Ce!) OPS "à, 9 National Museums National Museum of Canada of Natural Sciences Ottawa, 1974 Publications in Zoology, No. 9 Field and Laboratory Studies of Daphnia schodleri Sars from a Winterkill Lake of Alberta Chi-hsiang Lei and Hugh F. Clifford ee oe ae ek USE) | CALIFORNIA | | ACADEMY OF SCIENCES | fy - 2 1974 | | LIBRARY | Publications de Zoologie, n° 9 Musées nationaux Musée national des du Canada Sciences naturelles Digitized by the Internet Archive in 2011 with funding from California Academy of Sciences Library http://www.archive.org/details/publicationsinzo91 nati Field and Laboratory Studies of Daphnia schodleri Sars from a Winterkill Lake of Alberta National Museum of Natural Sciences Musée national des Sciences naturelles Publications in Zoology, No. 9 Publications de Zoologie, n° 9 Published by the Publié par les National Museums of Canada Musées nationaux du Canada Staff Editor Bonnie Livingstone Field and Laboratory Studies of Daphnia schodleri Sars from a Winterkill Lake of Alberta Chi-hsiang Lei and Hugh F. Clifford ©Crown copyrights reserved Available by mail from the National Museums of Canada Marketing Services Ottawa, Ontario K1A OM8 Catalogue no. NM95-10/9-1 National Museum of Natural Sciences National Museums of Canada 1974 PO987654321 Y70987654 Printed in Canada Contents List of Figures, vi List of Tables, ix Biographical Note, xi Résumé, xii Summary, xiii Acknowledgements, xiv Part | Development of parthenogenetic eggs in vitro and duration of embryonic stages Abstract, 1 Introduction, 1 Materials and Methods, 2 Results, 2 Discussion, 6 Literature Cited, 7 Part Il Vital statistic properties of laboratory populations Abstract, 13 Introduction, 14 Materials and Methods, 14 Preadult Instars, 15 Longevity, 17 Growth, 17 Reproduction, 20 Discussion, 25 Literature Cited, 27 Part Ill Periodicity, reproduction and growth of D. schodleri in the lake Abstract, 41 Introduction, 41 Materials and Methods, 42 Description of the Study Area, 43 Zooplankton of Big Island Lake, 43 Life History of D. schgdleri, 44 Interpretations of Length-Frequency Distributions, 47 Literature Cited, 48 vi List of Figures Part | Development of Daphnia sch@édleri eggs and embryos in vitro 1 Egg three hours after deposition, 8 2 Egg at six hours, 8 3 Egg at nine hours, 8 4 Egg at 12 hours, 8 5 Egg at 13 hours, 8 6 Egg at 15 hours, 9 7 Egg at 18 hours, 9 8 Embryo at 21 hours, 9 9 Embryo at 22 hours (dorsal view), 9 10 Embryo at 24 hours (dorsal view), 9 11 Embryo at 24 hours (ventral view), 10 12 Embryo at 27 hours (side view), 10 13 Embryo at 27 hours (dorsal view), 10 14 Embryo at 30 hours (dorsal view), 10 15 Side view of embryo at 30 hours, 10 16 Embryo at 33 hours (dorsal view), 11 17 Embryo at 33 hours (side view), 11 18 Embryo at 36 hours (dorsal view), 11 19 Embryo at 38 hours (side view), 11 20 Embryo at 39 hours (dorsal view), 11 List of Figures 21 Embryo at 42 hours (ventral view), 12 22 Side view of embryo at 43 hours, 12 23 Embryo at 57 hours, 12 Part Il 1 Survival curves for male and female D. schodieri in instars and days, 28 2 Mean growth curves for eight male and eight female D. schgdleri, 29 3 Absolute growth increment curves for the males and females of Figure 2, 30 4 Growth curves in days of female D. schodleri at 20°C and 5 C3 5 Growth curves in instars of female D. schgdleri at 20°C and AGH ey 6 Scatter diagrams of mean carapace length and height of the males and females of Figure 2, 33 7 Coefficients of variation for each linear dimension of the D. schodleri of Figure 2, 34 8 Size-frequency distributions during the first seven instars of males and females reared at 25°C, 35 9 Mean number of young produced during each adult instar, 36 10 Mean brood size in relation to age of females, 37 11 Length of liberated young in different broods, 38 12 Formation of an ephippium in D. schodleri, 39 vil VIII List of Figures Part lil 1 Water temperature and dissolved oxygen of Big Island Lake, 1966 and 1967, 49 2 Seasonal changes in population size of the major zooplankters of Big Island Lake, June 1966 to July 1967, 50 3 Seasonal variations in parthenogenetic egg production and body length of mature parthenogenetic females, 51 4 The relationship between size of sexual eggs and size of sexual females, and between size of parthenogenetic eggs and parthenogenetic females, 52 5 Population length-frequency histograms of D. schgdleri in Big Island Lake, 13 June 1966 to 16 October 1967, 53 List of Tables Part | 1 Mean and relative durations of Daphnia schodieri's embryonic stages in the brood chamber, 4 2 Number of D. schgdleri females from Big Island Lake carrying embryos, 4 3 Mean durations of brooding periods and instars for D. schogdleri primiparous in the sixth, seventh and eighth instars, at 5°C, 5 4 Mean durations of brooding periods and instars for D. schgdleri primiparous in fifth instar, at 20°C, 6 Part Il 1 Initial size and size in first adult instar of female Daphnia schodleri, 16 2 Length of young in first instar and instar in which they reached maturity at 20°C, 16 3 Relative mean growth increments of total length for D. schgdleri males and females of Figure 2, 18 4 Relationship between egg size and size of young, 20 5 Mortality of laboratory populations at 20°C and 25°C, 21 6 Female size in first adult instar and number and size of their young in first adult instar, 22 7 Reproductive features of seven females that produced at least one ephippium at 5°C, 23 8 Vital statistic properties of D. schodleri's laboratory population, 24 Part III 1 Limnetic zooplankton found in Big Island Lake, other than Protozoa, opp. p.42 List of Tables 2 Seasonal changes in percentage composition of life cycle stages, June 1966 to July 1967, 44 3 Seasonal variations in mean sizes of parthenogenetic and gamogenetic females, 45 4 Relationship between parthenogenetic egg production and body size of mature female D. schgdleri for different sampling dates, 46 Biographical Note Chi-hsiang Lei Chi-hsiang Lei, a native of Taiwan, received his B.Sc. in Fishery Biology from the National Taiwan University, Taipei, Taiwan, in 1960 and his M.Sc. in Zoology from the University of Alberta, Edmonton, in 1968. His primary research interest is in the biology and ecology of zooplankton. He is at present studying for his Ph.D at the University of Kansas, Lawrence. His current research is on the ecological energetics of Daphnia. Hugh F. Clifford Hugh F. Clifford received his B.Sc. and M.Sc. degrees from Michigan State University and his Ph.D. in 1965 from Indiana University, where he studied under Dr. David G. Frey. His special interest has been the ecology of aquatic invertebrates in seasonally dry and winterkill habitats. He is currently carrying out limnological studies on a subarctic, brown-water stream of Alberta. Dr. Clifford is at present an associate professor in the Department of Zoology at the University of Alberta. xi Xi Résumé En combinant les relevés faits sur le terrain avec les observations de laboratoire, les auteurs ont étudié les aspects biologiques de spécimens de Daphnia schgdleri Sars provenant d'un lac a destruction hivernale d’Alberta. A 18°C, l'ensemble de la période embryonnaire in vitro fut d'environ 57 heures. Trois stades de croissance caractérisaient le développement embryonnaire. Toute- fois, deux stades seulement s’accompagnaient du rejet d'une membrane d'oeuf. À 5°C, la période moyenne d'incubation variait en fonction de la durée moyenne des stades adultes respectifs. Après le 16° stade, à 20°C, la période de chaque incubation est demeurée sensiblement la même, alors que celle des stades adultes ultérieurs s'est prolongée progressivement. Les femelles élevées en laboratoire à 5°C, 20°C et 25°C présentaient de quatre à sept stades larvaires. Les mâles en avaient trois ou quatre. À 25°C, la longévité moyenne était de 36 jours (18 stades) pour les femelles, et de 41 jours (18 stades) pour les mâles. Chez les femelles, la croissance absolue la plus considérable s'est produite à la mue qui survient entre l'adolescence et le stade primipare. Chez les mâles, cette croissance s'est manifestée entre la pré-adolescence et l'adolescence. À 20°C, les femelles avaient réalisé la plus grande partie de leur croissance dès le 10% jour, alors qu’ à 5°C, la courbe de croissance ne se stabilisait qu’au 40° jour. A 25°C, le nombre moyen de jeunes par ponte atteignait son maximum au quatrième stade adulte. À 20°C, il y avait deux maxima de reproduction, l’un au huitième stade adulte et l’autre au 31°. Dans le laboratoire, D. schgd/eri a pondu des oeufs fertiles à 5°C. Toutes les femelles pondant de tels oeufs traversaient un stade de stérilité immédiatement après le stade éphippial. Dans le lac, la génération ex ephippio est apparue tard au printemps et, au mois de mai, on pouvait observer la parthénogénè - se. On a noté deux périodes de reproduction sexuée. La plus importante a eu lieu en juin et juillet, alors que la population était maximale; l'autre, de moindre importance, s'est produite en septembre. La première période fut attribuée à la génération ex ephippio. Le nombre moyen d'oeufs parthénogénétiques par ponte diminua immédiatement avant le commencement de la période de reproduction sexuée. La ponte moyenne de toute la période à l'étude a été de 7,9 oeufs, ce qui est de beaucoup inférieur à la ponte moyenne des populations observées en laboratoire. Summary We studied the biology of Daphnia schogdleri Sars from a winterkill lake in Alberta by combining field and laboratory data. At 18°C the total jn vitro embryonic period was about 57 hours. During development there were three stages of embryonic size increase, but the shedding of an egg membrane could only be associated with two of these stages. At 5°C the mean duration of each brooding period varied directly with the mean duration of the respective adult instar. After the sixteenth instar at 20°C, the duration of each brooding period remained about the same, whereas the time interval of the subsequent adult instars became progressively longer. Female D. sch¢gd/eri, when cultured in the laboratory at 5°C, 20°C, and 25°C, had four to seven preadult instars; males had three to four preadult instars. At 25°C the average longevity was 36 days (18 instars) for females and 41 days (18 instars) for males. For females, the largest absolute growth increment was at the molt between the adolescent and primiparous instars; for males this increment was between the preadolescent and adolescent instars. At 20°C most of the growth of females was achieved by day 10, whereas the growth curves at 5°C did not level off until day 40. At 25°C, the mean number of young per brood was greatest in the fourth adult instar; at 20°C there were two peaks of young production, one in the eighth adult instar and another in the thirty- first adult instar. In the laboratory, D. schgdleri produced sexual eggs only at 5°C. All females producing sexual eggs had a sterile instar immediately following the ephippial instar. In the lake, the ex ephippio generation appeared in late spring, and by May parthenogenetic reproduction was taking place. There were two periods of sexual reproduction, a major period in June and July, at which time the population was exhibiting maximum numbers, and a minor period in September. The entire first period of sexual reproduction was accounted for by the ex ephippio generation. The average number of parthenogenetic eggs per brood diminished just before the onset of sexual reproduction. The average brood size for the entire study period was 7.9 eggs, which was considerably lower than the average brood size of laboratory populations. xiil XI Résumé En combinant les relevés faits sur le terrain avec les observations de laboratoire, les auteurs ont étudié les aspects biologiques de spécimens de Daphnia schgdleri Sars provenant d'un lac à destruction hivernale d’Alberta. A 18°C, l’ensemble de la période embryonnaire in vitro fut d'environ 57 heures. Trois stades de croissance caractérisaient le développement embryonnaire. Toute- fois, deux stades seulement s’accompagnaient du rejet d'une membrane d'oeuf. A 5°C, la période moyenne d'incubation variait en fonction de la durée moyenne des stades adultes respectifs. Après le 16° stade, à 20°C, la période de chaque incubation est demeurée sensiblement la même, alors que celle des stades adultes ultérieurs s'est prolongée progressivement. Les femelles élevées en laboratoire à 5°C, 20°C et 25°C présentaient de quatre à sept stades larvaires. Les mâles en avaient trois ou quatre. À 25°C, la longévité moyenne était de 36 jours (18 stades) pour les femelles, et de 41 jours (18 stades) pour les mâles. Chez les femelles, la croissance absolue la plus considérable s'est produite à la mue qui survient entre l'adolescence et le stade primipare. Chez les mâles, cette croissance s'est manifestée entre la pré-adolescence et l'adolescence. A 20°C, les femelles avaient réalisé la plus grande partie de leur croissance dès le 10° jour, alors qu’ à 5°C, la courbe de croissance ne se stabilisait qu'au 40° jour. A 25°C, le nombre moyen de jeunes par ponte atteignait son maximum au quatrième stade adulte. A 20°C, il y avait deux maxima de reproduction, l’un au huitième stade adulte et l’autre au 31°. Dans le laboratoire, D. schgd/eri a pondu des oeufs fertiles à 5°C. Toutes les femelles pondant de tels oeufs traversaient un stade de stérilité immédiatement après le stade éphippial. Dans le lac, la génération ex ephippio est apparue tard au printemps et, au mois de mai, on pouvait observer la parthénogénè- se. On a noté deux périodes de reproduction sexuée. La plus importante a eu lieu en juin et juillet, alors que la population était maximale; l’autre, de moindre importance, s'est produite en septembre. La première période fut attribuée à la génération ex