoology at Harvard College, 70, 429-581. Biggs, D. C. (1977). Respiration and ammonium excretion by open ocean gelatinous zooplankton. Limnology and Oceanography, 22, 1OS117. Bishop, J. W. (1967). Feeding rates of the ctenophore, Mnemiopsis leidyi. Chesapeake Science, 8, 259-264. Bishop, J. W. (1968). A comparative study of feeding rates of tentaculate ctenophores. Ecology, 49, 996-997. Burrell, V. G. (1968). The ecological significance of a ctenophore, Mnemiopsis leidyi (A. Agassiz), in a fish nursery ground. M.A. Thesis, The College of William and Mary, Virginia. Burrell, V. G. and Van Engel, W. A. (1976). Predation by and distribution of a ctenophore, Mnemiopsis leidyi A. Agassiz, in the York River Estuary. Estuarine and Coastal Marine Science, 4, 235-242. Coonfield, B. R. (1938). Symmetry and regulation in Mnemiopsis leidyi, Agassiz. Biological Bulletin (Woods Hole), 72, 299-310. Cooper, L. H. N. (1939). Phosphorus, nitrogen, iron and manganese in marine zooplankton. Journal of the Marine Biological Association of the United Kingdom, 23, 387-390.

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Corner, E. D. S. and Davies A. G. (1971). Plankton as a factor in the nitrogen and phosphorus cycles in the sea. Advances in Marine Biology, 9, 101-204. Cosper, T. C. and Reeve, M. R. (1975). Digestive efficiency of the chaetognath Sagitta hispida Conant. Journal of Experimental Marine Biology and Ecology, 17, 33-38. Cronin, L. E., Daiber, J. C. and Hulbert, E. M. (1962). Qualitative seasonal aspects of zooplankton in the Delaware River Estuary. Chesapeake Science, 3, 63-93. Curl, H. (1962). Standing crops of carbon, nitrogen and phosphorus and transfer between trophic levels, in continental shelf waters south of New York. Rapports et Pr5ckS-Verbaux des Rdunions, Conseil International pour I’Exploration de la Mer, 153, 183-189. Flores,‘M. and Brusca, G. J. (1975). Observations on two species of hyperiid amphipods associated with the ctenophore PZeurobraehia bachei. Bufletin of the Southern California Academy of Sciences, 74, 10-15. Praser, J. H. (1962). The role of ctenophores and salps in zooplankton production and standing crop. Rapports et Procks- Verbaux des Rbunions, Conseil International pour 1’Exploration de la Mer, 153, 121-123. Fraser, J. H. (1970). The ecology of the ctenophore Pleurobrachia pileus in Scottish waters. Journal d u Conseil. Conseil Permanent International pour I’Exploration de la Mer, 33, 149-168. Gauld, D. T. (1951). The grazing rate of planktonic copepods. Journal of the Marine Biological Association of the United Kingdom, 29, 695-706. Greve, W. (1970). Cultivation experiments on North Sea ctenophores. Helgolander Wissenschuftliche Meeresuntersuchungen, 20, 304-3 17. Greve, W. (1971). okologische Untersuchungen an Pleurobrachia pileus. I. Freilanduntersuchungen. Helgolander Wissenschaftliche Meeresuntersuchungen, 22, 303-325. Greve, W. (1972). Okologische Untersuchungen an Pleurobrachia p i l e w . 11. Laboratoriumsuntersuchungen. Helgolander Wissenschaftliche Meeresuntersuchungen, 23, 141-164. Greve, W. (1975a). Verhaltensweisen der Rippenqualle Pleurobrachia pileus (Ctenophora). Institut fur den Wissenschaftlicher Film, Wissenschaftlicher Film C 1181/1975. Greve, W. (1975b). Die Rippenquallen der sudlichen Nordsee und ihre interspezifischen Relationen. Institut fur den Wissenschaftlichen Film, Wissenschaftlicher Film C 1182/1975. Greve, W., Stockner, J. and Fulton, J. (1976). Towards a theory of speciation in Beroe. I n “ Coelenterate Ecology and Behavior”, (G. Mackie, ed.), pp. 251-258. Plenum Publishing Company, New York. Harbison, G. R., Biggs, D. C. and Madin, L. P. (1977). Associations of Amphipoda Hyperiidea with gelatinous zooplankton. 11. Associations with Cnidaria, Ctenophora and Radiolaria. Deep-sea Research, 24, 465-488. Harbison, G. R., Madin, L. P. and Swanberg, N. R. (1978). On the natural history and distribution of oceanic ctenophores. Deep Sea Research, 25, 233-256. Heinle, D. R. (1974). An alternate grazing hypothesis for the Patuxent Estuary. Chesapeake Science, 15, 145-150. Herman, S. S., Mihursky, J. A. and McErlean, A. J. (1968). Zooplankton and environmental characteristics of the Patuxent Estuary. Chesapeake Science, 9, 67-82.

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Heron, A. C. (1972). Population ecology of a colonizing species: the pelagic tunicate Thalia democratica. I. Individual growth rate and generation time. Oecologia, 10, 269-293. Hirota, J . r ( 1972). Laboratory culture and metabolism of the planktonic ctenophore, Pleurobrachia bachei A. Agassiz. I n " Biological Oceanography of the Northern North Pacific Ocean ", (A. Y. Takenouti, editor-in-chief), pp. 465-484. Idemitsu Shoten, Tokyo. Hirota, J. (1973). Quantitative natural history of Pleurobrachia hachei A. Agassiz in La Jolla Bight. Ph.D. Dissertation, Scripps Institution of Oceanography, University of California. Hirota, J. (1974). Quantitative natural history of Pleurohraclhia bachei in La Jolla Bight. United Statea National Marine Fisheries Service Fishery Bulletin, 72, 295-335. Hopkins, T. L. (1966). The plankton of the St. Andrew Bay system, Florida. Publications of the Institute of Marine Science University of Texm, 11, 12-64. Horridge, G. A. (1965). Macrocilia with numerous shafts from the lips of the ctenophore Beroe. Proceedings of the Royal Society of London Biological Sciences, 162, 351-364. Hyrnan, L. (1940). " The Invertebrates : Protozoa Through Ctenophora". McGraw Hill, New York. Jawed, M. (1973). Effects of environmental factors and body size on rates of oxygen consumption in Archaeomysis grebnitzkii and Neomysis awatschensis (Crustacea : Mysidae). Marine Biology, 21, 173-179. Kamshilov, M. M. (1959). Interrelations between organisms and the part they play in evolution. Zhurml Ohschchei Biologii Union of Soviet Socialist Republics, 20, 370-378. Kamshilov, M. M. (1960a). Size of ctenophore Beroe cucumis Fabricius. Doklady Akademii Nauk Union of Soviet Socialist Republics, 131, 957-960. Kamshilov, M. M. (1960b). Feeding of ctenophore Beroe cucumis Fabricius. Doklady Akademii Nauk Union of Soviet Socialist Republics, 130, 1138-1 140. Kramp, P. L. (1913).Medusae, Siphonophora and Ctenophora. Zoology of Iceland, 11, 1-37. Kremer, P. M. (19758). The ecology of the ctenophore, Mnerniopsis leidyi in Narragansett Bay. Ph.D. Dissertation, University of Rhode Island. Kremer, P. M. (1975b). Nitrogen regeneration by the ctenophore Mnemiopsis Eeidyi. I n " Mineral cycling in southeastern ecosystems ", (F. G. Howell, J. B. Gentry and M. M. Smith, eds.), pp. 279-290. United States Energy Research and Development Administr&on Symposium Series, N.T.I.S. NO. CONF-740513. Kremer, P. M. (197th). Excretion and body composition of the ctenophore Mnemiopsis leidyi (A. Agassiz) : comparisons and consequences. I n " Proceedings of the 10th European Symposium on Marine Biology", (G. Persoone and E. Jaspers, eds.), Vol. 2, pp. 351-362. Universa Press, Wetteren, Belgium. Kremer, P. M. (197613). Population dynamics and ecological energetics of a pulsed zooplankton predator, the ctenophore Mnemiopsis leidyi. I n " Estuarine processes ", (M. L. Wiley, ed.), Vol. 1, pp. 197-218. Academic Press, New York. Kremer, P. M. (1978). Respiration and excretion by the ctenophore Mnemiop.& leidyi. Mariwe Biology, 44, 43-50.

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Kremer, P. M. (In press). Predation by the ctenophore Mnemiopsis leidyi in Narragansett Bay. Chesapeake Science. Lazareva, L. P. (1961). Absorption of oxygen by the ctenophore Pleurobrachia pileus 0. F. Miiller of different sizes in relation to the temperature and salinity of the environment. Trudy Karadahs’koyi Biolohichnoyi Stccntsiyi, 17, 85-96. Lebour, M. V. (1922). The food of planktonic organisms. Journal of the Marine Biological Association of the United Kingdom, 12, 644-677. Lebour, M. V. (1923). The food of planktonic organisms. 11.Journal of the Marine Biological Association of the United Kingdom, 13, 70-92. Main, R. J. (1928). Observation of the feeding mechanism of the ctenophore, Mnemiopsis leidyi. Biological Bulletin (Woods Hole), 55, 69-78. Manteufel, B. P. (1941). Plankton and herring in the Barents Sea. Trudy Polyarnyi Nauchno-Issledovatel’skiiI Proektnyi Institut Morskogo Rybnogo Khozyaistwa I OkeanograJi of Union of Soviet Socialist Republics, 7 , 125-218. Marshall, S. M. (1973). Respiration and feeding in copepods. Advances i n Marine Biology, 11, 57-120. Mayer, A. G. (1912). Ctenophores of the Atlantic coast of North America. Publications of the Carnegie Institution of Washington, 162, 1-58. McIntosh, W. C. (1926). Additions to the marine fauna of St. Andrews since 1874. Annals and Magazine of Natural Hhtory, Series 9, 18, 241-266. Miller, R. J. (1970). Distribution and energetics of an estuarine population of the ctenophore, Mnemiopsis leidyi. Ph.D. Dissertation, North Carolina State University, Raleigh. Miller, R. J. (1974). Distribution and biomass of an estuarine ctenophoro population, Mnemiopsis leidyi (A. Agassiz). Chesapeake Science, 15, 1-8. Miller, R. J. and Williams, R. B. (1972). Energy requirements and food supplies of ctenophores and jellyfish in the Patuxent River Estuary. Chesapeake Science, 13, 328-331. Mullin, M. M. and Evans, P. M. (1974). The use of a deep tank in plankton ecology. 11. Efficiency of a planktonic food chain. fimnology and Oceanography, 19, 902-911. Nagabhushanam, A. K. (1959). Feeding of a ctenophore, Bolinopsis infundibuluna (0.F. Miiller). Nature, London, 184, 829. Nelson, T. C. (1925). On the occurrence and food habits of ctenophores in New Jersey inland coastal waters. Biological Bulletin (Woods Hole), 48, 92-1 11. Oviatt, C. M. and Kremer, P. M. (1977). Predation on the ctenophore Mnemiopsis leidyi, by butterfish, Peprilus tricanthus, in Narragansett Bay, Rhode Island. Chesapeake Science, 18, 236-240. Parsons, T. R., von Brockel, K., Koeller, P., Reeve, M. R. and Holm-Hansen, 0. (1977). The distribution of organic carbon in a marine planktonic food web following nutrient enrichment. Journal of Experimental Marine Biology and Ecology, 26, 235-247. Pianka, H. D. (1974). Ctenophora. I n “ Reproduction of Marine Invertebrates ”, (A. C. Giese and J. S. Pearse, eds.), Vol. I, pp. 201-265. Academic Press, New York. Prosser, C. L. and Brown, F. A. (1961). “ Comparative Animal Physiology”. W. B. Saunders Company, Philadelphia. Rajagopal, P. K. (1963). Note on the oxygen uptake of the ctenophore, Pleurobrmhia globosa, Current Science, 32, 319-320.

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Ralph, P. M. and Kaberry, C. (1950). New Zealand coelenterates. Ctenophores from Cook Strait. Zoology Publications from Victoria University of Wellington, 3, 11 PP. Reeve, M. R. (1970). The biology of Chaetognatlia. I. Quantitativo aspects of growth and egg production in Sagitta hkpida. I n “ Marine Food Chains”, (J.H. Steele, ed.), pp. 168-189. Oliver and Boyd, Edinburgh. Reeve, M. R. and Baker, L. D. (1975). Production of two planktonic carnivores (chaetognath and ctenophore) in south Florida inshore waters. United States National Marine Fisheries Service Pishery Bulletin, 73, 238-248. Reeve, M. R. and Walter, M. A. (1976). A large-scale experiment on the growth and predation potential of ctenopliore populations. I n “ Coelenterate Ecology and Behavior ”,(G. Maclrie, ed.), pp. 187-199. Plenum Publishing Company, New York. Reeve, M. R., Cosper, T. C. and Walter, M. A, (1975). Visual observations on the process of digestion and the production of faecal pellets in t h e chaetognath Sagitb hispida Conant. Journal of Experimental Marine Biology and Ecology, 17, 39-46. Reeve, M. R., Grice, G. D., Gibson, V. R., Walter, M. A., Darcy, K. and Ikeda. T. (1976). A controlled environmental pollution experiment (CEPEX) and its usefulness in the study of larger marine zooplankton under toxic stress. .77~ “ Effects of Pollutants on Aquatic Organisms ”, (A. P. Lockwood, ed.), pp. 145-162. Cambridge University Press. Reeve, M. R., Walter, M. A. and Ikeda, T. (1978). Laboratory studies of ingestion and food utilization in lobate and tentaculate ctenophores. Limnology and Oceanography, 23, 740-751. Rowe, M. D. (1971). Some aspects of the feeding behavior of the ctenophore Pleurobrachia pileus. M.S. Thesis, University of Hawaii. Russell, F. S. (1931). The study of copepods as a factor in oceanic economy. Proceedings of 5th Pacific Scientific Congress, 2023-2032. Russell, F. S. (1935). The seasonal abundance and distribution of the pelagic young of teleostean fishes caught in the ring-trawl in offshore waters in the Plymouth area. Part 11.Journal of the Marine Biological Association of the United Kingdom, 20, 147-180. Scott, A. (1913). The mackerel fishery off Walney in 1913. Report of the Lancashire Sealfisheries Laboratories, 22, 19-25. Swanberg, N. (1974). The feeding behavior of Beroe ovab. Marine Biology, 24, 69-76. Walter, M. A. (1976). Quantitative observations on the nutritional ecology of ctenophores with special reference to Mnemiopsis mccradyi. M.S. Thesis, University of Miami. Weill, R. (1935). Le fonctionnement des colloblastes. Comptes RendzLs de E’Academie des Sciences Paris, 201, 850-852. Williams, R. B. and Baptist, J. P. (1966). Physiology of Mnemiopsis in relation to its role as a predator. Association of Southeastern Biologkts Bulletin, 13, 48-49. Winberg, G. G. (1971). “ Methods for the Estimation of Production of Aquatic Animals Academic Press, New York, London.

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PETROLEUM HYDROCARBONS AND RELATED COMPOUNDS

E. D. S . CORNER The Laboratory, Marine Biological Association, Plymouth, England I. Introduction

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I. INTRODUCTION Marine organisms, including plankton, having been exposed to petroleum hydrocarbons released from submarine seeps throughout geological time, are likely to have evolved physiological and biochemical mechanisms allowing them to adapt to the presence of small quantities of these compounds in their natural environment. Nevertheless, there is considerable current interest in understanding what might happen to planktonic organisms exposed to the additional and localized inputs of hydrocarbons and related compounds that result from accidental spillages arising from relatively recent industrial activities such as the off-shore production and transport of crude oil. Accordingly, consequent upon incidents such as the wrecking of the tanker " Torrey Canyon a vast and widely dispersed literature has arisen during the past ten years dealing with the effects of petroleum hydrocarbons on numerous marine organisms. The publications on plankton considered in the present review, most of which refer to laboratory studies, are discussed in the context of a simplified food-chain model that begins with sea water and proceeds through phytoplankton to zooplankton. Although such a frame-work serves to carry the main theme of the treatment, several additional but relevant topics have had to be included. For example, in dealing with hydrocarbons in sea water attention has had to be given to matters such as their spatial distribution and the relative amounts in solution and in particulate form. Again, in discussing the levels and types of hydrocarbons in plankton it has been necessary to consider compounds of recent biogenic origin, some of which can also occur in crude oil. Furthermore, as certain studies with zooplankton have shown that the animals do not exclusively accumulate hydrocarbons from phytoplankton diets, work is also described that deals with the direct uptake of these compounds from solution in sea water. Finally, although the simplified food-chain model is not extended to include fish and benthic animals, consideration is given to factors affecting the retention of hydrocarbons by zooplankton, particularly copepods, which is of key importance in the transfer of these compounds to fish ; as well as to the release of hydrocarbons in faecal pellets, a possible means by which such compounds originally present in the euphotic zone could be eventually transferred to animals that dwell in sediments. '))

11. HYDROCARBON LEVELSIN SEAWATER When studying the accumulation and fate of hydrocarbons in plankton, and the possible effects of these compounds on the organisms,

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it is necessary to bear in mind the levels of hydrocarbons that plankton normally encounter in various sea areas. Accordingly, a brief review of the available data is attempted by way of introducing the more detailed treatment of studies with plankton that are dealt with in later sections. Although numerous attempts have been made to ascertain the levels and types of hydrocarbons present in sea water under a variety of conditions, the methods used (reviewed by Farrington and Meyer, 1975) have usually provided data for only a particular fraction of the various kinds of hydrocarbons present. More comprehensive analyses have occasionally been made (Barbier, Joly, Saliot and Tourres, 1973; Brown, Searl, Elliott, Phillips, Brandon and Monaghan, 1973), but generally the data still refer to groups of hydrocarbons (e.g. monocyclic aromatics) rather than to individual compounds. Data for individual hydrocarbons do exist, but most deal with n-alkanes and the iso-alkanes pristane and phytane (see Figs 2 and 3). A. Studies primarily concerned with alkanes Swinnerton and Linnenbom (1967) detected the simplest n-alkane, methane, at concentrations ranging from 0.025 to 0.283 pg/l at various depths in sampling areas in the Gulf of Mexico and 0.047-0.060 pgll in the North Atlantic. Frank, Sackett, Hall and Fredericks (1970) found somewhat higher concentrations of methane, 0-06-1.25 pg/1, near oil seeps in the Gulf of Mexico: ethane and propane were also present, but at much lower levels. It is known from the work of Blumer (1970) that dissolved organic compounds in coastal waters include a variety of hydrocarbons. Thus, in a qualitative study he identified n-alkanes from C,, to C,, with maximum concentration at C,,-C,, : the compounds included those with odd and others with even numbers of carbon atoms in roughly equal amounts, a distribution different from that in recent marine sediments (where odd-numbered n-alkanes preponderate) but similar to that in marine algae (Clark and Blumer, 1967). Isoprenoid hydrocarbons were represented by pristane ((&), which is also found in marine algae (Clark and Blumer, 1967) and zooplankton (Blumer, Mullin and Thomas, 1963, 1964), as well as phytane (C2,,) which is not commonly detected in marine organisms. Olefinic hydrocarbons were also found, one being identified as squalene which is also present in copepods (Blumer et aZ., 1964) and the liver oils of various species of shark (Heller, Heller, Springer and Clark, 1957 ; Blumer, 1967 ; Corner, Denton and Forster, 1969). Some of the hydrocarbons detected by Blumer (1970) have been

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identified and estimated by Whittle, Mackie, Hardy and McIntyre (1973) in water samples collected from 13 stations off the Scottish coast. Using sub-surface samples (3 m depth) that had been filtered through a 20 pm mesh they found levels of 0.3-1.5 pg/l for total alkanes, 0.015-0-043 pg/l for pristane and < 0.001-0.014 pg/l for phytane. Similar to Blumer’s (1970) observations the peak levels for compounds. individual n-alkanes were usually obtained with C,,-C,, Hydrocarbon levels vary considerably with sea area. Thus, Mackie, Platt and Hardy (1978), using techniques similar to those of Whittle et al. (1973), found that sea-water samples from King Edward Cove, South Georgia, contained 5.8 pg/l of n-alkanes within the range n-C,5-n-C33, together with 0.18 pg pristane/l; Iliffe and Calder (1974), studying hydrocarbons in the Gulf of Mexico and Caribbean Sea, found an average level of 47 pg/l for non-polar hydrocarbons in the Florida Strait, 12 pg/l in the mid-Gulf region, 12 pg/l in the Yucatan Strait, 5 pg/l in the Cariaco Trench and 8 pg/l in the Caribbean Sea, the samples containing n-alkanes in the range C,, to C,, with peak concentrations in the C,, to C,, region ; Carlberg and Skarstedt (1972), using infrared spectroscopy, obtained values in the range < 50 to 120 pg/l for non-polar hydrocarbons a t ten stations in the Baltic and Kattegat. Hardy, Mackie, Whittle, McIntyre and Blackman (1977) have recently described further data for the amounts of n-alkanes (C15 to CS3)in samples of sea water from various regions surrounding the U.K. The lowest value for n-alkanes in the surface film (mean value 5.7 pg/m2) was found in samples from the open sea (Celtic Sea) ; the the mean value for off-shore samples from sites near urban areas (62.9 pg/m2) was close to that for samples taken near oil refineries (64.2 pg/m2)and greater than that for those collected close to North Sea oil fields (32.8 pg/m2). Mean values for n-alkanes in sub-surface (Im depth) samples ranged from 0.57 pg/l (Celtic Sea) to 4.6 pg/l (North Sea oil fields). Studies described later (Section VII) show that hydrocarbons can enter zooplankton in two different ways: first, by direct uptake from solution in sea water ; second, by assimilation from particulate diets. I n considering the quantities of hydrocarbons available to the animals in the sea it is therefore useful to know the relative amounts of the compounds that are present in solution and as particulate material. I n addition, as certain species of zooplankton feed near the surface of the sea it is necessary to consider the spatial distribution of hydrocarbons, especially evidence for the presence of high concentrations in the surface micro-layer. These topics are discussed in the next two sections.

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and particulate hydrocarbons

Spillage of Bunker C oil from the grounded tanker " Arrow " in Chedabucto Bay, Nova Scotia, led to several studies of oil levels in that area and along the coast to Halifax Harbour and beyond (Levy, 1971, 1972; Forrester, 1971). Quantitative data were obtained by Levy (1971) for the levels of petroleum residues in the open ocean off Nova Scotia and in the St Lawrence system. Water samples were filtered through a 0.45 pm millipore membrane and the hydrocarbon content of the retained material was determined as equivalents of Bunker C oil using U.V. fluorescence spectroscopy. Similar analyses were made of hydrocarbons that passed through the filter, these being described as " dissolved ". The fluorescence technique is a rapid way of detecting aromatic compounds and allows a large number of samples to be processed in ship-board experiments ; natur??- to occurring organic material can produce interference thatbut is difficul quantify (Gordon, Keizer and Dale, 1974), particularly highly conjugated alkenes (Farrington and Meyer, 1975). The total levels of petroleum residues found in Chedabucto Bay by Levy (1971) were in the range 1fj-41 pg/l (as Bunker C oil equivalents). At several stations substantially higher concentrations of dissolved than particulate compounds were detected. Thus, in surface samples (1 m depth) particulate levels ranged from 5 to 16 pg/l and dissolved from 15 to 90 pg/l. Zsolnay (1971) measured what he terms '' non-olefinic '' hydrocarbons and describes as saturated hydrocarbons and aromatic compounds with only one ring in the Gotland Deep, a Baltic basin. Thinlayer chromatography was used to separate the hydrocarbons which were then estimated as total carbon. Average concentrations, based on samples from all depths (20-200 m) and expressed as carbon equivalents, were 57-2 pg C/1 for the dissolved hydrocarbons and 1.1 pg C/1 for the particulate, dissolved material in this case being defined as that passing through a pair of Whatman GF/C glass filters. Another study using thin-layer chromatography to separate the hydrocarbons from other lipids was that of Jeffrey (1970), who measured unsaturated hydrocarbons in Baffin Bay (Texas) and found 180 pg/l as dissolved (passing through a 0.3 pm filter) and 70 pg/l as particulate material. The particulate material was mainly phytoplankton, Baffin Bay being a shallow, warm region of high primary production. Nevertheless, the distribution of hydrocarbons between dissolved and particulate forms does not always favour the soluble fractions. Sediments, for example, adsorb levels of these compounds far higher

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than those found in the associated sea water. Thus, Di Salvo and Guard (1975), studying the hydrocarbons attached to suspended sediments in San Francisco Bay, found them to contain alkanes and aromatic compounds in concentrations ranging from 190 to 6 188 mg/kg dry weight; by contrast the levels in the associated sea water were o d y 15-450 11.811. Marty and Saliot (1976) have shown that the relative amounts of n-alkanes in particulate and dissolved form depend upon whether the samples are taken from polluted or unpolluted areas. Thus, for coastal waters of the English Channel (Roscoff area) the concentrations of total dissolved (i.e. passing through a Whatman GF/C filter) C,, to C,, n-alkanes at 0.5 m depth was 0.11 pg/l compared with 0.28 pg/l for those in particulate form; by contrast, for off-shore waters near the West African coast (2 m depth), the total quantity in solution was 5.66 pg/l but that in particulate form only 0.32 pg/l. One would expect the hydrocarbons detected off the West African coast to be associated with the high primary production in a region of upwelling, for Zsolnay (1973) has described a close correlation between hydrocarbon and chlorophyll a levels in water samples from the same sea area. Likewise Parker, Winters, Van Baalen, Batterton and Scalan (1976) detected higher levels of n-alkanes in spring (0.64 pg/l) than at other seasons (0.13-0.23 pg/l) in sea water samples from the Gulf of Mexico.

C. Hydrocarbons in or near the surface of the sea The presence of high concentrations of hydrocarbons in the surface micro-layer of the sea was noted by Garrett (1967) in samples from various Atlantic and Pacific sites near North America, but the compounds were not identified. Swinnerton and Linnenbom (1967) measured n-alkanes of low molecular weight (mainly methane) by gaschromatography in water samples from the Gulf of Mexico (South of Mobile, Alabama) and North Atlantic (500 km west of Ireland). They found higher concentrations a t the surface than a t depth (500 m) in the Gulf of Mexico samples, although peak concentrations occurred a t 30-40m. No significant change in hydrocarbon level with depth was observed in the Gulf of Mexico survey by Frank et al. (1970). Iliffe and Calder (1 974) found higher levels of non-polar hydrocarbons at a depth of 1 m (24 pg/l) than at other depths in the Yucatan Strait, but in the Florida Strait the highest hydrocarbon concentration (75 pg/l) was a t a depth of 144 m. Whittle, Mackie and Hardy (1974),

POLLUTION STUDIES WITH MBRINE PLANKTON-I

295

analysing hydrocarbons at different depths in the Clyde, found only 3.21 pg/l in the surface film compared with 7-8 pg/l in the top 15 cm, although at middle depth (10 m) the value obtained was an order of magnitude lower (0.31 pg/l). Duce, Quinn, Olney, Piotrowicz, Ray and Wade (1972) detected three hydrocarbons, tentatively identified as CZ1.,, C,,., and C,,., at a concentration of 8.5 pg/l in the surface micro-layer (100-150 pm) compared with 5.9 pg/l at 20 cm depth. Wade and Quinn (1975) measured the total hydrocarbons present in samples of the surface micro-layer (100-300 pm) from the Sargasso Sea and found the levels to vary from 14 to 559 pg/l (average 155) compared with 13-239 pg/l (average 73) at 20-30 cm depth: n-alkanes from C,, to C,, accounted for 11% of the total hydrocarbons in combined micro-layer and subsurface samples, being present at an average level of 25.1 pg/l. The authors concluded that a major source of the hydrocarbons was particles of weathered pelagic tar with diameter ranging from 1.0 mm down to 0.3 pm located in the surface micro-layer. Earlier, Morris and Butler (1973) had reported the large amounts of pelagic tar that could be collected by neuston net from the surface of the Sargasso Sea, the average value being 9.4 mg/m2. By comparison, the mean level recovered in the same way from the surface of the North Sea was only 317 pg/m2 (Offenheimer, Gunkel and Gassmann, 1977). The accumulation and retention of floating material in the Sargasso Sea is well known. The average level for pelagic tar in the Mediterranean was even greater : thus, Morris and Butler (1973) gave a figure of 20 mg/m2. However, evidence from a more recent study (Morris, Butler and Zsolnay 1975) indicates that the average level of pelagic tar in the Mediterranean has now fallen to 9.7 mg/m2, a value much closer to that for the Sargasso Sea. Conover (1971) has shown that zooplankton are able to ingest small droplets of oil and it seems probable that zooplankton species such as Anomalocera patersoni Templeton that live near the surface of the sea could also ingest small tar particles. Hydrocarbons assimilated from these particles might then be available for transfer to higher trophic levels ; in addition, unassimilated material could eventually reach the benthos as faecal pellets (see p. 348). Tar particles represent a persistent legacy of spilt oil, probably taking years to be degraded because they contain large amounts of high-melting point waxes and asphaltenes (Morris and Bulter, 1973). Further observations on surface enrichment of n-alkanes have been made by Marty and Saliot (1976). The ratio between the concentration of dissolved compounds in the micro-layer (0.44 mm film)

296

E. D. S. OORNER

and that in the underlying water ranged from 6.3 :1 (Etang de Berre : Marseilles) to 161 :1 (Roscoff area) : the corresponding values in terms of particulate hydrocarbons were 170 :1 and 350 :1 respectively. It should be noted that these ratios, if calculated for a micro-layer of only 100 d thickness, would give enrichment factors 104-106 times greater. Marty and Saliot (1976) concluded that the n-alkanes present in the surface micro-layer were in general of biological origin as they possessed a distribution concentrating on n-C,, to n-C,, which was found by Clarke and Blumer (1967) to be characteristic of marine algae. However, qualitative differences occur between sea areas : thus, Ledet and Laseter (1 974) describe the alkanes at the air-sea interface from off-shore Louisiana and Florida as mainly branched and cyclic compounds. Ideally, to establish the biological origin of hydrocarbons in sea-water samples from a particular area i t is necessary to make a direct comparison of these compounds with those present in the plankton: however, no detailed study of this type seems to have been made. Concerning work with aromatic hydrocarbons Levy (1971), in his studies of oil pollution in Chedabucto Bay, found values of 15-90 pgll for dissolved compounds a t a depth of 1 m compared with 7-9 pg/1 at 20 m. On the other hand, Gordon and Michalik (1971)) working in the same sea area, found slightly increasing concentrations with depth : 1.2 pg/l at 5 m, 1.4 pgll at 6-25 m and 1.8pgll at 26-50m. Subsequently, however, in a detailed study of this aspect in the northwest Atlantic Ocean, Gordon et al. (1974)) using Venezuelan crude oil as a reference standard for U.V. fluorescence measurements, obtained concentrations at the surface (0-3 mm) averaging 20-4 pg/l compared with 0.8 pg/l at 1 in and 0.4 pgll a t 5 m. Studies that include measurements of total mineral oil hydrocarbons have given conflicting evidence. Thus, Carlberg and Skarstedt (1972), using samples from Gijteborg Harbour, found values of 0.71 mg/l at the surface compared with 0.47 pg/l at 6 m depth. However, Pavletid, Munjko, Jardas and Matoricken (1975), estimating mineral oil concentrations at different depths in the Adriatic off the Jugoslaviaii coast, found values of 1-40, 0.65, 1.56 and 10.98 mg/l at depths of 0, 2, 5 and 10 m at Monte Gargano ;but at another station (Pelegrin) surface samples were higher than those at depth, being 4.23, 2.39 and 0.82 mg/l at 0, 5 and 10 m respectively. The various hydrocarbon levels in the sea that have so far been discussed are summarized in Table I.

TABLEI. EX~D~PLES OF HYDROCARBON LEVELSIN

Type of hydrocarbon Methane

Concentration 0.025 to 0.283

Methane

0.047 to 0.060

Methane

0.06 to 1.25

THE

SEA

Geographic location

Reference

Various depths between 0 and 500 m : Gulf of Mexico Various depths between 0 and 500 m : North Atlantic Various depths between 0 and 3 742 m : Gulf of Mexico

Swinnerton and Linnenbom (1967) Swinnerton and Linnenbom (1967) Frank et al. (1970)

3 m depth : Scottish Coast 3 m depth : Scottish Coast 3 m depth : Scottish Coast 0 to 20 m : King Edward Cove, S. Georgia 0 to 20 m : King Edward Cove, S. Georgia 1 m depth: Celtic Sea 1 m depth : North Sea 0 to 500 m :Florida Strait, Gulf of Mexico 0 to 500 m : Mid-Gulf 0 to 500 m : Yucatan Strait 0 to 900 m : Carioco Trench 0 to 200 m : Caribbean Sea 0 to 100 m : Baltic and Kattegat 0 to 100 m : Baltic and Kattegat 0 to 31 m : Goteborg Harbour 0 to 31 m : Goteborg Harbour 1 m depth : Chedabucto Bay, Nova Scotia 20 m depth :Chedabucto Bay, Nova Scotia

Whittle et al. (1973) Whittle et al. (1973) Whittle et al. (1 973) Mackie et al. (1978) Mackie et al. (1978) Hardy et al. (1977) Hardy et al. (1977) Iliffe and Calder (1974) Iliffe and Calder (1974) Iliffe and Calder (1974) Iliffe and Calder (1974) Iliffe and Calder (1974) Carlberg and Skarstedt (1972) Carlberg and Skarstedt (1972) Carlberg and Skarstedt (1972) Carlberg and Skarstedt (1972) Levy (1971) Levy (1971)

(PLgIl) n-Alkanes Pristane Phytane n-Alkanes (C15 to C=) Pristane n-Alkanes (C15 to CJ n-Alkanes (C15 to C3J Non-polar Non-polar Non-polar Non-polar Non-polar Non-polar Total Non-polar Total Dissolved aromatic Dissolved aromatic

0.3 to 1-5 0-015 to 0.043 <0*001 to 0.014 5-8 0.18 0.57 4.5 47 12 12 5 8 <50 to 120 <50 to 170 <50 to 200 50 to 710 15 to 90 7 to 9

0

2 2

:E e3

8

8z ti 2 c cd

3

+I

1

H

t.a CD

l

t 9

a

TABLEI-( continued) Type of hydrocarbon

Particulate aromatic Particulate aromatic Dissolved non-olefinic Dissolved non-olefinic Dissolved non-olefinic Dissolved non-olefinic Dissolved non-olehic Particulate non-olefmic Particulate non-olehic Particulate non - olefinic Particulate non-olefinic Particulate non-olehic Dissolved unsaturated Particulate unsaturated Dissolved n-alkanes Particulate n-alkanes Dissolved n-alkanes (C14 to C8,) Particulate n-alkanes (C14 to CS7) Dissolved n-alkanes Particu1at.e n-alkanes Dissolved n-alkanes (Cia to C3,) Particulate n-alkanes (C14 to C37) Total hydrocarbons Tot,al hydrocarbons

Concentration

(WlO 5 to 16

2 to 11 48 58 58 59 64 0.9 1.0 2.3 1.0 0.5 180 70 17.7

98.0 0.1 1 0-28 1144 3-34 5.66 0.32 1 4 to 559 13 to 239

Geographic location 1m depth : Chedabucto Bay, Nova Scotia 20 m depth : Chedabucto Bay, Nova Scotia 20 m depth : Gotland Deep, Baltic Basin 70 m depth :Gotland Deep, Baltic Basin 110 m depth : Gotland Deep, Baltic Basin 150 m depth :Gotland Deep, Baltic Basin 200 m depth : Gotland Deep, Baltic Basin 20 m depth :Gotland Deep, Baltic Basin 70 m depth :Gotland Deep, Baltic Basin 110 m depth :Gotland Deep, Baltic Basin 150 m depth : Gotland Deep, Baltic Basin 200 m depth :Gotland Deep, Baltic Basin (depth not given) : Baffin Bay, Texas (depth not given) : Baffin Bay, Texas Surface micro-layer : Roscoff, English Channel Surface micro-layer : Roscoff, English Channel 0.5 m depth : Roscoff, English Channel 0.5 m depth : Roscoff, English Channel Surface micro-layer : West African Coast Surface micro-layer : West African Coast 2m depth : West African Coast 2 m depth : West African Coast Surface micro-laver : Sareasso Sea 20 to 30 cm depth : Sarg&o Sea

0,

Reference Levy (1971) Levy (1971) Zsolnay (1971) Zsolnay (1971) Zsolnay ( 1971 ) Zsolnay (1971) Zsolnay (1971) Zsolnay (1971) Zsolnay (1971) Zsolnay (1971) Zsolnay (1971) Zsolnay ( 1971 ) Jeffrey (1970) Jeffrey (1970) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Marty and Saliot (1976) Wade and Quinn (1975) Wade and Quinn (1975)

pl

P P Q

0 Ld

3a

Total aromatic Total aromatic Total saturated Total a.romatics Total hydrocarbons Total hydrocarbons Volatile hydrocarbons (C, to C,) Volatile hydrocarbons (C, to C,) Non-volatile hydrocarbons Non-volatile hydrocarbons Total dissolved hydrocarbons Total dissolved hydrocarbons Total dissolved hydrocarbons Total dissolved hydrocarbons Total dissolved hydrocarbons Total oil Total oil Total oil n-Alkanes n-Alkanes n -Alkanes Total hydrocarbons Total hydrocarbons Total hydrocarbons

20.4 0.4 to 0.8 1 to 21 < 1 to 3 0.8 to 5 0.3 to 2 0.33 0.10 14 to 270 25 137 10 19 37 43 1400 to 4 230 1560 to 2 390 820 to 10 980 0.64 0.23 0.13 17 to 625 2.7 to 14.9 0.95 to 30.5

Surface : Chedabucto Bay Gordon et al. (1974) 1-5 m depth : Chedabucto Bay Gordon et al. (1974) Oto lOmdepth :NorthAtlantictankerroutesBrown et al. (1973) 0 to lOmdepth :NorthAtlantictankerroutes Brown et al. (1973) Brown and Searl (1976) Surface : Pacific tanker routes Brown and Searl (1976) 3 to 10 m depth: Pacific tanker routes Surface : Pacific tanker routes Koons (1977) 10 m depth : Pacific tanker routes Koons (1977) Searl et al. (1977) Surface : New York Harbour 0 to 10 m : Tokyo Harbour Brown et al. (1976) Surface : Brest Harbour Barbier et al. (1973) 2 000 m depth : West African Coast Barbier et al. (1973) 500 m depth : West African Coast Barbier et al. (1973) 4 500 m depth : West African Coast Barbier et al. (1973) 50 m deuth : West African Coast Barbier et al. (1973) Pavletid et al. '(1975) Surface :*Adriatic Pavleti6 et al. (1975) 5 m depth : Adriatic Pavleti6 et al. (1975) 10 m depth : Adriatic Parker et al. (1976) Spring: Gulf of Mexico Summer : Gulf of Mexico Parker et al. (1976) Parker et al. (1976) Winter : Gulf of Mexico Offenheimer et al. (1977) 10 m depth : North Sea Data from Hertz (1974) cited Surface : Gulf of Alaska by Myers and Gunnerson (1976) Data from Hertz (1974) cited Subsurface : Gulf of Alaska by Myers and Gunnerson (1976)

cd 0

2 82

m

! s I 1 ci

8

E;1 td

*

E

!I 8 I H

300

E. D. 8. UORNER

D. Comprehensive analyses Few data exist covering the whole range of hydrocarbons in seawater samples ; which is not surprising bearing in mind the difficulties encountered in analysing such a wide range of compounds. Frequently, the concentrations are so low that analytical equipment must be operated at maximum sensitivity ; moreover, the samples can easily be contaminated during collection. A further complication is that hydrocarbons do not maintain a steady concentration. They are constantly being removed or modified by processes such as microbial degradation, accumulation and metabolism by plankton and larger marine organisms, chemical and photochemical oxidation, volatilization, dissolution and adsorption on particulate material : at the same time they are being renewed by processes such as atmospheric transport, oil spills, submarine seeps and release from organisms. Furthermore, in coastal areas, industrial effluents, sewage and rivers make a further contribution which can sometimes be substantial. Thus, Hites and Biemaiin (1972), studying organic compounds in the Charles River (Boston), detected the aromatic hydrocarbon naphthalene at a maximum concentration of 3-4mg/l. Other examples of high levels of oil in the sea are the discharges from off-shore production facilities, such as those in the North Sea, which on average contain a 26 mg/l dispersion of oil in water (C.U.E.P. Pollution Paper No. 6, 1976). Not unexpectedly, high concentrations of oil are also found in the immediate vicinity of oil slicks : for example, Cormack and Nichols (1977) give values of 0.79-3.95 mg/l for oil concentrations a t a depth of 2 m beneath the centre of a small slick of “ Ekofisk ” oil. I n the study by Brown et al. (1973) ocean water samples were collected by tankers operating along the U.S. Gulf coast to East coast and the Caribbean to East coast. Two samples ( 3 1) were taken daily a t 12 h intervals, one from the surface using a bucket and the other through the bottom of the ship using a sanitary line (10 m depth) : special precautions were taken to avoid contamination. Approximately 400 samples were examined, concentrations of saturated hydrocarbons (alkanes and 1- t o 6-ring naphthenes) varying in the range 1 to 21 pg/l and those of aromatic compounds from 1 to 3 pg/l. As in several studies mentioned earlier, the concentrations in surface samples were greater than in those taken from 10 m depth. Average percentages of the total quantities of hydrocarbons accounted for by the various fractions, based on all the data, are shown in Table 11. Values obtained for aromatic substances showed that the simpler compounds (benzenes, indanes and indenes) were well repre-

301

POLLUTION STUDIES WITR MARINE PLANKTON-I

sented and that the levels of tetra-cyclic aromatics were relatively low. This is a distribution similar to that found in many crude oils, but Brown et al. (1973) suggested that sources other than this, such as organic materials released from sediments, could also have contributed. Their data demonstrate the low levels of particular groups of aromatic hydrocarbons found in Atlantic Ocean water. Taking the naphthalenes as an example: the highest value for total hydrocarbons was 50 pg/l and, if it be assumed (Table 11)that naphthalenes accounted for 4.3%, these compounds had a maximum concentration of only 2.2 pg/l. On many occasions total hydrocarbons amounted to only 1.0 pg/l; which gives a concentration of naphthalenes as low as 0.043 pg/L TABLE11.

