Effects of prehatching salinity and initial larval biomass on survival and duration of development in the zoea 1 of the estuarine crab, Chasmagnathus granulata, under nutritional stress

Effects of prehatching salinity and initial larval

biomass on survival and duration of development in

the zoea 1 of the estuarine crab, Chasmagnathus

granulata, under nutritional stress

Luis Gimenez*

Seccion Oceanologıa, Facultad de Ciencias, Igua 4225, 11400 Montevideo, Uruguay, and Biologische Anstalt

Helgoland, Stiftung Alfred-Wegener-Intitut fur Polar-und Meeresforschung, 27498 Helgoland, Germany

Received 19 July 2001; received in revised form 9 October 2001; accepted 9 January 2002

Abstract

The effects of individual larval biomass, and salinity experienced during embryogenesis (i.e.,

prehatching salinity) on starvation tolerance and growth of zoea 1 of the estuarine crab

(Chasmagnathus granulata) were evaluated in laboratory experiments. Freshly hatched zoeae 1

were obtained from broods maintained at three salinities (15x, 20xand 32x), and cultured at

20xunder different initial feeding periods and subsequent food deprivation (‘‘point of reserve

saturation’’ experiment: PRS) or under initial periods of food deprivation and subsequent feeding

(point of no return experiment: PNR). Another group of larvae were used for determination of

biomass (dry weight, carbon, and nitrogen) of zoea 1. Larval survival and duration of development

depended on the length of feeding period: no larvae reached the second instar under complete

starvation; survival was higher and duration of development shorter as the feeding period

lengthened. After different initial feeding periods (PRS experiment), zoeae 1 that hatched from eggs

incubated at the prehatching salinities of 15xand 20xshowed higher survival and shorter

duration of development than those at 32x. Prehatching salinity also affected the amount of

reserves accumulated during the first 2 days after hatching, with larvae from 15xand 20xshowing the highest percentage of total accumulation of carbon and nitrogen. Initial larval biomass

did not affect survival, but it had a slight effect on duration of development, with larger larvae (in

terms of biomass) developing faster. After different initial starvation periods (PNR experiment),

prehatching salinity did not affect survival, but it affected duration of development: larvae from

15xand 20xreached the second instar earlier. Variability in survival and duration of development

0022-0981/02/$ - see front matter D 2002 Elsevier Science B.V. All rights reserved.

PII: S0022 -0981 (02 )00012 -6

* Seccion Oceanologıa, Facultad de Ciencias, Igua 4225, 11400 Montevideo, Uruguay. Tel.: +598-2-

5258618; fax: +598-2-5258617.

E-mail address: [email protected] (L. Gimenez).

www.elsevier.com/locate/jembe

Journal of Experimental Marine Biology and Ecology

270 (2002) 93–110

was explained in part by among-brood variability in initial larval biomass: larvae with higher

biomass showed higher survival and shorter duration of development. Thus, C. granulata, survival

and duration of development under food stress depend on the interaction between environmental

conditions experienced before and after hatching (pre- and posthatching factors, respectively).

D 2002 Elsevier Science B.V. All rights reserved.

Keywords: Crab; Larval development; Life history; Population ecology

1. Introduction

Food availability in the plankton was formerly considered as one of the main factors

that account for the fluctuations in abundance of planktotrophic larvae (Thorson, 1950).

The positive relationship between the abundance of barnacle larvae and food availability

that was found by Barnes (1958) supported that hypothesis. Recent evidence suggests that

fluctuations in food availability in the field are rather of low importance for survival of

larvae of polychaetes, molluscs, and echinoderms (Fenaux et al., 1994; Olson and Olson,

1989; Hansen, 1999). However, food availability seems to be important for survival of

crustacean larvae (Olson and Olson, 1989; Boidron-Metairon, 1995), depending on larval

capability to capture, ingest, and convert food (Anger, 2001).