ephippio. Le nombre moyen d'oeufs parthénogénétiques par ponte diminua immédiatement avant le commencement de la période de reproduction sexuée. La ponte moyenne de toute la période à l'étude a été de 7,9 oeufs, ce qui est de beaucoup inférieur à la ponte moyenne des populations observées en laboratoire. Summary We studied the biology of Daphnia schgdleri Sars from a winterkill lake in Alberta by combining field and laboratory data. At 18°C the total jn vitro embryonic period was about 57 hours. During development there were three stages of embryonic size increase, but the shedding of an egg membrane could only be associated with two of these stages. At 5°C the mean duration of each brooding period varied directly with the mean duration of the respective adult instar. After the sixteenth instar at 20°C, the duration of each brooding period remained about the same, whereas the time interval of the subsequent adult instars became progressively longer. Female D. schgdleri, when cultured in the laboratory at 5°C, 20°C, and 25°C, had four to seven preadult instars; males had three to four preadult instars. At 25°C the average longevity was 36 days (18 instars) for females and 41 days (18 instars) for males. For females, the largest absolute growth increment was at the molt between the adolescent and primiparous instars; for males this increment was between the preadolescent and adolescent instars. At 20°C most of the growth of females was achieved by day 10, whereas the growth curves at 5°C did not level off until day 40. At 25°C, the mean number of young per brood was greatest in the fourth adult instar; at 20°C there were two peaks of young production, one in the eighth adult instar and another in the thirty- first adult instar. In the laboratory, D. schgdleri produced sexual eggs only at 5°C. All females producing sexual eggs had a sterile instar immediately following the ephippial instar. In the lake, the ex ephippio generation appeared in late spring, and by May parthenogenetic reproduction was taking place. There were two periods of sexual reproduction, a major period in June and July, at which time the population was exhibiting maximum numbers, and a minor period in September. The entire first period of sexual reproduction was accounted for by the ex ephippio generation. The average number of parthenogenetic eggs per brood diminished just before the onset of sexual reproduction. The average brood size for the entire study period was 7.9 eggs, which was considerably lower than the average brood size of laboratory populations. Xill Acknowledgements We are indebted to Messrs. L.T. Chen, G.R. Daborn and C.C. Lin for assistance in various aspects of the field and laboratory work, and to Dr. J.K. Lauber for suggesting the microphotographic technique. The study was supported by grants from the National Museums of Canada and the National Research Council of Canada. XIV Part | Development of Parthenogenetic Eggs in vitro and Duration of Embryonic Stages Abstract In vitro development of D. schodleri Sars eggs was observed hourly throughout the embryonic period. At 18°C the total embry- onic period was about 57 hours. During jn vitro development there were three stages of embryonic size increase, but the shedding of an egg membrane could only be associated with two of these stages. Development of parthenogenetic eggs was also observed in the brood chamber of females by separating embryonic development into eight stages. Each of the eight embryonic stages at 5°C had about the same relative duration as those at 20°C. Study of various samples taken from Big Island Lake revealed that the females, at almost all times, were carrying either a larger or smaller number of embryos of a particular stage than would have been predicted from random ovulation and the relative duration of each embryonic stage at 20°C. At 5°C the mean duration of each brooding period varied directly with the mean duration of the respective adult instar. At 20°C, where more instars were observed, the mean duration of the brooding period varied directly with the mean duration of the adult instar until the sixteenth instar; thereafter the duration of each brooding period remained about the same, whereas each of the subsequent adult instars continued to exhibit a progressively longer time interval. Introduction In western North America, Daphnia schodleri is found from Texas to the Arctic Circle. In the northern part of its range, e.g. the prairies of Alberta and Saskatchewan, D. schgdleri is usually found in temporary ponds and small winterkill lakes, often occurring in large numbers during the ice-free season. D. schogdleri also occurs sporadically in large northern oligotrophic lakes, e.g. Great Slave Lake; Anderson (1968) found D. sch¢dleri in high elevation mountain lakes, which may possibly become depleted of oxygen in the wintertime. Very little is known about the biology of D. schgdleri from any habitat in its northern range. By combining field and laboratory data, we studied the life history of D. schgdleri from a winterkill lake in central Alberta. This lake, Big Island Lake, is ice-covered for six months of the year, and winter stagnation occurs before the ice breaks up in April or May. The study is to be reported in three parts: | laboratory study of external embryological features and embryonic stages; Il laboratory study of reproduction; Il field study of seasonal abundance, sea- sonal variation in egg production, and cyclic reproduction. Part | deals with the in vitro development of parthenogenetic eggs, duration of embry- onic stages in the brood chamber, and the duration of the brooding period. growth and Development of Parthenogenetic Eggs Materials and Methods D. schodleri used in the laboratory experi- ments were the third generation of a single non-ephippial female taken from Big Island Lake on 3 July 1967. In the laboratory the animals were cultured in a diluted medium of Banta’s manure-soil stock (Banta 1921). Three hours after being deposited in the brood chamber, eggs were dissected out of the female under a dissecting microscope, using fine needles. The eggs were then transferred to a depression slide filled with filtered aquarium water at 18°C and ob- served hourly throughout the embryonic period. Several series of eggs developed jn vitro using this method. Duration of each embryonic stage in the brood chamber was based on 10 broods at 20°+° 1°C ‘and’ 46>" broods “ati-5>21/sG. Hereafter these temperatures are referred to as 20°C and 5°C respectively. Duration of the total brooding period at 20°C and 5°C was based on a varying number of broods, depending on the instar number (Tables 3 and 4, pp.5 and 6). Results In vitro development At three hours the parthenogenetic egg of D. schgdleri is opaque, appears yellowish green, and has a large, yellow, fat globule in its centre (Figure 1). By the six-hour stage, the egg is translucent and a transparent edge can be seen. At nine hours an invagination, which later separates the second antennae from the body proper, appears near the future anterior end of the embryo. During the next three hours, the invagination becomes more distinct; a vertical constriction appears at the extreme posterior end of the egg at 12 hours, marking the future bilaterally sym- metrical plan of the embryo. At 18 hours the abdominal appendages appear. Body segmentation becomes more promi- nent at 21 hours. At this time the embryo, having just cast off the outer egg membrane, shows a slight increase in size. The second antennae are present as short, forked stubs, closely appressed to the body. At 22 hours the prospective brain is distinguishable, and a mid-dorsal, longitudi- nal fold begins to thin out laterally and posteriorly to form the carapace. At 24 hours the head bulge becomes distinguishable, and the second antennae increase in length. A few feeble heartbeats were observed in 30-hour embryos. A grayish material ap- pears antero-dorsad to the prospective brain at 27 hours. This material differentiates into two, very small, pink bodies (eyes) by 30 hours but because of their minute size, they are difficult to demonstrate. These pink eyes can be easily distinguished in 33-hour em- bryos; also at this stage, the prospective ocellus appears. At 30 hours the caudal spine is visible, extending out from the posterior end of the prospective carapace. With subsequent de- velopment, the spine continues to increase in length and eventually will curl ventrad and anteriorly between the posterior edges of the carapace. The joints of the second antennae first become visible in the 27-hour embryo, with Development of Parthenogenetic Eggs the three terminal setae of the second anten- nae becoming distinguishable in the 36-hour embryo. Intestine and hepatic caeca are seen at 39 hours, although not in dorsal view. The embryo, although casting off the outer egg membrane at the 21-hour stage, remains enclosed in an inner egg membrane tending to restrict the movement of the embryo. By the 42-hour stage, the inner egg membrane is cast off; the embryo increases greatly in body length and is capable of moving about by using the second antennae. At this time the distal portion of the second antennae separates from the body; the two rami, originally appressed to each other, also separate. The two small pink eyes gradually increase in size, becoming completely fused at 43 hours. From 43 hours through 56 hours, no significant changes in external morphology were observed. During the 57-hour stage, the caudal spine extends and the three terminal setae of the second antennae become longer than their rami. Although the embryo rapidly increased in size at this stage, we did not observe the casting off of another membrane. With the extension of the caudal spine, embryonic development is considered com- pleted. At 18°C the total embryonic period is about 57 hours. All eggs of the /n vitro study developed into females. Embryonic stages in the brood chamber Parthenogenetic eggs were observed being deposited into the brood chamber on 246 occasions. At 20°C, eggs were deposited into the brood chamber from two to 49 minutes after molting, the average time being 14 minutes after molting. On only seven occa- sions were the eggs deposited 30 minutes or more after the female had molted. When the young were released from the brood cham- ber, they had usually attained a development similar to that of the jn vitro stage at 53 hours; the caudal spine was still curled and the three terminal antennal setae were still shorter than their rami. To study embryonic development in the brood chamber, eight recognizable stages of embryonic development were selected. The stages follow those defined by Green (1956), but, because of information available from the in vitro study, are slightly modified and more definitive. Stage /. Eggs opaque or translucent with transparent edges. At first eggs are gray or grayish green, but later they become yellow- ish green and a clear zone begins to form around the periphery (Figures 1 and 2). Stage //. Eggs with granular transparent edges and an invagination, indicating the prospective cephalic region. Later the invag- ination becomes more prominent, and a constriction appears at the posterior end (Figures 3-7). Stage III. Embryo apparent, but head not yet defined; segmentation of body promi- nent. In latter part of stage, antennae are present as short, forked stubs closely ap- pressed to the body; and a thick, mid-dorsal, longitudinal fold begins to grow out forming the carapace (Figures 8-11). Stage IV. Embryo with head bulge; antennae longer but still appressed to the body; abdominal appendages appear as stubby blocks of tissues (Figures 12-15). Stage V. Embryo with two small pink eyes; antennae longer, still appressed to the body, but with distinct rami joints (Figures 16-17). Stage V/. Embryo with two brown eyes; three terminal antennae setae visible; body completely enclosed in carapace (Figures 18-20). Stage VII. Embryo with two large black eyes very close to each other; distal portion of antennae separated from body (Figure 21). Stage VIII. Embryo with one large black eye (Figures 22 and 23), and all subsequent development while in the brood chamber. Mean duration of each embryonic stage for animals cultured at 5°C and 20°C is shown in Table 1. Low temperatures increased the duration of each embryonic stage and hence the entire embryonic period. Each of the eight embryonic stages at 5°C had about the same relative duration as those of 20°C, the main difference being a relatively shorter Stage VIII at 5°C. Green (1956) noted that female Daphnia magna Straus collected from ponds had larger numbers of embryos in particular Development of Parthenogenetic Eggs Table 1 Mean and relative durations of Daphnia schodleri's embryonic stages in the brood chamber, based on 46 broods at 5°C and 10 broods at 20°C 5c 20°C Sim Sis 6 $& fe t= = + Cc + = + gf $2) 88 #8 = 3 xcs Z © cf Stage (hrs) (%) | (hrs) (%) | 64.1 16 10.1 19 Il 71.0 18 8.1 15 Il 45.8 11 6.1 11 IV 64.1 16 6.9 13 Vv 24.0 6 372. 