RELATIVE AMOUNTS OF HYDROCARBON FRACTIONS EXTRACTED FROM ATLANTIC SEAWATER ~

Praction Paraffis Naphthenes Benzenes Indanes Indenes Naphthalenes Acenaphthenes Fluorenes Phenanthrenes Tetra-aromatics Benzothiophenes Dibenzothiophenes

Percentage of total Range Mean 10 to 0 to 2 to 4 to 5 to 3 to 4 to 3 to 0 to 2 to 0 to 0 to

27 25 13 13 13 8 9 12 15 7 24 9

18.5 9-8

9.5 9.8

8.7 4.3 6.7 8.5 8.3 3-7 7.3 4.8

Adapted from Brown et al. (1973).

Recently, Brown and Searl (1976) have measured the total hydrocarbons, both dissolved and particulate, in sea-water samples along tanker routes in the Pacific Ocean. Concentrations had average levels of 2 (0.8-5 pg/l) for surface waters and 0.8 (0-3-2 pg/l) for subsurface (3m and 10m depth) samples. I n nearly all cases the hydrocarbons were complex mixtures of paraffins, cyclo-alkanes, and 1,- 2- and 3-ring aromatics. Along the Singapore to San Francisco route aromatics accounted for 36% of the total; but on all other routes the value ranged from 15 to 22%. Taking the values for all samples, both surface and sub-surface, aromatic hydrocarbons in the Pacific range from 0.21 to 0.50 pg/l. Complementingthe work of Brown and Searl (1976) are the measure-

302

E. D. 8. CORNER

ments by Koons (1977) of volatile hydrocarbons along tanker routes in the Pacific Ocean. The hydrocarbons were in the range C, to C, and included saturated compounds such as n-pentane, cyclopentane, n-hexane, methylcyclopentane, n-heptane, methylcyclohexane and n-octane, as well as the aromatic compounds benzene, toluene and xylenes. The average concentration found in samples taken near the surface was 0.33 pg/l compared with 0.10 pgll for those from a depth of 10 m. Another comprehensive analysis of the hydrocarbons in sea water was that of Barbier et al. (1973) who examined samples from the English Channel (Brest and Roscoff ), Mediterranean (Villefranche) and off the west coast of Africa. All the samples were filtered through a 0.45 pm millipore membrane and so the data refer to " dissolved " hydrocarbons only. These were extracted from each sample (100 1) with chloroform, the extracts then being dried and saponified. Kydrocarbons were separated from the unsaponifiable material by thin-layer chromatography, then analysed by gas-liquid chromatography and, as in the study by Brown et al. (1973)) by mass spectrometry. Coastal waters, as well as surface waters, had hydrocarbon contents greater than those found in deep-water samples (500-5 400 m depth). The values ranged from 10 pgfl (open ocean off West Africa) to 137 pg/l (Creek of Poulmic: Brest Harbour) with an average of 40 pg/l. Further evidence for high levels of hydrocarbons in estuarine waters was found by Searl, Huffman and Thomas (1977) in their study of non-volatile hydrocarbons in New York Harbour. Quantities ranged from 14 to 270 pg/l with an average of 39 pg/1, this value being an order of magnitude higher than that found in open Atlantic Ocean waters TABLE111. RELATIVE AMOUNTS OF HYDROCARBON FRACTIONS EXTRACTED FROM BREST HARBOUR SEAWATER ___

Fraction n- and iao-alkanes

1-ring naphthenes 2-ring naphthenes 3-ring naphthenes 4- and > 4-ring naphthenes Mono-cyclic aromatics Bi-cyclic aromatics Poly-cyclic aromatics Data from Barbier et al. (1973).

Percentage of total 51.5 5.5 9.5 6.5 4-0 18.0

3-5 2.5

POLLUTION STUDIES WITH MARINE PLANKTON-I

303

(Brown et al., 1973) but close to the average of 25 pg/l found for Tokyo Harbour (Brown, Sear1 and Koons, 1976). The Brest sample, when analysed in detail by Barbier et al. (1973) using U.V. spectrometry and mass spectrometry, was found to have the percentage composition shown in Table 111. The n-alkanes in both coastal and open-sea waters ranged from C,, to C,,, the most abundant being in the C2, to C,, region, as found in other sea areas by Blumer (1970) and Whittle et al. (1973). There was no predominance of odd-numbered carbon compounds and generally the pattern of distribution resembled that found for marine algae by Clark and Blumer (1967). However, in coastal water the presence of aromatic compounds, such as those found in the Brest sample, indicated pollution. Only the Brest sample was analysed in detail, but assuming the data to apply generally and, again taking naphthalenes (or bi-cyclic aromatic compounds) as an example, the levels of this group of compounds varied within the range 0.35-4-9 pg/l, compared with values of 0-043-2.2 pg/l found by Brown et al. (1973). Bearing in mind that the samples analysed by Barbier et al. (1973) and by Brown et al. (1973) were taken from different sea areas, that there were certain differences in the analytical methods used and that Barbier et al. measured dissolved hydrocarbons whereas Brown et al. determined both these and the particulate fraction, the data are sufficiently close, at least at the higher end of the range, to provide a reasonable guide to the background levels of aromatic compounds that should be used in designing laboratory studies concerned with problems such as the uptake and retention of these compounds by plankton and their possible effects on the organisms.

111. HYDROCARBONS IN PLANKTON Determining whether planktonic organisms are contaminated with petroleum hydrocarbons in sea areas prone to oil pollution is complicated by the need to recognize that man-made pollution, such as an accidental discharge of crude oil, is not the only source of such compounds in these plants and animals : marine organisms are themselves capable of biosynthesizing hydrocarbons. A challenging problem for the analytical chemist has therefore been that of distinguishing between compounds such as hydrocarbons from fossil fuels and those of recent biogenic origin in the organisms. Several studies, discussed in detail in the next two sections, have shown that the hydrocarbons native to marine planktonic organisms

304

E. D. 9. CORNER

include only a few representatives of any one group of compounds. Crude oil, however, contains a much more complex mixture (Posthuma, 1977). For example, marine phytoplankton have only a restricted range of n-alkanes, whereas these usually occur in crude oil in a continuous homologous series from C, to C4,,. Likewise, zooplankton contain only a few branched alkanes (iso-alkanes), the major one being pristane ; crude oil, on the other hand, includes a wide range of these Alkanes(n-andiso-o-)

Tetra lins

Cycloalkanes

No phthalenes

Biphenyls

Benzololpyrene

FIG. 1. Structural formulae of various hydrooarbone found in crude oil. IE represents several types of alkyl substituent.

compounds. Cyclo-alkanes (naphthenes), particularly cyclopentane and cyclohexane derivatives with both substituted and unsubstituted rings, aromatic hydrocarbons including 1- to 5-ring compounds, together with their alkylated forms, and naphthenoaromatics such as the tetralins are all well represented in crude oil but are not normally found in planktonic organisms (Koons and Monaghan, 1976). By contrast, the alkenes (olefins),representatives of which have been found in both phytoplankton and zooplankton, are generally absent from crude oil (although they can occur in refinery products).

POILUTION STUDIES WITH MARINE PLANKTON-I

306

The structural formulae of some of the hydrocarbons found in crude oil are shown in Fig. 1. A. Phyloplankton The first investigation of hydrocarbons in phytoplankton using modern analytical methods was that of Clark and Blumer (1967). Cultures of three species of phytoplankton were used : Syracosphacra earterae, now H y m e n o m m carterae (Braarud e t Fagerl.) Braarud, Skeletonem costatum and an undetermined cryptomonad. Analyses of all the n-alkanes within the range C84H30 to C,,H,, showed that the total amounts varied from 34 to 121 mg/kg dry weight; also that in each species one particular n-alkane predominated, i.e. n-C,, in Skeletonem and the unknown cryptomonad and n-C,, in Syracosphaera. The predominance of n-C,, in the latter species was particularly marked in that it accounted for 45.5% of the total n-alkanes. The carbon preference index (i.e. the ratio of compounds containing an odd number to those with an even number of carbon atoms) was 1.1-1.2, so there w&s little evidence of the marked odd-carbon predominance ” found in marine sediments (CPI values of 2 4 4 . 5 : Cooper and Bray, 1963) ; which suggests that the source of a large proportion of the n-alkanes in sediments may not be marine phytoplankton. The isoprenoid hydrocarbon pristane (2, 6, 10, 14-tetrameth~lpentadecane)~ present in recent marine sediments (Blumer and Snyder, 1965), in petroleum (Bendoraitis, Brown and Hepner, 1963) and in zooplankton (Blumer et al., 1963, 1964) was also detected in all three species of phytoplankton. No studies of the mechanisms by which hydrocarbons are synthesized in marine unicellular algae seem to have been made. However, as far as the n-alkanes are concerned, work with higher plants indicates that the main mechanism is likely to be one of elongation from palmitic acid (C16)by the addition of C, units from malonyl-CoA, followed by decarboxylation (Kolattukudy, 1976), although recent work by Murray, Thomson, Stagg, Hardy, Whittle and Mackie (1977) indicates that in marine phytoplankton this process of chain-elongation may be limited. Thus, Murray et al. (1977) measured the radioactivity in aliphatic hydrocarbons present in various species of phytoplankton cultured with W-labelled Na,C03, and in mixed zooplankton feeding on the plant cells. Generally, only a few specific hydrocarbons were found to be labelled, compared with the wide array present in the plants and animals. I n particular, there was little evidence that long-chain hydrocarbons (C2a to C,.J were synthesized by either micro-algae or zooplankton ; which implies that such compounds, which have been detected in natural plankton samples, are exogenous in origin. A.H.B.-~S

13

306

E. D. 9. CORNER

Work by Lee, Nevenzel, Paffenh6fer, Benson, Patton and Kavanagh (1970) identified the C,lH,, olefinic hydrocarbon all-cis-3, 6, 9, 12, 15, 18-heneicosahexaene (HEH) in Skeletonema costatum (see Fig. 2) ; and Blumer, Mullin and Guillard (1970) investigated its distribution in numerous species of marine phytoplankton (Table IV). The presence of the C,, fatty acid docosa-all-cis-4, 7, 10, 13, 16, 19-hexaenoic acid in these algae led Blumer et aE. (1970) to suggest that HEH might arise by decarboxylation of this compound. However, later work (Youngblood and Blumer, 1973) showed that HEH was also present in three species of brown benthic algae that did not contain the C,, fatty acid ; which suggests that the hydrocarbon may also be derived in other ways. TABLEIV. H~~NEICOSAHEXAENE (HEH) CONTENTSor U r m UNICELLULAR ALUAJ3 ~~

No. of teat species Bacillariophyceae Dinophyceae Cryptophyceae Haptophyceae Euglenaphyceae Prasinophyoeae Cyanophyceae Rhodophyceae Xanthophyceae Chlorophyceae

2 2 2 3

1 1 1 1 1 1

HEH aa

yo

wet weight

0.00036 to 0.0027 0.0037 to 0.0040 10.0006 to 0.008 0.0015 to 0.010 0.0035 <0-0009 <0*00001 <0.00004 <0~00008

<0~000016

Summarized data from Blumer et al. (1970).

I n a later study (Blumer, Guillard and Chase, 1971) 22 species of marine planktonic algae belonging to nine algal classes were analysed (see Table V). Trace amounts of n-alkanes within the range n-C14H,, t o n-CZ5H5,were found in all the species, most of which also contained small quantities of pristane. However, among the groups Bacillariophyceae, Dinophyceae, Cryptophyceae, Haptophyceae and Euglenaphyceae the predominant hydrocarbon was HEH, the only exception being Rhizosolenia setigera, in which the predominant hydrocarbon was n-heneicosane (C,lH44); another centric diatom Tlzalassiosira Jlzcviatilis contained, in addition to HEH, a C,, tetra-olefin, but only as a minor component. Further observations, using species representing the Cryptophyceae (Blumer et al., 1970) and the Dinophyceae (Blumer et al., 1971), showed that HEH was most actively synthesized during

POLLUTION STUDIES WITH MARINE PLANKTON-I

307

the logarithmic growth phase : cultures harvested during the stationary phase contained greater amounts of C,, to C,, n-alkanes. By contrast, five algal species representing the Rhodophyceae, Xanthophyceae and Chlorophyceae did not contain HEH : instead, the predominant hydrocarbons were either n-C,,H3, or n-Cl7H3,, or olefins such as an unclassified pentadecene or 7-heptadecene. Two blue-green algae of the class Cyanophyceae were also studied. In one species, Oscillatoria woronichinii, the predominant hydrocarbon was n-C1,H3,, with traces of other n-alkanes ;in the other, Synechowccus bacillaris, the olefin 5-heptadecene predominated, with n-C15H32, n-C,,H,, and n-Cl,H3, also abundant and C,, and C,, mono-olefins present in low amounts. Tornabene, Kates and Volcani (1974), in studies using the nonphotosynthetic diatom Nitzschia alba Lewin et Lewin, found that aliphatic hydrocarbons accounted for about 0.1yo of the total lipids. Pristane, phytane and several long-chain n-alkanes (C,, to C2,) were detected. The presence of phytane in Nitzschia is interesting as this compound is normally regarded as non-biogenic in origin, its source being fossil fuels. The olefin HEH, characteristic of photosynthetic diatoms, was not found : instead, the predominant hydrocarbons were even-numbered C,,, c1, and C,, olehs, in contrast to the odd-numbered compounds found by Blumer et al. (1971). Possibly photosynthetic and non-photosynthetic diatoms have different pathways for the biosynthesis of hydrocarbons. Compared with those of n-alkanes, branched alkanes and olefins, analyses of aromatic hydrocarbons in phytoplankton are few. Smith ( 1954) reported that cycloalkanes and aromatic compounds accounted for more than 0.2% by weight of a dried sample of phytoplankton collected near Woods Hole, Massachusetts. More recent information mainly concerns levels of the carcinogen benzo[a]pyrene (BP), but there are doubts about some of the analytical methods used (Farrington and Meyer, 1975). Mallet and Sardou (1965) detected B P in amounts up to 400 pg/kg dry weight in samples of mixed plankton from the Bay of Villefranche, compared with a value of 5.5 pg/kg dry weight for a sample collected off the west coast of Greenland (Mallet, Perdriau and Perdriau, 1963), an area less prone to man-made pollution. The detection of B P in phytoplankton from another remote area, Clipperton Lagoon in the East Pacific, has been reported by Niaussat (1970) and Ehrhardt (1972). The synthesis of B P and other polynuclear aromatic hydrocarbons (PNAH) by bacteria-free cultures of the freshwater species Chlorella vulgaris Beij. has been demonstrated by Borneff, Selenka, Kunte and

308

1.D. 9. CORNER

TABLEV. HYDROCARBONS IN MARINEUNICELLULAR ALGAE Predominant hydrocarbon

Class and species

Trace hydrocarbons

Bacillariophyceae Cyclotella nana* Ditylum brightwellii (West) Van Heurck Lauderia borealis Gran Rhizosolenia setigera Brightw. Skeletonema costatum (Grev.) Cleve Thalassiosira%uviatilis Hust.

HEH

Pristane, n-alkanes

HEH HEH HEH, n-C,,, n-C,,

n-alkanes n-alkanes Pristane, n-alkanes

HEH HEH

Thalassiosira sp.

HEH

Pristane, n-alkanes Pristane, n-alkanes, n-C,, : 4 Pristane, n-alkanes, n-C,, : 4

Dinophyceae Gonyaulax polyedra Stein Gymnodinium splendens Lebour Peridinium trochoideum (Stein) Lemm. Peridinium trochoideum (old culture)

HEH HEH

n-alkanes n-alkanes

HEH

n -alkanes

HEH, n-C,,, n-C,,

n-alkanes

HEH HEH

Pristane, n-alkanes Pristane, n-alkanes

HEH HEH

Pristane, n-alkanes Pristane, n-alkanes

HEH

Pristane, n-alkanes

Euglenaphyceae Eutrepiella sp.

HEH, n-C,,, n-C,,

Pristsne, n-alkanes

Cyanophyceae Oscillatoria woronichinii Anissimova Synechoccus bacillaris Butch.

n-c,, n-C,, alkene, n-C,,

n -alkanes n-alkanes, other olefins

n-c17

n-alkanes

n-C,, alkene, n-C,,,

n-alkanes, other olefins

Cryptophyceae Cryptomonas (Rhodomonas?) Cryptomonas (old culture) Haptophyceae Coccolithus huxZeyi (Lohm.) Kampt. Isochrysis galbana Parke Phaeocystis pouchetii (Hariot) Lagerh.

Rhodophyceae Porphyridium sp. Xanthophyceae Tribonema aequa2e Pascher

"-el, Undetermined species

n-c,,, n-c,,, n-C,, alkene

n-alkanes, other olefins

POLLUTION STUDIES WITH MARINE PLANKTON-I

309

TABLEV (continued) Class and species Chlorophyceae Dunaliella tertwlecta Butch. Crouan frat. Derbesia tenuksima (De Not.)

Predominant hydrocarbon

Trace Hydrocarbons

n-C,, alkene n-C,, alkene, n-C,,

n-alkanes n-alkanes, other olefins

* Thalassiosira pseudonana Has10 et Heimdal [as Cyclotella nana Hust.]. HEH is 3, 6, 9, 12, 15, 18-heneicosahexaene (presumed all cis) : n-C, is a normal alkane with x carbon atoms : A-C, alkene is a normal alkene with x carbon atoms; nC,,:4 is a tetraoleiin. Adapted from Blumer et al. (1971). Maximos (1968). The alga was grown with 14C-labelledacetate added to the medium and radioactivity was eventually detected in the hydrocarbons, this technique being used to exclude the possibility that the compounds resulted from external contamination. No studies of this kind, however, seem to have been made with marine unicellular algae. It is to be hoped that more detailed data concerning the distribution and levels of aromatic hydrocarbons, particularly PNAH, in plankton will be obtained now that modern methods for analysing these compounds are being applied in the marine environment (Giger and Blumer, 1974).

B. Phytoplankton and crude oil as sources of hydrocarbons in the sea Interest in the quantitative importance of phytoplankton as a source of hydrocarbons in the sea prompted the following attempt t o compare the contribution from marine unicellular algae with that from crude oil. Note, however, that these are not the only sources of hydrocarbons in the sea : Peuerstein (1973) estimates that global emissions of hydrocarbons into the atmosphere total 90 x 106 metric tons per annum (mta) of which an average of 0.6 x lo6 mta, or roughly 0.7%, eventually reaches the sea. According to Grossling (1976) the total inputs of crude oil into the world’s oceans, based on 1972 levels of economic activity, are as shown in Table VI. The total, 3.77 x lo6 mta, does not include the contribution from on-shore oil seepage that may eventually reach the sea : nevertheless, it comes within the range of values (2.5-4.0 x los mta) previously given by others (e.g. Brummage, 1973 ; Charter, Sutherland

310

E. D. 9. OORNER

and Porricelli, 1973). Additional to these inputs is that from natural submarine seepage, for which Wilson (1973) gives a figure of 0.6 x los mta based on the average for high (Southern California)and low (western Canada) seepage areas. The overall total for oil from all sources is therefore about 4.4 x los mta.

TABLEVI. INPUTS OF OIL FOR

THE

WORLD’S OCEANS

~~

Ocean intake Source

( x 106 metric t o m

per annum) Industrial spent lubricants Automotive spent lubricants Aviation spent lubricants On-shore oil well accidents Off-shore oil well accidents Tanker cleaning operations Tanker accidents Off-shore pipe-line accidents On-shore pipe-line accidents

1.43 0.89 0.04 (0.53 0.33 0.35 0.19 0.01 0.001

Adapted from Grossling (1976).

Clark and Blumer (1967) found an average value of 72 mg/kg dry weight for hydrocarbons in marine phytoplankton, a figure that should be regarded as minimal in that it refers only to n-alkanes which are not always the predominant hydrocarbons in marine algae (see p. 308). A feasible estimate for primary production in the world’s oceans is that of Ryther (1969) who gives a value of 20 x los metric tons of organic carbon per year. According to the data of Parsons, Stephens and Strickland (1961), organic carbon accounts on average for 37% of the dry weight of marine phytoplankton : thus, combining this value with that of Clark and Blumer (1967), the average quantity of hydrocarbons in these organisms is 0.195 mg/g organic carbon. The total annual production of hydrocarbons as phytoplankton is therefore 3.9 x lo6 mta, which is similar to that of 4.4 x los mta contributed by crude oil. Such close agreement, bearing in mind the number of assumptions made, is probably fortuitous. Nevertheless, it seems reasonable to conclude that the annual quantity of hydrocarbons released into the sea aa crude oil and that produced as phytoplankton axe of the same order of magnitude.

POLLUTION STUDIES WITH HABINBI PLA?SKTON-I

311

C. Zooplankton Quantitatively, one of the most important hydrocarbons in zooplankton is pristane (2, 6 , 10, 14-tetramethylpentadecane)which is also present in the livers of basking sharks and sperm whales as well aa being a constituent of various crude oils (Blumer et al., 1964). The hydrocarbon was fist identxed in zooplankton by Blumer et al. (1963) who showed that it accounted for 0.46-0.90% of the dry weight and 0.86-2-9% of the total lipids in calanoid copepods collected from the Gulf of Maine. The highest values were obtained with the Borealarctic species Calanus hyperboreus and in a later study (Blumer et al., 1964) it was shown that when the animals were starved for 86 days, although all the weight loss was accounted for as a decrease in lipid coptent, pristane actually increased slightly, presumably being slowly formed from precursors. It would be interesting to know how pristane levels vary in calanoid species more active metabolically than C . hyperboreus which, during summer and autumn, enters a non-feeding " diapause " (Conover, 1962). Other species of zooplankton, including representatives of the chaetognaths, pteropods, ostracods, amphipods and euphausiids, were found to possess very little pristane in comparison with the copepods ; and even among these only the calanoids contained substantial quantities (Blumer et al., 1964: Bee Table VII). The pathways of biosynthesis of hydrocarbons in marine zooplankton have received little study. However, Avigan and Blumer (1968), using tracer isotope methods, showed that the pristane in calanoid copepods could be formed from phytol, a C,,-alcohol present in algal diets as a constituent of chlorophyll. Other phytol-derived hydrocarbons, detected in mixed zooplankton from the Gulf of Maine by Blumer and Thomas (1965a and b) and by Blumer, Robertson, Gordon and Sass (1969), are shown in Pig. 2. All are olefine and are present in amounts much smaller than those of pristane. Moreover, ufike pristane they do not occur in crude oils. Biochemical inter-relationships between phytol, pristane, phytane and various olefins are shown in Fig. 3. Blumer et al. (1963, 1964) noted that the copepod Rhincalanus ?uIcButus,although similar in feeding habits to Calanus spp., contained only traces of pristane ; and later work (Blumer et al.,1970) showed the main hydrocarbon in this species to be the C,, polyunsaturated olefin HEH. This hydrocarbon did not, however, occur in R.nusutus to the same extent as did pristane in other calanoid copepods. Thus the amounts of HEH in laboratory cultured animals were in the range

312

E. D; S. CORNER

TABLEVII. LEVELSOF PRISTANE IN VARIOUSSPECIES OF ZOOPLANKTON

Pristane content (Yototal (Yo dry wt) lipid)

Species

Group

Stage

Sagitta elegans Verrill Limacina retroversa (Fleming) Conchoecia sp. Paratherniato gaudichaudii (Guerin) Nematoscelis megalops Hansen Meganyctiphanes norvegica (M. Sara) Calanus finmarchicus Gunnerus Calanus finmarchicus Calanus finmarchicus Calanus finmarchieus Calanus finmarchicus Calanus ghcialis Jaschnor Calanus gbcialia Calanus hyperboreus Kr0yer Calanus hgperboreus Calanus hyperboreus Rhincalaniu nasutus Giesbrecht Rhincalanus nasutus Pareuchaeta norvegica (Boeck) Pareuchaela norvegica Pareuchaeta norvegica Pareuchaeto n.orvegica Metridia longa (Lubbock) Metridia lzrcens Boeck Pleuromamma robusta (F. Dahl) Euchirella Tostrata (Claus) Candacia armata Boeck

Chaetognath

ns

0.02

0.05

Pteropod Ostracod

ns ns

0.01 go.01

0.14 0.03

Amphipod

ns

t0.01

0.04

Euphauviid

ns

<0*01

0.09

Euphausiid

ns

g0.01

0.02

Copepod Copepod Copepod Copepod Copepod Copepod Copepod

IV V V Female Male V

0.85 0.73

1.47

0.77

1.45 1-69 1.31

Copepod Copepod Copepod

V

Copepod Copepod

0-46 0-68 0.45 0.47

0.86

V Female

0.92 0.84 0.90

1-62 2.94

Female Female

0.01 0.01

0.03 0.03

0.02 0.03 0.05

0.14 0.19

0.03 0.01

0.08 0.15

g0.01 0.01 I0.02

0.01 0.04 < 0.20

V

IV V

Copepod Copepod Copepod Copepod Copepod Copepod

Female Female Female Female

Copepod Copepod Copepod

V and adult V and female Female

Data from Blumer et al. (1964); ns

= not

stated.

0.01

-

POLLUTION STUDIES WITH MARINE PLANKTON-I

313

0.0006-0.22 pg/copepod and accounted for < 0.007 t o 0.47y0 total lipid : the levels in " wild " animals were greater, varying from 0.061 to 0.46 pg/copepod and 0.28 to 1.2% total lipid. Blumer et a2. (1970) suggested that the levels of HEH in R. nasutus might vary with the amounts in algal foods used by the animals; Pristane(2,6,10,i4-tetramethylpentadecane)detected by Blumer etul (1973)

I

1

] Neophytadiene

Cz0- phytadienes detected by

Blumer and Thomas ( 1 9 6 5 ~ ) isomeric phytadienes

1 -

-

-

c

-

-

CIs-di-and tri-olefins detected by Blumeretul (1969)

c~s-heneicosa-3,6,12,15,18hexaene (HEH) detected by Leeeta/ (1970)

FIG.2. Hydrocarbons detected in zooplankton.

they also pointed out that the species seemed exceptional in being able to accumulate HEH from plant diets. Thus, Eucalanus bungii Giesbrecht, belonging to the same taxonomic family as R. nasutus, contained little or no HEH when reared on algal cultures that provided R. nasutus with the olefin. Likewise, Lee et al. (1970) could not detect HEH in Calanus helgolandicus (Claus) fed on Skeletonema, which contains considerable amounts of HEH (Blumer et al., 1970). Possibly Note.

The top line of Fig. 2 should read Blumer et al. (1963).

314

E. D. 8. (IORNEB

these other species are less able to accumulate HEH from algal diets. On the other hand, compared with R. naswtw, they may be more successful in metabolizing the hydrocarbon (Lee et d., 1970). The levels of hydrocarbons, which are normally low, found in various samples of zooplankton are shown in Table VIII. I n the study by Lee, Nevenzel and Lewis (1974) with Euchaeta juponica Marukawa pristane was usually the major hydrocarbon, PHYTOL

\\I//

Metabolic chanqes hydrocarbons in zooplankton . giving .

Neo -phytadiene

A \

A H .....

.....A

.....

I

Phytenic acid

lsornerises

H

2

0

Di hydrophytol

L+ A .....

.....

(CIS and truns)

..... Norphytene L

I

..... Phytonic acid

Satul tion

1

Isornerises

I

Zooplankton: decarboxylation

I

accounting for 3&50y0 of the total; HEH was also detected in substantial quantities, representing 30-40% of the total in adults and copepodid V and 7% in the eggs; trace amounts of a series of n-alkanes and n-alkenes were also detected, ranging in chain length from C,, to C26. I n the samples examined by Whittle et al. (1974) pristane accounted for 79.7% of the total, most of the remainder being n-alkanes (18-19%) and squalene (0.33%). Pristane was also the main hydrocarbon found in the copepod " slick '' studied by Lee and Williams

1

+d

a

TABLEVIII. HYDROCARBON LEVELSIN ZOOPLANKTON

zz rj

Species Gnathophawk sp. (mysid) Acanthaphyra purpurea Milne Edwards (decapod) Nematobrachion sexspinosis Hansen (euphausiid) A . purpurea (female) A . purpurea (male) Ewhaeta j a p o n k Marukawa Mixed plankton samples from the Clyde Copepod " slick " from N. West Pacific Surface zooplankton from E. Mediterranean Mixed zooplankton from E. Gulf of Mexico (Summer) Mixed zooplankton from E. Gulf of Mexico (Autumn) Mixed zooplankton from E. Gulf of Mexico (Winter)

* Data calculated using wet weight: dry weight ratio of 7-0:l.

Hydrocarbon levels As yo lipid As yo wet weight 1-2 2 3

3 33 0.43 0.36 1-44

0.0148 to 0.0299 0.170 0.0294 0.02 to 0.046 0.038 to 0.052
Reference Morris and Sargent (1973) Morris and Sargent (1973) Morris and Sargent (1973) Morris (1974) Morris (1974) Lee et al. (1974) Whittle et al. (1974) Lee and Williams (1974) Morris (1974) Calder (1976) Calder (1976) Calder (1976)

a

Elm (I)

8

f

2 m

#

k

5 0 x

I

H

316

E. I). 9. OORNER

(1974), accounting for 80% of the total; the remainder was n-alkanes ranging from C,, to C,, with a peak at CZ5. The relatively high levels of hydrocarbons detected by Morris (1974) in surface samples of zooplankton from the eastern Mediterranean probably reflect petroleum pollution in this area. The compounds consisted mainly of n-alkanes in the range C,, to C,, with C,, predominating, these being present in greater amounts than those of pristane. Polyunsaturated C,, and C,, hydrocarbons and squalene were also found. Interestingly, compared with the other samples of zooplankton investigated (see Table VIII), those from the Gulf of Mexico (Calder, 1976), a sea area with a relatively long history of oil exploration, had the lowest levels of hydrocarbons. A further finding of interest was that whereas total lipid levels did not vary much with season, that of the hydrocarbons was much greater in winter than in summer or autumn (Table IX). Although no comparative details are given, neither dissolved hydrocarbons nor those associated with particulate material apparently bore any relation to the hydrocarbons in the zooplankton, which therefore do not appear to have arisen from exogenous sources such as oil pollution. On the other hand, evidence of an association between the levels of dissolved hydrocarbons and those present in plankton from the same sea areas has recently been reported by Whittle, Mackie, Hardy, McIntyre and Blackman (1977). Thus, the average total of n-alkanes in plankton collected near oil refineries was 270 pg/g dry weight compared with 71 in samples taken from the open sea (Celtic Sea) : values for dissolved hydrocarbons collected from similar areas were 4.5 pg/l and 0.57 pg/l respectively (Hardy et aE., 1977: see Table 1). The relative amounts of the individual n-alkanes were also determined, but the data provided no clear indication of whether the compounds were endogenous or had been accumulated from the environment. Further evidence that zooplankton from the Gulf of Mexico (South Texas Outer Continental Shelf) possess a hydrocarbon pattern characteristically biogenic is that of Parker et al. (1976), included in Table IX, who found particularly high levels of C,, n-alkanes and pristane. Parker et al. (1976) noted the marked difference between the hydrocarbon patterns in samples of zooplankton and neuston collected simultaneously from the same sea area : a third of the neuston samples had n-alkane patterns typical of petroleum, which was attributed to the presence of micro-tarballs in the surface. Concerning the possible biological function of the naturally occurring hydrocarbons in zooplankton, Blumer et al. (1964) proposed that pristane might be used by calanoid copepods as a means of achieving

317

POLLUTION STUDIES WITH MARINE PLANKTON-I

TABLE Ix. SEASONAL

CHANGES I N

HYDROCARBON LEVELSIN ZOOPLANKTON Spring

Summer

Autumrz

Winter

(Data for total hydrocarbons : Gulf of Mexico. Summarized from Calder (1976)) Zooplankton biomass (mg dry wt/m3) 91 18 13 Total lipid content (mg/g dry wt) 49.9 37.7 135 212 135 719 Total hydrocarbons (pg/g dry wt) 19.3 2-4 9.4 Total hydrocarbons (pg/m8) (Data for individual compounds (pg/g dry wt) : South Texas Continental Shelf, Gulf of Mexico. Summarized from Parker et al. (1976)) 6.0 0.8

39.6 2.0

2.0 1.2 0.6 3.0 49.1 0.1 1-0

1.8 0.2 8.8 1.1

1.6 0.6 1.3 3.3 17.8

3.2 1-0 18.6 3.6 3.0 2.0 0.7 8.1 73.9

0.05

0.7

1.4

4.7

buoyancy ; and Youngblood, Blumer, Guillard and Fiore (1971) suggested that HEH might influence sex ratio, drawing attention to the correlation between the percentage of males produced and the degree of predominance of HEH in the algal diets used by the younger stages of C. helgolandicus in studies by Paffenhbfer (1970). However, heavier mortality occurred in the experiments in which fewer males were produced and this may have selectively affected the male animals (Paffenhofer, 1970). The influence of environmental factors on the sex ratio of calanoid copepods and the possible importance of hydrocarbons in this context are topics that obviously deserve further study (see Sections VIII and IX).