In particular, decapod larvae are able to cope with short periods of lack of food

provided that certain critical points are not exceeded (Anger and Dawirs, 1981; Anger et

al., 1981a; Dawirs, 1984; Anger, 1987a; Staton and Sulkin, 1991; Anger, 2001). In

general, the moulting cycle can be successfully completed if larvae are fed for enough time

to reach the ‘‘point of reserve saturation’’ (PRS) at the D0 stage (Anger and Dawirs, 1981;

Anger et al., 1981a; Anger, 1987a). Within that period, larvae accumulate biomass to be

used as energy source during subsequent development (Anger and Dawirs, 1982; Dawirs,

1986). Accumulation of sterols (precursors of ecdysone) is important for completion of the

moulting cycle (Gore, 1985; Spindler and Anger, 1986; Anger and Spindler, 1987). Larvae

are also able to tolerate periods of lack of food without being previously fed, provided they

are fed from earlier than or from the ‘‘point of no return’’ (PNR) to the end of the moulting

cycle (Anger and Dawirs, 1981; Anger et al., 1981a; Dawirs, 1984). If the PNR is

surpassed, larvae die as a consequence of irreversible damage to the hepatopancreas

(Storch and Anger, 1983). That damage may prevent the accumulation of lipids and

production of ecdysone.

In the field, natural variability in environmental factors, including food stress, may

interact to explain larval survival. Furthermore, environmental conditions prevailing

during the embryonic development may affect larval capacity to face periods of food

limitation. Organisms living in estuarine environments are under variable conditions of

salinity and food availability. This is the case in the crab Chasmagnathus granulata, that

occurs in dense populations in estuarine marshes of the South American Atlantic coast of

Brazil, Uruguay and Argentina (Boschi, 1964). C. granulata experiences variable salinity

conditions during embryonic, larval, and juvenile–adult phases (Anger et al., 1994;

Spivak et al., 1994). This species exhibits an export strategy: the first zoea is transported to

the open ocean, develops through additional three or four stages and reinvades as

L. Gimenez / J. Exp. Mar. Biol. Ecol. 270 (2002) 93–11094

Fig. 1. C. granulata. Designs for PRS and PNR experiments for zoeae 1 that hatched from embryos incubated at

different salinities. Before the experiments, samples were taken for measurement of initial dry weight (DW),

carbon (C) and nitrogen (N) content. Each rectangle is a treatment consisting of 20 individuals that were fed

(striped areas) and deprived of food (white areas).

L. Gimenez / J. Exp. Mar. Biol. Ecol. 270 (2002) 93–110 95

megalopae the nursery habitat (Anger et al., 1994). Laboratory experiments showed that:

(1) negative effects of very low (5–10x) salinities on larval survival can be reduced if

embryonic development occur at moderately low salinities (15–20x; Charmantier et al.,

in press), and (2) larval survival under ad libitum feeding conditions is positively

correlated with initial larval biomass (Gimenez, 2000). Initial individual larval biomass

is affected by salinity experienced during embryogenesis (Gimenez and Anger, 2001). The

acclimatory effect as well as the initial individual larval biomass could interact with food

availability to determine survival and growth of zoea 1.

Fig. 2. C. granulata. Mean mortality of zoeae 1 from three prehatching salinities, and maintained under different

initial feeding (PRS experiment: a, c) and starvation (PNR experiment: b, d) periods. (a and b) Comparison of

mortalities; (c and d) adjusment of sigmoidal curves to estimate PRS0, PRS50, PNR50 and PNR100 (see arrows).

See Table 1 for values of PRS50 and PNR50. Error bars: standard deviation.


I evaluated the tolerance of zoea 1 of C. granulata to limited starvation periods, and the

effect of prehatching salinity and initial egg biomass on tolerance to food stress. I

addressed the following questions:

(a) How do zoeae 1 respond, in terms of survival and duration of development, to

different periods of food deprivation?

(b) What are their PRS and PNR?