6 VI 42.7 int 4.7 9 Vil 24.0 6 3.0 6 Vill 65.5 16 113 22) Total 401.2 53.4 stages than would have been expected con- sidering the calculated duration of each stage; he suggested that either egg laying was synchronized for the whole population or that females in the same stage of a reproduc- tive instar tended to congregate in a particu- lar area of the pond. For each of seven samples from Big Island Lake, taken at various times during the study, we deter- mined the number of D. schodleri females carrying embryos of each of the eight embry- Table 2 onic stages, and then compared these actual frequencies with the expected frequencies (assuming random ovulation) of the 20°C laboratory animals, using Chi-square analy- sis (Table 2). Only the 26 June 1967 sample had an actual frequency that was not signif- icantly different (92 per cent level) from the expected frequency. The tendency was for the actual number of one or sometimes two stages to be greatly out of proportion with the expected number, thus influencing the Chi- square analysis. This supports Green’s (1956) suggestion that egg laying in Daphnia can be synchronous. Brooding period Brooding period is the length of time that developing individuals spend in the brood chamber (Anderson and Jenkins 1942). It is not necessarily the same as the embryonic period. Occasionally the developing young are released before the embryonic period has been completed. For example: in our study, although the young were usually released in a stage similar to the in vitro stage at 53 hours, occasionally the embryos would be released in a stage resembling the /n vitro stage at 42 hours. These latter embryos usually survived, completing their develop- ment in the culture medium. Low temperature increased the brooding period of D. schgdleri (Tables 3 and 4) just Number of D. schodleri females from Big Island Lake carrying embryos of a particular embryonic stage for each of the sampling dates Expected Observed Frequencies Frequency 1966 1967 | 20°C July July July May June June July Lab Stage 13 19 27 27 13 26 3 Animals | 4 14 3 18 2 9 6 19 Il 4 3 10 19 10 iy/ 31 15 Ill 26 6 8 9 19 5 14 11 IV 22 11 re 17 39 8 8 13 V 2 7 1 6 2 4 0 6 VI 13 9 2 7 9 4 2 9 VII 1 1 3 2 3 4 1 6 VIII 8 6 1 (0) 21 18 2 72] Total 80 57 41 78 105 69 64 Development of Parthenogenetic Eggs Table 3 Mean duration of brooding periods and instars for Daphnia schodleri primaparous in the sixth, seventh, and eighth instars, at 5°C Sixth Instar Seventh Instar Eighth Instar § 2 5 2 S 2 z 8 $ 3 ee = Me = D Se © D 5 © D D & re) oe D & re) A UG a ® c © eS ONE c © c 2 13) (6 c © - 2 8 2 SE ot ors SRE 5 + 2 9 o © 6 © 6 9 = © o = ss Sick a 3 = (5 =a 0 6 = Sa a6 days no days days no 15.9 10 18.2 6 W710) 16.5 2 1922 6 18.5 18.0 2 20.0 1 170 17.0 1 as it increased the embryonic period. At 5°C, the mean duration of the brooding period varied with the mean duration of the adult instars, the time interval of adult instars increasing progressively with age. At 20°C, where more instars were followed, the brood- ing period duration progressively increased in time during the early reproductive instars, as did the duration of the adult instars. How- ever, by the sixteenth instar the mean duration of the brooding periods had levelled off, exhibiting no further correlation with the progressively increasing time intervals of the adult instars. Since we observed that only rarely were the eggs deposited into the brood chamber 30 minutes or more after a molt, females in very late instars must have fairly long barren periods between releasing the broods and molting again, this barren period increasing as the females become older. In field studies, Daphnia populations are often separated into categories depending on re- productive features, one category being females of adult size but without eggs in the brood chambers; these females are usually considered to be temporarily or permanently sterile. For D. schgdleri, the barren females are more likely to be old, slowly reproducing females instead of sterile females. Development of Parthenogenetic Eggs Table 4 Mean durations of brooding periods and instars for Daphnia schodleri primiparous in fifth instar, at 20°C Mean Mean duration brooding Broods of instar period observed Instar number hours hours no. 5 53.0 51.8 26 6 53.5 Ses 24 7 53.3 50.8 24 8 56.0 52.3 24 9 56.4 53.0 24 10 55.6 51.8 24 11 57.8 5325 24 12 59.3 54.0 24 13 597 5872 24 14 60.4 54.8 22 15 63.3 55.8 22) 16 60.9 54.9 20 17 62.2 56.0 20 18 61.9 55.4 19 19 62.9 55.0 20 20 64.7 5579 19 21 66.4 56.5 18 22 66.3 557 16 23 65.7 55.9 12 24 66.6 56.4 13 25 68.1 56.9 9 26 70.9 56.3 12 27 70.9 56.2 13 28 69.6 54.8 13 29 7872. 5572 14 30 67.9 54.5 9 31 69.6 55.5 11 32 68.1 54.9 6 33 68.6 53.9 6 34 68.2 525i 3 35 69.4 52.4 2 36 72.8 54.5 6 37 WN83 54.8 5 38 79.6 56.6 5 39 UT 55.3 4 40 125.7 55.1 2 Discussion The origin and significance of Daphnia's egg (or embryonic) membranes are not well understood. Shiino (1968) states that cla- docerans have one egg membrane, a vitelline membrane. Lebedinsky (1891) describes a second membrane for D. similis Claus, the second membrane being called a chorion. Obreshkove and Fraser (1940), studying the in vitro development of D. magna partheno- genetic eggs, report two egg membranes, a thin, inner, vitelline membrane and a thicker, outer, egg membrane (equivalent to Leb- edinsky’s chorion); these workers do not mention the casting off of either membrane. Several other workers have observed both inner and outer membranes (sometimes called larval and ovular membranes respec- tively) surrounding cladoceran eggs without describing the shedding of either membrane. During in vitro development of D. magna eggs, Davis (1968) observed the process of size increase and consecutive shedding of two membranes. Esslova (1959) studied the in vitro embryonic development of D. pulex (de Geer) parthenogenetic eggs and found three membranes called the egg membrane (outer) and first and second (inner) embry- onic membranes. All three membranes were observed being cast off, the second embry- onic membrane being cast off during late embryonic development when the caudal spine extends and the embryo increases rapidly in size. Esslova equated the casting off of all three membranes to three prenatal molts. During the development of D. schgdleri eggs, we observed two membranes, both of which were cast off (the cast-off outer membrane is seen in Figure 8); the embryo subsequently increased in size after each cast. Although the exact nature of these membranes is not known, we consider the casting off of the membranes to be part of the hatching process and not true prenatal molts. D. schodleri embryos also increased rapidly in size after extension of the caudal spine, but we did not observe the casting off of a membrane at this time. # Development of Parthenogenetic Eggs Literature Cited Anderson, R.S. (1968). The zooplankton of five mountain lakes in southwestern Alberta. Nat. Mus. Can., Natur. Hist. Pap. 39. Anderson, B.G., and J.C. Jenkins (1942). A time study of events in the life span of Daphnia magna. Biol. Bull.(Woods Hole) 83:260-72. Banta, A.M. (1921). A convenient culture medium for Daphnia. Science(New York), n.s. 53:557-58. Davis, C.C. (1968). Mechanisms of hatching in aquatic invertebrate eggs. Oceanogr. Mar. Biol. Annu. Rev. 6:325-76. Esslova, M. (1959). Embryonic development of parthenogenetic eggs of Daphnia pulex. (in Czech) Acta. Soc. Zool. Bohemoslov. 23(1):81-87 Green, J. (1956). Growth, size and reproduction in Daphnia (Crustacea: Cladocera). Proc. Zool. Soc. London 126:173-204. Lebedinsky, J. (1891). The development of Daphnia from summer ovum. Zool. Anz. 14:149-52 (cited from Obreshkove and Fraser, 1940). Obreshkove, V., and A.W. Fraser (1940). Growth and differentiation of Daphnia magna eggs /n vitro. Biol. Bull.(Woods Hole) 78:428-36. Shiino, S. M. (1968). Crustacea. Pages 333-80 in Matozo Kumé and Katsuma Dan, eds. Invertebrate embryology. (Transl. by Jean C. Dan.) Nolit, Belgrade. Development of Daphnia schédleri eggs and embryos in vitro All pictures were taken under a compound microscope at a magnification of 100 Figure 1 Egg three hours after deposition. Figure 2 Egg at six hours, with a transparent periphery. Figure 3 Egg at nine hours, showing at upper-right surface the first invagination. Figure 4 Egg at 12 hours; the first demarcation of the future cephalic region. A constriction (a transparent spot) at the extreme posterior end marks the beginning of the bilaterally symmetrical plane of development. Figure 5 Egg at 13 hours. Figure 6 Egg at 15 hours, showing a clear constriction at the posterior end and further demarcation of the cephalic portion. Figure 7 Egg at 18 hours; further demarcation of the cephalic and abdominal appendages. Figure 8 Embryo at 21 hours; further demarcation of the cephalic and abdominal appendages and a more prominent bilaterally symmetrical development. The embryo has just cast off the outer egg membrane but it is still enclosed in the inner egg membrane. The cast-off egg membrane is at the side of the embryo. A blastodermic thickening begins to appear at the cephalic region, giving the first external evidence of brain development. Figure 9 Embryo at 22 hours (dorsal view); further demarcation of abdominal appendages and development of prospective brain. The antennae appear as short, forked stubs, closely appressed to the body. The thick, mid-dorsal, longitudinal fold has begun to thin out laterally and posteriorly to form the carapace. Figure 10 Embryo at 24 hours (dorsal view); further development of brain mass, carapace and other structures. 10 Figure 11 Embryo at 24 hours (ventral view), showing the formation of labrum and mandibles. The labrum appears as a round lobe in the cephalic region; the two mandibles appear as two smaller lobes, one on each side of prospective labrum. Figure 12 Embryo at 27 hours (side view), showing the abdominal appendages appearing as stubby blocks of tissue. Brain mass can be seen clearly in the cephalic region. A grayish mass of granular substance appears anterodorsally to the prospective brain mass. Figure 13 Embryo at 27 hours (dorsal view), showing a definite head bulge, and further development differentiations. Figure 14 Embryo at 30 hours (dorsal view); further development of carapace; the caudal spine has now appeared at the posterior end of carapace. Joints of antennal rami are also distinguishable but not in focus. Figure 15 Side view of embryo at 30 hours (ventral side up). Figure 16 Embryo at 33 hours (dorsal view); the appearance of the two small pink eyes, and, posteriorly, the ocellus (the black body in front of the brain mass). Figure 17 Embryo at 33 hours (side view, ventral side up). The ocellus appears as a small pigment body ventral to the brain mass, and two eyes are located just antero-dorsal to the brain mass, although difficult to locate in this picture. Figure 18 Embryo at 36 hours (dorsal view), showing two distinct brown eyes. Figure 19 Embryo at 38 hours (side view), showing an infolding of the cephalothorax. The carapace now covers nearly all the body. The caudal spine is curled ventrad and anteriorly. Figure 20 Embryo at 39 hours (dorsal view); further differentiation of the two eyes. Embryo is still enclosed in the inner egg membrane. 11 172 Figure 21 Embryo at 42 hours (ventral view), after casting the inner egg membrane. The two eyes are nearly fused. Distal portion of antennae have completely separated from the body. Figure 22 Side view of embryo at 43 hours, showing the fused compound eye. The curled caudal spine adheres closely to the postabdomen. The three, terminal, antennal setae are still shorter than their rami. Figure 23 Embryo at 57 hours, representing a fully developed young. The caudal spine has extended, and the three terminal, antennal setae have become longer than the terminal ramus. Part Il Vital Statistic Properties of Laboratory Populations Abstract The female offspring of Daphnia schodleri Sars when cultured in the laboratory at 5°C, 20°C, and 25°C, had four to seven preadult instars; males had three to four preadult instars. At 25°C the average longevity was 36 days (18 instars) for females, and 41 days (18 instars) for males; at 20°C the average longevity for females was 52 days (21 instars). As the initial size (total length in first instar) of neonate females increased, they tended to become mature in earlier instars. For females, the largest absolute growth increment was at the molt between the adolescent and primip- arous instars; for males this increment was between the preadolescent and adolescent instars. The greatest relative growth incre- ment for females was prior to the preadoles- cent instar; for males the increment was between the preadolescent and adolescent instars. At 20°C most of the growth of females was achieved by day 10, whereas the growth curves for 5°C females did not level off until day 40. The coefficients of variation for carapace length, height, and total length were highest in the preadult instars for both males and females; there was no evidence that carapace length or height was a less variable measurement than total length. Fitting data of these linear dimensions to the allometric equation indicated that both male and female carapace length and female height increased in size relatively more rapidly than total length; the relationship between male height and total length was isometric. The first four instars of both males and females were easily determined from size—frequency distributions (when size-class limits were narrow), but the association between instars and size modes was not so clear-cut in later instars. At 25°C, the mean number of young per brood was greatest in the fourth adult instar; at 20°C there were two peaks of young production, one in the eighth adult instar and another in the thirty-first adult instar. For a 110 day unit of time, mean total young production was 48 at 5°C, 378 at 20°C, and 234 at 25°C. Larger females produced more and larger young during the first adult instar than smaller females. There was a tendency for the size of the young to increase with increasing age (in instars) of females when the population was followed through 12 instars at 20°C; but for the one female followed for 34 instars, there was a sharp decline in average size of young after the twenty-third instar. In the laboratory D. schgdleri produced sexual eggs only at 5°C. All females produc- ing sexual eggs had a sterile instar immedi- ately following the ephippial instar. 