STUDIESWITH PHYTOPLANKTON IV. TOXICITY An important factor influencing the toxicity of an oil is the size and chemical composition of the water-soluble fraction (WSF), which includes a number of low-boiling aromatic hydrocarbons. Some of these, such as benzene and toluene, are rapidly lost by weathering (Frankenfeld, 1973), but others, notably bi-cyclic aromatic hydrocarbons such as naphthalene and its alkylated derivatives (e.g. 1- and

318

E. D. 9. CORNER

2-methylnaphthalene, dimethylnaphthalenes) are more persistent. Studies by Boylan and Tripp (1971) and by Anderson, Neff, Cox, Tatem and Hightower (1974) have shown that the high proportions of bi-cyclic and tri-cyclic aromatic hydrocarbons in the WSFs of oils TABLEX. HYDROCARBON CONTENTSOF WATER-SOLUBLE FRACTIONS OF FOUR TESTOILS Hydrocarbon colztent (msll) Compound

S. Louisiana Kuwait crude. oil crude oil

No. 2 fuel oil

Bunker 0 residual

oil

Alkanes It- and wo-alkanes, C, to C, n-alkanes, c,, to C,, Cyclopentane and 2-methylpentane Methylcyclopentane Methylcyclohexane

8-94 0.089

10.76 0.004

0*424* 0.047

0.058 0.012

0-380 0-230 0.220

0.590 0.190 0.080

0.020 0.019 0.030

0.005 0.004 0.002

Aromatics Benzene Methylbenzenes Naphthalene Methylnaphthalenes Biphenyl Methylbiphenyls Fluorene Methylfluorenes Dibenzothiophene Phenanthrene Methylphenanthrenes

6-75 6.85 0.12 0.178 0.001 0.002 0.001 0.002 0.001 0.001 0.003

3.36 6.60 0.02 0-051 0.001 0.002

0.550 3.28 0.84 1.09 0.011 0.017 0.009 0.011 0.004 0.010 0.010

Total saturates Total aromatics Total hydrocarbons

9.86 13.91 23.77

0.001

0.002 0.001 0.001 0.002 11.62 10.04 21.66

0-54 5.73 6-27

0.040

0.310 0.210 0.690 0.001 0.002 0.005

0.006 0.001 0.009 0.014 0-081 1.29 1-37

Fractions prepared from 1 pert oil layered on 9 parts 20%, Instant Ocean. * Unresolved GC peaks, probably includes some olefins. Summarized data from Anderson et aZ. (1974).

such as No. 2 fuel oil and Bunker C (Table X) could be responsible for the relatively high toxicities of these oils to marine animals. Crude oils, their total WSFs and individual components of them, have all been used in toxicity studies with phytoplankton.

POLLUTION STUDIES WITH MARINE PLANKTON-I

319

A. Studies using crude oils and their water-solublefractions The first laboratory study of the effects of crude oil on phytoplankton seems to have been that of Galtsoff, Prytherch, Smith and Koehring (1935) who found that a heavy layer of Lake Pelto crude oil over a culture of Nitzschia closterium (Ehrenb.) W. Sm. began to inhibit growth after one week. The WSF of the oil, prepared by dialysis through a collodion membrane, also inhibited growth when used at high concentrations (25 and 50% in sea water) over a period of 13 days.

Days

FIG.4. A. Development of Ditylum brightwellii in sea water containing different concentrations of fuel oil: 1, 0.001 ml/l; 2, 0.01 ml/l; 3, 1.0 ml/l; pecked line = oontrol. B. Development of Meloei~amonilijormia; 1 , O . O O l ml/l; 2,O.l ml/l; 3, 10 ml/l; dashed line = control (After Mironov and Lanskaya, 1966.)

Russian work on the effects of crude oil on many species of marine phytoplankton colleoted from the Black Sea (summarized by Mironov, 1968, 1972) showed that species differed considerably in their sensitivities. The effects, however, appeared to vary with the oil concentration used. For example, Mironov and Lanskaya (1966) found that a level of 0.001 ml/l, over a period of three days, stimulated cell division by Ditylum brightwellii (West) Grun. ex Van Heurck but slightly inhibited that of Melosira moniliformis (0. F. Miill.) Agardh; on the other hand, whereas 1.0 ml/I caused a 100% reduction in cell number over 24 h with 10 mlp used over three days did not significantly affect the original number of cells in a oulture of Melosira (Fig. 4). ~~~~~~~~

320

E. D. 9. UORNER

The oil “concentrations” described in studies such as that of Mironov and Lanskaya (1966) represent oil added to but not necessarily dissolved in the sea water ; in fact, suspensions of oil were used and not solutions. Prouse, Gordon and Keizer (1976) refer to oil as being ‘ I accommodated ” in sea water, earlier work (Gordon, Keizer and Prouse, 1973) having shown that oil agitated with an aqueous phase does not all pass into solution : a large fraction (ca. 90%) is present in particulate form. It is necessary to check the extent to which the amount of oil originally added to sea water might vary during a toxicity experiment. Thus, Gordon and Prouse (1973), studying the effects of three oils (Venezuelan crude, No. 2 fuel oil and No. 6 fuel oil) on the photosynthesis of natural phytoplankton from Bedford Basin, Nova Scotia, measured the amounts of oil (both dissolved and particulate) directly before and after the incubation period. The method used was fluorescence spectroscopy (Keizer and Gordon, 1973) which detects aromatic compounds only, but the results were expressed in units of total oil used as a standard. All three oils inhibited photosynthesis, measured by I4CO, uptake, when present in amounts ranging from 50-300 pg/l, No. 2 fuel oil having a greater effect than the others. However, when lower amounts of oil were used (50 pg/l) Venezuelan crude stimulated photosynthesis. The quantities used in the experiments included the average value of 20 11.811found at a depth of 1 m in Bedford Basin a t the same time (Keizer and Gordon, 1973), although much higher levels could occasionally be observed (e.g. 800 pgll at a depth of 25 cm beneath a 2-day old slick of crude oil). By using quantities of oil that included those normally found in field situations the authors were able to conclude that the 1973 levels of oil contamination in Bedford Basin would have had no serious effect on photosynthesis by the natural phytoplankton community. Studies of the toxicities to phytoplankton of the WSPs of crude oils have been made by Lacaze (1969) who detected a 10% reduction in growth of the diatom Phaeodactylum tricornutum Bohlin in a medium containing water-soluble components of Kuwait crude oil used at a level of 10 ml/L I n addition, Nuzzi (1973) showed that the WSFs of three different oils varied considerably in toxicity to phytoplankton, that of No. 2 fuel oil being much more toxic than that of either No. 6 fuel oil or an outboard-motor oil when tested with either an axenic culture of Phaeodactylum or a natural population of phytoplankton. I n further tests, three algal species showed different susceptibilities to the No. 2 fuel oil, Chlamydomonas sp. being the most resistant and Xkeletonema costatum the least. Certain findings indicate that the effects of petroleum hydrocarbons

POLLUTION STUDIES WITH MARINE PLANKTON-I

321

on phytoplankton vary with season. Thus, Gordon and Prouse (1973) found that the effect of Venezuelan crude oil was much more marked in spring than in autumn; and Fontaine, Lacaze, Le Pemp and Villedon de Nayde (1975) observed that the effects of the WSF of Kuwait crude oil on 14C-uptake by natural phytoplankton populations in the Gulf of St Malo (English Channel) were more marked in summer than in spring. At 12"C, the spring temperature, 14C-uptake increased by over 100% at a hydrocarbon concentration of 15 pg/l; but at 17"C, the summer temperature, it was inhibited by over 90%. Changes in species composition could account for the differences in sensitivity to hydrocarbons with season. Another possibility, mentioned by Fontaine et al. (1975), is that auxins present in crude oil (Gudin and Harada, 1974) might particularly affect spring populations. Temperature effects seem to vary markedly with species, however, for in further experiments by Fontaine et al. (1975), using the single species Phaeodactylum tricornutum, the inhibition of W-uptake at 7-14°C was much greater than that a t 16-25°C. Further studies of the varying degrees of sensitivity to crude oil shown by different phytoplankton species have been made by Pulich, Winters and Van Baalen (1974). Six unialgal species were used: Agmenellum quadruplicatum (Menegh.) BrBb., Nostoc sp, Thalassiosira pseudonana (Hust.) Hasle et Heindal, Dunaliella tertiolecta Butch., Chorella vulgaris var. autotrophica (Shihira et Krauss) Fott et Novakova and ~lenodiniumhallii Freudenthal et Lee (referred t o as Gymnodinium halli). Growth-rate data were expressed in terms of doubling time and any lag in initiation of growth was measured by comparing the times needed by control and oil-treated samples of algal cells to reach the same point on the growth curve. Photosynthesis was measured as oxygen production (Van Baalen, 1968). Two crude oils (Kuwait and Southern Louisiana) and No. 2 fuel oil were used, a WSF of the oil itself and of various distillates formed a t different temperatures being prepared in each case. Differences in sensitivities of algal species were demonstrated by the finding that the growth of Chlorella was severely inhibited by watersoluble components of the low-boiling fractions (195-270°C) : these, however, had little effect on the growth of either Thalassiosira or Agmenellum which were more susceptible to water-soluble extracts of high-boiling fractions (285-385°C). Experiments using No. 2 fuel oil equilibrated with sea water (15 mg total extractables/l) in various dilutions (0.0075-7-5 mg/l) showed that these had no effects on growth measured as mean generation times for the six test species. However, there was an occasional

322

E. D. 9. CORNER

lengthening of the lag phase before growth began, particularly in the experiments with Qlenodinium, Thlassiosira and Agmenellum, the lag phases being substantially increased by exposure to a concentration of 1.5 mg total extractables/l. Studies of the effects of the water-soluble components of a No. 2 fuel oil on photosynthesis (Fig. 5) showed further interesting differences in susceptibility between species : for example, photosynthesis in Thalassiosira was much more readily affected than that in Chlorella, which in turn was more susceptible than that in Agmenellum.

Minutes

FIG.6. Effect of water-soluble fraction of No. 2 fuel oil on photosynthesis by 3 species of marine unicellular algae. A, Agmenellum qwdrmplicatum: pecked line = control containing sea water plus growth medium; continuous line = 60% oil: water v/v (i.e. 1-0ml sea water containing oil solubbs plus 1.0 ml algal suspension). B, Chl'hlorelka autotrophica: pecked line = control; continuous line = 20% oil: water v/v. C, Thalaseiosira p8ewEonana: dashed line = control; continuous line 5 12% oil: water v/v. Algal concentrations for all 3 test species approximately 1 x 107 cellslml growth medium. Temperature, 3OOC. (After Pulich et al., 1974.)

Relating to the work by Pulich et aZ. (1974) involving distillate fractions of oils is that of Parsons, Li and Waters (1975) using three different mixtures of hydrocarbons : aromatics (benzene, toluene, m-xylene, o-xylene and p-cymene), n-alkanes (C12to C16) and n-alkenes (Clo to C14). Laboratory studies were made with natural phytoplankton populations, one dominated by Bkeletonema costatum and the other by Nitzschia sp. Hydrocarbons in low concentrations enhanced photosynthesis by the population dominated by Nitzschia, the effect being greater with aromatic compounds than with either n-alkanes or n-alkenes : thus,

POLLUTION STUDIES WITH MdRINE PLANKTON-I

323

at the 5 pg/l level aromatic hydrocarbons enhanced photosynthesis by 70% whereas the corresponding value with n-alkanes was less than 50% and for n-alkenes less than 40%. These effects diminished as the concentrations of hydrocarbons increased, a particularly rapid fall-off being observed with the aromatic compounds. Different trends were observed, however, using the population dominated by Skeletonema : enhancement of photosynthesis by the aromatic compounds was less than 20% at the 5 pgll level but increased to 60% at 500 pg/l ; low levels of n-alkanes slightly suppressed photosynthesis but higher levels (> 100 pg/1) enhanced it ; suppression of photosynthesis by n-alkenes occurred at all concentrations in the range 5 to 500 pgll, higher amounts causing greater effects. Further studies, using unialgal species, have recently been made by Prouse et al. (1976) who, as in their earlier work with natural populations of phytoplankton, paid particular attention to the need to study the toxic effects of crude oil using concentrations similar to those found in the environment. I n addition they took care t o monitor changes in hydrocarbon composition and level during the experiments, using fluorescence spectroscopy and gas chromatography. During the course of the experiments ( 1 6 1 8 days) they found that the composition of the hydrocarbons " accommodated " in the sea water changed markedly with time, compounds predominant a t the end being, not unexpectedly, the least volatile, most soluble and most resistant to biological alteration : that is, aromatic compounds of medium molecular weight. The presence of algae had a marked effect on the levels of oil in the test media, which fell by over 90% in 12 days. One new feature of the work was that growth data for all five test species ~ D u ~ l i ~ ltertiolecta, la Fmgilaria sp., Monochvysis sp., Skeletonema sp. and Chaetoceros sp.) cultured under axenic conditions were only obtained after the lag phase had finished and the plants were growing exponentially ; another was that the experiments lasted much longer (average 11 days). An initial concentration of 50 pg/l of a No. 2 fuel oil stimulated the growth of Fragilaria and initial levels of 55 and 106 pg/1 of Kuwait crude oil enhanced that of Dunaliella. However, contrary to previous findings (Gordon and Prouse, 1973), no strong inhibitions were observed and any minor ones that occurred in occasional experiments were short-lived (Fig, 6). Furthermore, the experiments did not show any consistent differences in response between the five test species. Consistent with the evidence that petroleum hydrocarbons can stimulate photosynthesis by certain species of phytoplankton are data by Dunstan, Atkinson and Natoli (1975) who measured the growth of

324

E. D. 5. CORNER

4 phylogenetically different marine algae exposed to a wide range (0.1 to 100 mg/l) of concentratous of the volatile, aromatic hydrocarbons benzene, toluene and xylene. The growth rate of Dunaliella tertiolecta was markedly stimulated by all 3 compounds ; smaller effects were observed with Amphidinium carterae Hubert and HymenomonaS cartrterae (Braarud et Paged.) Braarud as Cricosphaera carterae ; no enhancement of growth rate was found with Skeletonema costatum. The

Days

FIG.6. Growth of Dunaliella tertiolecta in the presence of No. 2 fuel oil. Oil concentrations: 50 (initial) falling t o 2 (final)pg/l (open triangles) :380 (initial) falling to 45 (final)pg/l (open circles). Dashed line = control. (After Prouse et al., 1976.)

growth rate of Dunaliella was also stimulated and that of Skeletonema reduced by No. 2 fuel oil, both effects depending upon the presence of the volatile fraction. Winters, O'Donnell, Batterton and Van Baalen (1976) have continued the work of Pulich et al. (1974) with further studies of the effects of WSFs of fuel oils on the growth of individual phytoplankton species. The work was less concerned than that of Prouse et al. (1976) with using oil concentrations close to those found in the environment. Instead, the main emphasis was on analysing the numerous chemical components of the water-soluble fractions of the oils and attempting to identify those that are particularly toxic. The fuel oils used, referred to by refinery location, were Baytown (Texas), Baton Rouge (Louisiana), Billings (Montana) and Luiden (New Jersey): the algal species were

325

POLLUTION STUDIES WITH MARINE PLANKTON-I

Agmenetlum quadruplicatum, Coccochloris elaabeus (Brkb.) Dr. et D., Dunaliella tertiolecta, Chlorella vulgaris var. autotrophica, Cylindrotheca sp. and Amphora sp. About half the water-soluble components of each oil were identified by gas-chromatography and mass spectrometry. Included were compounds such as naphthalene, alkyl-naphthalenes, benzene and alkylbenzenes identified earlier by others (e.g. Boylan and Tripp, 1971): particularly interesting, however, was the detection of phenols, methylanilines (o-, m- and p-toluidine) and indoles, the methyl-, dimethyland trimethyl- derivatives of which were present in relatively high amounts. Phenols accounted for more than half the total identified organic compounds in the WSF of the Baytown fuel oil and were also well represented in that of the Montana fuel oil (Table XI). TABLEXI. MAJORCONSTITUENTS OF WATER-SOLUBLE FRACTIONS OF FUELOILS

Montana (PgP) Total identified organics Methylnaphthalenes Dimethylnaphthalenes Phenols Anilines

8.07 0.53 0.31 2.33 2.57

Baytown New Jersey (Pg/l)

(PgP)

7.90 0-81

5.66 1.30 0.55 1-96 0.27

0.24 4.12 0-72

Baton Rouge 3-52 0.76 0.41 1-08 <0.02

WSFs prepared by adding 1 part oil to 8 pads culture medium. Summarized data from Winters et al. (1976).

Growth data indicated the high toxicities to Agmenellum and Coccochloris of the Baytown and Montana fuel oils; in addition, the water-soluble fraction of New Jersey fuel oil lengthened the lag phase with'each test species although the final growth rates attained by Dunatietla, Chtorella, Cylindrotheca and Amphora were close to those of the controls. Compared with the other three fuel oils, that from Baton Rouge, which had relatively low concentrations of phenols and anilines, was much less toxic. Individual compounds identified in the water-soluble fractions of the oils were tested for their effects on algal growth using the " aIgal lawn " technique (Pulich et al., 1974) in which measurements were made of the zone of inhibition around a disc treated with the test material set in agar gel containing the algal cells. The main finding was the

326

E. D.

9. UORNER

relatively high toxicities of the substituted anilines, particularly p-toluidine, when used against the coccoid blue-green alga Agmenellum (see Table XII). Additional observations showed that other species of blue-green algae are also affected by p-toluidine which may well possess a selective toxicity for these organisms. TABLEXII. EFFECTS OF HYDROCARBONS AND RELATED COMPOUNDS GROWTHOF Agmenellum qzcadruplicatum MEASUREDBY THE " ALGAL LAWN" TECHNIQUE

ON THE

Compound P- Toluidine

2, 4, 6-Trimethylphenol

Dimethylquinoline 2, 6-Dimethylphenol Dimethylnaphthalene Biphenyl 1, 2, 4-Trimethylbenzene p-Cresol Di-iso-propylbenzene Methylnaphthalene Naphthalene Phenanthrene Tri-ethylbenzene 1, 2, 4, Ei-Tetramethylbenzene Fluorent,

Amount used in teat (mg) 0.5 0.1 0.01 36* 36 36 10 36 36 36 36 20-22 36 36 36 6 3 1 0

2

1

36 36 32 23 21 16 16 12 10 2 4 2 0

36 27 11 7-8 4 10 8 6-7 4 0 4 0 0 0

0

* Numbers are zones of inhibition in mm measured from edge of filter pad.

Complete

kill of test organism gives 36 mm zone of inhibition. Where 0 - , m- and p-derivatives tested isomer with highest toxioity taken. Data summarized from Pulich et al. (1974) and Winters et al. (1976).

Recent work by Winters, Batterton and Van Baalen (1977) has shown that the hydrocarbon derivative phenalen-1-one (perinaphthenone) is present in the water-soluble fraction of No. 2 fuel oil and is almost as toxic to green algae as p-toluidine is to blue-green algae. Further evidence of differences in response between algal species was provided by the finding that toluidines were present in water-soluble fractions of Baytown and Montana fuel oils in quantities sufficient to kill Agmenellum and Coccochloris, whereas the benthic diatoms Cylindrotheca and Amphora were not much affected by the watersoluble components of any of the oils tested.

POLLUTION STUDIES WITH MARINE PLANKTON-I

327

B . Studies using naphthalene Work already described has included studies with specific distillate fractions of oils (Pulich et al., 1974) and individual constituents of the water-soluble fractions (Pulich et al., 1974; Winters et al., 1976, 1977). I n addition, detailed studies have been made using the bi-cyclicaromatic compound naphthalene, which has been shown to be present in considerable amounts in aqueous extracts of oils (Boylan and Tripp, 1971) and is fairly toxic to a wide range of marine organisms. Thus, as shown in Table XII, in terms of its toxicity to the micro-alga Agmenellum, naphthalene is more active than either phenanthrene or fluorene, but less so than its own alkylated derivatives (methyl- and dimethylnaphthdene) or those of phenol and aniline. The main difficulty in working with naphthalene is loss of the hydrocarbon by evaporation. For example, Vandermuelen and Ahern (1976), studying its effects on the growth of FTagilaria sp., found that the concentration in the culture medium fell by over 90% in 18 days. This made interpretation of the findings difficult. Thus, a 50% saturated solution of naphthalene in sea water caused a marked inhibition of growth over 14 days, after which the cell population showed a rapid increase: the cells might have recovered through overcoming the inhibitory effects of naphthalene, but there was also the possibility that such recovery was related to loss of the hydrocarbon from the culture medium. Further studies on the toxicity of naphthalene to algae were those of Soto, Hellebust, Hutchinson and Sawa (1975~)and Soto, Hellebust and Hutchinson (1975a). This work, having been done with the freshwater species Chlamydomows angulosa Dill, lies outside the scope of the present review but deserves brief mention because of its relevance to the important question whether aromatic hydrocarbons can be metabolized by algae. The data from one of several experiments are shown in Fig. 7. Cells treated with 14C-l-naphthalene for several days and then transferred to fresh medium containing no hydrocarbon lost radioactivity rapidly over two days during which there was no increase in the number of cells. Such losses could have been the result of passive diffusion out of the cells, or metabolism, or both. On the other hand, cells pre-treated with W-1-naphthalene and then left in the medium containing the hydrocarbon did not lose radioactivity until they began to divide, the apparent coincidence in time between resumption of cell division and loss of radioactivity per cell suggesting that dilution by cell division alone was probably the main factor affecting the level

328

E. D. 9. CORNER

of naphthalene in the cells. This observation, together with the further finding that no non-volatile radioactive compounds were detected in the medium, was taken to imply that the plant cells were unable to metabolize the hydrocarbon. The question whether aromatic hydrocarbons remain in the plant cells unchanged or in the form of metabolites is important in the context of the transfer of the compounds to higher trophic levels : it therefore deserves detailed investigation using various unialgal species, both freshwater and marine, as well as with natural populations of

I

Days

FIG.7. Uptake and release of naphthalene by C ~ Z a r n ~ d o angulosa, rn~~ together with growth data. Cells initially incubated in naphthaIene-saturated growth medium in e closed system for 6 days (A) or 7 days (B) and then either washed and transferred t o fresh medium (open circles) or left in the naphthalene-saturated system (open squares). (After Soto et aZ., 1975a.)

phytoplankton. Many marine micro-organisms such as bacteria and fungi have the ability to degrade petroleum hydrocarbons. So far, however, the only degradation study with an alga seems to be that of Walker, Colwell and Petrakis (1975) who showed that the achlorophyllous species Prototheca zopjii Kriiger can degrade motor oil and South Louisiana crude oil, in which respect the plant was as efficient as various species of bacteria but not so efficient as the fungi. The effect of naphthalene on photosynthesis by three marine

POLLUTION STUDIES WITH MARINE PLANKTON-I

329

unicellular algae has been recently studied by Vandermuelen and Ahern (1976). Suppression of photosynthesis was found to be concentration-dependent : for example, with Pavlova Zzctheri (Droop) Green as Monochrysis 1zLtheri Droop, a concentration of 1 mg naphthalene/l reduced photosynthesis by 40% whereas a concentration of 7 mgfl reduced it by 90%. Cells transferred to uncontaminated medium after pre-incubation for 4 h in a naphthalene concentration of 5 mg/l quickly recovered the ability to photosynthesize, the normal rate being restored within 5 h. Treatment with the hydrocarbon did not reduce levels of chlorophyll a in the cells, but did cause a marked decrease in ATP levels both in the light and the dark. Vandermuelen and Ahern (1976) take this to mean that suppression of photosynthesis could have arisen from a blockage of oxidative phosphorylation. However, although this could well be the case with naphthalene, whole oils seem to have a direct effect on chlorophyll a levels (Mills and Ray: unpublished observations quoted by Anderson, 1975).

V. MECHANISMS OF PRYTOTOXICITY Although numerous studies have been made of the effects of petroleum hydrocarbons on growth and photosynthesis by marine phytoplankton, little work has been done concerning the mechanisms by which the compounds exert their toxic action. Certain conclusions regarding the effects of hydrocarbons on terrestrial plants, freshwater micro-algae and marine multicellular algae (reviewed by O’Brien and Dixon, 1976) do, however, have sufficient relevance to justify inclusion in the following brief account. Van Overbeek and Blondeau (1954) suggested that hydrocarbon molecules disrupt the plasma membrane by displacing those of other lipid compounds, so affecting its semi-permeability ; in addition, that the inhibition of photosynthesis could result from hydrocarbons dissolving in the lipid phase of the grana of chloroplasts and increasing the distance between individual chlorophyll molecules. Baker (1970) has proposed that a similar disruption could occur in mitochondria1 membranes with inhibition of the tricarboxylic acid cycle and oxidative phosphorylation, as noted by Vandermuelen and Ahern (1976) in their studies of naphthalene toxicity to Monochrysis lutheri. Distortion of the lipid in cell membranes by kerosene, with subsequent penetration by toxic agents, was found with different species of marine red algae by Boney and Corner (1959). The importance of physical factors was emphasized by the work of Currier (1951) who found that the toxicities of benzene, toluene,

330

E. D. S. UORNER

xylene and trimethyl-benzene in aqueous solution were inversely related to solubility. Data for the partition coefficients of these compounds between water and paraffin oil indicated that penetration into plant cells would increase with the number of methyl substituents in the benzene ring. Kauss, Hutchinson, Soto, Hellebust and Griffiths (1973) found that toxicity to Chlorella vulgaris increased along the series benzene, toluene, xylene and naphthalene, with water-solubilities increasing in the reverse order. However, it should be noted from the work of Pulich et al. (1974) and Winters et al. (1976), summarized in Table XII, that increasing the number of substituent methyl groups in benzene from 3 to 4 causes a marked reduction in toxicity to the micro-alga Agmenellum quadrqlicatum. Stimulation of both growth and photosynthesis by low concentrations of oil have been noted by various workers using unialgal cultures (Galtsoff et al., 1936; Xronov and Lanskaya, 1966; Kauss and Hutchinson, 1975; Soto et al., 1975a) and natural populations of phytoplankton (Gordon and Prouse, 1973; Parsons et al., 1975). It has been suggested that such stimulation may result from the oil components being used as metabolic substrates by the plant cells (Soto et al., 1976a); in addition, that it could be caused by the presence in oil of growth regulating compounds (Gordon and Prouse, 1973). Growth stimulation induced by contact with individual PNAH, including carcinogens found in fossil fuels, has been found with sporelings of multicellular red algae (Boney and Corner, 1962 ; Boney, 1974): however, the effects of such compounds on growth or photosynthesis by unicellular algae seems not to have been studied. Another area of investigation has been concerned with the effects of oil components on the chemical composition of marine algae. Thus, Soto, Hellebust and Hutchinson (1975b) studied the effect of naphthalene on the levels of pigments, lipid, protein, carbohydrate and total carbon in the freshwater alga Chlamydomonas angulosa and found that high concentrations of the hydrocarbon decreased cellular protein by 34% in seven days. This loss, however, was nearly all recovered within one day after the cells had been transferred to fresh medium. Changes in lipid levels followed the reverse pattern, more than doubling during the seven-day exposure to naphthalene, but falling by roughly the same amount when the hydrocarbon was removed from the growth medium. Carbohydrate also showed a marked increase during naphthalene treatment, this change probably being associated with thickening of the cell walls: after transfer of the culture to fresh medium, carbohydrate levels fell ~ E Jthe cell walls regained their normal thickness. The observation that a petroleum hydrocarbon may induce an

POLLUTION STUDIES WITH MARINE PLANKTON-I

331

increased conversion of protein into lipid in plant cells has implications in terms of primary and secondary production that deserve more detailed study, particularly with marine species : it is significant, however, that even when high concentrations of the hydrocarbon are used the effects on plant cells are reversible. The studies of Soto et ab. (1975b) were carried out with a single hydrocarbon. However, Davavin, Mironov and Tsimbal (1975) investigated the effects of whole crude oil, using emulsified suspensions in sea water covering the range 0-1-10 ml/l, and found that it inhibited the biosynthesis and modified the polymerization of DNA and RNA in multicellular algae from the Black Sea. No similar study with unicellular algae seems to have been made.

VI. CONTROLLEDEGO-SYSTEM EXPERIMENTS I n the previous sections most of the experimental work described has been carried out in the laboratory with a restricted number of constituents of the marine eco-system. However, because of interactions between species in nature it is necessary t o discover how whole eco-systems react to pollutants. A recent important development has therefore been to carry out pollution studies using large enclosures containing whole eco-systems set up in inshore areas and subject to natural light and temperature conditions (Takahashi, Thomas, Siebert, Beers, Koeller .and Parsons, 1975). Some of these enclosures are polluted with substances such as crude oil, or mixtures of hydrocarbons, or heavy metals: others serve as controls. Physical, chemical and biological variables are monitored during a preliminary period of stabilization, after which the pollutants are added and their long-term effects on the system followed, usually over several weeks. This type of approach was first used by Lacaze (1974) in studying the effect of Kuwait crude oil on primary production in an experimental eco-system set up in the Rance Estuary (North French coast). Each enclosure, made of rilsan (impermeable to hydrocarbons), contained 560 1 of sea water from which larger zooplankton had been excluded by filtration. I n the experiment using Kuwait crude oil 100 ml was added to the enclosure, giving a suspension equivalent to 180 mg/l. Over a period of four weeks during September and October the temperature fell from 17 to 14OC in the open water and primary production steadily decreased : in the control enclosures, however, it fell much more rapidly during the first week and then stabilized at a, level much lower than that of the open water, probably because inside the enclosures the nutrients could not be renewed. Primary production

332

E. D. 9. CORNER

in the samples treated with crude oil underwent an immediate reduction of about 50% during the first day after the oil was added, but by the third day it had regained the same level as that in the controls (see Fig. 8), the rapid initial drop being caused by the presence in the oil of toxic volatile fractions that were quickly lost. Over the next four days primary production in the oil-treated sample fell far more rapidly than that in the controls and stabilized a t a substantially lower level throughout the remainder of the experiment. Clearly the oil caused a

September

October

FIG.8. Primary production in an eco-system polluted by crude oil (open circles, continuous line) compared with that in an untreated system (sled circles, dashed line). Oil added on 25 September. (After Lacaze, 1974.)

significant inhibition of primary production by the natural phytoplankton population : however, the effects could have been unnaturally enhanced if the plants were, in fact, in a state of nutrient deficiency. A further complication in this and other long-term studies using whole marine eco-systems exposed to natural conditions of light and temperature is that chemical and photo-chemical oxidations of crude oil components could proceed rapidly a t the surface of the sea water and lead to the formation of compounds such as hydroperoxides which have considerable biological activity (Burwoodand Speers, 1974 ;Frankenfeld, 1973; Larson, Blankenship and Hunt, 1976). Lacaze and Villedon de Naide (1976) examined this possibility using P h u e o ~ ~ ~ y l u m tricornutum as the test organism and Kuwait crude oil. Compared with oil suspensions kept in the dark, those exposed to 10 000 lux for

POLLUTION STUDIES WITH MARINE PLANKTON-I

333

40 h or more under fluorescent light had more than twice the toxicity as measured in terms of inhibition of photosynthesis. A controlled eco-system experiment of a more extensive kind was that of Lee,' Takaha.shi, Beers, Thomas, Seibert, Koeller and Green (1977). I n this study the WSF of No. 2 fuel oil was added to an enclosure of 60000 1 capacity containing sea water from Saanich Inlet (British Columbia), giving an initial hydrocarbon concentration of 40 pgll. The levels of individual compounds were monitored at different depths throughout the duration of the experiment (19 days) and showed a substantial decrease : e.g. those of naphthalene, methylnaphthalenes and dimethylnaphthalenes combined amounted to 12 pg/l the first day after the WSF was added to the enclosure, but after 3 days had fallen to 5 pgll and by day 16 were below detectable limits. Microbial degradation and metabolism by zooplankton both contributed to the losses of these hydrocarbons. So did removal by sinking particles, concentrations of hydrocarbons increasing in the sediment from the oil-treated enclosure, which included both phytoplankton and faecal pellets. When the addition of the WSF took place the standing stock of diatoms in the control enclosure dominated by Cerataulina bergonii (Perag.) Schiitt, now Cerataulina pelagica (Cleve) Hendey, was much higher than that in the oil-treated sample (Fig. 9a). This large diatom population in the control fell sharply during the next four days, being replaced by a micro-flagellate bloom (Fig. 9b) : it then rose again to a peak after ten days. I n the treated sample the relatively small diatom standing stock diminished still further after the WSF was added and did not recover : however, micro-flagellates showed a sharp increase (Fig. 9b), though not so great as that in the control enclosure; the population at first being dominated by Chrysochromulina kappa Parke et Manton and later by Ochromonas sp. Associated with the micro-flagellate levels in the treated sample were sharp increases and reductions in the micro-zooplankton populations, particularly tintinnids such as Helicostomella subulata (Ehrenberg) (Fig. 9c). However, this close association was not observed in the control enclosure, presumably because diatoms were also available as a food for the herbivores. The dominant zooplankton species was Pseudocalanus minutus (Kroyer); other copepod species were also present and, to a lesser extent, larvaceans, ctenophores and medusae. No major differences were observed between the zooplankton standing stocks of the treated and control enclosures, presumably because even the initial concentration of hydrocarbons used (40 pg/l) was far less than the LC,, values

I

l

2

l

l

4

l

l

6

l

l

8

l

10

l

l

12

l

l

14

l

l

16

l

l

~

~

18

Days

FIQ.9. Effects of a water-soluble fraction of No. 2 fuel oil on the production of diatom (a),micro-flagellates ( b ) , and tintinnids ( c ) in a CEPEX study. Control enclosure; filled circles, continuous line. Enclosure to which WSF added on day 6; filled triangles, dashed line. (After Lee et al., 1977.)

POLLUTION STUDIES WITH MARINE PLANKTON-I

335

(590-1 350 pg/l) found for copepods in laboratory bioassays (see Table

XV). There was, however, some indication of a slower rate of growth by Pseudocalanus in the enclosure treated with hydrocarbons. As Fisher and Wurster (1974) have emphasized, aquatic communities consisting of herbivores selectively feeding on plants are so interlinked that toxic compounds directly affecting one component of the community can indirectly alter the species composition of the other. The controlled eco-system experiment carried out by Lee et al. (1977) provides a good example of how pollutants causing changes in the composition of a phytoplankton population can lead to alterations in the structure of the micro-zooplankton community representing the next trophic level.

VII. FATEOF HYDROCARBONS IN ZOOPLANKTON A. Uptake and release Mention has been made earlier of the finding by Blumer et al. (1964) that the biogenic hydrocarbon pristane is not metabolized by C. hyperboreus. More recent studies have been mainly concerned with the fate in zooplankton of hydrocarbons, such as PNAH, which are found in crude oil and include the carcinogen B P (although the amounts of this compound are small, 0-029-44 mg/kg with an average of 2.0: Pancirov and Brown, 1975).

Days

FIG.10. Net uptake and release of radioactivity, expressed as equivalents of benzo[ulpyrene, by Culunus plumchrua. Filled circles, copepods exposed t o 1.0 p g hydrocarbon/800 ml; open triangles, copepods exposed to this same concentration for three days and then transferred to clean sea water. (After Lee, 1976.)

W W Q,

TABLEXIII. RETENTION OF RADIOACTFJITY BY ZOOPLANKTON EXPOSED TO LABELLED HYDROCARBONS

Test species Calanus plumchrus Calanus plumchrua Calanus plumchrus Calanus helgolandicus Calanus helgolandicus Calanus helgolandicus Euchaeta japonica Euchaeta japonica Parathemisto paci$ca Cyphocaris challengeri Stebbing

Hydrocarbon

BP 20-MC 1-OD

BP BP BP 20-MC N

BP BP

Conc’n (P9POO ml)

Exposure period (days)

Depuration period (days)

yo Radioactivity

1.0 0.2 5.0 1.0 1.0 1.0 0.2 80 15 15

3 1 4 2 4 4 1 4 2 2

17 8 3 9 7 28 8 8 14 6

0-31 8.0

retained

40.0 2.0 0.50 0.23 7.7 2.5 0.28 6.7

BP, 8H-benzo[a]pyrene;20-MC, 3H-20-methylcholanthrene;1-OD,14C-l-octadecane;N, 14C-l-naphthalene.Data from Lee (1975).

Y

U

P Q

0

s

POLLUTION STUDIES WITH MARINE PLANKTON-I

337

The first of these studies was by Lee (1975) who examined the net uptake and release of various hydrocarbons by several groups of zooplankton animals collected off the coasts of California, British Columbia and in the Arctic. Copepods were mainly used-although a few observations were also made with euphausiids, amphipods, crab zoeae, ctenophores and jellyfish-and the hydrocarbons were 14C-lnaphthalene, 14C-BP, 3H-BP, 3H-20-methylcholanthrene and 14C-loctadecane. Typical data are shown in Fig. 10 for the species Calanus plumchrus Marukawa exposed to 3H-BP in sea water at 1.25 pgll, together with 50 pgll of water-soluble hydrocarbons from No. 2 fuel oil. During the first three days there was an approximately linear increase in the net uptake of hydrocarbon, but no further increase was detected when exposure was continued for another seven days. Animals transferred after three days to clean sea water (containing no added hydrocarbons) lost radioactivity gradually over a further 17-day period, at the end of which a residue equivalent to 0.31% of that originally accumulated still remained in the animals. I n studies with various species of copepod different times of exposure and depuration were used, but in every case a small residue of the original level of radioactivity remained in the animals after the depuration period (Table XIII). Most notable among these experiments was that with Calanus hyperboreus in which 0.23% of the original level of radioactivity was still present in the animals after a period of depuration lasting 28 days. Studies by Corner, Harris, Kilvington and O’Hara (1976b), using the copepod Calanus helgolandicus and the bi-cyclic aromatic hydrocarbon naphthalene, showed that the net uptake of 14C-l-naphthalene from solution in sea water varied with the concentration of hydrocarbon used (Fig. 11). Adult female Calanus showed a daily net uptake of the hydrocarbon from sea water containing very low levels : e.g. 17.8 pglanimal from a sea water concentration of only 0-5 pg/l; this level being an order of magnitude lower than that of 4.8 pg/l calculated from the data of Barbier et al. (1973) for total dissolved bi-cyclic aromatic hydrocarbons in a Channel harbour area (see p. 303). Using an apparatus designed to provide Calanus with a known ration of Biddulphia sinensis Grev. (Corner, Head and Kilvington, 1972),14C-1-naphthaleneincorporatedin the algal cells was administered to the animals and the subsequent rate of depuration compared with that observed using animals that had accumulated a similar level of hydrocarbon from solution alone. It was found that when the hydrocarbon was taken up by way of the food the subsequent rate of depuration was notably slower (Fig. 12). A.I.B.-IS

14

t I

I

I 1 1 o . * n 1

lo-'

1

I

#1,,,.1

'

100

I I I I 1 J

10'

I

'

,111111

'

' J * ~ ~ d

lo3

I02

Naphthalene concentration in sea water (&I)

Fra. 11. Net uptake of radioactivity, expressed as equivalents of naphthalene, in 24 h by adult female Calanw, helgolandicw, exposed t o various concentrations of 1%-I-

+

naphthalene in sea water. Relationship defined by y = 1 . 0 6 6 ~ 1.67; correlation coefficient = 0.990. (From Corner el al., 197Gb with permission of the Council of the Marine Biological Association.)