(c) Do prehatching salinity and initial larval biomass affect PRS, PNR, and duration

of development?

(d) Does PRS depend on rates of reserve accumulation during the initial feeding

period?

2. Methods

Juvenile and adult C. granulata were collected in Mar Chiquita coastal lagoon,

Argentina (37�33VS), brought to the Helgoland Marine Station, and maintained in the

laboratory at constant temperature (21 �C) and salinity (15xor 32x) until females laid

eggs.

Experiments with C. granulata were run under controlled conditions of temperature

(18 �C), and photoperiod (12:12). Seawater was filtered (Orion, mesh size: 1 mm) and

directly used for experiments or previously diluted in appropriate amounts of desalinated

seawater to obtain the required salinities. Larvae were fed ad libitum with Artemia sp.

nauplii when appropriate (see below).

Table 1

C. granulata. Mean, standard deviation (Std), and range of variation of PRS50 (in days) from zoeae 1 hatched

from different prehatching salinities (Ex)

Ex Mean Std Range

15 1.41a 0.14 1.30–1.63

20 1.39a 0.07 1.30–1.46

32 2.00b 0.55 1.43–2.65

Columns with different letters denote significant differences ( p< 0.05) after SNK test.

Table 2

C. granulata. Two-way ANOVAs to test the effects of different initial feeding (PRS) and starvation (PNR) periods

on survival of zoeae 1 from different prehatching salinities (Ex)

ANOVA Factor dff MSf dfe MSe F p

(1) Ex 2 0.27 99 0.03 8.18 <10�4

PRS 8 4.28 99 0.03 127.56 <10�6

PRS�Ex 16 0.04 99 0.03 1.13 0.34

(2) Ex 2 17 99 256 0.07 0.93

PNR 8 15,413 99 256 60.09 <10�6

PNR�Ex 16 236 99 256 0.92 0.55

dff, MSf, dfe, MSe: degrees of freedom and mean square of factors and error respectively.



2.1. PRS and PNR experiments

Three groups of five ovigerous females were maintained from egg laying until hatching

of zoea 1, at the salinities 15x, 20xor 32x, respectively. Thus, embryos were

maintained in vivo (i.e., the brood was not removed from the female), and each brood was

assigned to one of the previously named salinities. In order to obtain larvae from embryos

at 15x, females laid eggs at 15x(see Gimenez and Anger, 2001); ovigerous females

maintained at 20xand 32xlaid eggs at 32x. Freshly hatched larvae of each brood

were used for determination of biomass and for PRS and PNR experiments (Fig. 1).

Biomass was measured as dry weight (DW), carbon (C) and nitrogen (N): five replicates of

40 larvae per brood were rinsed in distilled water for few seconds, dried on filter paper,

transferred to tin cartridges, and dried for 48 h in a vacuum drier (Finn-Aqua Lyovac

GT2E), weighed on a microbalance (Mettler UMT2, precision: 0.1 mg), and analysed in a

Carlo Erba Elemental Analyser (EA 1108).

Experiments with larvae were run at 20x, simulating an estuarine condition. Larvae

were divided assigned into groups of 20 individual per vial (vol.=80 ml) and fed or

deprived from food. Every day, water and food were changed and dead animals were

eliminated from the experiments.

For the PRS experiment larvae were, in eight parallel treatments, initially fed for

different time periods (1–8 days) and then left without food until all larvae died or

moulted to the second zoea (Fig. 1). For the PNR experiment (also eight treatments),

larvae were initially deprived from food and then fed. Besides, there were two control

treatments of larvae fed or starved permanently. In both experiments, mortality and mean

duration of development was estimated. Data for mortality were adjusted to a sigmoid

dose–response curve

MðtÞ ¼ M0 þ ðMf �M0Þ=ð1þ 10P50�tÞ

where M(t) is the number of dead larvae and t is the initial feeding (for PRS) or starvation

(for PNR) period, respectively (Paschke, 1998). The adjusted parameters were the

estimated mortality of the control group (M0), the asymptotic mortality as the initial

feeding or starvation period increases (Mf) and the period estimated to obtain a mortality of

50% (P50) (sensu Anger, 1987a). P50 will be called PRS50 or PNR50 depending on the

experiment. We thus obtained a value of PRS50 and PNR50 for each brood (replicate).