13 Vital Statistic Properties of Laboratory Populations Introduction Part Il of the study of Daphnia schgdleri Sars, deals with the laboratory study of various life history phenomena (vital statistics) of growth, reproduction and related features. We were interested in comparing vital statistic proper- ties of females with those of males, and also D. sch¢dleri’s properties with those of other species. In addition, we felt that certain aspects of the laboratory study might be valuable for interpreting aspects of D. schodleris biology in the lake. This last- mentioned aspect is covered in Part Ill. Part | covered the /n vitro development of parthe- nogenetic eggs, duration of embryonic stages in the brood chamber, and the duration of the brooding period. 14 Materials and Methods Laboratory experiments were carried out under three temperature conditions: 5°+1°C, 20°+1°C, and temperatures fluctuating between 22°C and 29°C with a mean tem- perature of 25°C; the three temperature conditions are hereafter referred to as 5°C, 20°C and 25°C. The male and female D. schogdleri used in the culture experiments at 25°C were descendants of a single partheno- genetic female collected from Big Island Lake on 22 June 1966. This field female, shortly after being brought into the laboratory, released five female neonates; the subse- quent offspring (the second generation of the field female) of these five females were used in the laboratory experiments at 25°C. After releasing the five female neonates, the field female released 31 eggs into the brood chamber; these eggs were dissected out and allowed to develop in filtered aquarium water. All of the 24 eggs that hatched developed into males. These were the males used in the laboratory experiments at 25°C. D. schodleri females used in the experiments at 20°C and 5°C were the third generation of a parthenogenetic female taken from Big Island Lake on 3 July 1967. The young daphnids were isolated a few hours after being released from the brood chambers and cultured individually in 25 ml of the diluted medium of Banta’s manure-soil stock (Banta 1921). On alternative days, the total volume of the medium was changed. Measurements were made at the time of isolation and daily thereafter by placing the individuals in a depression slide with a drop of culture medium. One drop of saturated chloretone solution was added to impede activity of the 25°C animals; this was not needed at 20°C and 5°C. Measurements were made with a calibrated ocular microme- ter. Total length was the distance from the apex of the head to the base of the spine; carapace length was the greatest length of the carapace exclusive of the spine; height was the shortest distance between the two lines tangential to the carapace. D. schodleri cultured at 25°C and 5°C were examined and appropriate observations made at least once a day. The animals kept at Vital Statistic Properties of Laboratory Populations 20°C were observed hourly from O800 to midnight each day from the time the animals passed as eggs into their mother’s brood chamber to the end of their adult life. Preadult Instars The number of preadult instars for female D. schgdileri varied from four to seven. There was a tendency for females to become primiparous in earlier instars at higher tem- peratures (Table 1). When first primiparous, females ranged in size from 1.44 to 2.41 mm; those becoming primiparous in early instars (e.g. fifth) were generally smaller than females becoming sexually mature in later instars (e.g. eighth). For the same tempera- ture and primiparous instar, the seventh, the mean size of ephippial females was smaller than that of parthenogenetic females. At 20°C (the temperature at which the majority of animals were cultured) there was a relationship between size in the first instar and the instar number in which sexual maturity was reached (Table 2). The larger the female in the first instar, the fewer the number of preadult molts. Also, for a given primiparous instar, there was a positive correlation between size in the first instar and size in the first adult instar. For example, for the 57 females primiparous in the fifth instar at 20°C, the correlation coefficient was 0.7807 and is significant at the 99 per cent level. Males were followed from the first instar to sexual maturity only at 25°C, 21 males becoming sexually mature in the fourth instar and two in the fifth instar. Sexual maturity in males was determined by the presence of sperm in the testes, the shape of the ventral margin of the carapace, and the size of the hook on the first thoracic leg. Vital Statistic Properties of Laboratory Populations Table 1 Initial size, and size in first adult instar of female Daphnia schodleri Number of First clutch Mean length in Mean length in animals of eggs laid first instar first adult instar Temperature instar mm mm 25°C 22) 5 0.548 1.640 8 6 0.544 1.739 20°C 57 5 0.640 1.798 27 6 0.587 1.805 1 7 0.488 1.755 5G 6 6 0.700 2.075 10 7 0.619 2.199 3 ie 0.644 1.911 2 8 0.634 2.308 *Produced ephippia Table 2 Length of young in first instar and instar in which they reached maturity, at 20°C Number mature in 5th instar 6th instar 7th instar Length in first Number instar (mm) reared no. % no. % no. % 0.455-0.488 1 1 100.0 0.488-0.520 3 2 66.7 1 Sons 0.520-0.553 7 2 28.6 5 71.4 0.553-0.585 2 8 66.7 4 33.3 0.585-0.618 8 5 62.5 3 3719 0.618-0.650 30 22 73.3 8 26.7 0.650-0.683 9 6 66.7 3 3373 0.683-0.715 8 7 87.5 1 125 0.715-0.748 2 2 100.0 0.748-0.780 1 1 100.0 0.780-0.813 4 4 100.0 16 Vital Statistic Properties of Laboratory Populations Longevity At 25°C, males and females had approx- imately the same life-span and went through almost the same number of instars. (Fig. 1). In respect to instars, mortality of females was fairly constant throughout the life-span; for males, mortality was greatest between the fifteenth and twentieth instars. Forty-six females were also followed at 20°C. When compared to the population at 25°C, females at 20°C lived much longer and went through many more instars, the mean life-span in days and instars being 52 and 21 respectively, with maximum longev- ity of 113 days and 41 instars. In addition, one female at 5°C survived for 207 days. The 41 instars (for two females) of D. schgdleri at 20°C possibly represent the highest number of instars recorded for a species of Daphnia. Possibly the large num- ber of instars observed in the laboratory was due to a lack of crowding in our cultures, each animal having been cultured individu- ally in a small dish. Growth Growth of females and males at 25°C Figure 2 shows the mean growth curves of total length, carapace length, and height for eight females primiparous in the sixth instar at 25°C. Growth curves for the three morph- ometric dimensions were of the same shape. D. schgdleri females in the experiment had an average total length of 0.52 mm immedi- ately after being released from the brood chambers; for the remainder of the life-span, females increased 2.41 mm in mean total length, of which 1.57 mm, or 65 per cent was attained by the eighth instar. For females, absolute growth increments for total length increased through the fifth instar, then gradually decreased until the fifteenth instar, and fluctuated for the re- mainder of the life-span (Fig. 3). The maximum absolute growth increment there- fore was at the molt between the adolescent (fifth) and primiparous (sixth) instars for D. schodleri females. However, relative growth increments of total length were greatest in earlier instars, between instar two and three and between instar three and four (Table 3); relative growth increments then decreased rapidly for the remainder of the females’ life- span. Anderson, Lumer and Zupancic (1937), by calculating coefficients of correlation, determined possible relationships for D. pulex females between (a) initial size (i.e. total length in first instar), (A) final size (total length in twentieth instar), (t) duration of growth (the number of instars required to attain a length of 0.8A), and (i) initial velocity of growth (increment between first and second instars). They found possible inverse relationships between initial size and initial velocity of growth, and between duration of growth and final size. We performed similar calculations using data for the eight female D. schgdleri that lived for at least 20 instars at 25°C, and obtained the following coeffi- cients of correlation: ran = 0.1792, r,=0.5965, 1,4, = 0.3338, fa = -0.5157, and ra,=0.6601. Vital Statistic Properties of Laboratory Populations Table 3 Relative mean growth increments of total length for D. schodleri males and females of Figure 2 Mean relative Number increment (%) of animals Instars females males females males 1-2 26.9 11.8 8 8 2-3 28.8 24.7 8 8 3-4 30.6 Se 8 8 4-5 25.9 8.9 8 8 5-6 2077 52 8 8 6-7 10.1 375 8 8 7-8 Ihre 2.8 8 8 8-9 6.2 27 8 8 9-10 5.4 372 8 8 10-11 3.0 322 8 8 11-12 2.9 2/5 8 8 12-13 2.4 1.8 8 8 13-14 2.4 128 8 8 14-15 273 2.3 8 8 15-16 el sill 8 8 16-17 ileal a7 8 8 17-18 1.8 let 8 8 18-19 1.8 1.6 8 7 19-20 0.7 1.6 8 6 20-21 0.1 1. 6 4 21-22 1.4 1.6 6 4 22-23 0.2 i158) 5 4 23-24 -0.1 -1.5 5 2 24-25 0.3 145 4 2 25-26 0.1 15 4 1 26-27 1.6 3 None was significantly different from zero (t test, 95 per cent level), indicating no evident relationships among these characteristics in D. schgdleri females. The mean growth curves for the three morphometric dimensions of males were all of the same shape (Fig. 2). The male growth curves were less sigmoid in shape than were the growth curves of females. The male D. schgdleri had a mean total length of 0.76 mm in the first instar; by the eighth instar they had a mean total length of 1.49 mm, which was 73 per cent of their final mean total length. The inflection point of each curve came earlier for males than for females. It was found, in measuring total length, that both the maximum absolute growth incre- ment and the maximum relative growth increment came between the preadolescent (second) and the adolescent (third) instars (Fig. 3 and Table 3). The degree of interdependency, if any, between initial size, final size, duration of growth, and initial velocity of growth was also tested for males, using data for the six males that lived for 20 or more instars at 25°C. The following coefficients of correlation were obtained: ran = 0.5787, r;=-0.2697, r,,=-0.0623, fa = -0.2501, and r,, = 0.5237. The results were the same as those described above for the females—none of the male coefficients of correlation was significantly different from zero. Growth of females primiparous in different instars at 20°C and 5°C High temperature increased the growth rate of D. schgdleri by shortening the duration of each instar, growth per day being much more rapid at 20°C than at 5°C (Fig. 4). At 20°C most of the growth was achieved by day 10, whereas the growth curves for females at 5°C did not level off until about day 40. At the same temperature, 5°C, the growth curves for females becoming primiparous in different instars were similar, except for females becoming primiparous in the seventh instars and producing ephippial eggs instead of parthenogenetic eggs. For these ephippial females, growth was slower, especially from the adolescent instar (instar six at day 36) on. Although there was much more growth per day at 20°C, growth at the end of the eleventh instar was slightly greater at 5°C (Fig. 5). Observations at 5°C were termi- nated when the females were in the eleventh and twelveth instars; hence their final size is not known. By extrapolating from the growth curves of Figure 4, it is estimated that the 5°C females, with the exception of ephippial females, would have to live for approximately 170 days to attain a final size equal to the final size of the 20°C animals. Such a life- span for D. schgdleri at 5°C is certainly possible (See ‘‘longevity’’, p.17), at least for laboratory populations. The slower growth of ephippial females from the adolescent instar Vital Statistic Properties of Laboratory Populations on, both per day and per instar, would seem to support Berg’s depression hypothesis (1934); i.e. ephippial females are in a state of depression. Relationships and variability of linear dimensions Total length is the most common linear measurement used to determine size-fre- quency distributions of Daphnia; total length is also usually used for most of the other ecological and physiological work of Daphnia, where an index of body size is needed. Because body shape will vary between species and may vary within the same species (obviously so for those species exhibiting cyclomorphosis), total length may not in all cases be the best index of size increase for Daphnia. Carapace length and height (and body weight) can also be used as indices of Daphnia’s body size. Figure 2, p.29 gives no indication that the growth of the carapace and increase in height for both male and female D. schodleri are different from increase in total length. The relationships between carapace length and total length and between height and total length were nearly linear when data of Figure 2 were plotted logarithmically (Fig. 6). Using the least-square method, data for these relationships were fitted to the power, or allometry, equation, using the log form log. Y =log,b + klog,X where b is the value of Y (carapace length or height), when X (total length) equals unity, and k is the ratio of the specific growth rates of Y and X. The values of log ,b and k (and k’s 95 per cent confidence intervals) are shown in Figure 6. The relation between male height and total length is isometric (k = 1); the other relationships are nearly but not exactly isometric, both male and female carapace length and female height increasing in relative size slightly more rapidly than total length. The above constants are based on the