Doys

Fra. 12. Release of radioactivity by adult female Calanw, helgolalzdicw, that had accumulated 14C-1-naphthalene either from a sea-water solution (filled circles) or from a diet of Biddulphia cells (open circles). Levels expressed as percentages of the radioactivity originally present in the animals. (From Corner el al. 1976b with permission of the Council of the Marine Biological Association.)

POLLUTION STUDIES WITH MARINE PLANKTON-I

339

In a further study (Harris, Berdugo, Corner, Kilvington and O’Hara, 1977a) the daiIy net uptakes of 14C-1-naphthalene were measured using seven species of copepod representing oceanic, neritic and estuarine forms. The levels of hydrocarbon used, which ranged from 0.2 to 1000 pg/l, included those for bi-cyclic aromatic hydrocarbons present in the sea under a wide variety of conditions (see Section 11). Combined data from experiments with all the test species showed that total lipid content was a good indicator of the net uptake by copepods of an aromatic hydrocarbon such as naphthalene from solution in sea water during short-term exposures, the regression equation being log y = 0.974 log x +0.61, with r = 0.98 and n = 153, y being pg hydrocarbon/pg copepod body lipid, x the concentration of hydrocarbon in sea water in pg/l, r the correlation coefficient and n the number of determinations. Harris et al. (1977a) confirmed an earlier observation by Lee (1975) that surface adsorption of the hydrocarbon was only a minor factor influencing its net uptake by the animals. They also showed, however, that temperature and degree of starvation were of major importance and that both were negatively correlated with net uptake. Further experiments by Harris et al. (1977a) verified earlier findings by Lee (1975) that small amounts of hydrocarbons accumulated by copepods can still be detected in the animals after prolonged periods of depuration. I n particular they showed that when nauplius I of the estuarine copepod Eurytemora afinis Poppe were immersed in sea water solutions of 14C-l-naphthalene for 24 h, transferred to fresh sea water and reared in the laboratory to adults over a period of 34 days, radioactivity accounting for 10% of the original amount in the nauplius could still be detected in the adults. Such persistence of small quantities of aromatic hydrocarbons, or their derivatives, in zooplankton over long periods implies that although natural processes such as volatilization, photo-oxidation and microbial breakdown of these compounds may occur soon after an oil spill, several weeks later the transfer of an aromatic hydrocarbon like naphthalene from zooplankton to a higher trophic level, such as fish, could still be taking place. B. Quantitative importance of the dietary pathway Corner et al. (1976b) found that, in terms of providing the same level of radioactivity in C. helgolandicw, the quantity of 14C-l-naphthalene needed in solution was much greater than that required as particulate food and concluded that the dietary pathway of uptake

340

1.D. 5. CORNER

was more important quantitatively. Harris et al. (1977a) obtained more direct and detailed evidence of this in 24 h experiments in which levels of radioactivity were measured in animals that simultaneously accumulated lac-1-naphthalene from solution alone and from solution supplemented by a known quantity present in algal food. Compared with the amount of labelled hydrocarbon represented by the suspension of algal cells, the amount in solution needed to provide the same increase in radioactivity in the copepods was three orders of magnitude greater in experiments with females and even two orders of magnitude greater in those with male animals that capture a relatively small rakion. Further work showed that the quantitative importance of the dietary route was not affected by the levels of hydrocarbon in the sea water or present as food : it did however depend upon the level of food available, greatly increasing at lower cell concentrations. Earlier it was noted (Section 11) that in some sea areas the amounts of hydrocarbon “dissolved” in sea water are greater than those present in particulate form. What is not known from such studies is the extent to which the particulate material could be used as a food by zooplankton. Assuming, however, that only a small fraction was present as phytoplankton that could be captured by these animals, this, compared with the much higher levels of hydrocarbon in solution in sea water, could still be a more important source of a compound such as naphthalene in herbivorous copepods.

C. Long-term exposure experiments The work by Corner et al. (1976b) and Harris et al. (1977a) dealt only with short-term exposure of zooplankton to aromatic hydrocarbons, such as might occur immediately after an oil spill. However, it is also important to know what happens when animals are subjected to long-term exposure to low concentrations of these compounds, a condition characteristic of sea areas subjected to regular small inputs of industrial effluents or natural oil seeps. The amounts of hydrocarbons accumulated by zooplankton after long-term exposure to these compounds is of particular interest because these quantities could be critical in producing sub-acute effects: they are also the amounts likely to be transferred to higher trophic levels. So far, only two laboratory studies of this problem have been made: one by Lee (1976), already described, and a more recent investigation by Harris, Berdugo, O’Hara and Corner (1977b). I n the latter study adult female Calanus Fvelgolandicus and Eurytemora afinis were exposed to 14C-1-naphthaleneover periods of 10-15 days,

341

POLLUTION STUDIES WITH MARINE PLANKTON-I

the animals being maintained in the laboratory on algal diets and the hydrocarbon therefore being taken up from solution and from the food. Concerning the transfer of hydrocarbons to higher trophic levels, Harris and co-workers found that after a 10-day exposure to 14C-1naphthalene present at a low concentration (I ,ug/l) a much higher level of radioactivity was accumulated per unit body weight by E . afinis than by C. helgolandicus (Fig. 13) ; and using feeding data for the herring (Blaxter and Holliday, 1958) they calculated that the weekly intake of hydrocarbon by a young fish would be 50 times greater if it fed on E. affinis.

2

4

6

Days

c

FIQ.13. Net uptake of radioactivity, expressed as naphthalene equivalents, by Calanue helgolnndicus (filled circles), and Eurytemora aflnis (opencircles), in terms of dry body weight. (After Harris et al., 1977b.)

As found by Lee (1975) the amounts of I4C-l-naphthalene accumulated by both species used in the study by Harris et al. (197713) reached a maximum after multiple-day exposure; in addition most, but not all, of the radioactivity accumulated by the animals during prolonged exposure was rapidly lost after they were transferred to fresh sea water. The maximum level of radioactivity reached during long-term exposure to low levels of I4C-l-naphthalene probably represents a steady-state level in the animals, with hydrocarbon uptake balanced by hydrocarbon release. The higher levels of radioactivity found in animals exposed t o higher concentrations of 14C-1-naphthalene also showed

342

E. D. 9. CORNER

signs of eventually reaching a maximum, however (Fig. 14), and Harris et al. (1977b) suggested that even under conditions where large quantities of hydrocarbon were entering the animals, enzymes involved in the metabolism of hydrocarbons could still be induced or activated sufficiently to restore the balance between uptake and metabolic loss. Evidence for the metabolism of hydrocarbons by zooplankton is considered in the next section. A

A

I

/-

L /%-8

0

5

10

Days

FIG.14. Net uptake of radioactivity, expressed as naphthalene equivalents, b y Eurytemora afinis (A) and Calanus helgolandicus (B) after multiple-day exposure to hydrocarbon concentrations of 0.2 pg/l (filled circles), 1.0 pg/l (open circles), 10 pg/l (open triangles),50 pg/l (filledsquares),177 pg/l (open squares)and 992 pg/l (filledtriangles). (After Harris el al., 1977b.)

D. Metabolism There is evidence from both in vivo and in vitro studies that a mechanism for hydroxylating certain aromatic hydrocarbons is possessed by several species of marine fish (Lee, Sauerheber and Dobbs, 1972b ; Payne, 1976 ; Roubal, Collier and Malins, 1977b ; Stegeman and Sabo, 1976) and crustacean (Corner, Kilvington and O'Hara, 1973; Cox, Anderson and Parker, 1975; Burns, 1976; Lee, Ryan and Neuhauser, 1976; Singer and Lee, 1977). On the other hand, at least one group of marine invertebrates, the bivalve molluscs, do not seem able to metabolize hydrocarbons (Carlson, 1972 ; Lee, Sauerheber and Benson, 1972a)but release them from the tissues unchanged (Stegeman and Teal, 1973; Neff and Anderson, 1975).

POLLUTION STUDIES WITH MARTNE PLANKTON-I

343

Although only a few studies have so far been made of hydrocarbon metabolism in zooplankton, there is evidence that certain groups of these animals are able to metabolize several types of hydrocarbon. Thus Lee (1975) found that all the micro-crustaceans used in his study with zooplankton-including copepods, amphipods, crab zoeae and euphausiids-could metabolize naphthalene, BP, 20-methylcholanthrene and octadecane. Of the different species tested the amphipod Parathemisto paci$ca Stebbing showed the most rapid degradation of ingested hydrocarbons, over 50% of each of the four compounds studied being metabolized in 24 h. The main metabolites were hydroxylated derivatives of the hydrocarbons, but more polar compounds were also tentatively identified (e.g. octadecanoic acid as a metabolite of octadecane). It is worth noting, however, that not all the species tested possessed the ability to metabolize hydrocarbons : BP was released unchanged by the ctenophore Pleurobrachia pileus (0.F. Miiller) and by an unidentified species of jellyfish. I n his experiments with 3H-BP Lee (1975) showed that Calanus plumchrus which had accumulated this hydrocarbon during prolonged exposure retained metabolites in its tissues for several days; and in similar experiments with Euchaeta japonica, in which 14C-1-naphthalene was used, 88% of the radioactivity retained by the animal after 24 h was still in the form of metabolites. A study of 14C-lnaphthalene metabolism in Calanus helgolandicus by Corner et al. (197613) added to the work of Lee (1975)in two ways : first, by providing evidence for metabolism using animals that had accumulated the hydrocarbon through the diet; secondly, by taking special care to exclude the effects of bacteria, which are known to degrade PNAH (Gibson, 1976; Lee and Ryan, 1976; Harris et al., 1977b),by incorporating the hydrocarbon in autoclaved nauplii of the barnacle Elminius modestus Darwin used as a sterile diet. The ration of autoclaved nauplii captured by the copepods was much smaller than those observed using untreated na,uplii (Corner, Head, Kilvington and Pennycuick, 1976c) but sufficient were taken to ensure a measurable level of radioactivity in the animals after 24 h. The animals were then given a depuration period of 24 h in fresh sea water, after which 8 6 1 0 0 % of the radioactivity retained in them could still be accounted for as naphthalene, whereas only 25-38% of that excreted into the surrounding sea water during this same period of depuration was present as the unchanged hydrocarbon. Thus, although both studies showed that aromatic hydrocarbons were metabolized by copepods, Lee (1975) found most of the metabolites to be retained by the animals, and therefore available for transfer to a higher trophic level, but Corner et al. (1976b)

344

E. D. 9. CORNER

observed that the major fraction was rapidly excreted. To explain these different findings Corner et al. (1976b) suggested that as Lee’s (1975) animals were exposed to the hydrocarbons for a much longer

n

1

IOOOq

-

Hours

WIG. 15. Accumulation (A) and depuration (B) of naphthaleilo and its metabolites (exproused as naphthol equivalents) by Stage V Pandalua platyceroa exposed to 14C-l-naphthaleneat a concentrationin sea water of 8-12 pg/l. (After Varanasi and Malins, 1977.)

period before depuration began this might favour the retention of metabolites in the tissues. Subsequent work, using multiple-day exposure periods (Harris et al., 1977b) confirmed this view in that 66--77y0 of the radioactivity detected in the tissues of C. helgolandicw

POLLUTION STUDIES WITH MARME PLANKTON-I

345

exposed to 14C-1-naphthalene over periods of 4-6 days could no longer be accounted for as the unchanged hydrocarbon. Attention is drawn later (Section VIII) to the extreme sensitivity of larvae of the spot shrimp Pandulus phtyMr08 Brandt and the Dungeness crab Cancer rnagieter Dana to the hydrocarbon naphthalene (Sanborn and Malins, 1977). Because of the likelihood that hydrocarbons are complexed with organic macro-molecules (e.g. proteins) in the sea, these workers used naphthalene both in the free state and as a complex with bovine serum albumin (BSA). Experiments with Stage V spot shrimp showed that the hydrocarbon was accumulated much more rapidly when used in the free state, but that naphthalene metabolites (hydroxylated derivatives measured as naphthol equivalents) accumulated in the animal whether the hydrocarbon was taken up in the free state or as the complexed form. The metabolites were formed fairly slowly, about 4-50/, of the radioactivity being present in forms other than naphthalene after 10 h (Fig. 16A). Short-term depuration experiments (24 h) showed that there was a rapid initial loss of radioactivity as unchanged hydrocarbon during the first 12 h ; but that radioactivity in the form of metabolites was retained (Fig. 15B). Varanasi and Malins (1977) draw attention t o the possibility that the toxic effects of low levels of naphthalene to Pandaha larvae may be related to the inability of these animals to release toxic metabolites of the hydrocarbon. The observation by Lee (1975) that micro-crustaceans included among the zooplankton are able to convert aromatic hydrocarbons into hydroxylated derivatives implies that these animals, like fish (Stegeman and Sabo, 1976) and large marine crustaceans (Burns, 1976 ; Philpot, James and Bend, 1976; Singer and Lee, 1978), possess the enzymes known as mixed function oxygenases (MFO), which work with mammals has shown to be involved in the metabolism of steroids and numerous drugs, as well as compounds such as petroleum hydrocarbons. The enzymes are NADPH (reduced pyridine nucleotide) dependent and are sometimes referred to as aryl-hydrocarbon hydroxylase (AHH) when working with a specific substrate such ;t9 BP. They operate in conjunction with the electron-transport system cytochrome P-450 and NADPH-cytochrome c reductase and are inhibited by cytochrome c and carbon monoxide. The overall reaction catalysed is : AH

+ 2e- + 2H+ + O,+AOH + H,O.

With B P as the substrate the assay ie based on measurements of 3-hydr oxybenzo [alpyrene . A full and well-presented account of the distribution of these

346

E. D. 9. CORNER

enzymes in marine animals is given by Varanasi and Malins (1977). I n the context of the present review, however, it is worth stressing that aquatic animals can possess a highly active AHH system: for example, trout liver-microsomes metabolize B P at a rate 5-10 times higher than male rat liver-microsomes when measured per mg of microsomal protein (Ahokas, Pelkonen and Karki, 1975). Furthermore, work by Clark and Diamond (1971) has shown that these high efficiencies for the metabolism of BP are maintained over a wide range of temperatures (5-30°C). A large number of substrates, including " foreign " organic compounds (xenobiotics), can induce high levels of MFO in mammals (Conney, 1967) and fish (Payne and Penrose, 1975; Payne, 1976): however, related studies with zooplankton species have not so far been made, although micro-crustaceans, being more metabolically active, might possess MFO systems less sluggish than those normally found in larger crustacean species (Philpot et al., 1976). Payne and Penrose (1975) suggest that the existence of inducible AHH in fish may provide a convenient means of assessing previous exposure of the animals to PNAH although, as they point out, more needs to be known about the decay of this induced activity. There is the further complication that AHH may also be induced by other compounds such as chlorinated hydrocarbons and aromatic pesticides. At present, nothing is known about the induction of MFO, including AHH, in zooplankton. The metabolic changes undergone by PNAH in marine animals have been reviewed elsewhere (Corner, 1975; Corner et al., 1976a; Varanasi and Malins, 1977). Briefly, the first stage in the process is the formation of an epoxide which is subsequently converted either into a dihydro-diol or, by conjugation with glutathione, into a premercapturic acid (see Fig. 16). The enzymes involved, epoxide hydrase and glutathione-8-transferase, have been detected in marine fish and invertebrates (James, Fouts and Bend, 1976: cited by Varanasi and Malins, 1977) but no studies have yet been made with planktonic organisms. Compared with the parent hydrocarbon, hydroxylated derivatives such as the dihydro-diol are more water-soluble and, either in the free state or as conjugates with sulphuric acid and either glucuronic acid (mammals and fish) or glucose (Arthropoda), are released by the animals. Thus, these metabolic changes would help to reduce the levels of hydrocarbon in the animals and could be regarded as a " detoxifying " process. Recent work with mammals has shown that dihydro-diols can themselves undergo further oxidation to form " diol-epoxides " (Booth and Sims, 1974) and, in the case of the carcinogenic compound BP, it

341

POLLUTION STUDIES WITH MARINE PLANKTON-I

is this derivative (7,8-&hydro-7,8-dihydroxybenzo[a]pyrene-9,lO-oxide) and not the parent hydrocarbon which is mainly responsible for carcinogenic activity (Sims, Grover, Swaisland, Pal and Hewer, 1974). It therefore seems that although some metabolic changes may facilitate the removal of hydrocarbons from the animal, others can increase the carcinogenic potential of these compounds. Stegeman and Sabo (1976) Dihydro- diol

Reactions with cell constituents(eg protein and nucleic acids)

1

Underqo conjugation reactions giving giucuronides (or glucosides) and sulphates

Epoxide

NADPH

+ H+

NADPH

+H-0

NHCOCH,

Premcrcapturic acid

FIG.16. Metabolism of an aromatic hydrocarbon in mctrine animals. Enzymes involved in main reactions: (1) mixed-function oxygenase, (2) epoxide hydrase, (3) glutathione-8-transferase.

have commented on the possible linkage between the activation of potential carcinogens by metabolism and the greater incidence of neoplasia in fish from contaminated regions. It would be interesting to know whether the toxic effects of PNAH in zooplankton are in any way related to metabolic changes undergone by these compounds in the animals, particularly as work with shrimp and crab larvae (Sanborn and Malins, 1977)has shown that naphthalene, which is particularly toxic to

348

E. D. 9. UORNER

these animals, is converted by the spot shrimp into metabolites which are retained in the tissues.

E. Release of hydrocarbons in faecal pellets Studies by Freegarde, Hatchard and Parker (1 971) with Calanus Jinmarcliicus and by Conover (1971), using this species and Ternora longicornis (0.F. Muller), have shown that when the animals feed in the presence of fine suspensions of crude oil they are able to ingest oil particles and release them in faecal pellets. As these are slightly heavier than sea water they sink out of the surface layer and thus provide a means for transferring a substantial fraction of an oil spill from this region to the benthos: indeed, Conover (1971) concluded that perhaps 20% or more of the particulate Bunker C oil released off the Atlantic coast of Canada by the " Arrow '' disaster was sedimented in zooplankton faeces. Likewise, Elder and Fowler (1977) have estimated that faecal pellets released by the euphausiid Meganyctiphanes norvegicca (M. Sars) are an important means by which other organic compounds, in this case polychlorinated biphenyls, are contributed to sediments in the Ligurian Sea. The transfer of hydrocarbons not only occurs when zooplankton ingest oil particles : water-soluble hydrocarbons directly taken up from solution by algal cells which are then only partially digested by herbivorous zooplankton could also be released in faecaI pellets and transferred t o the benthos in this way. Although the sinlung rates of the faecal pellets ase not so great as to exclude the possibility of the material being removed as a food by animals during its descent (mean values range from 66 to 240 m/day for the faecal pellets released by species of copepods and euphausiid : Turner, 1977) faecal pellet production by these animals can still contribute substantial amounts of material to the benthos in certain sea areas (Seki, Tsuji and Hattori, 1974; Davies, 1975): in the study by Davies (1975), for example, it was found t h a t 27% of the primary production as carbon in a Scottish sea loch was transferred to the benthos as faecal pellets released by herbivorous zooplankton. Accordingly, in studying the fate of hydrocarbons in zooplankton it is important to consider the quantities of these compounds that are released in faecal pellets. So far, however, the only study of this problem is that by Harris et al. (1977a) who determined the amounts of radioactivity released in faecal pellets by Calanus helgolandicw ingesting a known ration of Biddulphia previously exposed t o 14C-l-naphthalene. The data, summarized in Table XIV, show that quite a high proportion of the ingested radioactivity \

TABLEXIV. ASSIMILATION OF NAPHTHALENE FROM A BY Calanus helgolandicus

Animals

Females Males Females Females

Dietury constituent

Naphthalene Naphthalene Nitrogen Phosphorus

Diet used: Biddulphia sinen-&.

PLANTDIET

yo Ration cuptured A

f

Rejected in faeces 41.9 39.3 65.9 59.6

Assimilated 58.1 60.7 34-1 40.4

Summarized data from Harris et al. (1977a).

=

Retained in animal 31.2

26-8 7.5 0

-~

-

Released in soluble form 26.9 23.9 26.6 40.4

m

z

i

m

dx

E

i 2 3

W

t P

(D

350

E. D. 9. CORNER

(39.3-41-9y0) was present in the faecal pellets, such radioactivity, in the absence of any evidence for the metabolism of naphthalene by micro-algae (see p. 328), being assumed to represent unchanged hydrocarbon. Many studies have been made of the assimilation efficiencies of herbivorous zooplankton in terms of basic dietary components (e.g. carbon, nitrogen and phosphorus) and the values usually range from about 50 to 90% (see review by Corner and Davies, 1971). The assimilation efficiency obtained by Harris et al. (1977a) for 14C-1-naphthalene (68*1-60-7y0) is near the lower end of this range: however, the Biddulphia used as a diet, compared with other species of diatom, is much less readily digested (Corner et al., 1972) and further data included in Table XIV show that assimilation efficiency in terms of naphthalene was substantially higher than that found for either dietary nitrogen or phosphorus using this particular plant food. It is also worth noting from Table XIV that most of the nitrogen and all the phosphorus assimilated from the diet by the animals was released in soluble form, whereas an average value of only 51.2% was obtained with the hydrocarbon. Thus, compared with natural dietary constituents, naphthalene is not only more readily assimilated by the animals: it is less readily lost in soluble form. As pointed out by Parker, Freegarde and Hatchard (1971), the extent to which copepods can contribute to the immobilization of an oil slick depends upon the amount of oil dispersed as fine droplets suitable for capture, the daily volume of sea water swept clear and the number of animals present in the sea. For a concentration of dispersed oil droplets equivalent to 1.5 mg/l, which can persist near an oil slick for a considerable time in a choppy sea (Parker et al., 1971),and assuming a maximal value of 750 ml/day for the volume swept clear by a single Calanus (Paffenhofer, 1971; Corner et al., 1972), the total quantity of oil ingested daily is 1-125 mg. Making the further assumption that the value obtained with naphthalene by Harris et al. (1977a) is typical of oil hydrocarbons in general, roughly 40% of this ingested ration, or 0-45 mg, wilI be rejected as faecal pellets. Thus, one female Calanus in one litre of sea water could transfer 30% of the dispersed oil into faecal pellets daily. Another example of the quantitative importance of the amount of hydrocarbon released as faecal pellets can be assessed from further data by Harris et al. (1977a) presented in Fig. 17. This shows the quantity of radioactivity (as naphthalene equivalents) represented by a suspension of 20 000 Biddulphia cells/l exposed for 24 h to a low concentration of I*C-l-naphthalene (1.37 pg/l) in sea water; the daily

351

POLLUTION STUDIES WITH MARINE PLANKTON-I

ration captured by one female Calanus feeding on this cell suspension for one day ; and the amounts retained and released, either in soluble form or as faecal pellets, by the animal. Radioactivity in the faecal pellets represented 172.8 pg naphthalene/copepod and that present as suspended cells 3 620 pg hydrocarbon/l. Thus, under the experimental conditions, one female Calanus incorporated 4.8% of the total hydrocarbon available as phytoplankton in a litre of sea water into faecal pellets each day. Naphthalene concentration in sea woter (1 37pg/0

~

Naphthalene present in suspension of Biddulpim cells ( 3620 pg/l)

Napht ha lene captured by oneCa/anus

~

(3178pgl

)

(Retained (688~9)

Naphthalene released in faecal pellets (172 8 p g l

(Soluble release) (762pg)

FIG.17. Quantitative aspects of the transfer of an aromatic hydrocarbon from solution in sea water to faeclal pellets released by zooplankton.

VIII. TOXICITYSTUDIESWITH ZOOPLANKTON The toxicities of crude oils, of their total water-soluble fractions and of individual hydrocarbons, to a wide variety of marine animals have been referred t o in several reviews (Nelson-Smith, 1970 ;Butler, Berkes and Powles, 1974; Moore and Dwyer, 1974; Anderson, 1975): most of the work described, however, deals with larger marine animals and relatively few studies have been made with zooplankton. Hyland and Schneider (1976), in considering the possible effects of oil pollution on planktonic communities in the open sea, conclude that although these organisms may be affected by physical coating in a floating oil slick, or by poisoning in the toxic plume immediately beneath, the effects may only be temporary, population densities and age-distributions being rapidly restored because of high reproduction rates and effective dispersal mechanisms such as the immigration of organisms from unaffected areas. Be that as it may, however, they emphasize the much greater susceptibility t o oil contamination of local breeding populations of marine organisms, particularly the larval forms of certain fish (ichthyoplankton) and crustaceans and molluscs (meroplankton) in confined coastal areas where recovery from the effects of pollution may take several years. Toxicity data for both ichthyoplanktonic and meroplanktonic animals are therefore included

352

E. D. 9. CORNER

in Table XV, which summarizes the various toxicity studies reviewed in this section.

A. Crudeoil In early toxicity work, using whole oil, unstable suspensions of this material were prepared by adding it to sea water and shaking the mixture. Detailed studies involving multiple-day exposure were carried out by Mironov (1969) with Acartia clausi Giesbrecht, Paracalanus parvus (Claus), Penilia avirostris Dana, Centropages ponticus Karavajev and Oithona nana Giesbrecht collected from the Black Sea. The materials tested were crude oil (Malgobek), Bunker fuel oil F-12 and " solar " oil. I n the experiments with adult Acartia a concentration of 0.001 ml crude oil/l (ca. 1 mg/l) produced a slight toxic effect, 50% of the treated population dying in 4-5 days compar6d with 6.5 days for the controls; similar small effects were noted with fuel oil and solar oil used a t the same concentration. However, when animals in better condition were used, 50% of the control samples dying in nine days, no significant effects were observed with either crude oil or solar oil at the concentration of 1 mg/l ; moreover, that found with Bunker fuel oil was only slight, 50% of the test animals dying in eight days compared with nine days for the controls. Definite effects were found with all three oils used a t a concentration of 10 mg/l: at 100 mg/l all three oils killed the whole population of test animals in one day. Slight toxic effects were found with Bunker fuel oil at a concentration of 1 mg/l in experiments with Penilia, Centropages and Oithona ; but a more marked effect was observed with Paracalanus. As in the tests with Acartia, this oil at a concentration of 10 mg/l produced definite toxic effects with all four species ; and at 100 mg/l all populations were killed within one day. Mironov (1969) also carried out studies using the naupliar stages of Acartia and Oithona. Slight toxic effects were detected using Bunker fuel oil at a level of 1 mg/l, 50% of the treated population surviving only 3-5 days compared with 4.5 for the controls (Acartia) and 1.5 days compared with 3 (Oithona). The high mortality rates of the controls suggest that the animals were under stress, which may have increased their sensitivity to the oil. Studies by Barnett and Kontogiannis (1975) using the tidal pool copepod Tigriopus californicus (Baker) showed that crude oil was much less toxic to this species than to those studied by Mironov (1969). I n an earlier study, Kontogiannis and Barnett (1973) determined the concentrations of crude oil and its various fractions that represented

POLLUTION STUDIES WITH MARINE PLANKTON-I

353

the critical level of toxicity (i.e. above which the survival of the animal was reduced below that of untreated controls) and found those for diesel oil and kerosene to be 87 and 83 mg/l respectively. As might be expected, the harpacticoid copepod Tigriopus is much more resistant than a pelagic species such as Acartia or Oithona to the effects of crude oil : thus Kontogiannis and Barnett (1973) found that a concentration of 25 ml/l took three days to kill a population of Tigriopus, whereas Mironov (1969) found that 0.1 ml/l killed those of Oithona and Acartia within 24 h. I n none of the studies described so far was an even suspension of oil in the test medium maintained throughout the period of the experiment : for example, Barnett and Kontogiannis (1975) initially dispersed the oil using an ultrasonic probe, but the suspensions lasted for only 30 min. However, Spooner and Corlfett (1974) to some extent overcame this problem by placing the test samples on a vertically rotating wheel and by mixing the oil (Kuwait 250°C Residue oil: equivalent to one-day weathered) with the dispersant BP 11OOX. They studied the effects of the mixture on faecal pellet production, as a measure of feeding rate, by Calanus helgolandicus feeding on Platymonus suecica Kylin and found that a marked inhibition occurred during a 20h exposure to oil used at a concentration of 10 mg/l. Nevertheless, after the animals were removed from the oil suspension and transferred to fresh sea water the normal feeding rate was recovered within seven days I n a recent study by Wells and Sprague (1976) larvae of the American lobster Homarus americanus Milne-Edwards were used as test organisms and care was taken to monitor the levels of Venezuela crude oil during prolonged exposure, using the U.V. method of Zitko and Carson (1970). There were considerable losses of hydrocarbons during the experiments, some of which lasted 30 days, but the toxicity data refer to initial concentrations only, which therefore represent maximal values. Oil concentrations causing the death of 50% of the test population in four days (4-day LC,,) were 0-86 mg/l for first-stage larvae and 4.9 mg/l for third- and fourth-stage larvae ; by comparison the 30-day LC,, value for first-stage larvae was 0-14 mg/l. As observed in the study by Spooner and Corkett (1974) the presence of oil reduced food consumption. Vaughan (1973) used methods similar to those of Wells and Sprague (1976) to study the effects of three crude oils on larvae of the rock crab Cancer productus Randall: 4-day LC,, values were 3.2 mg/l for No. 2 fuel oil, 220 mg/l for Louisiana crude and 250 mg/l for Kuwait crude. I n addition, Wells and Sprague (1976) have calculated from the data of Percy and

354

1.D. 8. OORNER

Mullin (1975) using Venezuela crude oil that 4-day LC,, values for adults of two cold-water species, the amphipod Onisimus afinis H. J. Hansen and the copepod Calanus hyperboreus, were 29 and 82 mg/l respectively. A further contribution from the work of Percy and Mullin (1975)was the use of cell-free homogenates of Onisimus in order to study the effects of petroleum compounds on metabolic processes. Respiration rates of homogenates prepared from animals pre-exposed for 24 h to Norman Wells crude oil were found to be 10-46% greater than those of controls. Testing the effects of petroleum compounds on respiration by whole animals is complicated by the fact that changes in the rate of oxygen consumption could simply reflect differences in locomotory activity : the use of cell-free extracts overcomes this difficulty. The work of Percy and Mullin (1975)has the additional value that it is one of the few studies made using Arctic species. B. Water-soluble hydrocarbons The acute effects of the WSF of No. 2 fuel oil (Exxon, Baytown) on both coastal and oceanic zooplankton have been studied by Lee and Nicol (1977)using mixtures of animals collected from the Gulf of Mexico and off the coast of Texas. The 2-day LC,, obtained with coastal zooplankton was 9-5 mg/l (50% WSF). However, oceanic animals were much more sensitive in that the time to kill 50% of the sample a t the same concentration was only 9 h. Among the copepod species calanoids were more sensitive than cyclopoids ; and members of the meroplankton (e.g. barnacle nauplii) were more resistant than holoplanktonic forms (e.g. copepods). I n a subsequent study (Donahue, Wang, Welch and Nicol, 1977), a wide range of aromatic compounds found in petroleum were tested for toxicity to nauplii of the barnacle Balanus amphitrite niveus Darwin. Estimated from the effects on larval activity, alkylated benzenes and naphthalenes were more toxic than the parent compounds ; in addition, phenalen- 1-one, which is particularly toxic to green microalgae (Winters et al., 1977), and naphthalene were found to affect phototactic response. Evidence for the higher toxicities t o zooplankton of alkylated napthalenes compared with that of the parent hydrocarbon has also been found in a recent study by Ott, Harris and O’Hara (1978). Using Eurytemora afinis as the test organism they found 24h LC,, values of 0-316,0.852, 1.499 and 3.798 mg/l . for 2,3,5-trimethylnaphthalene, 2,6-dimethylnaphthalene, 2methylnaphthalene and naphthalene respectively. Further studies with meroplanktonic animals are those of Byrne

POLLUTION STUDIES WITH MARINE PLANKTON-I

355

and Calder (1977) who tested the toxic effects of the WSFs of six crude oils using larvae of the quahog clam Mercenaria sp. Two-day LC,, values varied considerably with the oil used, that for the WSF of Kuwait crude being 25 mg/l and that for the WSF of used motor oil only 0.10 mg/l. I n addition, longer-term exposures (10 days) caused increased toxicities (see Table XV). Compared with those of adults, the susceptibilities of the larval, meroplanktonic stages of marine crustaceans to the WSF of a crude oil can be markedly greater. Thus, Brodersen, Rice, Short, Macklenberg and Karinen (1977) found that 4-day LC,, values for Stage I larvae of the King crab Paralithodes camtschatica (Tilesius) vaned from 0.78 to 1-12 mg/l whereas the corresponding values for adults were in the range 3.6-5.0 mg/l: likewise, in experiments with the kelp shrimp Eualus suckleyi (Stimpson) the 4-day LC,, values for Stage I larvae varied from 0.87 to 1.4 mg/l compared with 1.8-2.1 mg/l for adults. The toxic effects of the WSF of a high aromatic heating oil have been tested by Berdugo, Harris and O'Hara (1977) using the estuarine copepod Eurytemora afiniis. Animals exposed for 24 h to the dissolved hydrocarbons equivalent to a total concentration of 520 pg/l ingested algal food at a rate only 62y0that of untreated controls. By contrast, the single hydrocarbon naphthalene used a t a higher concentration (1.0 mg/l) caused only an 11% reduction in feeding rate, indicating that compounds other than naphthalene in the WSF were mainly responsible for its toxicity. Important developments in toxicity work with zooplankton were introduced by Sanborn and Malins (1977) in their studies with larval stages of Pandalus platyceros and Cancer magister. First, a continuousflow method was used in which the concentration of hydrocarbon was constantly maintained ; secondly, the hydrocarbon was used both in the free state and as a complex with BSA (see p. 345). 1%-l-Naphthalene, used either alone or as the BSA complex, was extremely toxic to the animals, a concentration of only 8-12 pg/l causing 100% mortality in 24-36 h. A continuous-flow system has also been used by Forns (1977) in studying the effects of unweathered South Louisiana crude oil on larvae of the lobster Homarzts americanzts. A threshold of sensitivity was observed between crude oil concentrations of 0.1 and 1-0 mg/l. Thus, animals exposed to 0-1 mg/l were active feeders with consistent locomotive behaviour and displaying strong aggressiveness : those exposed to 1.0 mg/l were lethargic, active motions were minimal, feeding was depressed and frequently the animals appeared to be dead. I n addition, animals exposed to 1 mg oil/l had a slower rate of

356

E. D. S. OORNER

development. Mortality of these animals reached 50% after nine days, whereas survival for the control larvae and those treated with 0.1 mg oil/l exceeded 50% throughout the 17-day period of the experiment. Compared with static systems, those involving continuous flow have decided advantages : for example, replenishment of dissolved oxygen and nutrients, removal of metabolic waste products, reduction of bacterial contamination and maintenance of constant levels of volatile and unstable test substances such as hydrocarbons. Continuous-flow methodology therefore provides a better approximation to conditions prevailing in the natural environment and is finding increased application in toxicity studies with corttponents of crude oil (see, for example, Hyland, Rogerson and Gardner, 1977 ; Roubal, Bovee, Collier and Stranahan, 1977a).

C. Possible eflects of hydrocarbons

on reproduction by zooplankton A recent study of the enzyme AHH in the blue crab Callinectes sapidus Rathbun (Singer and Lee, 1977) has focussed attention on the possible interference of xenobiotics, such as aromatic hydrocarbons, with steroid metabolism. Thus, Singer and Lee (1977) noted that the levels of activity of the enzyme, measured by the method of Whitlock and Gelboin (1974) in the green gland, were inversely related to the production of the steroid hormones, ecdysones, which control moulting in these animals (Faux, Horn, Middleton, Fales and Lowe, 1969). It is well known that crustaceans do not carry out de novo sterol biosynthesis, but convert dietary compounds, such as the phytosterols present in plants, into desmosterol and subsequently cholesterol, the latter conipound then undergoing further changes to give steroids such as the mammalian sex hormones (see review by Goad, 1976). In mammals a further hydroxylation of these sex hormones involves the enzyme system androgen hydroxylase, the activity of which can be greatly stimulated by pre-treatment of the animal with certain drugs (Conney and Klutch, 1963; Lu, Kuntzman, West, Jacobson and Conney, 1972). Desmosterol and cholesterol have both been detected in the copepod Euchaeta japonica by Lee et al. (1974) and the recent use of a radioimmunoassay technique has indicated the probable presence of the sex hormone oestradiol-l7-/3 in Calanus helgolandicus (O’Hara, Corner and Kilvington, 1978). Furthermore, work by Lee (1975), referred to earlier (p. 343), has shown that hydroxylated derivatives are formed from BP in copepods, which implies that AHH is present in these animals.