Statistical analyses for PRS50, PNR50, and other variables were run following Day and

Quinn (1989), Zar (1996) and Underwood (1997). We evaluated if prehatching salinity and

the initial larval biomass affected PRS50 and PNR50 with a one-way ANOVA (three levels:

15x, 20xand 32x, and five replicates per level) using the initial larval biomass as

a covariate. The role of biomass on PRS50 and PNR50 was also analysed by Pearson

correlation. Variance homogeneity was checked with Cochran test and normality with

normal plots of residuals. Only for PRS50 variances were heterogeneous, so data were

Fig. 3. C. granulata. Correlation between PNR50 and individual dry weight, carbon and nitrogen content in

freshly hatched zoeae 1. For C content, the point within the circle has been taken out of the analysis. ns: Non-

significant correlation.


transformed (reciprocal transformation); variances of transformed data were homoge-

neous and data followed normal distribution.

For the duration of development and mortality rate, the effects of prehatching salinity,

feeding/starvation period, and initial larval biomass were analysed with two-way ANOVA

with biomass as a covariate. The role of biomass in explaining variability in duration of

development was additionally evaluated with Pearson correlations.

2.2. Growth of zoea 1

The PRS50 of zoea 1 was affected by the prehatching salinity but not by the initial larval

biomass (see Results). This could have occurred because the salinity affected the rate of

reserve accumulation during the first days of feeding. To evaluate this hypothesis an

additional set of broods (five per prehatching salinity; total = 15) were used to determine

biomass of 2 days old and premoult zoea 1. Larvae were mass-reared in bottles at 20xwith ad libitum food and used for determination of DW, C, and N as described above. I

also determined biomass at premoult zoea 1 (n = 14 broods) and estimated the percentage

of accumulated biomass after 2 days with respect to that accumulated from hatching to

premoult (called thereafter total accumulated biomass). The effect of prehatching salinity

on increments in biomass after 2 days was evaluated with a one-way ANOVA. An

additional ANCOVA was performed on increments in biomass after 2 days, using total

accumulated biomass as a covariate.

3. Results

3.1. Mortality

No larva maintained under complete starvation was able to moult to the second zoea

(Fig. 2a). For the PRS experiment a very small group of larvae (ca. 2%) was able to moult

when fed for only 1 day: the PRS0 (the longest initial feeding time that did not allow a

successful development) was 1 day (Fig. 2c). If the initial feeding period was equal or

longer than 3 days, the mortality was low and independent of the length of feeding period

Table 3

C. granulata. Two-way ANOVAs to test the effects of different initial feeding (PRS) and starvation (PNR) periods

on duration of development of zoeae 1 from different prehatching salinities (Ex)

ANOVA Factor dff MSf dfe MSe F p

(1) Ex 2 3.93 84 0.34 11.39 <10�4

PRS 6 1.68 84 0.34 4.86 <10�3

PRS�Ex 12 0.16 84 0.34 0.45 0.94

(2) Ex 2 3.92 48 0.48 8.21 <10�3

PNR 3 47.11 48 0.48 98.58 <10�6

PNR�Ex 6 0.32 48 0.48 0.67 0.67

Symbols are as in Table 2.


(Fig. 2a, c). The PRS50 ranged from 1.3 to 2.6 days (Table 1); there was a significant effect

of prehatching salinity on PRS50 (F2,12 = 7.71; p = 0.01), and on survival in almost all

treatments (Table 2). Larvae from the prehatching salinities 15xand 20xhad an earlier

PRS50 and lower mortality under food-limited conditions, i.e. a higher tolerance to

starvation if they were initially fed (Table 1, Fig. 2a, c). Besides, some larvae were able

to moult after only 1 day of feeding. There were no significant correlations between PRS50and initial larval DW (r =�0.31; p = 0.26), C (r =�0.40; p = 0.14) or N (r =�0.37;

p = 0.18) content.