assumption that simple allometry, for the linear dimensions tested, holds throughout the life-span of D. schodleri. There is some indication from Figure 6 that there are deviations from simple allometry, especially during the early instars. However this is not treated further in the report. We also examined the relative variability of the three linear dimensions for the eight female and eight male D. schgdleri of Figure 2, p.29, calculating the linear dimensions’ coefficients of variation (CV) for each instar (Fig. 7). There was no evidence that either carapace length or height was a less variable measurement than total length; in fact for the preadult instars, especially males, total length was the least variable of the three measurements. CV values of all measure- ments were highest in the preadult instars for both males and females, and it was in the preadult instars that both the maximum absolute and maximum relative growth increments occurred. Are any of these relatively high CVs of the early instars statistically significant? Lewontin (1966) showed that the variance of the logarithms (natural or common) of measure- ment gives a measure of relative variability that can be used for statistical tests; he also pointed out that for CVs of 30 or less, the square of the CV closely approximates the variance of natural logarithms. Squaring each CV value of each linear dimension used to construct Figure 7, we tested the following hypothesis for each sex of D. schodleri: within a particular instar none of the linear dimension’s squared CVs is significantly larger (one-sided ‘’F’’ test, 95 per cent level) than the smallest squared CV of that stage. Significant variability was found only in the first two instars of males, i.e. height (vs. total length) in the first instar, and height (vs. total length) and carapace length (vs. total length) in the second instar. Therefore, of the three linear dimensions, total length would be the most satisfactory measurement for docu- menting changes in D. schgdleri’s body size. Size-frequency distributions In field studies of Cladocera life histories, information is often obtained by analyzing size—frequency distributions, i.e. plotting the number of individuals against size, usually total length, with the resulting graph exhibit- ing a number of size modes, which are taken as growth stages or instars. We compared the 19 Vital Statistic Properties of Laboratory Populations results obtained by size-frequency distri- butions, where discrete instars are not known, with size-frequency distributions of individuals of discrete instars. For each of the first seven instars, total lengths of 32 females reared at 25°C were measured to the nearest 0.003 mm (0.1 micrometer unit) and the females were grouped into 0.06 mm (two micrometer units) size classes by instar, resulting in a size-frequency distribution by discrete instars. Then the 32 total-length values for each of the seven instars were lumped together. This resulted in a ‘‘com- posite’’ size—frequency distribution, which would be similar to that obtained from field samples where discrete instars are not known. The same procedure was used for 24 males reared at 25°C. Results are shown in Figure 8. The modes of the composite distribution did not correspond exactly to the mean total length of the instars. However, for the first four instars of both females and males, each instar is easily recognized from a correspond- ing distinct mode in the composite distri- bution; in later instars this relationship is not so clear-cut, male instars five through seven being almost completely indistinguishable in the composite distribution. Overlapping of the values of the discrete instars tends to increase with instar number. If the size—class limits were to be expanded, e.g. from 0.06 mm to 0.10 or 0.25 mm, overlapping would be accentuated and the composite curve would be further smoothed; consequently it would be very difficult, if not impossible, to associate distinct instars with modes of the composite curve. And, of course, this would be the case if the sexes were not separated, assuming males made up a substantial part of the field population. In short, for D. schgdleri, size-frequency distributions are most indicative of early instars when the size—class limits are rather narrow. 20 Reproduction Natality throughout the life-span of D. schgdleri Natality of D. schgd/eri was based on the number of young released from the brood chamber during each instar. The number of young released may be smaller than the actual number of eggs produced per instar, since occasionally non-viable eggs were found, especially during the later instars. But it was impossible to count accurately the number of eggs in the brood chambers of living D. schgdleri; and since non-viable eggs should not be included in the count it was felt that term young production, instead of egg production, was more appropriate for this section. Also it was established that the size of the young, when released from the brood chamber, was proportional to the size of the eggs. This was done by dissecting out the eggs from 10 females, determining individual egg volumes, and allowing the eggs to develop in filtered aquarium water; the total length of each of the newly hatched young was then measured, and the results, given in Table 4, indicate that the size of the young is correlated with the size of the egg. The calculated coefficient of correlation was 0.970 and is significant at the 99 per cent level. Generally, the mean number of young produced per brood at 25°C increased progressively during the early adult instars Table 4 Relationship between egg size and size of young for each of 10 D. schgdleri females Number Number Mean egg Mean length of eggs of young volume (mm) of newly measured measured (mm°x10%) hatched young 13 13 6.175 0.639 12 11 5.540 0.631 9 5 5.423 0.599 12 12 7.672 0.683 7/ 5 5.691 0.618 10 9 8.440 0.703 10 9 6.006 0.615 6 6 11.012 0.771 14 13 7.440 0.677 10 6 5.138 0.563 Vital Statistic Properties of Laboratory Populations and then gradually decreased in the later instars (Fig. 9). The mean number of young per brood was greatest in the fourth adult instar; the mean number per brood then fluctuated for the following 10 instars and thereafter rapidly decreased. The rapid decrease in mean number of young per brood coincided with the initial and thereafter continuing mortality of the laboratory popu- lation (Table 5). The sequence was different for the population that was followed at 20°C, and which lived for many more instars. In that case there were two peaks of young production, one in the eighth adult instar and another in the thirty-first adult instar, when the laboratory population was quite old. The Table 5 Mortality of laboratory populations at 20°C and 25°C used to estimate mean number of young per brood (Figure 9) Adult instar Number of number survivors 20°C population 1 31 2 30 3-9 29 10 28 11-12 27 13-14 25 15 23 16-17 20 18-20 19 21 18 22-24 1) 2/ 25 16 26-27 15 28 14 29-30 12 31-32 11 33 10 34 8 35 36 2 S37/ 1 38 O0 25°C population 1-15 8 16-17 6 18 5 19-20 4 21 3 22 3 23 (0) reason for the second peak is unexplained. Of the original 31 females, there were 11 still living and reproducing at the time of the second peak (Table 5). An examination of fecundity records for each of the original 31 females gives no indication that the peak was an artifact due to the dying off of less reproductive females, which, had this been the case, would have negatively influenced the mean brood size for a particular instar. The peak was real in the sense that the surviving females had larger individual broods at this time. The experiment at 20°C extended over 15 weeks and, although there was no change in ‘‘crowding’’ since the females were cultured individually, the culture medium was changed every other day. Perhaps, in some way unknown to us, the constituents of the culture medium varied at the time of the second peak, changing the food level and resulting in increased fecundity. The mean total young production per female (including all females, not just the mean total young of surviving females) at 25°C and 20°C was 234 and 378 respec- tively—the mean number of young per brood being 12.5 at 25°C and 16.3 at 20°C. Females at 25°C produced fewer young per brood than females at 20°C and because they did not live as long as those at 20°C, their mean total egg production was consid- erably less than that for the 20°C females. Broods occurred on the average every 2.7 days at 20°C and 18.1 days at 5°C (Fig. 10). Low temperature considerably delayed the onset of reproduction. At 20°C, the females, on the average, released their first brood seven days after being released from the brood chamber themselves; at 5°C the average time for this process was 64 days. Average total young production per unit time was, of course, greater at 20°C than at 5°C. For example, females 110 days old at 20°C had an average total young production of 378. For the same unit of time but at 5°C, average total young production was 67 for those females primiparous in the sixth instar, 44 for those primiparous in the seventh instar, and 34 for females primiparous in the eighth instar. These values for 5°C females also suggest that at the same temperature 21 Vital Statistic Properties of Laboratory Populations there is possibly greater total production for females becoming primiparous in early instars. In short, 20°C seems to be the optimum temperature (of the three tested) for young production. At 5°C, because of less frequent molts, young production per unit time was considerably less than at 20°C. At 25°C, broods per unit time were not recorded, but the longevity data for 25°C females (Fig. 1, p.28) indicate only slightly more instars per unit time (2.3 days per instar) than for those at 20°C. And, as indicated above, 25°C females produced fewer young per brood and did not live as long as 20°C females. Sizes of females in first adult instar and number and size of young produced There were variations in the number of young produced in the first adult instar and these variations were positively correlated with the size of the female in the first adult instar (Table 6). The correlation coefficient for the 55 females primiparous in the fifth instar at 20°C was 0.732 and is significant at the 99 per cent level. The correlation holds for other primiparous instars: for 27 females primipa- rous in the sixth instar (20°C) and 28 females primiparous in either the fifth or sixth instar (25°C) the correlation coefficients were 0.866 and 0.653 respectively, both being significant at the 99 per cent level. Not only do larger females produce more young per brood in the first adult instar than smaller females, they also produce larger young. The correlation coefficient between the size of females in the first adult instar and the size of young produced by these females was 0.685 and is significant at the 99 per cent level. There was a tendency for the size of the young to increase with increasing age (in instars) of the female (Fig. 11). This would be expected since the older females are larger. However this relationship did not hold for the one female that was followed through 34 instars. For this female, there was a sharp decline in average size of young after the twenty-third brood (twenty-seventh instar), even though the female continued to increase slightly in total length during its life-span. 22 Table 6 Female size in first adult instar, and number and size of their young in first adult instar* . Mean Mean Mean Number length number length of (mm) of of (mm) of females females young young 1 1.44 3.0 0.55 2 1.56 DA5 0.54 1 157 4.0 0.57 1 1.59 8.0 0.53 1 1.60 8.0 0.54 3 1.63 6.7 0.55 3 1.66 4.3 0.61 1 1.68 2.0 0.62 1 1.69 3.0 0.60 1 1.70 5.0 0.57 6 1772 4.7 0.60 1 1573 4.0 0.59 2 1.76 4.5 0.62 5 1.79 5.8 0.59 1 1.80 4.0 0.63 1 1.81 6.0 0.62 7 1.82 5.6 0.59 1 1.83 5.0 0.61 5 1.85 5.0 0.62 1 1.88 7.0 0.62 1 1.89 9.0 0.56 1 1.93 10.0 0.59 2 1.98 95 0.59 1 1.99 9.0 0.58 1 2.20 8.0 0.67 1 2.21 8.0 0.67 2 2.24 11.0 0.66 1 2.28 14.0 0.65 *Based on 55 females primiparous in the fifth instar at 20°C. Production of sexual eggs In the laboratory, D. schgd/eri produced ephippia only at low temperatures; seven females produced sexual eggs at 5°C without the presence of males. These ephippial females were followed through a varying number of instars (Table 7). Four females, in addition to producing sexual eggs, also produced at least one brood of parthenoge- netic eggs (all of which subsequently devel- oped into females), while the other three females produced only sexual eggs. The unfertilized sexual eggs were followed for two months at 20°C, but they did not hatch. All seven females had a sterile instar immedi- Vital Statistic Properties of Laboratory Populations Table 7 Reproductive features of seven females that produced at least Animals 6 7/ 8 1 8 E O 2 13 {177 3 10 E 4 10 E 5 E 0 6 E 0 7 E 0 one ephippium at 5°C* Instars mmmoownm|o = J | i O A sterile instar; neither sexual nor parthenogenetic eggs were produced. E The production of an ephippium and ephippial eggs. dead The death of the mother animal. = No further observations. *The numbers shown in each column represent the number of parthenogenetic young produced in the indicated instar. ately following the ephippial instar. This is different from observations of other workers, e.g. Berg (1931), where no sterile instars were found, the daphnids producing either sexual or parthenogenetic eggs in the instar immediately following the ephippial instar. In short, in the laboratory, individual D. schgdleri producing sexual eggs may or may not also produce parthenogenetic eggs sometime during the life-span; even for those ephippial females that do subsequently (or did previously) produce parthenogenetic eggs, there is no strict alteration of the two types of reproduction in the sense of being predictable. The significance of the sterile instar is unexplained. Since, for many aspects of Daphnia's biology, it is desirable to know if the females are about to produce or have in the past produced ephippia, we include here the description of ephippial formation for D. schgdleri. Females that are to produce sexual eggs (and an ephippium) in the next instar usually can be recognized by the presence of a compact dark mass of small fat globules in the ovaries. The mass at first is very small and consists of closely packed small fat globules surrounded by larger fat globules. Near the end of the pre-ephippial instar, the compact dark mass has grown quite large, and it occupies nearly all of the ovary. The female soon undergoes ecdysis and the new exoskeleton of the female, which is now in the ephippial instar, has an indention on the dorsal margin at the head-carapace junction (Fig. 12A). The dorsal part of the carapace, which will eventually develop into the ephip- pium, now is light brown or gray in color, and it is separated from the other part of the carapace by an irregular line. Very shortly, two fully developed sexual eggs are depos- ited into the modified dorsal part of the carapace (Fig. 12B). The modified dorsal part of the carapace now becomes darker and is gradually pushed upwards (Fig. 12C). Its separation from the remainder of the cara- pace becomes evident and eventually the modified dorsal part is completely freed, by the mechanics of molting, from the rest of the carapace. The female, after molting and discarding the ephippium, still possesses an indention on the dorsal margin (Fig. 12D). 