POLLUTION STUDIES WITH MARZNE PLANKTON-I

357

No i n vitro studies have yet been carried out to examine the possibility that PNAH might interfere with steroid metabolism in copepods, thereby influencing moulting, sex ratio and reproduction, although there is evidence from the recent i n vivo study by Berdugo et al. (1977) that petroleum hydrocarbons might inhibit the rate of egg-production by Eurytemora afinis. The test solution was the WSF of a high aromatic heating oil, with the volatile components benzene and toluene excluded, and ovigerous females immersed in this solution for 4 h showed a subsequent daily rate of egg-production approximately 30% that of untreated controls. This work has recently been extended by Ott, Harris and O’Hara (1978) who found that multiple-day exposure (maximum 29 days) of Eurytemora to each hydrocarbon at a concentration of 10 pg/l in sea water reduced the length of adult life, the total number of nauplii produced, the mean brood size and the rate of egg-production. However, in contrast to the LC,, data (see Table XV), increased alkylation of the hydrocarbon did not lead to greater inhibition of fecundity : thus, dimethylnaphthalene had less effect than the parent hydrocarbon on the total number of eggs produced; and both methyl-and dimethylnaphthalene had less effect than naphthalene on the rate of egg-production. As pointed out by the authors, the hydrocarbons did not necessarily have a specific effect on fecundity, inhibition of which could have resulted from a reduction in feeding rate. Viability of the eggs is another important factor in reproduction and unpublished work by Lewis and Lee (cited by Lee, 1975), using Euchaeta juponicu, has shown that the hydrocarbons 1- and 2-methylnaphthalene at a concentration of 80 pg/l cause a 60% reduction in the number of eggs reaching nauplius 11.

D.

Summary and general comments

Results from the various toxicity studies with zooplankton are summarized in Table XV. A proper assessment of the possibility that the levels of hydrocarbons in different sea areas are likely to affect the dominant species of zooplankton inhabiting the same regions cannot be made at present : much more work is needed. However, comparison of the data in Table XV and Table I at least indicates that the amounts of crude oil found in the Adriatic (1-40-10.98 mg/l) would affect several species of zooplankton, including the larval stages of certain fish, and that the maximal levels of total hydrocarbons reported for the North Sea (625 pg/l), the Sargasso Sea (559 pg/l) and Goteborg Harbour (710 pg/1) are sufficiently high to affect clam larvae and adult copepods.

w

cn

TABLEXV. TOXICEFFECTS ON ZOOPLANKTON OF CRUDE OILS, WATER-SOLUBLE FRACTIONS AND INDMDUAL HYDROCARBONS

Test material

Temp*

("1'

Critical level of toxicity (mq/l .. or p.p.m.*)

Bunker fuel oil F-12

NS

1.o

Bunker fuel oil F-12

NS

1.0

Bunker fuel oil F-12

NS

1.0

Bunker fuel oil F-12

NS

1.o

Bunker fuel oil F-12

NS

1.0

Diesel oil

10

87

Kerosene

10

83

Venezuela crude

20

0-86

Venezuela crude

20

4.9

Venezuela crude

20

0.14

Venezuela crude

15-20

1.ot

8

3.2

No. 2 fuel oil

Toxic effect

Species

00

Reference

Holoplankton and meroplankton Treated animals died slightly Paracalanw parvwr (adults) faster than controls Treated animals died slightly Centropageaponticwr (adults) faster than controls Treated animals died slightly Penilia aviroatria (adults) faster than controls Treated animals died slightly Oithona nanu (adults young faster than controls stage) Treated animals died slightly Acartia clauSii (adultsf young faster than controls stage) californicw ~ (adults) Treated animals died slightly T i g r i o p ~ faster than controls Treated animals died slightly Tigriopw californicw (adults) faster than controls H o m a w americanw (stage I 4-day LC,, larvae) 4-day LC,, Homarw americanus (stage 111-IV larvae) Homarw americanus (stage 1 30-day LC,, larvae) Slower rate of development; Nerwnariu sp. (stage I-IV feeding inhibited ; 9-day LC,, larvae) Cancer productwr (larvae) 4-day LC,,

+

Mironov (1969) Mironov (1969) Mironov (1969)

P

Mironov (1969)

0

rn

Mironov (1969) Kontogiannis and Barnett (1973) Kontogiannis and Barnett (1973) Wells and Sprague (1976) Wells and Sprague (1976) Wells and Sprague (1976) Forns (1977) Vaughan (1973)

8

2

0

Kuwait crude Louisiana crude Venezuela crude Venezuela crude Kuwait crude B.P. ll0OX WSF of Kuwait crude WSF of S. Louisiana crude WSF of Bunker C WSF of No. 2 fuel oil WSF of Florida J a y crude WSF of used motor oil WSF of Kuwait crude WSF of Louisiana crude WSF of Bunker C WSF of No. 2 fuel oil WSF of Florida J a y crude WSF of Cook Inlet crude

4-day LC,, 4-day LC,, 4-day LC,, 4-day LC,, Inhibition of feeding

Cancer productus (larvae) Cancer productus (larvae) Onisimus a&& (adults) Calanus hyperboreus (adults) Cdanim helgolandicus (adults)

8 8 8 5 12

250 220 29 82 10

25 25

>25 6.0

2-day LC,, 2-day LC,,

Mercenaria sp. (larvae) Mercenaria sp. (larvae)

Vaughan (1973) Vaughan (1973) Percy and Mullin (1975) Percy and Mullin (1975) Spooner and Corkett (1974) Byrne and Calder (1977) Byrne and Calder (1977)

25 25 25

3.2 1-3 0.25

2-day LC,, 2-day LC,, 2-day LC,,

Mercenuria sp. (larvae) Mercenaria sp. (larvae) Mercenaria sp. (larvae)

Byrne and Calder (1977) Byrne and Calder (1977) Byrne and Calder (1977)

25 25 25 25 25 25

0.10 2.0 2.1 1.6 0.53 0.05

2-day LC,, 10-day LC,, 10-day LC,, 10-day LC,, 10-day LC,, 10-day LC,,

Mercenaria Mercenaria Mercenaria Mercenaria Mercenaria Mercenaria

Byrne and Calder (1977) Byrne and Calder (1977) Byrne and Calder (1977) Byrne and Calder (1977) Byrne and Calder (1977) Byrne and Calder (1977)

3-5

1.1

WSF of Cook Inlet crude

3-5

0.96

WSF of No. 2 fuel oil WSF of No. 2 fuel oil WSF of high aromatic heating oil WSF of high aromatic heating oil Naphthalene Naphthalene 2-Methylnaphthalene

25-26 25-26 15

9.5 9.5 0.52

+

*

2 c!

sp. sp. sp. sp. sp. sp.

(larvae) (larvae) (larvae) (larvae) (larvae) (larvae)

(failure to swim) Eualus suckleyi (Kelp shrimp larvae) 4-day LC,, (failure to swim) Paralithodes camtschaticn (King crab larvae) 2-day LC,, Mixed coastal plankton 9-hoW LC,, Mixed oceanic plankton 62 ?& reduction in feeding rate Eurytemora afinis (adults) 4-day LC,,

Brodersen et al. (1977) Broderaen et al. (1977) Lee and Nicol (1977) Lee and Nicol (1977) Berdugo et al. (1977)

U

2 Z

M

8

8E

' 1

H

15

0.52

15 15 15

1.0 3.798 1.499

30% inhibition of egg production 11% reduction in feeding rate 24h-LC,, 24h-LC5,

Eurytemora afinis (adults)

Bardugo et al. (1977)

Euryternora a$& (adults) Eurytemora a&& (adults) Eurytemora a@& (adults)

Berdugo et al. (1977) Ott et al. (1978) Ott et al. (1978)

a:

5a 0

TABLEXV (continued)

Test material

Temp. ("C)

Critical level of toxicity (mgllor p.p.rn.*)

Toxic effect

Species

Reference

2,B-Dimethylnaphthalene 2,3,5-Trimethylnaphthalene Naphthalene

15

0.852

24h-LC5,

Euytemora afinis (adults)

Ott et al. (1978)

15

0.316

24h-LC5,

Eurytemora afinis (adults)

Ott et al. (1978)

10

100%mortality in 24-36 h

Pandalus plutycero8 (larvae)

Naphthalene

10

100% mortality in 24-36 h

Cancer naugister (larvae)

I-Methylnaphthalene

NS

0.0080.012t 0.008 0.012t 0.080

45% reduction in eggs reaching Euchaeta japonica (adults)

Sanborn and Malins (1977) Sanborn and Malins (1977) Lee (1975)

2-Methylnaphthdene

NS

0.080

46% reduction in eggs reaching Euchueta japonica (adults)

nauplius 11 Lee (1975)

nauplius I1

Icthyoplankton Prudhoe Bay crude

5-11.5

88-110

Prudhoe Bay crude

5-11.5

14-16.0

4-day LC,, Avoidance effects

Onwrhynchw gorbwchia (Wdbaum) (Pink salmon fry) Onwrhynchw gorbwchia (Pink salmon fry)

Rice (1973) Rice (1973)

Prudhoe Bay crude

4-5

0.73

Ekofisk crude

6-7.5

0.1

‘‘ Oil

”

16.5-18

0.1-1.0

‘I

Oil ”

16.5-18

0.01-0.1

“

oil

”

16.5-13

1.0

<‘

oil

”

165-18

10-100

Iran crude

1s

10000 (ca. 10

Venezuela crude

9

p.p.m. in solution) 100

Benzene

10-17.5

40-45

Benzene

10-17.5

20-25

Benzene

16.519.2

20-25

Decrease in growth

Oncorhynchus gorbuschia (Pink salmon fry) 30% decrease in egg hatching Mallotus villosus (Miiller) (capelin) Rhombus meeoticus (Pall.) Death of eggs (Black Sea Turbot) Only 5 5 4 9 % of eggs hatch Rhombus maeoticus (Black Sea Turbot) 3-4 fold increase in abnormal Rhombus maeoticus larvae (Black Sea Turbot) Larvae killed Rhombus maeoticus (Black Sea Turbot) After 100 h exposure, twice the Gadus morhua L. number of dead eggs as in untreated controls. Large number of malformed embryos Large number of malformed embryos 4-day LC,, for mortality a t hatching 2-day LC,, for mortality a t larvae day 7 2-day LC,, for mortality a t larvae day 3

Rice, Moles and Short (1975) Johannessen (1976) Mironov (1972) Mironov (1972) Mironov (1972)

z

*

0

Mironov (1972) Ktihnhold (1972)

Clupea harengus membras L. Linden (1976) (eggs and larvae) Clupea pallasi Val. (PaciGc Struhsaker, Eldridge and Echeverria (1974) herring) Clupea palluai (Pacificherring) Struhsaker, Eldridge and Echeverria (1974) Engraulis mordax Girard Struhsalrer, ElWdge and Echeverria (1974) (Northern anchovy)

* Values refer to amounts of oil originally added to sea water, or to initial levels of hydrocarbons measured in WSFs. t Hydrocarbon levels maintained under continuous flow.

m

3E B

b td

[ i

H

NS = Not stated.

w

WJ

c,

362

E. D. 9. CORNER

I n addition Table XV draws attention to the enhanced sensitivities of lobster and crab larvae compared with those of adult amphipods and copepods ; and to the high toxicities of the two alkylated naphthalenes. Concerning this latter finding, mention has already been made of the numerous water-soluble components of crude oils used in the important studies by Winters et al. (1976) which demonstrate the toxicities of a wide range of individual compounds to marine unicellular algae (p. 326). Comparable studies with zooplankton have not been made : yet, in the context of oil pollution there is an obvious need to extend toxicity work with hydrocarbons to include studies with related compounds, particularly the alkylated phenols that can occur in significant quantities in the WSFs of crude oils and which, in comparison with commonly used hydrocarbons such as naphthalene and its alkyl-derivatives, are less volatile (Winters and Parker, 1977) and could therefore be more persistent in the sea.

IX. CONCLUSIONS Whether further inputs of petroleum compounds into various sea areas will eventually have serious effects on plankton populations is a matter of conjecture. A strong likelihood, however, is that these compounds will continue to be released into the sea throughout the foreseeable future. It therefore seems worthwhile to conclude by summarizing suggestions for further work, basing these on gaps in knowledge already identified in earlier sections.

A. Chemical analyses As studies with planktonic organisms have shown that certain groups of hydrocarbons (e.g. PNAH and their alkylated derivatives) and related compounds (alkylated phenols and anilines) present in crude oil possess considerable toxicity, future analyses of petroleum compounds in sea water, both in the dissolved and particulate forms, and especially from areas affected by chronic oil spills, should be primarily concerned with identifying and measuring the levels of these more toxic components. Such levels should then be compared with those needed to produce measurable biological effects in short- and long-term toxicity studies. I n addition, there is a need to analyse these compounds and their derivatives in phytoplankton and both herbivorous and carnivorous zooplankton and young fish, particularly in the vicinity of oil spills, in order to assess the extent to which such compounds may be concentrated in the food web.

POLLUTION STUDIES WITH MARINE PLANKTON-I

363

Concerning the spatial distribution of hydrocarbons in the sea, many previous studies have demonstrated the high levels of these compounds that occur at the surface. Work in various sea areas has shown, however, that large concentrations of phytoplankton occur at frontal areas and in the thermocline region in stratified waters (Lorenzen, 1967 ; Lasker, 1975 ; Savidge, 1976; Pingree, Holligan, Mardell and Head, 1976). The basic importance of phytoplankton in marine production makes such regions especially important : accordingly, in determining the levels of petroleum hydrocarbons in the sea, either to obtain background information or to follow what happens immediately after an oil-spill, test samples should be taken from these regions as well as from the surface. Furthermore, because of the importance of zooplankton faecal pellets in transferring plant material from the euphotic zone to the benthos there is a need, should an oil-spill occur simultaneously with a spring diatom increase, to extend studies concerned with the spatial distribution of petroleum compounds to include analyses of benthic fauna, especially those used as a food by demersal fish.

B. Toxicity studies Although much of value has already been learned about the effects of individual petroleum compounds on planktonic organisms, more needs to be known about the toxicities of mixtures of these compounds, especially those present in the WSFs of crude oils from areas of relatively recent oil exploration such as the North Sea. Components of these mixtures may behave synergistically in terms of toxic effects : moreover, the uptake and retention of a particular component could also be influenced by the presence of others. I n addition, more work is needed on the toxicities of compounds such as hydroperoxides and thiacyclanes (sulphoxides) which result from oxidation processes during the weathering of crude oil on the surface of the sea. Recent toxicity studies with zooplankton have incorporated continuous-flow methods, or variations thereof, together with monitoring of hydrocarbon levels throughout the experiments. Such techniques, although time-consuming, should also be applied in future work concerned with sub-acute long-term effects. Here the main emphasis should be on using mixtures of petroleum compounds at naturally occurring levels ranging from those encountered in the immediate vicinity of an oil-spill, and at different depths in the toxic plume beneath, to those found in estuaries and in-shore waters subject to small but frequent inputs of oil. The possible influence of petroleum

364

E. D. 9. OORNER

compounds on secondary production in the sea should then be assessed in terms of their effects on rates of feeding, moulting, growth, eggproduction and sex ratio, using animals maintained over multiplegenerations in laboratory cultures. There is also a need for short-term studies using animals maintained over a single generation, or even individual stages (particularly with meroplanktonic and ichthyoplanktonic species), to investigate the effects of petroleum compounds on further factors influencing the phytoplankton/zooplankton relationship : important among these are the efficiencies with which plant constituents are assimilated and converted into animal tissue, the process of nutrient regeneration, and food selection by zooplankton in the presence of mixtures of diets occurring naturally in the sea. Finally, because of interactions between species in nature, there is a need for further studies on the effects of petroleum compounds on complete eco-systems, particularly those found in Arctic regions where, as Percy and Mullin (1975) point out, a marked reduction in the rate of loss of the components of crude oil is likely to produce more prolonged toxic effects. The use of large plastic enclosures for work of this kind has already led to some interesting findings and it seems desirable for future developments to include additional studies along these lines.

C . Biochemical work Studies have already been made of the effects of petroleum compounds on photosynthesis and the gross chemical composition of unicellular algae (although the algal studies still need t o be done with marine species). Work with fish (Stegeman and Sabo, 1976; Sabo and Stegeman, 1977) has shown that lipid metabolism is affected in animals exposed for long periods to petroleum compounds. The importance of lipids in zooplankton (Lee, Nevenzel and Paffenhbfer, 1971) emphasizes the need for similar studies to be made with these animals, especially the influence of petroleum compounds on the synthesis of particular lipid fractions such as triglycerides, wax esters, phospholipids and sterols. Also needed are studies of the effects of petroleum compounds on the relative amounts of lipid and protein metabolized by planktonic organisms, both plant and animal. Another biochemical problem is that of ascertaining the extent to which detoxification mechanisms may occur in marine unicellular algae. I n addition, present indications that zooplankton animals such as micro-crustaceans possess the enzyme systems involved in convert-

POLLUTION STUDIES WITH MARINE PLANKTON-I

366

ing PNAH into hydroxylated derivatives should be followed up with studies, both in vivo and in vitro, of the induction of these enzymes in animals exposed to various xenobiotics, the rate of decay of any induced activity and the relative levels of such activity in animals from contaminated and uncontaminated areas. Related to which is the additional need to establish whether the enzymes epoxide hydrase and glutathione-8-transferase are present in planktonic organisms ; and to carry out detoxification and depuration studies, which so far have been made with hydrocarbons only, with related compounds such as phenols and anilines. A further area of biochemical study is that of establishing mechanisms for the biosynthesis and metabolism of steroids, particularly sex hormones and moulting hormones, in zooplankton and to assess the possible influence of the structurally related PNAH on these processes.

X. ACKNOWLEDGEMENTS I am particularly grateful to Dr M. F. Spooner for her many helpful criticisms of the manuscript; and to Drs R. P. Lee and D. C. Malins for sending me advance copies of work that was still in the process of publication and for allowing me to quote it in preparing this review. I am also indebted to several of my colleagues at the M.B.A. : particularly to Mr D. S. Moulder who supplied me with many publications dealing with all aspects of oil pollution, to Dr G. T. Boalch and Miss Elizabeth Roberts for ascertaining the authorities of numerous plant and animal species, to Mr C. C. Kilvington for checking the bibliography, to Miss Linda Carpenter and Miss Marsha Rapson for typing the manuscript and to Mr G. A. W. Battin for re-drawing the figures. XI. REFERENCES Ahokas, J. T., Pellronen, 0. and Karki, N. T. (1975). Metabolism of polycyclic hydrocarbons by a highly active aryl hydrocarbon hydroxylase system in the liver of a trout species. Biochemical and Biophysical Research Communications, 63, 635-641. Anderson, J. W., ed. (1975). " Laboratory Studies on the Effects of Oil on Marine Organisms : an Overview Report to the American Petroleum Jnstitute Division of Environmental Affairs. (A.P.I. Publication No. 4249), 70 pp. Washington, D.C. Anderson, J. W., Neff, J. M., Cox, B. A., Tatem, H. E. and Hightower, G. M. (1974). Characteristics of dispersions and water-soluble extracts of crude and refked oils and their toxicity to estuarine crustaceans and fish. Marine Biology, 27, 75-88.

".

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Avigan, J. and Blumer, M. (1968). On the origin of pristane in marine organisms. Journal of Lipid Research, 9, 350-352. Baker, J. M. (1970). The effects of oils on plants. Environmental Pollution, 1, 27-44. Barbier, M., Joly, D., Saliot, A. and Tourres, D. (1973). Hydrocarbons from sea water. Deep-sea Research, 20, 305-314. Barnett, C. J. and Kontogiannis, J. E. (1975). The effect of crude oil fractions on the survival of a tidepool copepod, Tigriopus caliJornicus. Environmental Pollution, 8, 45-54. Bendoraitis, J. G., Brown, B. L. and Hepner, L. S. (1963). Isolation and identification of isoprenoids in petroleum. Paper 15 presented at Sixth World Petroleum Congress, 1963. Section 5, pp. 13-29. Berdugo, V., Harris, R. P. and O’Hara, S. C. M. (1977). The effect of petroleum hydrocarbons on reproduction of an estuarine planktonic copepod in laboratory cultures. Marine Pollution Bulletin, 8 , 138-143. Blaxter, J. H. S. and Holliday, F. G. T. (1958). Factors influencing the feeding behaviour of herring. Proceedings of the Royal Physical Society of Edinburgh, 27, 15-28. Blurner, M. (1967). Hydrocarbons in digestive tract and liver of a basking shark. Science, New York, 156, 3 9 6 3 9 1 . Blumer, M. (1970). Dissolved organic compounds in sea water: saturated and olefinic hydrocarbons and singly branched fatty acids. I n “ Symposium on Organic Matter in Natural Waters ”, held a t the University of Alaska September 2-4, 1968. (D. W. Hood, ed.), pp. 153-167. Institute of Marine Science, Occasional Publication No. 1. Umversity of Alaska. Blumer, M. and Snyder, W. D. (1965). Isoprenoid hydrocarbons in recent sediments : presence of pristane and probable absence of phytane. Science, New York, 150, 1588-1589. Blumer, M. and Thomas, D. W. (1965a). Phytadienes in zooplankton. Science, New York, 147, 1148-1149. Blumer, M. and Thomas, D. W. (1965b). “ Zamene ”, isomeric C,, monooleha from marine zooplankton, fishes and mammals. Science, New York, 148, 370-371. Blumer, M., Mullin, M. M. and Thomas, D. W. (1963). Pristane in zooplankton. Science, New York, 140, 974. Blumer, M.,Mullin, M. M. and Thomas, D. W. (1964). Pristane in the marine environment. Helgolander Wissenschaftliche Meeresuntersuchungen, 10, 187-201 Blumer, M., Robertson, J. C., Gordon, J. E. and Sass, J. (1969). Phytol-derived C,, di- and triolefkic hydrocarbons in marine zooplankton and fishes. Biochemistry, 8 , 4067-4074. Blumer, M., Mullin, M. M. and Guillard, R. R. L. (1970). A polyunsaturated hydrocarbon (3, 6, 9, 12, 15, 18-heneicosahexaene) in the marine food web. Marine Biology, 6, 226-235. Blumer, M., Guillard, R. R. L. and Chase, T. (1971). Hydrocarbons of marine phytoplankton. Marine Biology, 8 , 183-189. Boney, A. D. (1974). Aromatic hydrocarbons and the growth of marine algae. Marine Pollution Bulletin, 5, 185-186. Boney, A. D. and Corner, E. D. S. (1959). Application of toxic agents in the study of the ecological resistance of intertidal red algae. JournaE of the Marine Biological Association of the United Kingdom, 38, 267-275.

POLLUTION STUDIES WITH MARINE PLANKTON-I

367

Boney, A. D. and Corner, E. D. S. (1962). On the effects of some carcinogenic hydrocarbons on the growth of sporelings of marine red algae. Journal of the Marine Biological Association of the United Kingdom, 42, 579-585. Booth, J. and Sims, P. (1974). 8,g-Dihydro- 8,9-dihydroxybenz(a)anthracene 10,ll-oxide: a new type of polycyclic aromatic hydrocarbon metabolite. Federation of European Biochemical Societies Letters, 47, 30-33. Borneff, J., Selenka, F., Kunte, H. and Maximos, A. (1968). Experimental studies on the formation of polycyclic aromatic hydrocarbons in plants. Environmental Research, 2, 22-29. Boylan, D. B. and Tripp, B. W. (1971). Determination of hydrocarbons in seawater extracts of crude oil and crude oil fractions. Nature, London, 230, 44-47. Brodersen, C. C., Rice, S. D., Short, J. W., Mecklenburg, T. A. and Karinen, J. F. (1977). Sensitivity of larval and adult Alaskan shrimp and crabs to acute exposures of the water-soluble fraction of Cook Inlet crude oil. I n " Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) " held a t New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 575-578. American Petroleum Institute, Washington, D.C. Brown, R. A. and Searl, T. D. (1976). Nonvolatile hydrocarbons in the Pacific Ocean. I n " Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment ". Proceedings of the Symposium held a t Washington, D.C., 9-1 1 August, 1976, pp. 240-255. American Institute of Biological Sciences, Arlington, Virginia. Brown, R. A., Searl, T. D., Elliott, J. J., Phillips, B. G., Brandon, D. E. and Monaghan, P. H. (1973). Distribution of heavy hydrocarbons in some Atlantic Ocean waters. I n " Proceedings of the Joint Conference on Prevention and Control of Oil Spills " held at Washington, D.C., March 13-15, 1973, pp. 505-519. American Petroleum Institute, Washington, D.C. Brown, R. A., Searl, T. D. and Koons, C. B. (1976). " Measurement and interpreNOAA Data Report ERL tation of hydrocarbons in the Pacific Ocean MESA-19, 246 pp. Report prepared for U.S. Department of Commerce National Oceanic and Atmospheric Administration, Boulder, Colorado. Brummage, K. G. (1973). The sources of oil entering the sea. I n " Background Papers for a Workshop on Inputs, Fates, and Effects of Petroleum in the Marine Environment ", Vol. 1 , pp. 1-6. National Academy of Sciences, Washington, D.C. Burns, K. A. (1976). Hydrocarbon metabolism in the intertidal fiddler crab Uca pugnax. Marine Biology, 36, 5-11. Burwood, R. and Speers, G. C. (1974). Photo-oxidation as a factor in the environmental dispersal of crude oil. Estuarine and Coastal Marine Science, 2, 117-135. Butler, M. J. A., Berkes, F. and Powles, H. (1974). " Biological Aspects of Oil Pollution in the Marine Environment : A Review ". 133 pp. Manuscript Report No. 22A of Marine Sciences Centre, McGill University, Montreal. Byrne, C. J. and Calder, J. A. (1977). Effect of the water-soluble fractions of crude, refined and waste oils on the embryonic and larval stages of the quahog clam Mercenaria sp. Marine Biology, 40, 225-231.

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E. D. 9. CORNER

Calder, J. A. (1976). Hydrocarbons from zooplankton of the Eastern Gulf of Mexico. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held at Washington, D.C., 9-1 1 August, 1976, pp. 160-183. American Institute of Biological Sciences, Arlington, Virginia. Carlberg, S. R. and Skarstedt, C. B. (1972). Determination of small amounts of non-polar hydrocarbons (oil) in sea water. Journal d u Conseil international pour Z’Exploration de la Mer, 34,506-515. Carlson, G. P. (1972). Detoxification of foreign organic conipounds by the quahaug, Mercenaria mercenaria. Comparative Biochemistry and Physiology, 43B, 295-302. Charter, D. B., Sutherland, R. A. and Porricelli, J. (1073). Quantitative estimates of petroleum to the oceans. I n “ Background Papers for a Workshop on Inputs, Fates, and Effects of Petroleum in the Marine Environment ”, Vol. 1, pp. 7-30. National Academy of Sciences, Washington, D.C. Clark, H. F. and Diamond, L. (1971). Comparative studies on the interaction of benzopyrene with cells derived from poikilothermic and homeotherinic vertebrates. 11. Effect of temperature on benzopyrene metabolism and cell multiplication. Journal of Cellular Physiology, 77, 385-392. Clark, R. C., Jr. and Blumer, M. (1967). Distribution of n-paraffins in marine organisms and sediment. Limnology and Oceanography, 12, 79-87. Conney, A. H. (1967). Pharmacological implications of microsomal enzyme induction. Pharmacological Reviews, 19, 317-366. Conney, A. H. and Klutch, A. (1963). Increased activity of androgen hydroxylases in liver microsomes of rats pretreated with phenobarbital and other drugs. Journal of Biological Chemistry, 238, 1611-1617. Conover, R. J. (1962). Metabolism and growth in Calanus hyperboreus in relation to its life cycle. Rapport et Procks-verbaux des Rkunions, Conseil permanent international pour I’Exploration de la Mer, 153, 190-197. Conover, R. J. (1971). Some relations between zooplankton and Bunker C oil in Chedabucto Bay following the wreck of the tanker “ Arrow ”. Journal of the Fisheries Research Board of Canada, 28, 1327-1330. Cooper, J. E. and Bray, E. E. (1963). A postulated role of fatty acids in petroleum formation. Geochimica et cosmochimica Acta, 27, 1113-1 127. Cormack, D. and Nichols, J. A. (1977). The concentrations of oil in sea water resulting from natural and chemically induced dispersion of oil slicks. I n “ Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) ” held at New Orleans, Louisiana, March 8-10. (A.P.l. Publication No. 4284), pp. 381-385. American Petroleum Institute, Washington, D.C. Corner, E. D. S. (1975). The fate of fossil fuel hydrocarbons in marine animals. Proceedings of the Royal Society of London, B 189, 391-413. Corner, E. D. S . and Davies, A. 0. (1971). Plankton as a factor in the nitrogen and phosphorus cycles in the sea. I n “Advances in Marine Biology”, Vol. 9 (F. S. Russell and C. M. Yonge, eds.), pp. 101-204. Academic Press, London and New York. Corner, E. D. S., Denton, E. J. and Forster, G. R. (1969). On the buoyancy of some deep-sea sharks. Proceedings of the Royal Society of London, B 171, 415-429.

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POLLUTION STUDIES WITH lMrlRINE PLAXKTON-I

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Corner, E. D. S., Head, R. N. and Kilvington, C. C. (1972). On the nutrition and metabolism of zooplankton. VIII. The grazing of Biddulphia cells by Calanus helgolandicw. Journal of the Marine Biological Association of the United Kingdom, 52, 847-861. Corner, E. D. S., Kilvington, C. C. and O’Hara, S. C. M. (1973). Qualitative studies on the metabolism of naphthalene in Maia spuinado (Herbst). Journal of the Marine Biological Association of tke United Kingdom, 53, 819-832. Corner, E. D. S., Harris, R. P., Whittle, K. J. and Mackie, P. R. (197th). Hydrocarbons in marine zooplankton and fish. I n “ Effects of Pollutants on Aquatic Organisms (A. P. M. Lockwood, ed.), pp. 71-105. Society for Experimental Biology Seminar Series, Vol. 2. Cambridge University Press. Corner, E. D. S., Harris, R. P., Kilvington, C. C. and O’Hara, S. C. M. (1976b). Petroleum compounds in the marine food web : short-term experiments on the fate of naphthalene in Calan.us. Journal of t?&eMarine Biological Association of the United Kingdom, 56, 121-133. Corner, E. D. S., Head, R. N., Kilvington, C. C. and Pennycuick, L. ( 1 9 7 6 ~ ) .On the nutrition and metabolism of zooplankton. X. Quantitative aspects of Calanus helgoladicus feeding as a carnivore. Journal of the Marine Biological Association of the United Kingdom, 56, 345-358. Cox, B. A., Anderson, J. W. and Parker, J. C. (1975). An experimental oil spill: the distribution of aromatic hydrocarbons in the water, sediment, and animal tissues within a shrimp pond. I n “ Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution ” held at San Francisco, California, March 25-27, pp. 607-612. American Petroleum Institute, Washington, D.C. Currier, H. B. (1951). Herbicidal properties of benzene and certain methyl derivatives. Hilgardia, 20, 383-406. Davavin, I. A., Mironov, 0. G. and Tsimbal, I. M. (1975). Influence of oil on nucleic acids of algae. Marine Pollution Bulletin, 6, 13-15. Davies, J. M. (1975). Energy flow through the benthos in a Scottish sea loch. Marine Biology, 31, 353-362. Di Salvo, L. H. and Guard, H. E. (1975). Hydrocarbons associated with suspended particulate matter in San Francisco Bay waters. I n “ Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution ” held at San Francisco, California, March 25-27, pp. 169-173. American Petroleum Institute, Washington, D.C. Donahue, W. H., Wang, R. T., Welch, M. and Nicol, J. A. C. (1977). Effects of water-soluble components of petroleum oils and aromatic hydrocarbons on barnacle larvae. Environmental Pollution, 13, 187-202. Duca, R. A., Quinn, J. G., Olney, C. E., Piotrowicz, S. R., Ray, B. J. and Wade, T. L. (1972). Enrichment of heavy metals and organic compounds in the surface microlayer of Narragansett Bay, Rhode Island. Science, New York, 176, 161-163. Dunstan. W. M., Atkinson, L. P. and Natoli, J. (1975). Stimulation and inhibition of phytoplankton growth by low molecular weight hydrocarbons. Marine Biology 31, 305-310. Ehrhardt, M. (1972). Petroleum hydrocarbons in oysters from Galveston Bay, Environmental Pollution, 3, 257-27 1. Elder, D. L. and Fowler, W. S. (1977). Polychlorinated biphenyls : penetration into the deep ocean by zooplankton faecal pellet transport. Science, New Yo&, 197, 459-461.

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E. D. 5. CORNER

Farrington, J. W. and Meyer, P. A. (1975). Hydrocarbons in the marine environment. I n “ Environmental Chemistry ”, Vol. 1, pp. 109-136. Chemical Society, London. Faux, A., Horn, D. H. S., Middleton, E. J., Fales, H. M. and Lowe, M. E. (1969). Moulting hormones of a crab during ecdysis. Chemical Communications, 175-176. Feuerstein, D. L. (1973). Input of petroleum to the marine environment directly from the atmosphere. I n “ Background Papers for a Workshop on Inputs, Fates, and Effects of Petroleum in the Marine Environment ”, Vol.1, pp. 31-38. National Academy of Sciences, Washington, D.C. Fisher, N. S. and Wurster, C. F. (1974). Impact of pollutants on plankton communities. Environmental Conservation, 1, 189-190. Fontaine, M., Lacaze, J.-C., Le Pemp, X. and Villedon De Nayde, 0. (1975). Des interactions entre pollutions thenniques et pollutions par hydrocarbures. I n ‘‘ IIes JournBes d’Etudes sur les Pollutions Marines ” (Monaco, 6-7 DBcembre 1974), pp. 115-122. Commission Internationale pour 1’Exploration Scientiflque de la Mer MBditerranBe, Monaco. Forns, J. M. (1977). The effects of crude oil on larvae of lobster Homarus americanus. I n “Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) ” held a t New Orleans, Louisiana, March 8-10 (A.P.I. Publication No. 4284), pp. 569-573. American Petroleum Institute, Washington, D.C. Forrester, W. D. (1971). Distribution of suspended oil particles following the grounding of the tanker Arrow. Journal of Marine Research, 29, 151-170. Frank, D. J., Sackett, W., Hall, R. and Fredericks, A. (1970). Methane, ethane, and propane concentrations in Gulf of Mexico. Bulletin of the American Association of Petroleum Geologists, 54, 1933-1938. Frankenfeld, J. W. (1973). Factors governing the fate of oil at sea; variations in the amounts and types of dissolved or dispersed materials during the weathering process. I n ‘‘ Proceedings of the Joint Conference on Prevention and Control of Oil Spills ” held at Washington, D.C., March 13-15, 1973, pp. 485-495. American Petroleum Institute, Washington, D.C. Freegarde, M., Hatchard, C. G. and Parker, C. A. (1971). Oil spilt a t sea: its identification, determination and ultimate fate. Laboratory Practice, 20, 35-40. Galtsoff, P. S., Prytherch, H. F., Smith, R. 0. and Koehring, V. (1935). Effects of crude oil pollution on oysters in Louisiana waters. Bulletin of the Bureau of Fisheries, Washington, 48, No. 18, 143-210. Garrett, W. D. (1967). The organic chemical composition of the ocean surface. Deep-Sea Research, 14, 221-227. Gibson, D. T. (1976). Microbial degradation of carcinogenic hydrocarbons and related compounds. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held a t Washington, D.C. 9-11 August, 1976, pp. 225-238. American Institute of Biological Sciences, Arlington, Virginia. Giger, W. and Blumer, M. (1974). Polycyclic aromatic hydrocarbons in the environment : isolation and characterization by chromatography, visible ultraviolet, and mass spectrometry. Analytical Chemistry, 46, 1663-1671.

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POLLUTION STUDIES WITH MARINE PLANKTON-I

371

Goad, L. J. (1976). The steroids of marine algae and invertebrate animals. I n “ Biochemical and Biophysical Perspectives in Marine Biology ”, Vol. 3 (D. C. Malins and J. R. Sargent, eds.), pp. 213-318. Academic Press, London and New York. Gordon, D. C., Jr. and Michalik, P. A. (1971). Concentration of Bunker C fuel oil in the waters of Chedabucto Bay, April 1971. Journal of the Fisheries Research Board of Canada, 28, 1912-1914. Gordon, D. C., Jr. and Prouse, N. J. (1973). The effects of three oils on marine phytoplankton photosynthesis. Marine Biology, 22, 329-333. Gordon, D. C., Jr., Keizer, P. D. and Prouse, N. J. (1973). Laboratory studies of the accommodation of some crude and residual fuel oils in sea water. Journal of the .Fisheries Research Board of Canada, 30, 1611-1618. Gordon, D. C., Jr., Keizer, P. D. and Dale, J. (1974). Estimates using fluorescence spectroscopy of the present state of petroleum hydrocarbon contamination in the water column of the Northwest Atlantic Ocean. Marine Chemistry, 2, 251-261, Great Britain. Department of the Environment Central Unit on Environmental Pollution (1976). “ The Separation of Oil from Water for North Sea Oil Operations ”, 29 pp. (Pollution Paper No. 6). Her Majesty’s Stationery Office, London. Grossling, B. F. (1976). A n estimate of the amounts of oil entering the oceans. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held at Washington, D.C., 9-11 August, 1976, pp. 6-36. American Institute of Biological Sciences, Arlington, Virginia. Gudin, C. and Harada, H. (1974). Presence de substances de type auxinique dans le petrole. Comptes rendus hebdomadaires des Sbances de l’Acadbmie des Sciences, Paris, 278, 1229-1231. Hardy, R., Mackie, P. R., Whittle, K. J.,McIntyre, A. D. and Blackman, R. A. A. (1977). Occurrence of hydrocarbons in the surface film, sub-surface water and sediment in the waters around the United Kingdom. Rapports et Procbs-verbaux des Rkunions, Conseil International pour I’Exploration de la Mer, 171, 61-65. Harris, R. P., Berdugo, V., Corner, E. D. S., Kilvington, C. C. and O’Hara, S. C. M. (1977a). Factors affecting the retention of a petroleum hydrocarbon by marine planktonic copepods. I n “ Proceedings of Symposium on the Fate and Effects of Petroleum Hydrocarbons in Marine Ecosystems and Organisms ”held at Seattle. 10-12 November, 1976. (D. Wolfe, ed.), pp. 286304. Pergamon Press, New York. Harris, R. P., Berdugo, V., O’Hara, S. C. M. and Corner, E. D. S. (1977b). Accumulation of 14C-1-naphthaleneby an oceanic and an estuarine copepod during long-term exposure to low-level concentrations. Marine Biology, 42, 187-195. Heller, J. H., Heller, M. S., Springer, S. and Clark, E. (1957). Squalene content of various shark livers. Nature, London, 179, 919-920. Hites, R. A. and Biemann, K. (1972). Water pollution: organic compounds in the Charles River, Boston. Science, New York, 178, 158-160.