Fig. 4. C. granulata. Relationship between mean duration of development and initial feeding (a) and starvation (b)

periods for zoeae 1 from different prehatching salinities. Error bars: standard deviation. See Table 3 for significant

differences among treatments.


Fig. 5. C. granulata. Correlation between duration of development and carbon content at hatching in zoeae 1 from

different prehatching salinities (a) fed ad libitum or under starvation periods of (b) 1, (c) 2, and (d) 3 days.


In the PNR experiment most larvae survived and moulted to the second zoea when they

were fed permanently (Fig. 2b). With a day of food deprivation the groups hatched from

20xand 32xshowed also low mortality compared to those from 15x. The initial

starvation period increased larval mortality (Table 2) especially after 2 or more days of

food deprivation. When the period of starvation was 6 days, only 1 out of 300 larvae

survived and moulted to zoea 2 (Fig. 2b). The PNR100 (i.e., the shortest period of

starvation that produced 100% of mortality) ranged from 4 to 7 days depending on the

brood (Fig. 2d). The PNR50 also depended on the brood, and ranged from 1.6 to 3.3 days.

Prehatching salinity did not affect PNR50 (F2, 12=1.59; p> 0.05) or mortality (Fig. 2b, d).

The initial individual larval biomass explained part a of the variability among broods:

there was a significant positive correlation between PNR50 and N content per individual;

for C content the correlation was significant when one high PNR50 value was excluded

from the analysis (Fig. 3). Significant correlations showed that larvae with higher biomass

tended to tolerate starvation for a longer time.

3.2. Duration of development

The length of the feeding period and the prehatching salinity (Table 3) significantly

affected the duration of development of the zoea 1. After different initial feeding periods

(PRS experiment) the larvae fed 2 days moulted ca. 1 day later (i.e., with a delay of 8–

17%) than those fed for at least 3 days (Fig. 4a). Larvae from the prehatching salinity

32xmoulted ca. 1 day later (i.e., with a delay of 8–18%) than those from 15xor 20x(Fig. 4a). Duration of development was correlated with C content at hatching only for

larvae fed 2 days (r=�0.53; p < 0.05): larvae with higher C content moulted earlier.

Increasing initial starvation periods (PNR experiment) produced a considerable delay in

the timing of moulting (Fig. 4b). For starvation periods equal or shorter than 3 days there

was a delay of 1.0–1.8 days (i.e., 18–24%) per any additional day of food deprivation.

For longer starvation periods the delay was longer: the surviving larvae after starvation

periods of 4–7 days took 12–15 days to moult. The duration of development of larvae

from the prehatching salinity 32xwas longer than that from other salinities: they suffered

Table 4

C. granulata. Linear regression (Y=a+bX ) and correlation of duration of development (Y ) and initial starvation

period (X ), of zoeae 1 from different prehatching salinities

15x 20x 32x

a 5.56 5.74 5.93

b 1.21 1.33 1.56

S.E. (b) 0.09 0.13 0.16

r 0.95 0.92 0.91

r2 0.89 0.84 0.82

p (b=0) < 10�9 10�8 < 10�7

p (b=1) < 0.05 < 0.05 <0.01

S.E. (b): standard error of slope ; p (b= 0), p (b= 1): probability of slope being equal or lower than 0 or 1,

respectively.



a delay of 0.5–1.5 days (i.e., from 5% to 10%) compared to the larvae hatched from the

salinities 15 and 20x(Fig. 4b). Duration of development was correlated with C content at

hatching for larvae that suffered at least 1 day of starvation (Fig. 5): larvae with higher C

content tended to moult earlier. Correlations between duration of development and N

content were significant for larvae under 1 day of starvation ( p < 0.05), and marginally

significant ( p < 0.08) for those under longer starvation periods.