23 Vital Statistic Properties of Laboratory Populations Table 8 Vital statistic properties of D. schodleri s laboratory populations, and vital statistics of other species as gathered from selected reports D. schodleri, present study Other studies Number of preadult instars Maximum longevity Greatest absolute growth increment Greatest relative growth increment Relative growth Instar with greatest relative variability Adult instar with largest number of young or eggs Maximum number of young or eggs per brood Mean number of young or eggs per brood 24 4-7 (89), 3-4 (éd) 28 instars, 65 days (99, 25°C) 41 instars, 113 days (99, 20°C) 26 instars, 68 days (6¢,25°C) Between adolescent and primiparous instar (2%); between preadolescent and adolescent instar (dd). Prior to preadolescent instar (99); between preadolescent and adolescent instar (dd). Carapace length vs. total length, k= 1.059 (99), 1.098 (dé); height vs. total length, k= 1.079 (99), 0.972 (4). Carapace length: fourth instar (99), second instar (dd); height: third instar (22), second instar (dé); total length: third instar (2%), second instar (dé). Fourth (25°C), eighth and thirty-first (20°C) 24 (25°C), 39 (20°C) 1205) (25°C);, 116537 (20;C) 4 (99) D. galeata mendotae!, D. laevis2, 3-5 (99)D. obstusa3; 4-5 (22) D. pulex4, D. curvirostris3, D. schgdleri®; 4-6 (9%) D. thomsoni?; 5-6 (99) D. atkinsoni3; 4-8 (99) D. magna®*7 22 instars, 54 days (99 D. magna’, 25°C); 202 days (8°C), 150 days (10°C), 99 days (18°C), and 57 days (28°C)(22 D. magna®); 26 instars, 149 days (99 D. schgdleri®, 16°C); 179 days (8°C), 150 days (10°C), 92 days (18°C), and 46 days (28°C) (dé D. magna®). Between adolescent and primiparous instar (92 D. pulex*) (22 D. magna®); between preadolescent and adolescent instar $ D. curvirostris and others?; species with both the above and even prior to preadolescent instar?. Height vs. total length, k= 1.09, 1.03, and 0.315, for three growth stanzas, D. pulex?; k=1.13 and 1.05 for two growth stanzas, D. magna®. Seventh D. magna'®, fifth D. magna’, sixth D. laevis? 105, D. magna'®; 36, D. laevis? 6.3, D. schodleri® (16°C); 5.8-23.1, D. laevis? (depending on the medium); 6.7-25.9, D. laevis? (depending on the clone). Vital Statistic Properties of Laboratory Populations D. schgdleri, present study Other studies Magan. total 48 (5°C), 378 (20°C), 234 (25°C) number of young or eggs for 110-day period Hall 1962. Ingle et al. 1937. Green 1956. Anderson, Lumer and Zupancic 1937. LeSuer 1959. où À © NN — Discussion Table 8 summarizes the vital statistic proper- ties of the D. schodleri laboratory populations and also summarizes values for other species, as gathered from selected reports. One must use caution when comparing interspecific values of different studies, even at the same temperature, because of the culture medi- um's food level, which in most studies is unquantified, and which can affect most of the vital properties. Even when the laboratory culture conditions are adequately quantified, equating laboratory food levels with those of the field is difficult; hence one must also use the utmost caution in associating laboratory life history phenomena with life history phenomena of field studies. The instar in which the female becomes primiparous is important in influencing many other life history phenomena. For example, D. schgdleri females becoming primiparous in early instars were generally smaller than females becoming sexually mature in later instars; large female neonates in the first instar became primiparous in earlier instars than did smaller neonates. At 5°C, females becoming primiparous in early instars pro- duced more young during their life-span than did females becoming primiparous in late instars. Within a specific primiparous instar, the larger females produced more and larger young per clutch than did the smaller females. In short, by knowing the primipa- rous instar, much of the variability of life history properties can be accounted for. Determining and considering the primiparous instar should be as much a part of a 36 (8°C), 49 (18°C), 15 (28°C), D magna®; 150 (25°C), D. magna’; 138 (16°C), D. schodleri®; 1,072, D. magna 9 (one individual). Anderson 1932. Anderson and Jenkins 1942. MacArthur and Baillie 1929. Hersh and Anderson 1941. Kerhervé 1927 (cited in Hutchinson 1967:581). oo ON OD controlled laboratory experiment as control- ling the temperature or food level. There are not many studies dealing with the biology of male Daphnia. We found that male D. schodleri differed from the females mainly in respect to various growth proper- ties. The survivorship curves of males and females at the same temperature are similar, the males being as long-lived as the females and having about the same number of instars. But at the same temperature, females became primiparous from instars five to eight, while males became sexually mature in earlier instars, mainly in the fourth instar. These differences in time of sexual maturity influenced most of the other growth parame- ters studied and compared. The absolute growth curves for height, carapace length, and total length of females had, because of the later onset of sexual maturity, a later inflexion point; also the growth curves (in instars) of females were more typically sig- moid than those of males. Related to this and also influenced by the onset of sexual matur- ity, the absolute and relative growth incre- ment curves of males and females were different. For males, both the greatest abso- lute and relative growth increments came at the molt between the preadolescent and adolescent instars; for females, the greatest absolute increment was between the adoles- cent and primiparous instars, and the largest relative growth increment came even before the preadolescent instar. Pertaining to relative growth per se, the length of the male’s carapace increased relatively more rapidly than did total length, more so than the same phenomenon in 25 Vital Statistic Properties of Laboratory Populations females. But height of males, as might be expected considering the shape and smaller adult size of males, increased relatively less rapidly than did total length; in contrast the height of females increased relatively more rapidly than did total length. As would be expected considering the nature of coefficient of variation values, the relative variability of the values of the three linear dimensions — carapace length, height, and total length — was much greater for both males and females during the preadult instars. The relative variablility of the females’ dimensions ex- tended over a longer time (in instars) and this is probably accounted for by the later onset of sexual maturity of females. In the middle and late adult instars, the linear dimensions’ relative variability was quite similar for both sexes. 26 Vital Statistic Properties of Laboratory Populations Literature Cited Anderson, B.G. (1932). The number of pre-adult instars, growth, relative growth and variation in Daphnia magna. Biol. Bull.(Woods Hole) 63:81-98. Anderson, B.G., and J.C. Jenkins (1942). A time study of events in the life span of Daphnia magna. Biol. Bull.(Woods Hole) 83:260-72. Anderson, B.G., H. Lumer and L.J. Zupancic, Jr. (1937). Growth and variability in Daphnia pulex. Biol. Bull.(Woods Hole) 73:444-63. Banta, A.M. (1921). A convenient culture medium for Daphnia. Science(New York), n.s. 53:557-58. Berg, K. (1931). Studies on the genus Daphnia O.F. Miller, with special reference to the mode of reproduction. Vidensk. Medd. dansk Naturhist. Foren. 92:1-222. (1934). Cyclic reproduction, sex determination and depression in the Cladocera. Biol. Rev.(Cambridge) 9:139-74. Green, J. (1956). Growth, size and reproduction in Daphnia (Crustacea: Cladocera). Proc. Zool. Soc. London 126:173-204 Hall, D.J. (1962). An experimental approach to the dynamics of a natural population of Daphnia galeata mendotae. Ph.D. Diss., University of Michigan. 76 pp. Hersh, A.H., and B.G. Anderson (1941). Differential growth and morphological pattern in Daphnia. Growth 5:359-64. Hutchinson, G.E. (1967). A treatise on limnology. Vol. Il. Wiley and Sons, New York. 1,115 pp. Ingle, L., T.R. Wood, and A.M. Banta (1937). A study of longevity, growth, reproduction and heart rate in Daphnia longispina as influenced by limitations in quantity of food. J. Exp. Zool. 76(2):325- 52: Kerhervé, J.B. de (1927). La descendance d'une Daphnie (D. magna) ou ses millions de germes en une saison. Ann. Biol. Lacustre. 15:61-73. (Cited in Hutchinson 1967.) Lei, C. (1968). Field and laboratory studies of Daphnia schgdleri Sars from Big Island Lake, Alberta. M.Sc. thesis., University of Alberta. 149 pp. LeSuer, B.W. (1959). Life history and ecology of Daphnia pulex spp. pulicoides Woltereck 1932. M.Sc. thesis., Montana State College. 33 pp. Lewontin, R.C. (1966). On the measurement of relative variability. Syst. Zool. 15:141-42. MacArthur, J.W., and W.H.T. Baillie (1929). Metabolic activity and duration of life. J.Exp. Zool. 53(2,3):221-68. 27 100 90 80 70 2 o —~ 60 6 > 5 50 5 n 40 30 20 10 oO Instars 2 OF © > 2 2 n 5 10 15 20 25 30 35 40 45 50 55 60 65 70 Days Figure 1 Survival curves for male and female D. schgdleri, in instars and days. Data based on 20 males and 14 females at 25°C. 28 € ko 1.0 H 0.5 == MALES € =. 0 mec 6) 5 10 15 20 25 30 a = INSTAR CD. T Za Eanes e 2.0 H 1.5 1.0 T: TOTAL LENGTH C: CARAPACE LENGTH H: HEIGHT 0.5 FEMALES O (0) 5 10 15 20 25 30 INSTAR Figure 2 Mean growth curves based on data of eight females that were primiparous in the sixth instar and lived for at least 20 instars at 25°C and eight male D. schgdleri that lived for 18 or more instars at 25°C. 29 0.20 ——— TOTAL LENGTH oa CARAPACE LENGTH Se Rs HEIGHT 0 5 10 15 20 25 30 0725 0.20 O on INCREMENT IN LENGTH PER INSTAR (mm) © = 0 5) 10 15 20 25 30 INSTAR Figure 3 Absolute growth increment curves for the males and females of Figure 2. The increment values of a particular instar represent the absolute increase in size between that instar and the instar to follow, e.g. the value at instar 5 represents the increase between 5 and 6. 30 ost ott oot ‘S66a onaueb -oueyued jo peeysui eiddiyde Buisnpoid inq ‘9,g ye Je]SUI YJUBASS au} UI SNOJediwud sajewWay aaiy) ‘3 ‘2.8 ye 1eJSU1 UjyBie eu] ui snozeditunid sajeway om) ‘q (s4op) aby 06 os OL 09 os OF ‘9.G 12 12]SUI YJUaAeS ay} Ul SNOJediwid sajewa OL ‘2 ‘9,.G 12 Je}SUI YIXIS ay} UI SNOJediWd sajewa xIS ‘g ‘9,02 12 seysul yyy eu} Ul SNOJediwud sajeway LE ‘y ‘S4e}SuUl a}edIpul S}OP BY} “DG pue 7,07 ye WajpPyIs ‘Gg ajewaj jo SAep ul SaAIN2 YyMOIDH + anbiy 0€ 0G OT Buse] (ww) 31 (mm) Length Figure 5 Growth curves in instars of female D. schgdleri at 20°C and 5°C. A, six females primiparous in the sixth instar at 5°C; B, 10 females primiparous in the seventh instar at BSG: C, two females primiparous in the eighth instar at 5°C; D, 31 females primiparous in the fifth instar at 20°C; E, three females primiparous in the seventh instar at 5°C, but producing ephippia instead of partheno- genetic eggs. 32 Instar Æ 20 c k=1.059(1050- 1.068) = b=0.735 as H k=1.079 (1.066 ~1.093} = b= 0.525 C, k=1.098(1.078 ~1.118) © 1.0 b=0.741 _ H k=0.972(0.937~1.006) a b=0.510 O — 0.6 A = O . VA Ww oe! WwW ©) 10) 2 < a < (ave q 0! Eee PRE ee 9 | U OL.) Ole 1.0 20) 30 O41 Oe 1.0 2.0 FEMALES MALES TOTAL LENGTH (mm) Figure 6 Scatter diagrams of mean carapace length and height plotted against mean total length for the first 18 instars of the eight females and the eight males of Figure 2; both variates plotted logarithmically. The allometric constants b and k, and k’s 95 per cent confidence intervals (in parathesis) are shown for each relationship. C = carapace length, H = height. 33 er MALES ioc ena ene — TOTAL LENGTH D —-——- CARAPACE LENGTH BF MEN ee HEC ! \\ 6 4 2 0 Us FEMALES COEFFICIENT OF VARIATION 4 2 (0) 2 4 6 8 10 12 14 16 18 INSTARS Figure 7 The coefficients of variation (= 100 times standard deviation divided by the means) for each linear dimension for the first 18 instars of the eight females and the eight males of Figure 2. 