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1.D. 9. OORNER

Hyland, J. L. and Schneider, E. D. (197ej. Petroleum hydrocarbons and their effects on marine organisms, populations, communities, and ecosystems. In " Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment ". Proceedings of the Symposium held at Washington, D.C., 9-11 August, 1976, pp. 464-506. American Institute of Biological Sciences, Arlington, Virginia. Hyland, J. L., Rogerson, P. F. and Gardner, G. R. (1977). A continuous flow bioassay system for the exposure of marine organisms to oil. I n " Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) " held a t New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 547-550. American Petroleum Institute, Washington, D.C. Iliffe, T. M. and Calder, J. A. (1974). Dissolved hydrocarbons in the eastern Gulf of Mexico Loop Current and the Caribbean Sea. Deep-sea Research, 21, 481-488. Jeffrey, L. M. (1970). Lipids in marine waters. In. " Symposium on Organic Matter in Natural Waters ", held at the University of Alaska, September 2-4, 1968 (D. W. Hood, ed.), pp. 55-76. Institute of Marine Science, Occasional Paper No. 1. University of Alaska. Johannessen, K. I. (1976). Effects of seawater extract of Ekofisk oil on hatching success of Barents Sea capelin. International Council for the Exploration of the Sea: Committee Meetings, Papers and Reports. Fisheries Improvement Committee, No. E : 29, 12 pp. Kauss, P. B. and Hutchinson, T. C. (1975). The effects of water-soluble petroleum components on the growth of Chlorella vulgaris Beijerinck. Environmental POllutiOn, 9, 157-1 74. Kauss, P., Hutchinson, T. C., Soto, C., Hellebust, J. and Griffiths, M. (1973). The toxicity of crude oil and its components to freshwater algae. In " Proceedings of the Joint Conference on Prevention and Control of Oil Spills "held at Washington, D.C., March 13-15,1973, pp. 703-714. American Petroleum Institute, Washington, D.C. Keizer, P. D. and Gordon, D. C. Jr. (1973). Detection of trace amounts of oil in sea water by fluorescence spectroscopy. Journal of the Fisheries Research Board of Canada, 30, 1039-1046. Kolattukudy, P. E. (1976). Biogenesis of nonisoprenoid aliphatic hydrocarbons. I n " Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment ''. Proceedings of the Symposium held at Washington, D.C., 9-11 August, 1976, pp. 121-158. American Institute of Biological Sciences, Arlington Virginia. Kontogiannis, J. E. and Barnett, C. J. (1973). The effect of oil pollution on survival of the tidal pool copepod, Tigriopus caliifornicus. Environmental Pollution, 4, 69-79. Koons, C. B. (1977). Distribution of volatile hydrocarbons in some Pacific Ocean waters. I n " Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) " held at New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 589-591. American Petroleum Institute, Washington, D.C. Koons, C. B. and Monaghan, P. H. (1976). Input of hydrocarbons fromseeps and recent biogenic sources. I n " Sources, Effects and Sinks of Hydrocarbons Proceedings of the Symposium held at in the Aquatic Environment Washington, D.C., 9-11 August, 1976, pp. 85-107. American Institute of Biological Sciences, Arlington, Virginia.

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POLLUTION STUDIES WITH MARINE PLANKTON-I

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Kiihnhold, W. W. (1972). The influence of crude oils on fish fry. I n “ Marine Pollution and Sea Life ”-Proceedings of F.A.O. Technical Conference held at Rome, 9-18 December, 1970 (M. Ruivo, ed.), pp. 315-318. Fishing News (Books) Ltd., London. Lacaze, J. C. (1969). Effets d‘une pollution du type “Torrey Canyon” sur l’algue unicellulaire marine Phaeodactylum tricornutum. Revue internationale d’Oc6anographiemddicale, 13-14, 157-179. Lacaze, J. C. (1974). Ecotoxicology of crude oils and the use of experimental marine ecosystems. Marine Pollution Bulletin, 5, 153-156. Lacaze, J. C. and Villedon de Naide, 0. (1976). Influence of illumination on phytotoxicity of crude oil. Marine Pollution Bulletin, 7, 73-76. Larson, R. A., Blankenship, D. W. and Hunt, L. L. (1976). Toxic hydroperoxides : photochemical formation from petroleum constituents. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held a t Washington, D.C., 9-11 August, 1976, pp. 299-308. American Institute of Biological Sciences, Arlington, Vir,aiia. Lasker, R. (1975). Field criteria for survival of anchovy larvae : the relation between inshore chlorophyll maximum layers and successful first feeding. Fishery Bulletin of the National Oceanic and Atmospheric Administration (United States) Wmhington, D.C., 73, 453-462. Ledet, E. J. and Laseter, J. L. (1974). Alkanes a t the air-sea interface from offshore Louisiana and Florida. Science, New York, 186,261-263. Lee, R. F. (1975). Fate of petroleum hydrocarbons in marine zooplankton. I n “ Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution ” held a t San Francisco, California, March 25-27, pp. 549-553. American Petroleum Institute, Washington, D.C. Lee, R. F. and Williams, P. M. (1974). Copepod “ slick ” in the Northwest Pacific Ocean. Naturwissenschaften, 61, 505-506. Lee, R. F. and Ryan, C. (1976). Biodegradation of petroleum hydrocarbons by marine microbes. I n “ Proceedings of the Third International Biodegmdation Symposium ” held at the University of Rhode Island, Kingston, Rhode Island, 17-23 August 1975. (J.M. Sharpley and A. M. Kaplan, eds), pp. 119125. Applied Science Publishers : London. Lee, R. F., Nevenzel, J. C., Paffenhofer, G.-A., Benson, A. A., Patton, S. and Kavanagh, T. E. (1970). A unique hexaene hydrocarbon from a diatom (Skeletonema costatum). Biochimka et biophysica Acta, 202, 386-388. Lee, R. F., Nevenzel, J. C. and Paffenhofer, G.-A. (1971). Importance of wax esters and other lipids in the marine food chain :phytoplankton and copepods. Marine Biology, 9, 99-108. Lee, R. F., Sauerheber, R. and Benson, A. A. (19724. Petroleum hydrocarbons : uptake and discharge by the marine mussel, Mytilus edulis. Science, New York, 177, 34P346. Lee, R. F., Sauerheber, R. and Dobbs, G. H. (1972b). Uptake, metabolism and discharge of polycyclic aromatic hydrocarbons by marine fish. Marine Biology, 17, 201-208. Lee, R. F., Nevenzel, J. C. and Lewis, A. G. (1974). Lipid changes during life cycle of marine copepod, Euchaetajaponica Marukawa. L+ids, 9,891-898. Lee, R. F., Ryan, C. and Neuhauser, M. L. (1976). Fate of petroleum hydrocarbons taken up from food and water by the blue crab Callinectes sapidw. Marine Biology, 37, 363-370.

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Lee, R. F., Takahashi, M., Beers, J. R., Thomas, W. H., Seibert, D. L. R., Koeller, P. and Green, D. R. (1977). Controlled ecosystems : their use in the study of the effects of petroleum hydrocarbons on plankton. I n “ Physiological Responses of Marine Biota to Pollutants ”, (F. J. Vernberg, A. Calabrese, F. P. Thurberg and W. B. Vernberg, eds), pp. 323-342. Academic Press, London and New York. Lee, W. Y. and Nicol, J. A. C. (1977). The effects of the water soluble fractions of no. 2 fuel oil on the survival and behaviour of coastal and oceanic zooplankton. Enwironmental Pollution, 12, 2 79-2 9 2. Levy, E. M. (1971). The presence of petroleum residues off the East Coast of Nova Scotia, in the Gulf of St. Lawrence, and the St. Lawrence River. Water Research, 5, 723-733. Levy, E. M. (1972). Evidence for the recovery of the waters off the East Coast of Nova Scotia from the effects of a major oil spill. Water, Air and Soil Pollution, 1, 144-148. Linden, 0. (1976). The influence of crude oil and mixtures of crude oil/dispersants on the ontogenic development of the Baltic herring, Clupea harengm membras L. Ambio, 5, 136-140. Lorenzen, C. J. (1967). Vertical distribution of chlorophyll and phaeo-pigments : Baja California. Deep-sea Research, 14, 735-745. Lu, A. Y . H., Kuntzman, R., West, S., Jacobson, M. and Conney, A. H. (1972). Reconstituted liver microsomal enzyme system that hydroxylates drugs, other foreign compounds, and endogenous substrates. 11. Role of the cytochrome P-450 and P-448 fractions in drug and steroid hydroxylations. The Journal of Biological Chemistry, 247, 1727-1734. Mackie, P. R., Platt, T. S. and Hardy, R. (1978). Hydrocarbons in the Marine Environment. 11. Distribution of n-alkanes in the fauna and environment of the sub-antarctic island of South Georgia. Estuarine and Coastal Marine Science, 6 , 301-313 Mallet, L. and Sardou, J. (1965). Recherche de la presence de l’hydrocarbure polybenzbnique benzo 3-4 pyrhne dans le milieu planctonique de la region de la Baie de Villefranche. I n “ Pollutions Marines par les Microorganismes et les Produits Petroliers ” (Symposium de Monaco, Avril 1964), pp. 331-334. Commission Internationale pour 1’Exploration Scientifique de la Mer Mbditerranbe, Monaco. Mallet, L., Perdriau, L.V. and Perdriau, S. (1963). Etendue des pollutions par les hydrocarbures polybenz6niques du type benzo-3,4-pyritne en Mer du Nord et dans 1’0cban Glacial Arctique. Bulletin de Z’Accz&mie nationale de Midicine, 147, 320-326. Marty, J. C. and Saliot, A. (1976). Hydrocarbons (normal alkanes) in the surface microlayer of seawater. Deep-sea Research, 23, 863-873. Mironov, 0. G. (1968). Hydrocarbon pollution of the sea and its influence on marine organisms. Helgolander Wissenschaftliche Meeresuntersmhungen, 17, 335-339. Mironov, 0. G. (1969). The effect of oil pollution upon some representatives of the Black Sea zooplankton. 2oologicheski.l: zhurnal, 48, 980-984. (In Russian.) Mironov, 0. G. (1972). Effect of oil pollution on flora and fauna of the Black Sea. I n “ Marine Pollution and Sea Life ”-Proceedings of F.A.O. Technical Conference held at Rome, 9-18 December 1970. (M. Ruivo, ed.), pp. 222-224. Fishing News (Books) Ltd., London.

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Mironov, 0. G. and Lanskaya, L. A. (1966). The influence of oil on the development of marine phytoplankton. Paper presented at the Second International Oceanographic Congress, Moscow, 1966, pp. 161-164. Moore, S. F. and Dwyer, R. L. (1974). Effects of oil on marine organisms: a critical assessment of published data. Water Research, 8, 819-827. Morris, B. F. and Butler, J. N. (1973). Petroleum residues in the Sargasso Sea and on Bermuda beaches. I n “ Proceedings of the Joint Conference on Prevention and Control of Oil Spills ” held a t Washington, D.C., March 13-15, 1973, pp. 521-529. American Petroleum Institute, Washington, D.C. Morris, B. F., Butler, J. N. and Zsolnay, A. (1975). Pelagic tar in the Mediterranean Sea, 1974-75. Environmental Conservation, 2, 275-281. Morris, R. J. (1974). Lipid composition of surface films and zooplankton from the Eastern Mediterranean. Marine Pollution Bulletin, 5 , 105-108. Morris, R. J. and Sargent, J. R. (1973). Studies on the lipid metabolism of some oceanic crustaceans. Marine Biology, 22, 77-83. Murray, J., Thomson, A. B., Stagg, A., Hardy, R., Whittle, K. J. and Mackie, P. R. (1977). On the origin of hydrocarbons in marine organisms. Rapports et Procks-verbeaux des Rdunions, Conseil International pour I’Exploration de la Mer, 171, 8 6 9 0 . Myers, E. P. and Gunnerson, C. G. (1976). “ Hydrocarbons in the Ocean ”. MESA Special Report, 42 pp. Marine Eco Systems Analysis Program : U.S. Department of Commerce National Oceanic and Atmospheric Administration Environmental Research Laboratories, Boulder, Colorado. Neff, J. M. and Anderson, J. W. (1975). Accumulation, release, and distribution of benzo[a]pyrene-C14in the clam Rangia cuneata. I n “ Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution ” held a t San Francisco, California, March 25-27, pp. 469-471. American Petroleum Institute, Washington, D.C. Nelson-Smith, A. (1970). The problem of oil pollution of the sea. I n “ Advances in Marine Biology ”, Vol. 8 (F. S. Russell and C. M. Yonge, eds.), pp. 215306. Academic Press, London and New York. Niaussat, P. (1970). Pollution, par biosynthdse “ in situ ” d’hydrocarbures canc&ig&nes, d’une biocohose lagunaire reproduction “ in vitro ” de ce ph6nomhne. Revue internationale d’Ocdanographie mkdicale, 17, 87-98. Nuzzi, R. (1973). Effects of water soluble extracts of oil on phytoplankton. I n “ Proceedings of the Joint Conference on Prevention and Control of Oil Spills ” held at Washington, D.C., March 13-15, 1973, pp. 809-813. American Petroleum Institute, Washington, D.C. O’Brien, P. Y. and Dixon, P. S. (1976). The effects of oils and oil components on algae: a review. Britkh Phycological Journal, 11, 115-142. Offenheimer, C. H., Gunkel, W. and Gassmann, G. (1977). Microorganisms and hydrocarbons in the North Sea during July-August 1975. I n “ Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) ” held a t New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 593-609. American Petroleum Institute, Washington, D.C. O’Hara, S. C. M., Corner, E. D. S. and Kilvington, C. C. (1978). On the nutrition and metabolism of zooplankton. XII. Measurements by radioimmunoassay of the levels of a steroid in Calanw. Journal of the Marine Biological ASSOciation of the United Kingdom, 58, 597-605. Ott, F. S., Harris, R. P. and O’Hara, S. C. M. (1978). Acute and sub-lethal toxicity of naphthalene and three methylated derivatives to an estuarine copepod, Eurytemora afJinis. Environmental Pollution (In press).

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Paffenhofer, G.-A. (1970). Cultivation of Calanwr helgolandicus under controlled conditions. Helgoliinder Wissenschaf~l~cheM e e r e s u ~ ~ r s ~ h u n g e20, n, 346-359. Paffenhofer, G.-A. (1971). Grazing and ingestion rates of nauplii, copepodids and adults of the marine planktonic copepod Calanus helgolandicus. Marine Biology, 11, 286-298. Pancirov, R. J. and Brown, R. A. (1975). Analytical methods for polynuclear aromatic hydrocarbons in crude oils, heating oils, and marine tissues. In “ Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution ” held at San Francisco, California, March 25-27, pp. 103-113. American Petroleum Institute, Washington, D .C. Parker, C. A., Freegarde, M. and Hatchard, C. G. (1971). The effect of some chemical and biological factors on the degradation of crude oil at sea. In “ Water Pollution by Oil ”-Proceedings of a Seminar held at Aviemore, Scotland, 4-8 May 1970. (P. Hepple ed.), pp. 237-244. Institute of Petroleum, London. Parker, P. L., Winters, K., Van Baalen, C., Batterton, J. C. and Scalan, R. S. (1976). Petroleum pollution : chemical characteristics and biological effects. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held a t Washington, D.C., 9-11 August, 1976, pp. 257-269. American Institute of Biological Sciences, Arlington, Virginia. Parsons, T. R., Stephens, K. and Strickland, J. D. H. (1961). On the chemical composition of eleven species of marine phytoplankters. Journal of the Fisheries Research Board of Canada, 18, 1001-1016. Parsons, T. R., Li, W. K. W. and Waters, R. (1975). Some preliminary observations on the enhancement of phytoplankton growth by low levels of mineral hydrocarbons. In “ Annual Report to the National Science FoundationInternational Decade of Ocean Exploration : Controlled Ecosystem Pollution Experiment (CEPEX), May l , 1974-May l , 1975”. Section 13. 16 pp. U.S.A. Pavletib, Z., Munjko, I., Jardas, J. and Matoricken, I. (1975). Quelques observations sur la pollution verticale de la mer par les huiles min6rales et les phenols dans 1’Adriatique centrale et m6ridionale faites 8, l’automne 19711972. I n “ IIes Journees d’Etudes sur les Pollutions Marines ”, (Monaco, 6-7 Decembre 1974), pp. 47-51. Commission Internationale pour 1’Exploration Seientifique de la Mer Mediterranee, Monaco. Payne, J. F. (1976). Field evaluation of benzopyrene hydroxylase induction as a monitor for marine petroleum pollution. Science, New York, 191, 945-946. Payne, J. F. and Penrose, W. R. (1975). Induction of aryl hydrocarbon (benzo[a]Bulletin of Environmental pyrene) hydroxylase in fish by petroleum. Conturnination and Toxicology, 14, 112-116. Percy, J. A. and Mullin, T. C. (1975). “ Effects of crude oils on Arctic marine Beaufort Sea Technical Report No. 11, 167 pp. Beaufort invertebrates Sea Project, Department of the Environment, Victoria, B.C. Philpot, R. M., James, M. 0. and Bend, J. R. (1976). Metabolism of benzo[a]pyrene and other xenobiotics by microsomal mixed-function oxidases in marine species. I n “ Sources, Effects and Sinks of Hydrocarbons in the Proceedings of the Symposium held at WashingAquatic Environment ton, D.C., 9-11 August, 1976, pp. 185-199. American Institute of Biological Sciences, Arlington, Virginia.

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Pingree, R. D., Holliga.n, P. M., Mardell, G. T. and Head, R. N. (1976). The influence of physical stability on spring, summer and autumn phytoplankton blooms in the Celtic Sea. Journal of the Marine Biological Association of the United Kingdom, 56, 845-873. Posthuma, J. (1977). The composition of petroleum. Paper presented a t International Council for the Exploration of the Sea Workshop on: Petroleum Hydrocarbons in the Marine Environmentheld at Aberdeen, 9-12 September 1975, pp. 7-16. Prouse, N. J., Gordon, D. C. Jr. and Keizer, P. D. (1976). Effects of low concentrations of oil accommodated in sea water on the growth of unialgal marine phytoplankton cultures. Journal of the Fisheries Research Board of Canada, 33, 810-818. Pulich, W. M. Jr., Winters, K. and Van Baalen, C. (1974). The effects of a No. 2 fuel oil and two crude oils on the growth and photosynt,hesis of microalgae. Marine Biology, 28, 87-94. Rice, S. D. (1973). Toxicity and avoidance tests with Prudhoe Bay oil and pink salmon fry. I n " Proceedings of the Joint Conference on Prevention and Control of Oil Spills " hcld at Washington, D.C., March 13-15, 1973, pp. 667-670. American Petroleurn Institute, Washington, D.C. Rice, S. D., Moles, D. A. and Short, J. W. (1975). The efYect of Prudhoe Bay crude oil on survival and growth of eggs, alevins, and fry of pink salmon, Oncorhynchus gorbuschia. I n " Proceedings of the 1975 Conference on Prevention and Control of Oil Pollution " held at San Francisco, California, March 25-27, pp. 503-507. American Petroleum Institute, Washington, D.C. Roubal, W. T., Bovee, D. H., Collier, T. K. and Stranahan, S. I. (1977a). Flowthrough system for chronic exposure of aquatic organisms to seawatersoluble hydrocarbons from crude oil : construction and applications. I n " Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) " held at New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 551-555. American Petroleum Institute, Washington, D.C. Roubal, W. T., Collier, T. K. and Malins, D. C. (197713). Accumulation and metabolism of carbon-14 labelled benzene, naphthalene and anthracene by young coho salmon (Oncorhynchus kisutch). Bulletin of Environmental Contamination and Toxicology. (In press.) Ryther, J. H. (1969). Photosynthesis and fish production in the sea. Science, New Pork, 166, 72-76. Sabo, D. J. and Stegeman, J. J. (1977). Some metabolic cffects of petroleum hydrocarbons in marine fish. I n " Physiological R,esponses of inarine Biota to Pollutants " (F. J. Vernberg, A. Calabrese, F. P. Thrirberg and W. B. Vernberg, eds.), pp. 279-287. Academic Press, New York, London. Sanborn, H. R. and Malins, D. C. (1977). Toxicity and metabolism of na.phthalene : a study with marine larval invertebrates. Proceeding8 of the Society for Experimental Biology and Medicine, 154, 151-155. Savidge, G. (1976). A preliminary study of the distribution of chlorophyll a in the vicinity of fronts in the Celtic and Western Irish Seas. Estuarine and Coastal Marine Science, 4, 617-625.

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Searl, T. D., Huffman, H. L. Jr. and Thomas, J. P. (1977). Extractable organics and nonvolatile hydrocarbons in New York Harbor Waters. I n “Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) ” held a t New Orleans, Louisiana, March 8-10 (A.P.I. Publication No. 4284), pp. 583-588. American Petroleum Institute, Washington, D.C. Seki, H., Tsuji, T. and Hattori, A. (1974). Effect of zooplankton grazing on the formation of the anoxic layer in Tokyo Bay. Estuarine and Coastal Marine Science, 2, 145-151. Sims, P., Grover, P. L., Swaisland, A., Pal, K. and Hewer, A. (1974). Metabolic activation of benzo[a]pyrene proceeds by a diol-epoxide. Nature, London, 252, 326-328. Singer, S. C. and Lee, R. F. (1977). Mixed function oxidase activity in blue crab, Callinectes sapidus : tissue distribution and correlation with changes during molting and development. Biological Bulletin. Marine Biological Laboiatory, Woods Hole, Massachusetts, 153, 377-386 Smith, P. V. Jr. (1954). Studies on origin of petroleum: occurrence of hydrocarbons in recent sediments. Bulletin of the American Association of Petroleum Geologists, 38, 377-404. Soto, C., Hellebust, J. A. and Hutchinson, T. C. (197%). Effect of naphthalene and aqueous crude oil extracts on the green flagellate Chlamydonaonas angulosa. 11. Photosynthesis and the uptake and release of naphthalene. Canadian Journal of Botany, 53, 118-126. Soto, C., Hellebust, J. A. and Hutchinson, T. C. (197513). The effects of aqueous extracts of crude oil and naphthalene on the physiology and morphology of a freshwater green alga. Verhandlungen der Intermtionalen Vereinigung f u r theoretische und angewandte Limnologie, 19, 2145-2154. Soto, C., Hellebust, J. A., Hutchinson, T. C. and Sawa, T. ( 1 9 7 5 ~ ) . Effect of naphthalene and aqueous crude oil extracts on the green flagellate Chlamydomonas angulosa. I. Growth. Canadian Journal of Botany, 53, 109-117. Spooner, M. F. and Corkett, C. J. (1974). A method for testing the toxicity of suspended oil droplets on planktonic copepods used a t Plymouth. In “ Ecological Aspects of Toxicity Testing of Oils and Dispersants ”Proceedings of a Workshop . . held at the Institute of Petroleum, London. (L. R. Beynon and E. B. Cowell, eds.), pp. 69-74. Applied Science Publishers, Barking, Essex. Stegeman, J. 5. and Sabo, D. J. (1976). Aspects of the effects of petroleum hydrocarbons on intermediary metabolism and xenobiotic metabolism in marine fish. I n “ Sources, Effects and Sinks of Hydrocarbons in the Aquatic Environment Proceedings of the Symposium held a t Washington, D.C., 0-1 1 August, 1976, pp. 424-436. American Institute of Biological Sciences, Arlington, Virginia. Stegeman, J. J. and Teal, J. M. (1973). Accumulation, release and retention of petroleum hydrocarbons by the oyster Crassostreavirginica. Marine Biology, 22, 37-44. Struhsaker, J. W., Eldridge, M. B. and Echeverria, T. (1974). Effects of benzene (a water-soluble component of crude oil) on eggs and larvae of Pacific herring and northern anchovy. I n “ Pollution and Physiology of Marine Organisms” (F. J. Vernberg and W. B. Vernberg, eds.), pp. 253-284. Academic Press, London and New York.

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Swinnerton, J. W. and Linnenbom, V. J. (1967). Gaseous hydrocarbons in sea water : determination. Science, New York, 156, 1119-1120. Takahashi, M., Thomas, W. H., Seibert, D. L. R., Beers, J., Koeller, P. and Parsons, T. R. (1976). The replication of biological events in enclosed water columns. Archiv filr Hydrobiologie, 76, 5-23. Tornabene, T. G., Kates, M. and Volcani, B. E. (1974). Sterols, aliphatic hydrocarbons, and fatty acids of a nonphotosynthetic diatom, Nitzachia alba. Lipids, 9, 279-284. Turner, J. T. (1977). Sinking rates of fecal pellets from the marine copepod Pontella meadii. Marine Biology, 40, 249-259. Van Baalen, C. (1968). The effects of ultraviolet irradiation on a coccoid bluegreen alga : survival, photosynthesis, and photoreactivation. Plant Physiology, Lancaster, 43, 1689-1695. Vandermeulen, J. H. and Ahern, T. P. (1976). Effect of petroleum hydrocarbons on algal physiology : review and progress report. I n “ Effects of Pollutants on Aquatic Organisms (A. P. M. Lockwood, ed.), pp. 107-125. Society for Experimental Biology Seminar Series, Vol. 2. Cambridge University Press. Van Overbeek, J. and Blondeau, R. (1954). Mode of action of phytotoxic oils. Weeds, 3, 55-65. Varanasi, U. and Malins, D. C. (1977). Metabolism of petroleum hydrocarbons : accumulation and biotransformation in marine organisms. I n “ Effects of Petroleum on Arctic and Subarctic Marine Environments and Organisms, Vol. I1 :Biological Effects ” (D. C. Malins, ed.), pp. 175-270. Academic Press, New York, London. Vaughan, B. E., ed. (1973). “ Effects of oil and chemically dispersed oil on selected marine biota-a laboratory study ”. Battelle Pacific Northwest Laboratories Report (Contract 2 12B00439) for American Petroleum Institute Committee on Environmental Affairs. (A.P.I. Publication No. 4191), 91 p. Washington, D.C. Wade, T. L. and Quinn, J. G. (1976). Hydrocarbons in the Sargasso Sea surface microlayer. Marine Pollution Bulletin, 6, 54-67. Walker, J. D., Colwell, R. R. and Petrakis, L. (1975). Degradation of petroleum by a.n alga, Prototheca zopfli. Applied Microbiology, 30, 79-81. Wells, P. G. and Sprague, J. B. (1976). Effects of crude oil on American lobster (Homarus americanus) larvae in the laboratory. Journal of the Fisheries Research Board of Canada, 33, 1604-1614. Whitlock, J. P. Jr. and Gelboin, H. V. (1974). Aryl hydrocarbon (benzo[a]pyrene) hydroxylase induction in rat liver cells in culture. Journal of Biological Chemistry, 249, 261G2623. Whittle, K. J., Mackie, P. R., Hardy, R. and McIntyre, A. D. (1973). A survey of hydrocarbons in Scottish coastal waters. International Council for the Exploration of the Sea: Committee Meetings, Papers and Reports. Fisheries Improvement Committee, No. E : 30, 9 pp. Whittle, K., Mackie, P. R. and Hardy, R. (1974). Hydrocarbons in the marine ecosystem. South African JournaZ of Science, 70, 141-144. Whittle, K. J.,Mackie, P. R., Hardy, R., McIntyre, A. D. and Blackman, R. A. A. (1977). The alkanes of marine organisms from the United Kingdom and surrounding waters. Rapports et Procks-verbeaux des R6unions, Conseil International pour 1’Explorationde la Mer, 171, 72-78.

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Wilson, R. D. (1973). Estimate of annual input of petroleum to the marine environment from natural marine seepage. I n “ Background Papers for a Workshop on Inputs, Fates, and Effects of Petroleum in the Marine Environment ”, Vol. 1, pp. 59-96. National Academy of Sciences, Washington, D.C. Winters, K. and Parker, P. L. (1977). Water soluble components of crude oils, fuel oils, and used crank-case oils. I n “ Proceedings of the 1977 Oil Spill Conference (Prevention, Behavior, Control, Cleanup) ” held a t New Orleans, Louisiana, March 8-10. (A.P.I. Publication No. 4284), pp. 579-581. American Petroleum Institute, Washington, D.C. Winters, K., O’Donnell, R., Batterton, J. C. and Van Baalen, C. (1976). Watersoluble components of four fuel oils : chemical characterization and effects on growth of microalgae. Marine Biology, 36, 269-276. Winters, K., Batterton, 5. C. and Van Baalen, C. (1977). Phenalen-1-one: occurrence in a fuel oil and toxicity to microalgae. Environmental Science and Technology, 11,270-272. Youngblood, W. W. and Blumer, M. (1973). Alkanes and alkenes in marine benthic algae. Marine Biology, 21, 163-172. Youngblood, W. W., Blumer, M., Guillard, R. L. and Fiore, F. (1971). Saturated and unsaturated hydrocarbons in marine benthic algae. Marine Biology, 8, 190-201. Zitko, V. and Carson, W. V. (1970). “ The Characterization of Petroleum Oils and their Determination in the Aquatic Environment ”, 29 pp. Technical Report No. 217 of Fisheries Research Board of Canada. Zsolnay, A. (1971). Preliminary study of dissolved hydrocarbons and hydrocarbons on particulate material in the GotIand Deep of the Baltic. Kieler Meeresforschungen, 27, 129-134. Zsolnay, A. (1973). The relative distribution of non-aromatic hydrocarbons in the Baltic in September 1971. Marine Chemistry, 1, 127-136.

Adv. mar. Biol., Vol. 16, 1978, pp. 381-508.

POLLUTION STUDIES WITH MARINE PLANKTON PART II.

HEAVY METALS

ANTHONY G. DAVIES The Laboratory, Marine Biological Association, Plymouth, England

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I. Introduction .. .. .. .. .. 11. The Turnover of Heavy Metals by Phytoplankton . . . . .. . . A. The Kinetics and Mechanism of Metal Uptake by Phytoplankton B. The Effect of the Chemical Form of a Metal upon its Uptake by Phytoplankton . . .. .. .. .. .. .. C. The Role of Phytoplankton in the Biogeochemistry of Heavy . . .. .. .. .. .. .. Metals in the Sea 111. Laboratory Studies of the Toxic Effects of Heavy Metals upon Phytoplankton .. .. .. .. .. .. .. .. .. A. T h e Effects on the Growth of Phytoplankton . . .. .. B. Synergism and Antagonism of Mixtures of Heavy Metals towards Phytoplankton . . .. .. . * .. .. C. The Nature of Metal Toxicity in Phytoplankton . .. .. IV. Studies of the Toxic Effects of Heavy Metals upon Natural Populations of Phytoplankton .. .. .. .. .. .. .. .. A. The Effects on Primary Production Rates . . .. .. .. B. The Effects in Large Volume Sea Water Enclosures .. .. V. Heavy Metal Concentrations in Natural Populations of Marine Phytoplankton .. .. .. .. .. .. .. .. VI. The Turnover of Heavy Metals by Zooplankton . . .. .. A. Studies of Metal Fluxes through Zooplankton .. .. .. B. Food and Water as Sources of Metals for Uptake by Zooplankton C. The Effect of the Chemical Form of a Metal upon its Uptake by Zooplankton . . .. .. .. .. .. .. D. The Role of Zooplankton in the Biogeochemistry of Heavy Metals in the sea . .. .. .. .. .. .. VII. Laboratory Studies of the Toxic Effects of Heavy Metals upon Zooplankton A. The Effects on the Metabolic Activity of Zooplankton . B. The Effects on the Feeding and Ingestion Rates of Zooplankton .. C. The Effects on the Growth and Development of Zooplankton D. The Effects on the Fecundity of Zooplankton . . .. .. E. The Effects on the Phototactic Response of Zooplankton . . . . F. The Effects on the Swimming Activity of Zooplankton . . .. 6. The Combined Effects of Heavy Metals and Additional Environmental Stress upon Zooplankton . . .. .. .. .. VIII. Studies of the Toxic Effects of Heavy Metals upon Natural Populations of Zooplankton in Large Volume Sea Water Enclosures . . .. .. IX. Heavy Metal Concentrations in Natural Populations of Marine Zooplankton

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I. INTRODUCTION That heavy metals are poisonous towards microscopic marine life has been recognized for most of the present century. As early as the 1920's, paints containing the oxides of toxic metals were being tested for their capacity to prevent the fouling of surfaces immersed in the sea by deterring the settlement of algal spores and the planktonic forms of certain animals, particularly barnacles (Orton, 1929-30), and the continued development of anti-fouling coatings has provided the stimulus for measuring the concentrations of heavy metals necessary to kill both planktonic animal larvae and permanent members of the zooplankton (Corner and Sparrow, 1956, 1957 ; Wisely and Blick, 1967). Attempts have also been made to control the damaging outbursts of dinoflagellates known as " red tides '' by adding copper sulphate or copper ores to the affected areas, but with little success (Rounsefell and Evans, 1958 ; Marvin, Lansford and Wheeler, 1961). Thus, until 10 or 15 years ago, interest had centred upon the lethal aspects of metal toxicity. Since then, however, there has been an increasing awareness that the insidious build-up of low-level concentrations of metals in coastal and estuarine sea areas receiving industrial effluents and sewage could be having a deleterious effect upon the growth and development of the plankton leading to a decrease in the productivity of these regions. Further, it has been realized that the uptake of metals by the plankton provides an entry into marine food chains, the higher trophic levels of which are often used for human consumption. This has given rise to the upsurge in research into the longer term sub-lethal aspects of metal toxicity towards marine plankton and the way in which metals are accumulated at the first and second trophic levels which forms the basis of this review. The metals considered have been restricted mainly, but not exclusively, to the ten which appear to be most poisonous to marine life: mercury, cadmium, silver, nickel, selenium, lead, copper, chromium, arsenic and zinc. They are listed here in the order of decreasing toxicity given by Ketchum, Zitko and Saward (1975) though this particular ranking could obviously be a matter of some debate.

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The geochemical aspects of heavy metals in the sea have provided the subjects for several recently published articles and will not, therefore, be considered again here; information on the inputs of heavy metals to the sea, both due to natural processes and to pollution, may be obtained from reviews by Turekian (1971) and Goldberg (1976), and the chemical behaviour of metals in sea water has been discussed by, amongst others, Dyrssen and Wedborg (1974) and Stumm and Brauner (1975).

I n this review, the interactions of metals with phytoplankton and with zooplankton have, for the most part, been dealt with separately though there is inevitably some degree of overlap, especially in the sections dealing with metal uptake by zooplankton from their food supply. The portions concerned with the plants and with the animals progress in a similar manner. They begin with an examination of the rates and mechanisms of uptake and loss of heavy metals, including a discussion of the influence of the plankton upon the biogeochemistry of metals in the sea, then the toxic effects of the metals are considered using data obtained both in the laboratory and with natural populations and, finally, the heavy metal concentrations in natural populations of plankton are discussed. We start with the phytoplankton.