As shown in Table 4, the duration of development was linearly related to the length of

the initial feeding period, with a slope higher than 1. This slope reflects the sensitivity of

the larvae to an increment in the length of the starvation period (e.g., if the slope were 0,

the length of the starvation period would not affect the duration of development). This

slope tended to be higher for larvae hatched from 32xthan for 15xand 20x, but this

trend was not significant (see ANOVA (2) Table 3: the interaction term was not

significant). However, there was a significant correlation between the biomass at hatching

and the slope (Fig. 6), suggesting that larvae with higher biomass at hatching were less

sensitive to an increment in the length of the starvation period.

Fig. 6. C. granulata. Correlation between sensitivity to starvation, measured as the slope of the curve duration of

development– starvation period, and biomass of at hatching in zoeae 1 from different prehatching salinities.

Symbols are as in Fig. 4. The data points within the circles were taken out of the analyses.

Table 5

C. granulata. Changes in mean dry weight (DW), carbon (C) and nitrogen (N) of zoeae 1 from different

prehatching salinities

15x 20x 32x

DW

Biomass day 0 (mg/ind) 7.99 (1.03) 7.37 (0.63) 7.00 (1.04)

Biomass day 2 (mg/ind) 12.48 (1.18) 11.70 (0.56) 10.63 (1.19)

Increment (mg) 4.49 (0.45) 4.33 (0.25) 3.63 (0.71)

Increment (%) 56.2 58.8 51.9

SNK test a a b

C

Biomass day 0 2.73 (0.33) 2.31 (0.20) 2.27 (0.31)

Biomass day 2 4.63 (0.54) 4.12 (0.20) 3.85 (0.50)

Increment (mg) 1.90 (0.26) 1.81 (0.10) 1.59 (0.24)

Increment (%) 69.7 78.2 70.1

SNK test a a a

N

Biomass day 0 0.61 (0.07) 0.52 (0.06) 0.54 (0.07)

Biomass day 2 0.97 (0.11) 0.84 (0.07) 0.79 (0.10)

Increment (mg) 0.36 (0.05) 0.32 (0.03) 0.25 (0.04)

Increment (%) 58.1 60.8 45.4

SNK test a a b

Numbers in brackets are standard deviations; different letters denote separate significant differences among

prehatching salinities after SNK test.


3.3. Growth of zoea 1

Larvae from the prehatching salinities 15xand 20xhad always the highest

increment in biomass (Table 5). ANOVAs on increments in mg showed that increases

for larvae from the prehatching salinity 15xand 20xwere significantly different from

32xfor DW (F2,12=4.1; p < 0.05) and N (F2,12=8.8; p < 0.01), but only marginally

significant for C (0.10 < p < 0.05). Ranges of increment after 2 days were 51–59% for DW,

69–71 for C, and 45–61 for N (Table 1). The highest increments in DWand N were found

in larvae from the prehatching salinities 15xand 20x, while those from 15xshowed

the highest increments in C (Table 5). ANCOVAs on increments in biomass showed

marginally significant effect for DW (0.10 < p < 0.05), no significant effects for C

(F2,11 = 1.5; p = 0.27), and a significant effect for N content (F2,11 = 7.8; p < 0.01).

When increments in DW, C, and N after 2 days were expressed as percentage of

total accumulated biomass, larvae from 15 and 20xshowed the highest percentages of

accumulation (Table 6). The ANCOVAs showed significant effects for C (F2,10 =

4.37; p < 0.05) and N (F2,10=8.71; p < 0.01), and therefore confirmed a significantly

higher rate of C and N accumulation for zoeae 1 from the prehatching salinities 15xand

20x.