34 NUMBER OF INDIVIDUALS 0.4 0.6 0.8 1.0 We? 1.4 1.6 LENGTH (mm) nn oO on NUMBER OF INDIVIDUALS = 0.4 0.6 0.8 1.0 12 1.4 1.6 1.8 2.0 LENGTH (mm) Figure 8 Size-frequency distributions during the first seven instars of 24 males and 32 females reared at 25°C. Broken lines designate the discrete instars. The solid line is a composite curve for all instars (see text for further explanation). The vertical bars near the upper edge of the figure represent the mean total lengths for each of the first seven discrete instars. 35 ‘ueds-ayi] ‘2,07 12 Jeysul YY S,uonendod yoea Buunp Aujeuouw 10} G ejqej 22S eu} ui snozeditulid sajeway | ¢ UO Ajeniui peseq ‘Y ‘2,6C 1e sejsul ‘18JSUI yyxis ay} ul snoyediwund sajewa; }4Bie Ajjeniui ‘g npe yoea Buunp paonpoid BunoA jo saquinu ueay,y 6 anbi4 1D}sUu| GG? OF Ge O€ GG OG ST OT G pooiq sad Buno jo 1squnu uosw 36 25 Mean number of young per brood = © 60 70 80 90 10 20 30 40 50 Age of females (days) Figure 10 Mean brood size in relation to age (in days) of females. A, based initially on females primiparous in the seventh instar at 5°C; B, data based initially on females primiparous in the fifth instar at 20°C; C, data based initially on females primiparous in the sixth instar at 5°C; D, data based initially on females primiparous in the eighth instar at 5°C. 100 110 120 130 37 lst brood oO 40 2nd brood oO 40 3rd brood O 40 4th brood 0) 40) 5th brood (0) 40 6th brood vd Le) 19) = O = 40 = 7th brood > c o > O = 40 8th brood O 40 9th brood O 40 Ÿÿ 10th brood O 40 Ÿ MER breed O 40 Ÿ 12th brood 10) ——, — SS | 0.45 0.50 D 0.6 0.6 0.70 Length (mm) Figure 11 Length of liberated young in different broods, shown for the twelfth brood, all primiparous in the fifth as percentage of size-frequency distribution; based on instar at 20°C. The arrows indicate the mean length six females for the first 11 broods and three females of the young. 38 Figure 12 Formation of an ephippium in D. schgdleri. A, female at the beginning of ephippial instar, showing fully grown sexual egg in ovary and the modified dorsal part of carapace: B, the middle of ephippial instar, showing two sexual eggs, which have been deposited into modified dorsal part of carapace; C, near the end of the ephippial instar; D, female in the sterile instar, after molting and discarding ephippium. 39 iar teurs at % bow ot LA grat oH ite” Ga i i ‘ jar M a Dee ave tm de An bout où rss > he ami s- 2 fu tes Sale 7) "Ve qui Part III Periodicity, Reproduction, and Growth of D. schodleri in the Lake Abstract Daphnia schgdleri Sars from Big Island Lake overwintered in the resting egg stage. The ex ephippio generation appeared in late spring and by May parthenogenetic reproduction was taking place. There were two periods of sexual reproduction — a major period in June and July, at which time the population was exhibiting maximum numbers, and a minor period in September. The entire first period of sexual reproduction is accounted for by the ex ephippio generation, the ephippial females of the second period were probably of the third generation. There were, in all likelihood, between four and five reproducing genera- tions (either parthenogenetic or sexual) during the open season. According to Hutch- inson’s terminology (1967), D. schgdleri in Big Island Lake would be an aestival, more or less monoacmic, dicyclic species. Production of parthenogenetic eggs varied seasonally. The average number of eggs per brood diminished just before the beginning of the sexual periods, but the decrease in brood size was not due to a decrease in the average size of egg-bearing females at this time. The overall average brood size for the entire study period was 7.96 eggs, which is considerably lower than the average brood size of the laboratory populations. The size of resting eggs varied with size of the ephippial females; generally, ephippial females were smaller than parthenogenetic females. Introduction Part Ill of the study of Daphnia schodleri Sars is concerned with D. schgdleri’s biology in Big Island Lake, Alberta. Although Parts | and Il dealing with the embryology and vital Statistic properties of laboratory populations have been reported first, all three studies were carried out more or less concurrently. For this reason, it was not possible to design the field study to take advantage of what was learned from the laboratory populations. Nevertheless, the laboratory data were im- portant for intepreting aspects of D. schgdleri’s biology from the lake. 41 Periodicity, Reproduction and Growth Materials and Methods D. schgdleri was studied in Big Island Lake from 13 June 1966 to 3 July 1967. Twenty-four samples were taken from the limnetic region during this period, using a plankton net with a number 20 mesh size and 12.5 cm diameter opening. Plankton samples were preserved in five per cent formalin. In the laboratory each sample was diluted to a known volume (usually 100 to 200 ml), depending upon the number of plankters present in the sample. D. schogdleri and other plankton of several 1 ml sub- samples were counted in a Sedgwick-Rafter cell under low power (40x) of a microscope. Length measurements were made from the top of the head to the base of the spine; this was designated as body length. To facilitate the analysis of the population composition of D. schogdleri, individuals in each sample were grouped into five catego- ries as suggested by Green (1955): 1) Females with parthenogenetic eggs or embryos in the brood chamber, or females with large ovaries indicating that eggs were about to be laid. 2) Females of mature size (over 54.7 micrometer units = 1.78 mm) but without eggs or large ovaries, and possessing the long abdominal process by which eggs are retained in the brood chamber. 3) Females with ephippia, or females with the carapace showing signs of ephippial formation and the corresponding appearance of sexual eggs in the ovary; females with sexual eggs in the ovary, but not showing any signs of ephippial formation. These ovaries are composed of a compact dark mass, representing the developing sexual eggs. 4) Immature females. These are smaller than mature females (i.e. smaller than 1.78 mm) and do not yet have the long abdominal process that retains eggs in the brood chamber. 5) Males — either immature or mature specimens. The carapaces of category 1 animals were opened with fine needles, and the number of eggs or embryos in the brood chambers was 42 counted under low power of a dissecting microscope. The term ‘‘egg number” is used irrespective of whether eggs or embryos were counted. The mean egg number was calcu- lated from a sample of at least 25 females having eggs in their brood chambers. To estimate the volume of parthenogenetic eggs (egg size), samples of at least 50 eggs were dissected out of the brood chambers of females of various sizes, and measurements were made with the eggs covered by a film of water on a slide. The volume of each egg was then calculated using the formula: V=1/67gs? where g is the largest diameter and s is the least diameter of the egg. This formula was used by Green (1956) to calculate the volume of cladoceran eggs, which are not true spheres. Volumes were determined only for eggs at late Stage | or early Stage Il of embryonic development, developing em- bryos beyond early Stage II being rejected (see Part | for explanation of stages). Since newly deposited eggs tend to swell when first laid into the brood chamber, these also were rejected. The same methods were used to measure the length of ephippial females and to calculate the volume of sexual eggs. Table 1 Limnetic zooplankton found in Big Island Lake, other than Protozoa : 1966 1967 13 22 29 8 13 27 3 17 26 16 27 18 5 19 9 13 27 3 13 24 Samples collected June June June July July July Aug. Aug. Aug. Oct. Nov. Dec. Jan. Mar. Apr. May May June June June Daphnia schodleri Sars x x x x x x x x x Diaphanosoma leuchtenbergianum Fischer x x x x x x x Bosmina coregoni Baird x x x x x x Chydorus sp. . Ceriodaphnia sp. Cyclops varicans rubellus Lilljeborg Cyclops bicuspidatus thomasi Forbes Macrocyclops albidus (Jurine) Eucyclops agilis (Koch) Diaptomus siciloides Lilljeborg Nauplii Keratella cochlearis (Gosse) Keratella quadrata (O.F.M.) Brachionus sp. Rotaria neptunia (Ehrenberg) Filinia longiseta (Ehrenberg) Asplanchna sp. Polyarthra sp. Trichocerca sp. x x x x Dipleuchlanis sp. x x Lepadella sp. x x x x x x x x Kx x x x x x x x x x x KK KK x x x x x KK x x x MM EE MUR HT CCE, x X X KK KK x x x KK KK KK x x x x X KK x x x x x x KX xX x x x x x x KK KK KK x x x De ORS CAN EL DE 06 x x x x x x x x x x x x x x x x x x x x x x x me UO OK x x X X KK KK x x x x x X X KK KK x x x x x X KK KK KR KR xX x x x x x x Kx X x x x x x X x KKK KKK x x x x Present in sample. ase Fae Watts aoe] Does pint br ere 2 a ae san nm de ie Ag LE + eee La fi a a8 : al eevee LUS nos = ge mn evidemment , AS A om ‘ “artes Oo stg egal » # > Periodicity, Reproduction and Growth Description of the Study Area Big Island Lake is a shallow, unstratified, eutrophic lake located about 17 miles southeast of Edmonton, Alberta. It is one of several small kettle lakes, remnants of the Wisconsin glaciation, characteristic of the northern prairies. Eutrophication and senes- cence in a group of these lakes were described by Kerekes and Nursall (1966), and Nursall (1969). Big Island Lake has a surface area of about 121 hectares (300 acres), with a maximum depth of 2.5 meters; most of the lake is less than two meters in depth. An island, located in the middle of the lake, covers an area of nearly five hectares. Figure 1 shows water temperatures and dissolved oxygen values recorded during the study period. From November 1966 through April 1967, the lake was completely ice- covered, the average water temperature during this period being 1.5°C. Winter stagnation occurred from January through April; during this period there was no detectable dissolved oxygen, and hydrogen sulfide was present. The ice started breaking up in early May 1967. There are no fish in the lake. The dominant macro-invertebrates are the amphipods Gammarus lacustris Sars and Hyalella azteca (Saussure). The population ecology of G. lacustris in Big Island Lake was described by Menon (1969). Extensive algae blooms occurred sporadically during the summers of 1966 and 1967. Major bloom organisms were the blue-greens Microcystis flos-aquae Kirchn., M. aeruginosa Kuetz and Anabaena flos-aquae Breb. Other important algae were Pediastrum boryanum Menegh., P. duplex Meyen and species of Scenedesmus and Staurastrum. Zooplankton of Big Island Lake Table 1 lists the limnetic zooplankters, other than Protozoa, collected during the 14- month study period. The only zooplankters present in the lake throughout the entire winter were the cyclopoids C. varicans rubellus and C. bicuspidatus thomasi, and the rotifer Rotaria neptunia. Seasonal changes in the population densi- ties of the major zooplankters are shown in Figure 2. Population densities of the cope- pods include both adults and copepodites. The seasonal curve for Cyclopoida represents all species of this suborder found in the lake, C. varicans rubellus and C. bicuspidatus thomasi being the most abundant species. The 1966 cyclopoid population reached its peak in November. From December 1966 through 9 April 1967, all cyclopoids con- sisted of late copepodites and a few adult females; apparently reproduction does not take place during the winter months in Big Island Lake. In 1967, females with egg sacs were first collected on 13 May; males and a few nauplii were also present at this time. Diaptomus siciloides reproduced almost continuously during the summer months of 1966, females carrying egg sacs or sper- matophores being collected from June through mid-October 1966. The population reached its greatest density in late August; by 27 November, only a few D. siciloides were found in the lake. These specimens were adult males and females, but none of the females was carrying egg sacs or sper- matophores; also nauplii were absent at this time. By 18 December, D. siciloides had completely disappeared from the lake, and was not collected again until the next spring. Diaphanosoma leuchtenbergianum reached maximum numbers earlier in the ice-free season than did the copepods. Males, and females carrying resting eggs, were first collected in August 1966; on 16 October, females with resting eggs were still present in the lake. In 1967, Diaphanosoma first appeared in late May, at which time the entire population consisted of young females. Adult females carrying eggs and embryos were first collected on 13 June 1967. 