11. THE TURNOVER OF HEAVY METALSBY PHYTOPLANKTON

A. Kinetics and mechanism of metal uptake by phytoplankton Phytoplankton populations present surprisingly large surface areas to the sea water or culture medium in which they are growing. Cells of the common diatom Skeletonema costatum (Greville) Cleve, for instance, have been found to have areas in the range 235-369 pm2 (Smayda, 1970) so that a million cells would have a total surface exceeding 2 cm2. Under natural conditions, a million diatoms per litre of sea water is by no means unusual (Bainbridge, 1957) and tkis number can be greatly exceeded during the spring bloom (e.g. Butler, Corner and Marshall, 1970), while in laboratory cultures populations of a million cells per ml are commonplace. It is not surprising, therefore, that adsorption onto the outsides of cells represents an important aspect of metal uptake by phytoplankton. Little information is available on the physico-chemical nature of the surfaces of marine phytoplankton except for the work of Myers, Iverson and Harriss (1975) who showed, using a microelectrophoretic technique, that three estuarine species Nannochloris oczdata Droop, Pavlova (Monochrysis) lutheri (Droop) Green and Cyclotella mene-

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ANTHONY 0.DAVIES

ghiniana Kutzing all had negatively charged surfaces due, it was suggested, to the ionization of groups in the polysaccharide-glycoprotein matrices exposed to the sea water. Davies, Haydon and Rideal (1956) had previously demonstrated that bacterial surfaces are also negatively charged ; by following the variation in surface charge with changing pH in the solution bathing the cells, they established that the acid dissociation constant for the groups on the surface (pK = 2-9) was about the same as that of acidic polysaccharides (pK = 2.95) suggesting that the surface charge was associated with carboxylic rather than phosphatidic groups which have pK values of about 1.8. Davies et al. (1956) also found that the charges on bacterial surfaces could be neutralized and even reversed by heavy metal ions such as copper or lead. Similar experiments have not been carried out with marine phytoplankton, but the decrease in the net charge on their surfaces caused by increasing salinity observed by Myers et al. (1975) was presumably the result of interaction of the cations in sea water with the negative groups on the surfaces. On this basis and by analogy with more intensely studied cellular systems, e.g. erythrocytes (Bangham, Pethica and Seaman, 1958 ; Passow, 1970), it is possible to arrive at a tentative picture of the physico-chemical nature of the surfaces of phytoplankton as consisting of a mosaic of interspersed cationic and anionic exchange sites, provided by carboxylic, sulphydryl, phosphatidic, amino and other groups, the net charge on the surface being related to the degree to which the sites are occupied by protons and the other ions present in sea water, i.e. being a function of the pH and salinity. The initial uptake of a positively charged heavy metal ion can then be envisaged as occurring by the displacement of the cations already occupying the binding sites, the amount of metal finally bound onto the surface a t equilibrium being determined by the relative affinities of the sites for the metal and the sea water cations and also the concentrations of each remaining in solution, in accord with the principles of ion-exchange. The metal, once bound on to the surface, would be suitably placed for being transported, actively or passively, through the diffusion barrier presented by the cell membrane into the cytoplasm. From measurements of the amounts of 2osHgtaken up by dividing, non-dividing and formalin-killed cultures of Chaetoceros costatum Pavillard in the light and the dark, Glooschenko (1969) concluded that mercury was accumulated passively, and more recent studies of the mechanism by which metals enter marine phytoplankton, namely those on zinc uptake by Pheodactylum tricornutum Bohlin (Davies, 1973), and of mercury uptake by Isochrysis galbana Parke and Duna-

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POLLUTION STUDIES WITH MARINE PLANKTON-11

ZieZZa lcrtiolecta Butcher (Davies, 1976), also support the idea of passive uptake. I n the latter experiments, cultures were grown in a chelatorfree culture medium of enriched natural sea water containing very little of the metals being studied so that the plant cells would initially be virtually devoid of them ; in the case of zinc, this necessitated passing the sea, water used in the medium through a column of a chelating ion-exchange resin in order to remove as much as possible of the metal naturally present. When the populations were dense enough to make the measurements feasible, the metal, labelled with a radioactive tracer, was added to give a suitable concentration, and its incorporation by the cells followed by filtering samples of the culture a t regular intervals to determine the

t

(min)

.\/I (mi")+

.,

FIO.I. The uptake of inorganic mercury by A, ~sochryeisgalbanu and B, Dunaliella tertiolecta in cultures containing cell populations having approximately the same total surface areas. Initial mercury concentrations (pg/l): 0 , 10 ; A,20 ; 60. During the period ikstrated, the mercury transported across unit area of surfacein the Isochryaie galbana cultures was substantially greater than that in the Dunaliella tertiolecta cultures due t o differences in the permeabilities of their membranes to the metal ions. I n both cases, the weight of mercury taken up was a linear function of the square root of the elapsed time [C, D) indicating that, in the early stages, uptake was diffusion controlled. (From Davies, 1976. Reproduced by kind permission of the Council of the Marine Biological Association of the United Kingdom.)

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ANTHONY Q. DAVIES

radioactivity in the cells. I n order that the data could be interpreted in terms of uptake by a fixed number of binding sites, it was necessary that any increase in population during the period of the measurements be as small as possible ;it was, therefore, necessary to limit the analysis of the results t o those obtained during the first few hours after the EDTA added

I

06

O

L

b

I

I

2

I

I

I

4

Time (hours)

FIG. 2. The measurement of the

" extracellularly " bound zinc in Phaeodactylum tricornutum. At zero time, EDTA was added to a zinc containing culture to give a concentration of 3 x 10-SM in the medium and the change in the cellular zinc content followed over the period shown. Two typical examples are illustrated. The initial rapid decrease, presumed t o be due to desorption of zinc from the extracellular binding sites on the cells, was followed by a slower loss of more firmly bound metal. (After Davies, 1973.)

metal addition. It was found that, both with zinc and mercury, the metal accumulated by the cells increased linearly with the square root of the time elapsed after the addition of the metal to the culture, a relationship characteristic of a diffusion-controlled process (Crank, 1970) ; the results for mercury uptake by the two flagellates-plotted

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in this instance as the weight of mercury which had been taken up across unit area of cell surface-are shown in Fig. 1. Zinc uptake by Phueodactylum tricornutum differed from that for mercury uptake by the flagellates in that when the linear plot of cellular zinc content against d(time) was extrapolated back to zero time, it had a positive intercept on the ordinate. Davies' (1973) interpreted this as indicating that, immediately upon addition of the zinc to the medium, rapid equilibration took place between the solution and the exchange sites, some of the metal becoming bound to the exterior surfaces of the cells, and this was followed by the slower movement of the zinc inwards into the cell cytoplasm. This view was supported by the finding that when the chelating agent, disodium ethylenediaminetetraacetate (EDTA) was added at a concentration of 3 x lO-3M to cultures containing zinc-laden cells, there was an almost instantaneous decrease in the metal content of the cells due, it was presumed, to desorption from the extracellular binding sites, and a subsequent much slower loss of intracellular zinc (Fig. 2). The extracellularly bound zinc determined in this way increased hyperbolically with the concentration of zinc in solution in the medium in accord with the Langmuir adsorption isotherm indicating that there was a fixed number of zinc binding sites available on the cell surfaces. By subtracting the surface bound zinc from the total cellular zinc content, the weight of metal which had been bound intracellularly could be calculated and this too increased linearly with d(tirne). Similar detcrminations of t,he easily removed mercury in Isochrysis galbana (Davies, 1974) and Dumliella tertiolecta (Davies, 1976) using thioglycollic acid as the chelating agont gave rather different results. I n the case of Isochrysis galbana, the amounts of mercury extracted from the cells were greater than those taken up during the period illustrated in Fig. 1 and tended to be larger in older cultures. This suggested that the thioglycollic acid treatment had removed more than just the extracellularly bound metal and Cossa (1976) has since observed much the same effect on adding the sulphur-containing amino acid cysteine to cultures of Phaeodactylum tricornutum containing cadmium. Very little of the cellular mercury could be removed from Dunffiliella tertiolecta, however, probably indicating that most of the metal taken up by this species is precipitated in a refractory form-possibly as the sulphide (Davies, 1976). As a result, it was not possible to determine the concentrations of surface bound mercury on the two flagellates in this way; but it was obvious from attempts to determine the mercury binding capacities of their surfaces using parachloro-mercuribenzene sulphonate (PCMBS)

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ANTHONY 0 . DAVIES

which has a very low membrane penetrability and attaches only to exposed sulphydryl groups (VanSteveninck, Weed and Rothstein, 1965) that there were very few mercury binding sites on the exteriors of the cells, as the concentrations of PCMBS bound by the surfaces were too small to be measured accurately (Davies, 1976). The relationship between the amount of material (Mt) taken up by diffusion across unit surface area of a solid initially free from the material and the elapsed time (t)is, during the early stages of uptake, Mt = 2C (Dt/.rr)*where C is the concentration of the diffusing substance adjacent to the surface of the solid and D is its diffusion coefficient within the solid (Crank, 1970). Davies (1973) showed that the slopes of the plots of the intracellular zinc content of Phaeodactylum tricornutum against z/(time) were in reasonable agreement with the idea that the metal adsorbed on to the exterior cell surfaces provided the driving concentration, C, for inward diffusion. As explained earlier, the values of C in the case of mercury were very low and could not be determined directly. Davies (1976) found that if the driving concentration, C, was assumed to be that of the mercury in the culture medium, the values obtained for the diffusion coefficients for entry of mercury into the cells were too large. As this could only have arisen if the assumed driving concentration was too small, it suggested that a build up in the mercury concentration at the surfaces of the cells, presumably by adsorption, must have taken place. I n support of this, it was, in fact, found that the relative slopes of the linear plots of mercury taken up against z/(time) (Fig. lC, D) could be interpreted by assuming that the driving concentrations of mercury on the cell surfaces (C) were related to those in solution [Hg] by the adsorption isotherm

c = Cmax [Hgl/(k + [Hgl), C,,,

being the saturation value for C when [Hg] is large and k, the half saturation constant. Thus the early stages of uptake of both zinc and mercury by initially metsl-free phytoplankton could be explained as being due to rapid adsorption of the metals on to externally exposed binding sites on the cell surfaces followed by passive, diffusion controlled transport into the cytoplasm at rates proportional to the concentration of surface bound metal. An interesting feature of the results shown in Fig. lA, B was that the membrane of Dunaliella tertiolecta was markedly less permeable to mercury than that of Isochrysis galbana, which took up almost four times more of the metal across unit area of surface in the period of the experiment than the former species, Sick and Windom (1975) have

POLLUTION STUDIES WITH MARINE PLA'NKTON-II

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similarly found that mercury uptake by Dunuliella tertiolecta occurs considerably more slowly than by Carteria sp. and Cylindrotheca closterium (Ehr.) Reimann & Lewin as Nitzschia closterium Ehr. and this is believed to explain, at least in part, why Dunaliella tertiolecta has a greater mercury tolerance than most other species (Davies, 1976). Cadmium uptake rates were considerably slower than those of mercury for the same species, the rate of uptake by Dunaliella tertiolecta being intermediate between that for Carteria sp. (the slowest) and Cylindrotheca (Nitzschia)closterium (Sick and Windom, 1975) suggesting that the mechanisms of uptake of mercury and cadmium are different. At first sight, it seems surprising that a metabolically essential metal like zinc should be taken up passively. The work of Davies (1973) showed that metabolic control of zinc uptake is probably exercised indirectly by variations in the protein content of the cytoplasm. By continuing the measurements of the zinc content of Phaeodactylurn tricornutum for several days after the initial period of frequent sampling, Davies (1973) estabhhed that the cellular zinc levels in the growing cultures reached a maximum 12-15 hours after the metal was originally added and then decreased steadily despite the availability of further zinc for uptake in the culture medium. Similar reductions in the zinc content of Phaeodactylum tricornutum cells in growing cultures have been observed by Hayward (1969) and Cossa (1976). Davies ascribed the decrease to a gradual reduction in the intracellular binding capacity of the cells as the population developed and it was suggested that, as most of the zinc taken up by the cells is probably bound to protein, this could have been related to the switch from protein t o carbohydrate and lipid production in the cells as the nitrate in the culture medium was used up (Hobson and Pariser, 1971) as has been observed in other species of phytoplankton (Thomas and Dumas, 1970; Berland, Bonin, Daumas, Laborde and Maestrini, 1970). Recently, Cossa (1976) made the interesting discovery that the cadmium contents of the cells in cultures of Phaeodactylum tricornutum continued to increase during the same period in which the zinc content was decreasing ; that this could have been due to competition between the two metals for the binding sites in the cells, the cadmium being taken up more slowly and displacing the already bound zinc, is supported by the fact that in the stationary phase when the cells would have ceased to manufacture protein, there was a substantial decrease in the cellular cadmium contents. The changes in cadmium concentrations in the cultures studied by Motohashi and Tsuchida (1974) indicate that uptake of the metal during the growth phase followed by loss in the stationary phase also occurs in Skeletonem cosjatum.

390

ANTHONY

a. DAVIES

On the basis of the observations described, Davies (1973) proposed that the mechanism of zinc accumulation was along the lines illustrated in Fig. 3 ; i t was suggested that the rate of zinc uptake intracellularly was proportional to the concentration gradient across the cell membrane established by the quantities of zinc, q, and q,, adsorbed on the outside and inside of the diffusion barrier. The value of qowould, in accordance

90 =-qmco Z I

n

C

I

a

Z

n I

Z I

n

C

I

a

Z

n I

kO+G

I Ionic zinc in intracellular fluid

11 lntracellular zincprotein complexes

I

FIa. 3. Preliminary model for zinc uptake by Phaeodactylurn trkornu$urn. Initially, the zinc is rapidly adsorbed on to the extracellular binding sites outside the diffusion barrier presented by the cell membrane, displacing other sea water cations already occupying the positions. The quantity of zinc taken up in this way, q,, is related to the concentration in the sea water, C,, by the isotherm shown. Transport across the membrane then takes place a t a rate, AP/dT, which is proportional to the concentration gradient created by the difference in the quantities of zinc bound to unit surface area of the outside (so) and the inside (q,) of the membrane, that on the inside rapidly attaining equilibrium with the concentration of intracellular, unbound metal ion, CI. Most of the intracellular zinc is bound on to protein, the maximal binding regulates the value of C1and, in turn, of q, thereby controlling capacity of which (QM) the rate of zinc uptake (or loss when q,, - q, is negative).

with the experimental results, be related to the concentration of zinc in solution in the culture medium (C,) and q,, to the concentration of zinc in solution in the intracellular fluid (GI), in both cases through a Langmuir type adsorption isotherm. Inside tho cells, most of the zinc (Q) was expected to be associated with the binding sites on the protein and this would, therefore, control the value of C, and in turn of q,, equilibration intracellularly being a.ssumed to be rapid relative to the

POLLUTION STUDIES WITH NAFtINE PLANKTON-I1

391

rate of zinc diffusion through the membrane. The concentration gradient governing the diffusion would thus be an indirect function of the protein content of the cytoplasm; while in the early stages of a culture when the protein in the celIs was high, it would cause an inward movement of zinc, the gradual decrease in the cellular protein as growth continued would eventually release sufficient of the metal into the intracellular fluid to cause a reversal of the concentration gradient (qo-qi) and diffusion of zinc out of the cells giving rise to the maximum observed in their zinc content. Davies (1973) showed that a computerized stepwise version of this model which incorporated the idea of the reducing cellular protein content predicted variations in the zinc levels in Phaeodactylum tricornutum which were in very good agreement with those which had been observed experimentally.

B. The effect of the chemical form of a metal upon its uptake by phytoptankton A metal in sea water may be present in particulate forms andin solution. The particulate forms result from either the precipitation of an insoluble compound of the element, e.g. chromium hydroxide or lead carbonate, or adsorption of the metal ions on to other particulate material such as clay minerals and organic detritus in the water (Stumm and Brauner, 1975). The extent to which metals in these particulate forms are taken up by phytoplankton is difficult to judge, but adhesion of iron hydroxide-or more precisely hydrous ferric oxide-on to their surfaces has been observed (Davies, 1967) and probably represents the initial stage in iron assimilation by marine phytoplankton in chelatorfree culture media and under natural conditions (Davies, 1970). Once in the vicinity of the cells, localized decreases in pH and the presence of reducing and chelating agents in the cell surfaces can be envisaged as mechanisms by which the particulate metals might be remobilized for inward transport. As metal hydroxides act as scavengers for other elements in sea water (e.g. Krauskopf, 11956 ; Ishibashi, Fujinaga, Kuwamoto, Sugibayashi, Sawamoto, Ogino and Murai, 1968) this process provides an additional mechanism by which toxic metals could enter phytoplankton. Due to the lack of thermodynamic data for metals in sea water, the chemical nature of the dissolved forms remains, to a large extent, a matter of conjecture because it usually involves the extrapolation of stability constants for metal complexing obtained in solutions of a different ionic strength and composition. Predictions of the ionic speciation of the metals therefore depend on what are considered to be

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ANTHONY Q. DAVIES

the most reliable values for the stability constants and, as a result, often differ from one set of calculations to another (Stumm and Brauner, 1975). However, from an assessment of calculated and experimental data taken from the literature, Stumm and Brauner (1975) have listed the main inorganic species probably present in sea water ;for the metals covered by this review, they are: mercury 11, HgCli-; cadmium 11, CdCli ; silver I, AgC1,; nickel 11, Ni2+, NiCO:(?) ; selenium IV, SeOg-; lead 11, PbCO:, Pb(CO,)$; copper 11, CuCO:, &OH+ ; chromium 111, Cr(OH):, VI, CrOZ-; arsenic V, HASO:-; zinc 11, ZnOH+, Zn2+, ZnCO:. Many of these elements will also be complexed to some extent by the organic matter dissolved in sea water. Evidence has already been obtained for the existence of organically-bound mercuxy (Fitzgerald and Lyons, 1973), copper (Williams, 1969; Foster and Morris, 1971) and zinc (Fukai, Huynh-Ngoc and Murray, 1973), and as the proportions of the metal complexed in this way have been found to vary on a seasonal basis (e.g. Foster and Morris, 1971; Fukai et al., 1973), it seems likely that the organic matter is released by phytoplankton. Precisely which chemical species enter the phytoplankton is not known, but there is quite a lot of evidence to suggest that organicallybound metals are not taken up. The extraction of zinc and mercury from phytoplankton by chelating agents was discussed in the previous section (see Fig. 2, p. 386) ; this happens because the formation of the chelates reduces the concentration of the inorganic forms of the metals in solution in the culture medium thereby reversing the concentration gradient across the cell membrane causing the metal to diffuse out from the cells. If the metal chelate itself was taken up, there would have been either no removal of metal if the chelate entered the cells rapidly, or a gradual rise in the cellular metal content after the initial decrease if it entered slowly; as is evident from Fig. 2, neither of these possibilities took place. Other indirect evidence that organic metal complexes are not assimilated by phytoplankton has been obtained by Myers et al. (1975) who found that the presence of humic acids decreased the amount of mercury taken up by Cyclotellamenenghiniana, and by Cossa (1976) who showed that Phaeodactylum tricornutum accumulated less cadmium in the presence of 35 pM EDTA than in its absence. The most direct evidence that organically bound metals are not taken up by phytoplankton has been provided by Sunda and Guillard (1976) who grew Thalassiosira pseudonana (Hustedt) Hasle & Heimdal in a culture medium containing copper and a. range of concentrations of the chelating agent TRIS (trishydroxymethylamino methane). The copper content of the cells in 3-4 day old cultures was

POLLUTION STUDIES WITH MARINE PLANKTON-II

393

shown to be hyperbolically related, not to the total copper concentration present in the medium, but to the activity ( = concentration x activity coefficient) of the unchelated cupric ion (Fig. 4) ; as Sunda and Guillard used the same culture medium for each total copper concentration, the activity coefficient of the ions can be regarded as having remained constant, so that, in effect, the cellular copper levels were controlled by the free cupric ion concentrations in the culture media. It should however be remembered that although, in principle, the

-log

(Icu

FIG.4. The cellular copper content in 3-4 day old cultures of Thalassk&rapseudonuna as a function of acu, the activity of the uncomplexed cupric ion in the medium. The activity was varied both by changing the total amount of copper present and by adding t,he following concentrations of TRIS (a11 in mM) : A,1 ; A, 2 ; 0 , 3 ; 0, 6. The curve was calculated from the isotherm: copper/cell = 4-8 X a,./(a,, (After Sunda and Guillard, 1976.)

+

relationship obtained experimentally-copper (fg at)/csll r= 4.8 x acu/(acu 10-9'2)-depends only upon the ionic strength of the culture medium, variations in the biochemical composition of the phytoplankton at different nutrient concentrations or salinities could cause changes in the values of the two constants outweighing any activity coefficient corrections necessary to allow for differences in ionic strengths.

+

304

ANTHONY 0.DAVIES

Thus only the inorganic ionic species and possibly only the free ions in the case of the electropositive elements are taken up by phytoplankton. As the equilibration between the free hydrated metal ions and the complex ions formed with the anions present in sea water generally takes place very rapidly (Wilkins, 1975), the rate of uptake of a metal, in the absence of organic complexing, is unlikely to be limited by the concentration in the medium of the ionic species which enters the plant cells, as redistribution will quickly occur to replace the depleted species; in these circumstances, the rate of uptake will, in fact, be proportional to the total concentration of metal in the medium. However, the rates of dissociation of metal-organic complexes are often quite slow and if a large part of a metal is bound by an organic compound, its rate of uptake would, once the inorganic species have been incorporated, be limited by the rate at which the metal is released from the complex. This effect can be very important in studies of metal uptake using radioactive tracers, for if the label does not quickly exchange with the other chemical forms of metal present in the water due to the slow dissociation of the organically bound fraction, it could be taken up preferentially by remaining in the inorganic form. This situation appears to have occurred in the experiments of Bernhard and Zattera (1969) who were studying zinc uptake by Phaeodactytum tricornutum. They found that the s6Zn tracer added in an inorganic form was accumulated by the plant cells disproportionately to the stable zinc present in the sea water which formed the basis of the culture medium, and this resulted in a higher specific activity of 66Zn in the phytoplankton than in the water. Later work showed that, even after a year, ionic zinc added to sea water had not equilibrated with the naturally present metal (Piro, Bernhard, Branica and Verzi, 1973).

The presence in culture media or the sea of organic compounds which form complexes with heavy metals will thus not only compete for and thereby reduce the amount of the metals which are taken up by phytoplankton but will also decrease their rate of incorporation into the plant cells, both processes providing a degree of protection against the toxic effects of the metals. I n view of this, the amounts of metals taken up by phytoplankton from culture media containing high concentrations of chelating agents and heavy metals (e.g. Hayward, 1969; Riley and Roth, 1971) would be expected to bear little relation to the levels of metals present in natural populations. Surprisingly, many of the data obtained by Riley and Roth (1971) for the metal contents of cultured phytoplankton lie within the ranges found for metal levels in natural phytoplankton

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from different sea areas (Appendix 11),though as there appears to be a correlation between the amount of metal taken up by the cells and the concentration initially added to the culture medium (Riley and Roth, 1971), the agreement must, to a large extent, be fortuitous.

C. T h e role of phytoplankton in the biogeochemistry of heavy metals in the sea Because phytoplankton cells accumulate heavy metals, it would seem reasonable to expect,that there should be an inverse correlationpossibly on a seasonal basis-between primary production and the concentrations of the metals present in dissolved form in sea water. Further, the eventual sedimentation of dead or senescent plant cells provides a mechanism by which heavy metals introduced to the surface layers of the sea might be transferred to the bottom sediments. Direct evidence of the influence of phytoplankton on the geochemical cycling of heavy metals in the sea has proved difficult to obtain. Although Morris (1971) found that changes in the particulate concentrations of copper and manganese in the Menai Straits, North Wales, reached maxima which coincided with peaks in the flagellate counts during a bloom of Phaeocystis sp., and there was an inverse relationship between the dissolved and particulate levels of manganese, there was little sign that the Phaeocystis had had a.ny effect upon the concentrations of dissolved copper ; this finding was later confirmed by Foster and Morris (1971) who, over a two year period, were unable to detect any changes in the copper concentration in the same area which correlated with the seasonal growth of phytoplankton. Particulate copper concentrations in the sea off Florida had previously proved to be unrelated to chlorophyll a levels (Alexander and Corcoran, 1967) and Spencer and Brewer (1969) had failed to f h d any variations in the concentrations of copper, zinc or nickel in the Gulf of Maine or the Sargasso Sea which could be attributed to biological effects even though it was estimated on the basis of the phosphate utilization in the top 50 m and using a value of 6.5 x 10-3 for the copper/phosphorus ratio in phytoplankton, t h a t the copper should have been depleted by about 18% if all of the metal removed by the plant cells sedimented out and none was regenerated in the upper layers. Knauer and Martin (1973) used primary production rates and metallcarbon ratios obtained from their own data to estimate the amounts of cadmium, zinc, copper and manganese likely to be taken up by the phytoplankton population in Monterey Bay, California. They found that only in the case of cadmium would the quantity of metal accumulated by the plankton be

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ANTHONY U. DAVIES

large enough to produce a detectable change in its concentration, and the observed reduction in the dissolved cadmium was in quite good agreement with the calculated decrease. The validity of such calculations is strengthened by recent evidence on copper uptake by phytoplankton communities entrapped in large volume water enclosures where the sinking of phytoplankton occurs unhindered by turbulent mixing (Thomas and Seibert, 1977) for Topping and Windom (1977) showed that, up to values of about 10 mg carbon/m3/h, there is an approximately linear relationship between the rates of primary production and the rate of copper sedimentation in sinking plant cells. It seems likely, in fact, that it will be impossible to demonstrate directly variations in metal concentrations in the sea due to their uptake by phytoplankton until the methods for measuring metals in sea water are improved; for, as Topping (1974) has pointed out, the biologically induced variations in dissolved metal levels in oceanic or coastal areas of low to medium productivity are, in most cases, smaller than the errors associated with the analytical methods and are, therefore, unlikely to be detected. Indirect evidence of the influence of phytoplankton upon metal concentrations is now beginning to be provided by the findings that linear correlations exist between the concentrations of certain metals and the major nutrient ions, nitrate and phosphate. Boyle and Edmond (1975) took samples across the circumpolar current south of New Zealand where there were large horizontal gradients in the chemical properties of the water and found that the copper concentrations were directly proportional to the nitrate ion concentrations. Similarly, Martin, Bruland and Broenkow (1976) showed that, in the waters off Baja California, where upwelling causes sharp gradients in the concentrations of nutrients in the surface layers, cadmium concentrations were linearly related both to nitrate and to phosphate levels. Boyle and Edmond (1975) have rightly stated that such results do not in themselves establish a biological cause for the relationships between heavy metal and nutrient concentrations, but that this is a satisfactory explanation for them is supported by the finding that there was also it significant correlation (coefficient = 0.71) between the amounts of cadmium and phosphorus in the phytoplankton collected off Baja and analysed by Martin et al. (1976) indicating that the two elements were always taken up in a constant ratio (Cd/P = 4.9 x by atoms). The interpretation of trace element concentrations and ratios in particulate materials is increasingly yielding useful information about the biogeochemical cycling of heavy metals in the sea. Spencer and Sachs (1970) showed that, while zinc and copper concentrations in

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397

suspended material collected from deep water (> 150 m) in the Gulf of Maine were both linearly correlated with the aluminium content of the solids, the levels of both heavy metals in the particulates present in the surface waters were considerably higher than would have been expected on the basis of their aluminium content. Spencer and Sachs attributed these elevated zinc and copper contents to biological accumulation and pointed out that the much lower concentrations present in the suspended matter in the deep water implied that a large proportion of the heavy metals taken up by the solids in the surface layers must have been released back into solution as the particulates sank through the water column. The importance of biological accumulation in the recycling of heavy metals seems to depend on the particular element, however, for Chester and Stoner (1975) concluded from a study of trace element concentrations in suspended matter gathered from the surface waters of several oceans that, while lead and zinc are probably associated mainly with plankton, copper is distributed between plankton and other particulate materials. An interesting suggestion about the way in which phytoplankton might modify the geochemical cycling of heavy metals in certain locations was put forward by Schutz and Turekian (1965). It was found that the waters in upwelling areas tended to have higher than normal concentrations of the metals silver, cobalt and nickel and that, furthermore, the concentrations generally increased with depth. Schutz and Turekian postulated that the localized build-up of metals could be due to the metals being removed from the newly upwelled water by the abundance of phytoplankton resulting from the high nutrient levels and then carried downwards in the sinking plant cells, the metal-depleted water moving away from the area. As the cells sink downwards through the water, it was suggested that regeneration of the metals would elevate their concentrations in the inflowing subsurface currents which provide the source of the upwelling water ; thus, over a long period of time, there would be a gradual increase in the metal concentrations present in the water coming to the surface until, finally, a steady state situation would be established. The absence of normal mixing processes would, as a result, produce extended residence times for the metals in such situations. An increase in cadmium levels which might have resulted from such a process has been observed in the upwelling area off Baja California for concentrations of the metal in phytoplankton collected from the area were substantially higher than in oceanic plankton (Appendix 11) (Martin and Broenkow, 1975; Martin et al., 1976). Martin et al. (1976) found that the cadmium concentrations in the water

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ANTHONY Q. DAVIES

increased rapidly with depth from about 7ng/l at 25 m to almost 70 ng/l at 80 m ;similar increases occurred in the nitrate and phosphate concentrations as expected in upwelling situations. The observations support the view that heavy metals contained in sinking phytoplankton are returned to the water before t,he decaying cells reach the bottom and, 5 ~ sSchutz and Turekian (1965) have stated, there appears to be no evidence to suggest " that significant quantities of these elements are permanently removed as deep sea sediment by this mechanism ",

111. LABORATORY STUDIESOF THE TOXIC EFFECTS OF HEAVY METALS UPON PHYTOPLANKTON A. The effects on the growth ofphtJoplankton Numerous studies have been made of the effects of heavy metals upon the growth and metabolism of laboratory cultures of marine phytoplankton and most of the results currently available are summarized in Appendix I. An immediately noticeable feature of the data is the remarkable diversity of techniques and culture conditions used to obtain them; the phytoplankton cells have been grown in batch cultures-the most usual experimental method, continuous cultures (Rice, Leighty and McLeod, 1973 ; Kayser, 1976 ; BentleyMowat and Reid, 1977) and dialysis cultures (Jensen, Rystad and Melsom, 1974, 1976) under various conditions of temperature and illumination ; culture media have ranged from unenriched natural sea water (Erickson, 1972; Rice et al., 1973; Jensen et al., 1974, 1976) to artificial sea water containing high concentrations of nutrients and chelating agents (Nuzzi, 197.2)) and the parameters used as indices of the toxicities of the metals have included changes in growth rate (e.g. Davies, 1974; Sunda and Guillard, 1976)) the cell population after a certain period of growth (Erickson, Lackie and Maloney, 1970; Rosko and Rachlin, 1975) and effects on the rate of photosynthesis measured by 14CO, utilization (Tkachenko, Mortina and Lukankina, 1974; Zingmark and Miller, 1975) or oxygen production (Saraiva, 1973; Overnell, 1976). Each of the culture methods has advantages and disadvantages which should be borne in mind when considering the data obtained with them. Batch cultures are straightforward to set up and allow the simultaneous study of the effect of a wide range of metal concentrations upon cell populations all taken from the same stock culture and, therefore, initially in the same physiological condition ; however, the growth of the phytoplankton in batch cultures causes changes in the

POLLUTION STUDIES WITH MARINE PLANKTON-=

399

culture medium, and variations in the biochemical composition of the plant cells due to depletion of the nutrients, and the possible modification of the chemical form of the metal by the extracellular products could both influence the outcome of the experiments. Continuous cultures have the advantage of allowing the establishment of a physiologically stable population growing in an unchanging environment so that the effects of metals can be studied both in the short term as a single pulse of contamination, or in the longer term, by continued exposure to a chosen concentration of a metal ; to examine the effects of a range of metal concentrations takes a considerable time, however, due to the necessity of allowing the chomostat to reequilibrate when the concentration is changed and a further disadvantage of continuous cultures is that, for many metals, i t is not possible to use chelator-free culture media as adsorption of the metals would contaminate the complex system of pipe work associated with the apparatus. Dialysis cultures are useful as they permit the study, over an extended period, of the growth of a unialgal culture in natural unenriched sea water. The nutrient levels are maintained by inward diffusion from the water surrounding the membrane containing the culture and the build-up of extracellular products avoided as they diffuse out of the culture. Jensen et al. (1974, 1976) have demonstrated that it is possible to maintain a constant metal concentration in such cultures by the continuous feed of a stock solution. The main disadvantage of dialysis cultures is, as in batch cultures, that as the population increases, it eventually outstrips the nutrient supply and this is likely to cause changes in the biochemical status of the cells and hence their susceptibility to metals. Because of the wide range of techniques and conditions used, analysis of the data in Appendix I poses many problems. It is certainly impossible to extrapolate the results from most of the experiments to predict the effects of metals upon phytoplankton in the sea for, whereas the cultures have usually contained high densities of cells growing in nutrient-rich media under near optimal conditions, natural populations of phytoplankton are normally of low density and their growth, often in nutrient-depleted water, is regulated by a complex pattern of interacting and ever changing chemical, physical and meteorological factors. The experimental data summarized in Appendix I do, however, point to the factors which influence the toxicity of heavy metals towards phytoplankton; the most important ones appear to be: (1) the phytoplankton species, (2) the composition of the sea water supporting the plankton, (3) the cell population, (4) changes in the

400

ANTHONY

a. DAVIES

metal tolerance of the cells or in the chemical state of the metal during the period of growth and, of course, ( 5 ) the concentration of the metal. Although temperature and the level of illumination are also likely to influence the effects of metals because they regulate the metabolic activity of phytoplankton, they have been insufficiently investigated to warrant discussion a t the present time. 1. Metal toxicity in relation to the species of the phytoplankton Where workers have used the same culturing conditions and medium to study the effect of one metal upon a range of species of phytoplankton, it is possible to place the species in order of their metal tolerance ; examples for mercury and copper are given in Table I. In comparing the susceptibilities of different species to the metals, it must be remembered that because the measurements have been made in a standardized way, some species may have been growing under suboptimal conditions and may, therefore, have been less resistant to the effects of the metals than they might have been in a more favourable situation. The order of metal tolerance in Table I confirms previous experience with anti-fouling paints that green algae are generally more mercury tolerant than brown algae and diatoms, and that the situation, although less clear-cut, is largely reversed with copper (G. T. Boalch, private communication). Dunaliella tertiolecta is the exception to this being extremely resistant both t o mercury (Davies, 1976) and copper (Mandelli, 1969). It is significant, in this respect, that this species releases hydrogen sulphide during growth (Craigie, McLachlan, Majek, Ackman and Tocher, 1966) though Davies (1976), who found that the concentrations of sulphide which appeared in cultures of Dunaliella tertiolecta were insufficient to remove the mercury by precipitation until the populations were entering the stationary growth phase, has suggested that the metals are probably detoxicated intracellularly by the formation of their highly insoluble sulphides. It is interesting that while the production of dimethyl-@-propiothetin by Tetraselmis spp. (Craigie et al., 1966) seems t o confer mercury but not copper tolerance upon this genus, with Phaeodactylum tricornutum which is believed to produce dimethyl sulphide (Armstrong and Boalch, 1960), the reverse is true. The biochemical basis of heavy metal tolerance is, in general, very poorly understood. 2. Metal toxicity in relation to the composition of the culture medium The addition of soil extracts or synthetic chelating agents to marine culture media in order to maintain the availability of essential trace

TABLEI. ORDER OF HEAVY METALTOLERANCE IN PWTOPLANKTON SPECIES

t

K

~

1~ n d e ofz tolerance 01

Species in order of increasing metal tolerance

Reference

A. MERCURY Growth Isochry& galbana < Skeletonema costatum < Phaeodactylum tricornutum Dunaliella tertiolecta Oxygen evolution Pavlova (Monochrysis) lulheri (Droop) Green < Phaeodactylum tricornutum = Skeletonema coatatum = Attheya decora West < Brachiomonaa aubmarina Bohlin < Dunaliella tertiolecta Growth G y m d i n i u m splendens Lebour < Scrippsiella faeroeme (Paulsen) Balech & Soares < Prorocentrum micana Ehrenberg Growth Lauderia borealis Gran < Skeletonema wstatum < Thalmawairapaeudonana = Exuviaella mariaelebouriae Parke & Ballantinen < Amphidinium carterae Hulburt < Pavlova pinguia Green = Chaetoceros didymzrs Ehrenberg < Chlamydomonm palla Butcher = Fragillaria p h n a t a Ehrenberg = Phueodactylum triwrnnutum < Pavlova (Monochrysis)lutheri = Monallantw salina Bourelly = Porphyridium marinum Kylin < Prasinockadua marinua (Cienk.) Waern < Cylindrothecu closterium (Ehrenb.) Reimann & Lewin < Heterothrix sp. = Cryptomonas paeudobdtica Butcher < Tetraaelmis atriata Butcher

<

B. COPPER Coccachloris elabena (Breb.) Dr. & D. < Exuviaella ~ p <. alenodinium ~ foliaceum Stein < Cylindrothem (Nitzachia) doaterium < Skeletonema costaturn < Thulaaaioaira jluviatilia Hustedt < Thalaaaioaira paeudonana (Cyclotella nana) Dunaliella tertiolecta Growth O&9thdkCU8 luteua N. Carter < Iaochryaia gdbana < Amphidinium carteme < Thalaaaiosira paeudonanh (Cyclotella nana) < Skeletonema wstatum < Dundiella tertwlecta Oxygen evolution Brachhaonaa submarina < Skeletonema wstutum < Attheya dewra = Dunaliella tertwlecta < Pavlova ( M o n o c h r y k ) lutheri < Phaeodactylum tricornutum Growth Exuviaella mariae bbouriae" < Porphyridium marinum < Thulaasioaira paeudonana < Amphidinium carterm = Chaetoceros didymua < Lauderia borealis < Chlumydomonas palla < Prasinocladua marinua < Pavlova (Monochryma)lnutheri < Tetraaelmis atriata = Skeletonema coatatum = Cryptomonaa paeudobaltiea < Cylindrothecu closterium < Monallantua salina < Fragilarh pinnatu < Pavlova pinguis = Phaeodactylum tricornutum = Heterothrix sp.

Growth

<<

0-0

a. Now in Proorcentrum minimum (Pav.) J . Schiller. b. Now in Prorocentrum sp.

Davies (1974) Overnell (1976) Kayser (1976) Berland et al. (1976)

-

w

0

23

0

w

3E I

3 Mandelli (1969)

1 !M4

Eriokson et d. (1970)

Overnell (1976) Berland et al. (1976)

ii

T3 8 I

I3

rp

E

402

ANTHONY

a. DAVIES

metals-especially iron-by keeping them in solution is now routine, as is the use of organic pH buffers such as TRIS and glycylglycine which are also metal chelators (McLachlan, 1973). Most of the data given in Appendix I were obtained using culture media containing such compounds. The calculations of Spencer (1958) indicate that even at EDTA concentrations as low as 10 pM-the order of magnitude of that present, for example in the much-used f/2 culture medium of Guillard and Ryther (1962)-metals like copper and zinc are present largely in chelated form. The data of Sunda and Guillard (1976) similarly show that in a culture medium containing 2 mM TRIS, as used by Berland, Bonin, Kapkov, Maestrini and Arlhac (1976), only about 0.03y0 of the copper added to give a concentration of 1000 pg/l would be present as uncomplexed cupric ion (assuming an activity coefficient of 0.2). In view of the evidence discussed earlier (Section 11, B) that the quantity of a metal taken up by phytoplankton is related to the concentration of unchelated metal in the culture medium, the earlier observations that the toxicity of, for example, copper is moderated by the presence of chelating agents (Steemann-Nielsen and WiumAnderson, 1970; Davey, Morgan and Erickson, 1973) can be explained as being due to the lower cellular burdens of the metal accumulated by the phytoplankton in the presence of the chelators. The toxicity of a metal, determined a t a certain t o t d concentration in a culture medium, will therefore bear little relation to the effects caused by the same concentration of the metal in natural waters having considerably lower levels of chelating materials, and it is significant that the lowest inhibitory metal concentrations have been observed in cultures using either natural sea water or enriched sea water containing no added chelating agents. Another aspect of the composition of culture media used to study the effects of heavy metals upon marine phytoplankton which is often overlooked is the high concentrations of the nutrient ions, nitrate and phosphate, which are frequently present. Hannan and Patouillet (1972) have, in fact, suggested that the toxicity of metals towards phytoplankton may be inversely related to the available nutrient levels but as, in their experiments, t h e concentrations were varied by dilution of the culture medium with 3.5% sodium chloride, the concomitant changes in the cation balance may also have influenced their results. On the other hand, although it is not feasible to carry out extended growth studies at the nutrient levels normally present in sea water, the enrichments made to culture media have often been considerably in excess of those necessary to allow the cultures to develop for the

POLLUTION STUDLES WITH MARINE PLANETON-II

403

minimal period required-in batch cultures, 1 to 3 weeks depending on the growth rate of the plankton-to determine the effects of heavy metal additions. By keeping the nutrient levels as low as possible, the density of the cell populations is reduced and, as will be seen in the next section, this allows a more realistic appraisal of the toxicity of a metal to be made. 3. Metal toxicity in relation to the cell population Whilst it is possible to determine the effect of a heavy metal upon the growth of unialgal cultures containing cell populations similar in numbers to thoso which occur naturally by measuring their photosynthetic rates over short periods using 14C0, (e.g. Zingmark and Miller, 1975), in order to study the effects on the growth and physiology of cultures over extended periods, it is necessary to use cell densities which are considerably higher than those normally present in the sea in order to facilitate measurements of the changes in the population with time. However, as can be seen in Appcndix I, it has not been uncommon for extremely high cell densities to be used and this can give rise to an underestimate of the toxicity of a metal when determined as the concentration which must be added to the culture to cause growth inhibition. The reason for this lies in the fact that i t is the quantity of metal taken up by the cells which ultimately determines its effect upon their growth and, in dense cultures, more metal is required to produce a given cellular burden. The effect of different populations upon the amount of metal accumulated by individual cells may be illustrated by assuming that metals are bound by the cells in accord with the Langmuir adsorption isotherm qmc cl= k+c where g = weight of metal bound per cell, q, = maximal value of q when c, the concentration of metal in solution in equilibrium with the cells, is very large and k is the half-saturation constant. (Sunda and Guillard (1976) have shown that copper uptake by Thalassiosira pseudonana can be described by an expression of this form.) For the purpose of these calculations, qm is put equal to 100 fg/cell, as Davies (1976) found that the maximal mercury binding capacity of Isochrysis galbana was about 300fg/cell, and the data of Sunda and Guillard (1976) indicate that Thalassiosira pseudonanu can accommodate up to 30 fg copper/cell ; and the half-saturation constant k is given the value of 2 pg/l, the figure for any particular phytoplankton species depending

404

ANTHONY Q. DAVIES

upon the affinity of its binding sites for the metal. The values of q and c for a total metal concentration, T, may then be calculated by solving the equation

for different values of N, the cell population in numbers/ml. The results for two values of T are given in Table 11. The actual values obtained for q and c depend, of course, upon the constants q, and k but it can be seen that whereas, up to densities of lo4 cells/ml, the metal burdens of the cells would be little affected by changes in population, a t higher TABLE 11. AN ILLUSTRATION OF THE EFFECT OF PHYTOPLANKTON POPULATION DENSITY UPON THE METAL BURDEN ACCUMULATED BY INDIVIDUAL CELLS

N

T

0*060 5.0

T: q: c: N:

108

2-4 ca. 0.050

108 2.3 0.048 71 4.93

104

106

1.6

0.42 0.008 38 1-22

0.033

68 4-32

106

0.06 ca. 0 4.9 0.10

Total metal concentration (pg/l). Metal burden of each cell (fglcell). Concentration of metal in solution at equilibrium (pg/l). Population density (number of cells/ml).

cell numbers, because most of the metal is contained by the cells, the cellular metal content becomes approximately inversely proportional to the population density. Thus, if the response of the cells to the metal is a function of the amount they have taken up, there would be an apparent inverse relationship between the toxicity of the metal and the cell numbers. Although this effect has not been demonstrated using marine phytoplankton, it has been shown to occur in cultures of Chlorella pyrenoidosa Chick containing copper (Steemann-Nielsen, Kamp-Nielsen and Wium-Anderson, 1969) and mercury (KampNielsen, 19711. Quite a few of the data in Appendix I were obtained using cultures containing populations of more than 105 cells/ml and in those cases where changes in optical density or turbidity were used as a measure of growth, the cell numbers could have been substantially higher than this. Measurements of the effects of metals upon such dense cultures provide little ecologically useful information.

POLLUTION STUDIES WITH MARINE P-KTON-If

406

4. Metal toxicity in relation to changes in the metal tolerance of the celle or in the chemical state of the metal during the growth of the culture Stockner and Antia (1976) have suggested that short term investigations lasting only a few hours or days of the effects of pollutants upon marine phytoplankton take insufficient account of the possibility that the cell populations being studied might, given the time, adapt to the hostile conditions created by the presence of the contaminant in the culture. They have given several examples of how, after extended lag phases, cultures containing high levels of pollutants have grown at near normal rates ultimately producing populations comparable with those in uncontaminated controls. While the existence of strains of other microorganisms and higher plants with acquired metal tolerance is well documented (Ashida, 1965 ; Antonovics, Bradshaw and Turner, 1971), there is little information on the development of heavy metal resistance in marine phytoplankton although it is certainly possible to " train " them to withstand levels of metals which would normally be lethal (A. G. Davies, unpublished work). The resurgence of growth in polluted cultures may not always be of biological origin, however, because the detoxication of the contaminant due to its transformation chemically can cause a similar result. Stockner and Antia (1976) cited several examples of such changes but none were related to metals. There are, though, many cases in the literature where cultures have been found to recover from the toxic effects of mercury added in the inorganic form (e.g. Rice et at., 1973; Davies, 1974; Zingmark and Miller, 1975; Kayser, 1976). The sequence of events leading to a delayed outburst of growth in a mercury-conhining culture of Isochrysis galbana is shown in Fig. 5 . The concentration of mercury present in particulate form, i.e. in the plant cells, at first increased as the phytoplankton grew but although, on the fifth day, the cell numbers stopped increasing, the plankton continued to take up more of the mercury. During the same period, the total concentration in the culture (medium plus cells) decreased steadily due to the chemical reduction of the ionic mercury by the extracellular products from the cells, the resulting elemental form being removed by volatilization into the air being bubbled through the culture. By the fifteenth day of the experiment, all of the mercury remaining in the culture was contained by the plant cells and, at this time, the cells began to grow again eventually attaining a population density similar to that in the mercuryfree control. It is not clear why the disappearance of mercuric ions from the solution bathing the cells enabled growth to recommence but

406

ANTHONY 0 . DAVIES

it may be because the metal interferes with the uptake of the nutrients required for growth. The effective concentrations of metals in cultures may also be decreased by precipitation, adsorption on to the culture vessels and chelation by extracellular products. Studies of the effects of metals

r

lo

c c

o n

Days

FIQ. 6. The changes which took place in a mercury containing culture of Isochysis galbana. 0, total mercury concentration (pg/l) in culture (medium + cells); 0 , particulate mercury concentration (pg/1)in cells ; a, mean mercury concentration in cell material (fglpms); X, volume of cell material in culture (pms/ml o f medium). By day 5 , the mercury content of the cells W R Y sufficient to prevent growth but it continued to increase after this. Volatilization of metallic mercury from the culture caused the total concentration to decrease steadily until day 15 when the metal remaining was all contained in the phytoplankton. This led to a renewed outburst of exponential growth giving a final cell population, measured as the volume of cellular material present in the culture, approaching that attained in the mercury free control. (From Davies, 1974. Reproduced by kind permission of the Council of tho Marine Biological Association of the United Kingdom.)

upon phytoplankton based merely on observing changes in the cell populations with time obviously take no account of these possibilities. 5. Metal toxicity in relation to Concentration

I n view of the foregoing discussion, it will be appreciated that the concentration of metal added t o a culture medium a t the start of an experiment gives very little guide to the amount of metal incorporated by the cells and which will, therefore, influence their metabolic activity ; and although it was demonstrated some time ago that quantitative

407

POLLUTION STUDIES WITH MARINE PLANKTON-II

relationships exist between the growth rates of phytoplankton and the cellular contents of nutrients such as vitamin B,,, iron and silicate ion when they are rate-limiting (Droop, 1968 ; Davies, 1970 ; Paasche, 1973), few attempts have been made to establish similar expressions for growth-inhibiting toxic metals, most authors having been content to Experiment 1 Culture

' C D E F A

. .

V

X

X

X I

U

I\

xJx-

I

M

B. Experiment 2 Symbols as above

0.6 0.4

X

0.2

0

X

x

1

2

'

I

3

4

-

X

5

6

4r (fg H g l w 3 )

FIG. 6. The ratio of the specific growth rate (pH.) of mercury containing cultures of 18ochq& galbana to that in the mercury free control (p), plotted &B a function of the mercury content of the cell material (qt). The concentrations of mercury initially addedtothecultureswere (inpg/l):Experiment 1 ;B, 16S;C, 3.30;D, 6.08;E,7.60; F, 10.5. Experiment 2 ; B, 0.77 ;D, 3.30 ; E, 6.08 ;F, 7.60. Most of the data approximate to the line pEs/pm = 1 - q,/K,, K,, the intercept on the abscissa, representing the concentration of mercury in the cell material which just prevented growth. The points having q, values greater than K, correspond to the renewed growth which took place after loss of the mercury from the culture medium (see Fig. 6). (From Davies, 1974. Reproduced by kind permission of the Council of the Marine Biological Association of the United Kingdom.)

present their data simply as the reduction in growth rate or the cell population after a certain time as compared to the values in a metalfree control. That the growth rate of a culture is related to the heavy metal content of the phytoplankton cells has been shown in two cases-for mercury containing Isochrysis galbana (Fig. 6) (Davies, 1974) and copper containing Thalassiosira pseudonana (Fig. 7) (8unda and

408

ANTHONY G. DAVIES

Guillard, 1976). However, as most of the metal taken up by cells is likely to be bound on to structural rather than metabolic sites and the number of the former decreases as the nutrients are used up due t,o the switch from protein to carbohydrate and lipid production (Section 11, A), it might be expected that a given cellular burden of a heavy metal would prove to be more inhibitory in older cultures as more of the

Mean cellular copper content (fg at Cu/cell)

FIQ. 7. The specific growth rate (p) of copper containing cultures of Thalussiosira pseudonunu plotted as a function of the mean copper content of the cells. Concentra3; 0 , 5. The tions of TRIS present in culture medium were (mM) : A, 1 ; A,2 ; 0, shape of the plot suggests that there are at least two types of copper-sensitive metabolic sites in the cells. (After Sunda and Guillard, 1976.)

metal content of the cells would be free to attach to vulnerable enzyme systems. Davies (1974) found no evidence that this was the case for the effect of mercury upon Isochrysis galbana, but preliminary experiments with cadmium (A. G. Davies, unpublished data) have shown that as the cultures develop, the cells in them become less metal resistant. The effect of cadmium upon the growth of Isochrysis galbana is illustrated in Fig. 8. When the specific growth rates of the phytoplankton were plotted against the cadmium content of the cells using data taken from cultures containing differing levels of cadmium but having the same content of cell material and therefore the same remaining nutrient

POLLUTION STUDIES WITH MARINE PLANKTON-=

409

Fro. 8. The effect of cadmium upon the growth of Isoch.rysis galbana. Culture conditions were as given in Davies (1974), the population being measured, as previously, as the volume of cell material/ml of culture. Analysis of the experimental data in terms of the fractional reduction in specific growth rate caused by the cadmium w8.s carried out for the four cell densities designated A, B, C and D (see Fig. 9).

concentrations (A, B, C and D in Fig. 8 ) , it was found (Fig. 9) that they lay on a series of curves having the general form ! % = 1 - _ - - "qGd Pmax

b-qm

where pGa was the specific growth rate in the culture containing cadmium, pmaxwas that in the control, q,, was the cadmium content of the cell material (in ag cadmium/pm3 in Fig. 9) and a and b were constants for a given nutrient level. The increasing curvature of the plots supports the idea, as outlined earlier, that cells growing in batch

410

ANTHONY Q. DAVIES

cultures could become more susceptible to the effects of metals as the nutrients are utilized and this would obviously be quite important environmentally. Further investigations into relationships of this type are required, for once it is established that there is a definite link between the growth rates of phytoplankton and their heavy metal contents, variations in

A

20

40

ao

a 1.5

b

360

120

140

FIG.9. The ratio of the specific growth rate (pea) of cadmium containing cultures of Isoehrysis galbana to that in the cadmium free control (pmax)plotted as a function of the cadmium content of the cell ma.terial (qcd). The four sets of data A , B, C and D correspond to the four cell densities shown in Fig. 8, the curves being calculated from the expression pCd/pmsx= 1 - aqcd/(b-qca) using the values for the constants a and b given in the diagram. The increasing curvature of the plots and their decreasing intercepts on the abscissa as the populations in t,he cultures increased both reflect the reduction in cadmium tolerance of the older cells.

the concentrations and availability of metals in culture media will become of secondary importance to the amount of metal taken up by the cells which can easily be determined; and furthermore, as shown in Section V, it would become possible to utilize the data on the heavy metal contents of natural phytoplankton populations to assess the extent to which their growth might be affected by the metals present.

41 1

POLLUTION STUDIES WITH MARINE PWKTON-II

B. Synergism a d antagonism of mixtures of heavy metals towards phytoplankton The only studies of the effects of mixtures of metals upon marine phytoplankton seem to be those of Braek, Jensen and Mohus (1976) who measured the growth rates of Amphidinium mrterae Hulburt, Thalassiosira pseudomna, Skeletonem costatum and Phaeodactylum tricornutum in the combined presence of copper and zinc. The two metals were found to interact synergistically towards the first three species but antagonistically to the last. The results obtained with Amphidinium carterae are summarized in Table I11 which clearly illustrates that the reductions in growth rate caused by mixtures of the metals were greater than the sum of the effects due to each separately but at the same concentrations; further work on metal mixtures is required to discover whether other, possibly more serious, cases of synergism exist. TABLE111. THE SYNERGISTIC EFFECTOF COPPERAND ZINO UPON (IN RELATIVE UNITS) OF AMPEIDIHIUM OARTERAE

TEE

GROWTH RATE

Added metal 0On.c.

(ygll)

cu

0

60

75

100

160

200

260

1 .o 1 0-94 0.82 0.66 0.69 0.23 0.12

0.66

0.47 0-36 0-23 0

0.41 0.18

0.36

0.18

0

Zn 0 60 100 200 400 500 1 000 2 000

s o

0.47 0.47

0.35

The data are derived from Braek et al. (1976). Culture medium: 76% Sea water 0.18 mM NO, EDTA. I nutrients + 4.6 @ Illumination : 2.6 klux, 14 hours/day. Initial number of cellslml : (2-4) x 104.

+

+

7.26 pM HaPO;

+ micro-

Braek et at. considered that the decrease in the toxicity of copper towards Phaeodactylum tricornutum caused by the presence of zinc was due to competition of the two metals for the same binding sites, and they found that the toxicity of zinc towards the same species could similarly be moderated by increasing the levels of magnesium in the culture medium. It will be recalled that Cossa (1976) obtained evidence

412

ANTHONY Q. DAVIES

that seems to indicate that cadmium and zinc are also bound by the same groups in Phaeodactylum tricornutum supporting the view of Braek et al. that all divalent ions taken up by this species compete for the same binding positions.

C. The nature of metal toxicity in phytoplankton Some heavy metals, notably mercury, silver and copper, have very high affinities for sulphur and, as a result, it is generally believed that, in living material, they are bound mainly by the sulphydryl groups of proteins and enzymes. I n support of this view, Shaw (1954) demonstrated that there is a correlation between the toxicity of metals and their tendency to bind to sulphur expressed in terms of the solubility of their sulphides. The effect of metals upon phytoplankton might therefore be expected to be related to their disruption of the metabolic processes involving sulphur-containing cellular constituents and there are signs that this is, in fact, the case. The available evidence is concerned with changes in membrane permeability and the inhibition of cell division brought about by the presence of metals. 1. Membrane permeability

Rothstein (1959) has pointed out that the interaction of metals with the sulphydryl groups in a cell membrane, probably to form -S-metal-Sbridges, would produce considerable stress at the molecular level which could cause an increase in the permeability of the membrane leading to the loss of intracellular constituents as observed with several different types of microorganism ; mercury, for instance, has been found to cause potassium leakage from bakers’ yeast (Passow and Rothstein, 1960), human erythrocytes (Weed, Eber and Rothstein, 1962) and phytoplankton (Kamp-Nielsen, 1971 ; Shieh and Barber, 1973; Overnell, 1975). As it is necessary to use dense cell suspensions to allow the accurate determination of any changes in the levels of the intracellular constituents, the concentrations of heavy metals which have had to be added to cause membrane leakage have, in most cases, proved to be very high in order to reach the cellular burdens of metals necessary to produce the effect. However, Kamp-Nielsen (1971) using cultures of the freshwater phytoplankton Ghlorella pyrenoidom showed that the loss of cellular potassium occurred at the same concentrations of mercury or copper as those which caused a reduction in the rates of photosynthesis in the cultures. This suggested to Kamp-Nielsen that the damage to the membrane represented the primary cause for the decrease in growth ;

POLLUTION STUDIES WITH MARINE PLANKTON--II

413

but the more recent experiments of Overnell (1975) indicate that this may not always be the case for, while the range of copper concentrations which caused potassium leakage from Dunaliella tertiolecta was the same as that which inhibited oxygen evolution, the mercury levels which reduced the rate of oxygen production were an order of magnitude lower than those causing loss of potassium. Davies (1976) similarly found that the mercury concentrations which inhibited the growth of 1sochrysis galbana caused changes in the potassium contents of the cells which were no greater than those which occurred in Dunaliella tertiolecta, the growth of which WM unaffected by the presence of mercury. Thus, in the case of marine phytoplankton a t least, there need not always be a link between growth inhibition and membrane leakage due to metal uptake, though the fact that metals can cause severe disruption of phytoplankton membranes is shown by the observations of Erickson (1972) that large aberrant cells with cytoplasm exuding from them were present in copper-containing cultures of Thalassiosira pseudonana. Other morphological abnormalities in phytoplankton due to the presence of heavy metals have been reported by Nuzzi (1972) who found that phenylmercuric acetate caused the normally biradiate cells of Phaeodwtylum triwrnutum to become vacuolated or ovoid in shape, by Kayser (1 976) who noted that the thecae of Scrippsiella faeroense (Paulsen) Balech & Soares split open in the presence of mercury allowing the escape of a motile, naked form of the dinoflagellatethough this was thought to be the response of the organism to the unfavourable environment created by the metal, and by BentleyMowat and Reid (1977) who observed swelling of the cell contents of Ditylum brightwellii (T. West) Grunow ex Van Heurck. 2. Giant cells in metal-containing cultures of phytoplankton

An unexpected effect of some metals upon phytoplankton is that they cause the production of very large cells ; these have been observed both in copper and in mercury containing cultures (Erickson, 1972 ; Davies, 1974, 1976). Davies (1974) found that the size spectrum of cclls of Isochrysie galbana was very dependent upon the mercury concentration and that, at the highest sub-lethal concentration studied (initially 10.5 pg mercury/l), there was almost a doubling in the mean cell volume relative to that in the mercury-free control (Fig. 10). Giant cells were produced even in cultures of Dunaliella tertiolecta containing mercury at concentrations which had no effect upon its growth rate (Davies, 1976). It thus appears that metals inhibit the process of cell division independently of any effect they have upon the

414

ANTHONY

a. DAVIES

production of new cell material so that the phytoplankton increase in size. Shrift (1959) found that selenomethionine caused a similar uncoupling of growth from division in cultures of Chlorella vulgaria Beij. and it was shown that the resulting giant cells contained no methionine which reappeared in the cells only when they had recovered from the 75 Initial mercury concentration (jrgll)

A

65

B

E

F

0 1.55 7.5 10.5

rrr C

!i 35

15

0

4

8

12

16

20

24

28

Days

Bra. 10. The effect of mercury upon the mean cell volume in cultures of Isochrysis galbana. Even a t concentrations as low as 1.55 pg/l, mercury caused a temporary increase in mean cell size by inhibiting cell division without proportionally reducing the growth rate and the effect was especially evident at higher concentrations, almost a doubling in the mean cell volume relative to that in the control occurring in the culture initially containing 10.5 pg mercury/l. Here also, however, the cell population eventually reverted to a normal size distribution. (From Davies, 1974. Reproduced by kind permission of the Council of the Marine Biological Association of the United Kingdom.)

effect of the selenomethionine and had returned to a normal rate of division. Davies (1976) has suggested that mercury and copper might also prevent the production of methionine which appears to be necessary for celldivision to occur, for iodoacetamide, which like mercury and copper reacts with sulphydryl groups, inhibited the reduction of sulphate ions in Chlorella pyrenoidosa, an important product of the reduction being S-adenosyl methionine (Schiff, 1959). As the cultures eventually recovered from the uncoupling effects of selenomethionine

POLLUTION STUDIES WITH M I N E PLANKTON-XI

416

and meroury, it seems unlikely, even if the same effect did occur under natural oonditiom, that it would cause a permanent change in cell size distributions. From the foregoing, it appears that the reduction in the growth rate of phytoplankton caused by mercury is due to the inhibition of intraoellular metabolic processes which are quite separate from those controlling cell division. The data of Sunda and Guillard (1976) (Fig. 7 ) are interesting from this point of view, for the two-step curve which describes the decrease in the growth rate of Thalasaioeira pseudonana due to oopper suggests that inhibition results from the binding of the metal a t two independent sites of reaction each with a different affinity for copper ; only the decrease in growth rate associated with the higher levels of cellular copper was accompanied by increases in cell volume indicating the presence of even more highly copper sensitive processespossibly photosynthesis as suggested by Overnell (1976)-in the cellular metabolic system.

IV. STUDIESOF

THE TOXICEFFECTS OF HEAVY METALSUPON NATURAL POPULATIONS OF PHYTOPLANKTON

I n order to study the effects of heavy metals upon natural phytoplankton populations, they have to be contained in some way in order to prevent their dispersion and dilution by turbulence and diffusion. Two main approaches to the problem have been adopted. I n one, samples of water together with the indigenous population are placed in bottles and the photosynthetic rates of the phytoplankton measured, usually in terms of the rate of 14C uptake, in the presence of known concentrations of an added metal. By suspending the bottles either in eitu at the appropriate depth or m o d i f ~ gtheir transparency to simulate the light reaching the location from which the water was removed and placing them in a tank through which is flowing the sea water, it can be ensured that the temperature and illumination inside the bottles are approximately the same as those of the water and phytoplankton which the sample represents. The technique is relatively straightforward and allows the simultaneous study of a wide range of pollutant concentrations. The large area of container surface in contact with the sample will, however, usually cause a decrease in the concentration of the metal added to the water due to adsorption and may, especially if the experiments are prolonged, give rise to other problems such aa bacterial proliferation. Thus, another shortcoming of this method is that the measurements cannot be made over extended periods so that the results reflect only the immediate impact of the

416

ANTHONY Q. DAVIES

metals upon the phytoplankton. This weakness, to some extent, can be overcome by incubating the populations with the metals prior to the measurements in order to allow equilibration between the added metal and the plant cells, but there is a limit to the time over which this can be done before the unwanted effects described earlier become too prominent. This type of measurement cannot, as a result, give reliable information about the longer term changes brought about by metal pollution, such as alterations in the species composition or the development of resistant populations. Most of the difficulties can, at least in principle, be overcome by using large volume enclosures of sea water containing its natural populations of flora (and fauna) and floating in the water used to fill them so that the entrapped communities are subject to the same meteorological conditions as those outside. The quantity of water contained ( 2 500 m3) allows an extensive sampling programme to be carried out for monitoring a wide range of parameters of significance to the growth of the phytoplankton without serious depletion taking place. Preliminary studies indicated that the development of the enclosed populations in replicate containers was, provided they were filled simultaneously, similar both to each other and to that of the phytoplankton in the water surrounding the enclosures (Takahashi, Thomas, Seibert, Beers, Koeller and Parsons, 1975). More recently, however, it has been indicated that anomalies in the behaviour of the enclosed phytoplankton could arise due to shading and the absence of turbulence allowing the heavier diatoms to settle out (Tekahashi and Whitney, 1977). The chief difficulty associated with the use of these systems is their great expense so that the range of pollutant concentrations which can be used in a given experiment is limited by the number of operational enclosures. I n this section, the results obtained using both techniques will be examined.

A. The effects on primary production rates Knauer and Martin (1972) were probably the first to use the 14C uptake technique of Steemann-Nielsen (1952) for measuring the decrease of photosynthesis in natural populations of marine phytoplankton caused by a heavy metal. They found that up to 1 pg/l, inorganic mercury (added as mercuric chloride) actually caused a slight increase in the rate of primary production of phytoplankton taken from Monterey Bay, California, but at higher levels of the metal, photosynthesis was markedly inhibited ; methylmercury proved to be even

POLLUTION STUDIES WITH MARINE PLANKTON-II

417

more toxic than the inorganic form (Fig. 11). Later studies on populations taken from the Caspian Sea (Patin, Tkachenko, Ibragim and Fedotova, 1974), the Sargasso Sea and Gulf Stream. (Zingmark and Miller, 1976) and the Eastern Mediterranean and Red Sea (Ibragim and Patin, 1976) have all demonstrated similar effects with inorganic mercury though, in general, the degree of inhibition resulting from a

Mercury concentration (pg/l)

FIG. 11. The effect of mercury in the inorganic form (0) and as methylmercury (@) upon the photosynthesis of a natural population of phytoplankton collected from Monterey Bay, California. (After Knauer and Martin, 1972.)

given level of contamination ha8 tended to be substantially smaller in the more recent work (Table IV) ; the reason for this is not apparent. Patin et al. (1974) and Ibragim and Patin (1976) also measured the decreases in primary production rates caused by copper, cadmium, lead and zinc and their results (examples are given in Table IV) were in accord with the recognized decrease in toxicity in the metals in the order given in the Table. Additions of zinc, the least toxic metal studied, of up to at least 100 pg/1 in fact caused increases in the photosynthetio rates of the Caspian Sea phytoplankton. Ibragim and Patin (1976) extended their work to investigate the results of longer term exposure of the Mediterranean phytoplankton

418

ANTHONY Q. DAVIES

TABLEIV. INHIBITION OF PRIMARY PRODUCTIVITY OF PHYTOPLANKTON POPULATION^ TA-N FROM DIFFERENTSEA AREASBY METALS AT A CONCENR~ATIONOF 10 pggh

Metal ~

Mercury

Copper Cadmium Lead Zinc

(Productivity % of control)

Source of phytoplankton

Reference

~

Monterey Bay, California W. Caspian Sea Estuary, S. Carolina Sargasso Sea Gulf Stream Red Sea E. Mediterranean Sea W. Caspian Sea Red Sea E. Mediterranean Sea W. Caspian Sea Red Sea E. Mediterranean Sea Red Sea E. Mediterranean Sea W. Caspian Sea

12 28 76 49 55 28 50 46 * 56 54 79 65 82 86 92 ll5*

Knauer and Martin (1972) Patin et al. (1974) Zingmark and Miller (1976) Zingmark and Miller (1976) Zingmark and Miller (1976) Ibragim and Patin (1976) Ibragim and Patin (1976) Patin et al. (1974) Ibragim and Patin (1976) Ibragim and Patin (1976) Patin et al. (1974) Ibragim and Patin (1976) Ibragim and Patin (1976) Ibragim and Patin (1978) Ibragim and Patin (1976) Patin et al. (1974)

* Estimated from values a t lower and higher concentrations. to the metals by adding a range of concentrations to sea water containing the natural population and measuring the photosynthetic rates a t each metal concentration over the following five days. In general, the toxic effects of the metals were found to be less in the longer term experiments than in the short term studies where the productivity was measured soon after the metal additions had been made. At a concentration of 1 pg/l of copper, cadmium or lead, for instance, primary production rates were increased over that in the control even on the first day after the metals had been added, and on the second day, mercury added to give 1 pg/l and the other metals at 10 pg/l were either non-toxic or stimulatory. Mercury at 10 pg/l, though, permanently inhibited photosynthesis as did copper at 100 pg/l and cadmium and lead at 1000 pg/L Ibragim and Patin ascribed the reduction in toxicity in the longer-term experiments to " biological dilution ", i.e. the decrease in the metal burdens of the plant cells as they increased in number in the presence of a fixed amount of metal ; a decrease in the total metal concentrations due to losses on to the container would have reinforced this effect. Unfortunately, the metal contents of the phytoplankton were not determined. A further possible explanation of the apparent moderation in the effects of the metals could be that the phytoplankton which grew during the experiment were more metal resistant than those initially present. It is interesting to note that

POILUTION STUDIES WITH MARINE PLANRTON-II

419

while, at first, the phytoplankton community consisted mainly of Cylindrotheca (Nitzschia) closterium, Coscinodiscus granii Gough and Rhizosolenia alata Brightw., the final population was dominated by the first of these. As will be seen later, further evidence of the copper resistance of pennate diatoms has been obtained from the work with large scale water enclosures discussed in the next section.

B. The effects in large volume sea water enclosures

A series of experiments performed in the summer of 1974 with the CEPEX (Controlled Ecosystem Pollution Experiment) enclosures operated in Saanich Inlet (near Victoria, B.C., Canada) has provided the data to be discussed here on the longer term effects of copper on the growth and metabolism of natural phytoplankton populations and, in a later section, on the behaviour of natural zooplankton communities. Two separate sets of measurements were carried out. I n Experiment I, copper was added to two of the test enclosures on the second day after filling to give initial concentrations of 10 and 50 pg/l and, in Experiment 11, the copper additions of 5 and 10 yg/l were delayed until the ninth day after filling. The copper concentrations in the enclosures to which were initially added 10 pg/l (both experi.ments) and 50 pg/l gradually decreased due to loss by sedimentation and the 10 yg/1 enclosures were replenished several times in Experiment I, and once in Experiment I1 (Topping and Windom, 1977). No further copper was added to the enclosure initially containing 50 pg copper/] and at the end of the experiment after almost four weeks, it contained only about 20 pg/l (Topping and Windom, 1977). Nitrate, phosphate and silicate additions were made periodically to maintain the nutrients at predetermined concentrations. I n both experiments, regardless of the presence or absence of copper, the phytoplankton populations (measured as chlorophyll a and phytoplankton carbon concentrations in the top 10 m) decreased during the first four or five days after filling (Thomas, Holm-Hansen, Seibert, Azam, Hodson and Takahashi, 1977). It was suggested that this fall-off in the levels of phytoplankton might have been due to the absence of turbulent mixing in the enclosures; however, in a later series of experiments, Takahashi and Whitney (1977) noticed that the centric diatoms4huetoceros spp.-settled out from the top 20 m within 3 weeks, both in the enclosures and in the surrounding water. The copper additions, especially in Experiment I, caused an even greater decline in the phytoplankton levels in the test enclosures than

420

ANTHONY Q. DAVl3S

in the controls. This appeared to be due largely to the copper sensitivity of the centric diatoms-mainly of the genus Chaetoceros-which dominated the populations in the water used to fill the enclosures (Thomas and Seibert, 1977; Goering, Boisseau and Hattori, 1977); for, while in the controls, the proportion of the total phytoplankton carbon in the upper lOm represented by Chaetoceros spp. steadily increased during the four week period of Experiment I, that in the copper polluted enclosures decreased gradually for the first two weeks and then, during the third week, dropped almost to zero, simultaneously being replaced-roughly on a reciprocal basis-by the development of microflagellates. Pennate diatoms seemed to be more resistant to copper than the Chaetoceros spp. because, for a short period, there was a temporary increase in the numbers of Nitzschia delicatissima Cleve in the 10 pg/l enclosure and, in the last week of Experiment I, Navicula distans (W.Sm) Cleve became the dominant diatom in the 50pg/1 enclosure. I n Experiment 11, the proportion of the phytoplankton population represented by centric diatoms declined almost to zero over the first nine days after filling even in the copper free enclosure, the balance consisting maiilly of microflagellates (Fig. 12A) (Thomas and Seibert, 1977). From Day 9, however, the centric diatoms in the controls increased in number and, after four weeks, the population consisted mainly of these and dinoflagellates, some pennate diatoms and microflagellates also being present (Fig. 12A). (This succession of diatommicroflagellatdiatom was the converse of that observed by Takahashi et al. (1975) where the initial phytoplankton population consisted mainly of microflagellates which were temporarily displaced by Thalossiosira spp.) I n the copper polluted enclosures of Experiment 11, the centric diatoms failed to recover and were replaced by pennate diatoms (Fig. 12B, C) (Thomas and Seibert, 1977) providing further evidence of the greater resistance to copper of the pennate than the centric diatoms. It will be recalled that Ibragim and Patin (1976) had similarly found that the pennate diatom Cylindrotheca (Nitzschia) closterium eventually became dominant in Eastern Mediterranean phytoplankton which had been exposed for some time to heavy metals, in contrast to the earlier work of Maiidelli (1969) with cultures of Thalassiosira pseudonana (as Cyclotella nana) and Cylindrotheca (Nitzschia)closterium which indicated that the former was the more copper tolerant. At the end of both experiments, the phytoplankton crops in the polluted enclosures, as measured by the particulate nitrogen content or the concentrations of chlorophyll a, phytoplankton carbon and ATP

CEPEX Copper Experiment II,September 1974

A

Control

C

4

5pg Cu/l added

c

OpgC u l l added

FIG.12. The effect of copper on the proportions of phytoplankton carbon in the upper 10 m of the CEPEX enclosures represented by microfiagellates (light stippling), dinoflagellates (cross hatching), pennate diatoms (heavy stippling) and centric diatoms (white). The copper prevented the resurgenoe of the centric diatoms following the decrease which took place after the enclosures were filled and was lethal to the dinoflagellates, the pennate diatoms ultimately dominating the population. (After Thomas and Seibert, 1977.)

422

ANTHONY 0.DAVIES

in the top 10 m, exceeded those in the controls (Harrison, Eppley and Renger, 1977; Thomas et al., 1977). Due to the almost complete absence of herbivorous zooplankton in the copper-containing enclosures (Gibson and Grice, 1977), it was not, however, possible to attribute this positively either to the stimulatory effect of the copper or to the lower grazing pressure on the phytoplankton. Jt could also have been caused CEPEX Copper Experiment I.June 1974

Cu'added

Days afterfilling

FIG.13. The effect of copper on the mean productivity in the upper 5 m of the CEPEX enclosures. Control enclosures, J ( 0 ) and K ( 0) ; experimental enclosures, L with 10 pg copper/l (A)and M, initially containing 50 pg copper/l (0). The metal caused a marked reduction in the rates of photosynthesis, though after the fift,h day, due to the growth of microflagellates, it started to increase again eventually reaching values higher than in the controls. (After Thomas et al., 1977.)

by the communities-mainly microflagellates-present in the polluted enclosures having slower sinking rates than the chain-forming diatoms present in t-he controls (Harrison et al., 1977). The variations in the phytoplankton populations brought about by the copper were paralleled by changes in the rates of photosynthesis in the enclosures. On the day after the copper additions in Experiment I, for instance, primary productivity rates in the presence of copper were

POLLUTION STUDIES WITH MARINE PLANKTON-11

423

only 15-30% of those in the controls and, two days later, were even lower (Fig. 13) (Thomas et al., 1977). Thereafter the photosynthetic rates increased with the build-up of the copper resistant population of flagellated and diatoms eventually reaching values exceeding those in the controls, though the recovery of the population in the 50 pg copper/l enclosure was much slower than that growing in the presence of 10 pg copper/l. More detailed investigations of the effect of the copper additions on the metabolism of phytoplankton were carried out by Harrison et al. (1977) who studied nitrogen uptake and utilization, and Goering et al. (1977) who measured silicic acid uptake rates. Harrison et al. (1977) found that while the specific rates of uptake of both nitrate and ammonium ions by the enclosed phytoplankton communities were extremely variable, they were generally in accord with previously determined values for coastal phytoplankton. I n both experiments, reductions in the specific uptake rates occurred only during the days immediately following the copper additions, at other times the rates of nitrate and ammonium assimilation being regulated mainly by the concentrations of these ions present in the water. The formation of nitrate reductase in phytoplankton taken from either the control enclosures or the surrounding water was found t o be inhibited by copper additions of 10 and 20 pg/l while the resistant population which developed in the enclosure to which 50 pg copper/l had been added was relatively unaffected in its ability to produce the enzyme, even in the presence of an additional 20 pg copper/l. This observation may provide at least a partial explanation of the copper tolerance of some species of phytoplankton. Goering et al. (1977) found that the rates of silicic acid uptake by the phytoplankton communities present in the enclosures was decreased by concentrations of copper as low as 2.5 pg/l. Lewin (1954) had previously shown that the incorporation of silicic acid into diatoms required the presence of reduced sulphur-possibly sulphydryl groups on the cell membrane or in an enzyme-and that it could be prevented by the presence of sulphydryl inhibitors such as iodoacetamide, sodium arsenite and cadmium chloride; the effeot of copper which also has a high affinity for sulphur is therefore perhaps not surprising. It will also be seen later that Martin and Knauer (1973) found that an unusually large proportion of the copper content of natural phytoplankton was associated with the silica frustules. Goering et al. suggested that the greater copper tolerance of pennate diatoms as compared to the centric forms might be because the silicic acid uptake mechanism in the former was less susceptible to the effect of copper. At first sight, the results of

42