43 Periodicity, Reproduction and Growth Table 2 Seasonal changes in percentage composition of life-cycle stages of D. schodleri in Big Island Lake, June 1966 to July 1967 pete Fo CNT o 3 a ie : 2 2 i) 2 s 85 Soe 5 5 2 as $338 Ee à É ES £ Ê a = E2S Date LE DO QI ee = Es Lu 2 = ze3 1966 May 3 (0) (0) 0 O (0) (0) June 13 6.3 2.6 Sia O (0) 304 22 1872 14.1 29.2 42.9 0.6 Mille) 29 9.3 2178 28.0 40.9 (0) 193 July 8 7.4 0.7 87.4 4.1 0.4 269 15 11.4 272 85.9 0.5 (0) 185 19 44.2 10.9 41.0 SAS) (0) 129 27 17.4 2.9 78.3 1.4 (0) 69 Aug. 3 1-9 0) 98.1 (0) (0) 103 117 19.4 97 70.9 0) 0 31 26 20.0 20.0 60.0 0) (0) 5 Sept. 28 15.0 18.0 63.0 SZ 0.3 85 Oct. 16 O 3323 50.0 (0) 16.7 6 Nov. 27 O O O 0 0) (0) Dec. 18 O (0) O (0) 0 0 1967 Jan. 5 (0) (0) O (0) O (0) Mar. 19 O (0) O (0) (0) (0) Apr. 9 (0) (0) O 0 0) (0) May 3 O (0) 100.0 (0) 0 5 13 0) (0) 100.0 (0) (0) 8 27 16.2 0.8 83.0 (0) O 130 June 3 20.4 5.6 73.6 (0) 0.4 284 13 5.9 4.0 Te 18.4 0.1 1287 24 11859 1n=0 56.3 18.9 0.3 318 July 3 7.6 525 Us?4 3.8 237 Life History of D. schodleri D. sch@¢dleri overwinters in Big Island Lake in the resting egg stage. Soon after the resting eggs hatched in the spring (May) of 1966, the population rapidly increased in number via a series of parthenogenetic generations, the D. sch@gdleri population reaching its maximum density in June (Fig. 2). Partheno- genetic egg production started to decline at approximately the time maximum numbers were reached, and males and ephippial females appeared in the population (Table 2 and Fig. 3). There were two periods of sexual reproduction in 1966. The first started during the latter part of June and continued 44 through July, lasting about five or six weeks. The second period occurred during Septem- ber, at which time only a small number of ephippial females were collected. Between the two periods of sexual reproduction, the parthenogenetically reproducing population showed a slight increase in numbers. How- ever, as the lake was sampled only once between 26 August and 16 October, the extent of this recovery and the exact length of the second period of sexual reproduction is not known. The population continued to decline after the second period of sexual reproduction, and D. sch@gd/eri completely disappeared from the lake in early November. According to the Hutchinson's terminology Periodicity, Reproduction and Growth Table 3 Seasonal variations in mean sizes of parthenogenetic and gamogenetic females, mean number of parthenogenetic eggs, and mean volume of sexual eggs Mean length of Mean number of Mean length Mean volume parthenogenetic parthenogenetic of gamogenetic of sexual females with eggs eggs per female females eggs Date mm mm mm? x 103 1966 June 13 2/82 8.6 22 270 5.9 29 2.60 225 2.14 6.98 July 8 2.56 5.4 2.29 9.45 13 2.64 U2 2.28 8.21 June 19 2.07 3.9 1.83 4.23 27 2.04 4.6 2.26 4.03 Aug. 3 2.03 6.8 17 2107 7.4 26 27115 11 11210) Sept. 28 2.32 3.9 1967 May 27 2.62 2,979 June 3 2.30 Wo 13 2.63 6.7 2.20 8.07 24 2.50 6.3 2.38 8.12 July 3 2157 4.2 2/29 7.20 (1967), D. schgdleri of Big Island Lake would be an aestival, more or less monoac- mic, dicyclic species. Female daphnids of mature size but without eggs are often considered to be old, sterile females, which have temporarily or permanently stopped producing eggs. In Big Island Lake, mature females without eggs made up a considerable part of the popula- tion in late June 1966 and again in the autumn (Table 2). In the laboratory (Part 1), we found that females in late instars had a long barren period between releasing the brood and molting again (but after the molt another clutch of eggs was laid into the brood chamber), and hence what might, in analyz- ing field populations, be taken for either temporarily or permanently sterile females could, in fact, be old, slowly reproducing females. But we also determined in the laboratory (Part Il) that all females producing sexual eggs had a sterile instar immediately following the ephippial instar. Assuming these phenomena also hold for the field population, we suggest that the relatively large percentage of mature females without eggs in late June were mainly females that had produced ephippia (the first period of sexual reproduction) and were in the sterile instar. The large percentage of mature females without eggs in autumn was proba- bly due to both females being in the sterile instar following the ephippial instar (the second period of sexual reproduction) and also to a preponderance of old, slowly reproducing, but not sterile, females in the lake. Also paralleling our laboratory obser- vations, we found a positive correlation between the size (total length) of partheno- genetic females from Big Island Lake and the number of eggs in the brood chambers (Table 4). The degree of correlation between the size of females and the number of eggs in the brood chamber indicates a significant positive correlation (99 per cent level) for all sampling dates that were tested. The size of sexual eggs was positively correlated with the body size of ephippial females carrying these eggs (Fig. 4). The 45 Periodicity, Reproduction and Growth Table 4 Lake, temporary anoxia, especially at night Relationship between parthenogenetic egg production when the algae in the brood chamber are and body size of mature female Daphnia schgdleri for taking up oxygen, may be a major factor. different sampling dates Correlation Date Coefficient July 13, 1966 0.602 July 19 0.740 July 27 0.865 May 27, 1967 0.916 June 3 0.610 June 13 0.852 June 24 0.934 correlation coefficient was 0.754 and is significant at the 99 per cent level. It is often stated that sexual eggs of Daphnia are larger than the parthenogenetic eggs (e.g. Banta et al. 1939; Lack 1954). For D. schgdleri this is not the case, the sexual eggs being larger or smaller than parthenogenetic eggs, de- pending, at least in part, on the size of the females. Green (1956) found a similar relationship between sexual and parthenoge- netic eggs of D. magna. For the field population, there was no significant correla- tion between the size of parthenogenetic eggs and the size of the females carrying these eggs. Degenerate eggs were frequently observed in the brood chambers of D. schgdleri females from Big Island Lake. These eggs were dark gray-brown and appeared to be disintegrating; normal eggs were blue-green or yellow-brown and were of a firm texture. Degenerate eggs were excluded from the egg counts and no attempt was made to calculate the percentage of degenerate eggs in relation to total eggs present. On 22 June 1966 and again on 13 June 1967, when algae blooms of Microcystis and Anabaena were extensive, a large proportion of parthenogenetic females (about 20 per cent of all mature females) was carrying disintegrating embryos. These em- bryos were found entangled in the Microcys- tis and Anabaena present in the brood chambers. Brooks (1946) suggests that egg degeneration in Daphnia is due to inadequate nutrition. Hall (1964) suggests that degen- erate eggs may reflect a specific nutritional deficiency, a change in food level, temporary anoxia, or other conditions. In Big Island 46 Periodicity, Reproduction and Growth Interpretations of Length-Frequency Distributions Because of environmental factors, especially nutrition, age of Daphnia from field popula- tions cannot strictly be equated with size. Although both age and size are necessary to determine accurately factors affecting the growth of individuals, some understanding of the oscillatory nature of the various size (e.g. length) patterns of individuals comprising the population can be obtained by arbitrarily selecting size—class limits and following these size classes throughout the year. Also, by utilizing data available on population density (Fig. 2, p.50) and life history stages (Table 2, p.44) a more accurate interpretation of size—frequency distributions is possible. Total length of at least 100 females for each sampling date was measured; in sam- ples containing fewer than 100 females, all specimens were measured. The data were then grouped into histograms, each bar representing five eyepiece micrometer units, or 0.16 mm (Fig. 5). It was realized that these size-class limits were too wide to detect discrete instars (see Part Il for the relation between discrete instars and size— class groupings). We determined from lab- oratory work that the mean size of first instar females was 18.5 micrometer units (0.60 mm) and the mean size of females in the first adult instar was 54.7 micrometer units (1.78 mm). In the field study, the size of the smallest parthenogenetic female with eggs was 50.0 units (1.63 mm), and the size of the smallest ephippial female was 54.5 units (1.77 mm). Therefore, individuals larger than 55 units were considered adults, and those which were 55 units and smaller were considered immature animals. On 13 June 1966, the entire population was exclusively parthenogenetic and con- tained a large proportion of immature females (Fig. 5, and Table 2, p.44). Slobodkin (1954) has shown that in the early stages of population growth the few adult animals will have a high reproductive rate, resulting in a size—frequency distribution that is skewed towards the small end. For D. schgdleri, this was apparently the situation on 13 June, resulting in a rapidly reproducing population containing a large proportion of small ani- mals. On 29 June the size-frequency distribution had shifted to larger animals with most of the population consisting of adult females. This was due to a reduced repro- ductive rate and continuous growth of small animals. Also at this time sexually reproduc- ing females appeared in the population. These sexually reproducing females were of the ex ephippio generation. Mortality of the larger animals presumably caused the size-frequency distribution to shift in favour of small animals again, and by 8 July the population consisted mainly of small, immature females. On 13 July, the size—frequency distribution was still in favour of immature females, but they had grown and were now in medium size class ranges (35-45 units). These animals continued to grow, and by 19 July the length-frequency distribution was skewed to the right, the population containing mainly mature females of the second generation. But the nature of the histogram (supported by laboratory longevity data) suggests that the few remain- ing ephippial females were still those of the ex ephippio generation. After this date, and continuing for the remainder of the ice-free season, there was a tendency for the size— frequency distribution to be discontinuous but to be skewed towards smaller animals. The ephippial females of the second period of reproduction (Table 2, p.44) are most likely those of the third generation. Although the sampling intervals after 26 August were too far apart to determine accurately the subse- quent number of generations, there was certainly a fourth generation, and possibly a fifth. On 16 October, when the water tempera- ture was 4°C, very few daphnids were collected and none was carrying eggs. The largest animals collected at this time had a size of 60 units (1.95 mm). By 27 Novem- ber, free-swimming daphnids had com- pletely disappeared from the lake, and they were not found again until the following spring. The oscillatory nature of the length- frequency distributions was similar in 1967, and is not figured. 47 Periodicity, Reproduction and Growth Literature Cited Banta, A.M., T.R. Wood, L.A. Brown, and L. Ingle (1939). Studies on the physiology, genetics and evolution of some Cladocera. Carnegie Inst. Wash., Dep. Genetics, Pap. 39. 285 pp. Brooks, J.L. (1946). Cyclomorphosis in Daphnia. Part |: An analysis of D. retrocurva and D. galeata. Ecol. Monogr. 16:409- 47. Green, J. (1955). Studies on a population of Daphnia magna. J. Anim. Ecol. 24:84-97. (1956). Growth, size and reproduction in Daphnia (Crustacea: Cladocera). Proc. Zool. Soc. London 126:173-204. Hall, D.J. (1964). An experimental approach to the dynamics of a natural population of Daphnia galeata mendotae. Ecology 45(1):94-112. Hutchinson, G.E. (1967). A treatise on limnology. Vol. Il. Wiley and Sons, New York. 1115 pp. Kerekes, J., and J.R. Nursall (1966). Eutrophication and senescence in a group of prairie-parkland lakes in Alberta, Canada. Verh. Int. Verein. Theor. Angew. Limnol. 16:65-73. Lack, D. (1954). The evolution of reproductive rate. Pages 143- 56 in J.S. Huxley, ed. Evolution as a process. Allen and Unwin, New York. Menon, P.S. (1969). Population ecology of Gammarus lacustris Sars in Big Island Lake. Part |: Habitat preference and relative abundance. Hydrobiologia 33:14-32. Nursall, J.R. (1969). The general analysis of an eutrophic system. Verh. Int. Verein. Theor. Angew. Limnol. 17:100-15. Slobodkin, L.B. (1954). Population dynamics in Daphnia obtusa Kurz. Ecol. Monogr. 24:69-88. 48 © wv El a NS = Water temperature C i © 13 12 11 10 ppm =] Oxygen D F M 1966 1967 Figure 1 Water temperatures and dissolved oxygen of Big Island Lake, 1966 and 1967. 49 Cyclopoida A 5489 1688 101.2 60 50 40 30 20 10 60 | 50 Peleus 40 siciloides 30 20 10 O == 60 Diaphanosoma 50 leuchtenbergianum 40 30 20 10 60 50 Daphnia schodleri 40 30 20 10 oll aw 4 MJ JASONDJ FMAM) 4 1966 1967 NUMBER PER LITER Figure 2 Seasonal changes in population size of the major zooplankters of Big Island Lake, June 1966 to July 1967. 50 Mean egg number per brood D yp ND PV = Mean length of females with eggs (mm) § J J A S O N D J F M A M J J 1966 1967 Figure 3 Seasonal variations in parthenogenetic egg production and body length of mature parthenogenetic females. The periods when ephippial females occurred are indicated by black bars. 51 O = Parthenogenetic eggs 20 @= Resting eggs e x [à Es : 8 v O O : ae _yt---9°. 0° °° 2 aan ces 5° °° à à = %e . © 2.0 2.1 2.2 23 24 25 26 2.7 2.8 29 3.0 3.1 3.2 Mean length of females with eggs (mm) Figure 4 Figure 5 The relationship between size of sexual eggs and size Population length-frequency histograms of Daphnia of sexual females, and between size of partheno- schgdleri in Big Island Lake, 13 June 1966 to 16 genetic eggs and parthenogenetic females. Data for October 1967. The histograms represent both non- resting eggs based on a sample of 104 fresh ephippial and ephippial females, excluding male ephippial females collected on 24 June 1967. Data animals. Each individual histogram shows the for parthenogenetic eggs based on 136 fresh eggs percentage size distribution on the indicated date. The dissected out of parthenogenetic females collected on width of each bar represents 5 micrometer units 24 June 1967. The dotted line indicates the mean (0.16 mm), e.g. 16-20, 21-25 etc. volume of parthenogenetic eggs. 52 13 June 1966 29 June 1966 17 Aug. 1966 100 8 July 1966 cent 13 July 1966 26 Aug. 1966 Per 19 July 1966 16 Oct. 1966 27 July 1966 10 20 30 40 50 60 70 80 90 100 110 120 10 20 30 40 50 60 70 80 90 100 110 120 Body length in micrometer units 53 mn mole wae Ty en |= Ges, ¢ Los Ci OW eee eas met grues Lise ole i erEaEmins «et 2 cor} a dust Ge wie Com) € —yo —