4. Discussion

The survival of zoea 1 of C. granulata depended on the length of the feeding periods

but the larvae showed certain ability to tolerate starvation for a short time. As suggested by

Anger and Ismael (1997), C. granulata is an estuarine crab whose larvae resemble those of

marine crabs: they are strict planktotrophs with low C reserves and high growth rates. The

present results showed also how a prehatching environmental condition (salinity)

interacted with a posthatching factor (food availability) to affect larval mortality and

duration of development.

4.1. Mortality

The PRS50 (i.e., the feeding period necessary to reach a survival of 50%) of zoea 1

ranged between 1.4 and 2.0 days. Similar values (between 1 and 2 days) were found for

other decapods as Carcinus maenas (Dawirs, 1984), Sesarma cinereum (Staton and

Sulkin, 1991) and Crangon crangon (Paschke, 1998). Values of PRS50 for zoeae 1 of

Table 6

C. granulata. Accumulated biomass (DW, C and N) after 2 days as a percentage of total accumulated biomass

(i.e., from hatching to premoult), in zoeae 1 from different prehatching salinities

15x 20x 32x

DW 71 61 57

C 81 62 56

N 59 49 43

Symbols are as in Table 5.


C. granulata varied from 25% to 32% when expressed as percentage of the duration of

development of ad libitum fed larvae. In other marine decapods, those percentages are

higher (35–50% cf. Anger, 1987a; 55%: Pardo et al., 1997), and suggest a higher ability of

C. granulata to withstand starvation after shorter initial feeding periods. The PRS50 of the

zoea 1 of C. crangon reached 32% of duration of development if hatched from the small

summer eggs and 23% for those from the large winter eggs (Paschke, 1998). In C.

granulata, short PRS50 could be related to a high rate of biomass accumulation. Anger and

Ismael (1997) showed that C. granulata exhibited one of the highest rates of accumulation

of C and N measured in decapod larvae. The fraction of total biomass accumulated after 2

days was 55–80% for C and 40–60% for N (Table 6; see also Anger and Ismael, 1997).

Similar values C (71%) and N (60%) were found for C. maenas (Dawirs, 1986), while the

spider crab Hyas araneus had lower values (C ca. 25%, N ca. 18%: Anger and Dawirs,

1982). A high rate of accumulation of C suggests a rapid increase in lipid reserves to be

used under subsequent starvation periods (Storch and Anger, 1983; Dawirs, 1986).

Salinity experienced during embryonic development of C. granulata affected starvation

tolerance of zoeae 1. Mortality was lower and PRS50 was shorter in zoeae 1 of C.

granulata from low prehatching salinities (15xand 20x) most likely through a process

of acclimation to 20xduring the embryonic development. Acclimation must have

favoured the accumulation of biomass during the first days of development (Table 6).

An acclimatory effect has been observed in zoeae 1 of C. granulata when they were reared

at low salinities with ad libitum food (Gimenez, 2000). When zoeae 1 that hatched from

embryos at 32xwere reared at 5xand 10xvery few individuals reached the second

zoeal stage; however, in average more than 60% reached the second instar if they hatched

from 15xand 20x(Gimenez, 2000). Zoeae 1 that hatched from embryos maintained at

low salinities exhibit an enhanced osmoregulatory capacity (Charmantier et al., in press).

At 15xand 32x, no effects of acclimation on survival were observed in larvae reared

under ad libitum food conditions (Gimenez, 2000), but present results show effects under

limiting feeding conditions.

Larval tolerance to different initial starvation periods (measured as PNR50) was very

variable but always lower than 3 days, suggesting a low tolerance when compared with

other marine species such as C. maenas (PNR50 = 3.8 days: Dawirs, 1984) and C. crangon

(PNR50: 3–5 days: Paschke, 1998). The PNR50 of C. granulata was lower than 50% of

duration of development of ad libitum zoea 1, while for other species it is larger (e.g., C.

maenas: ca. 75%: Dawirs, 1984; H. araneus ca. 65%: Anger and Dawirs, 1981; C.

crangon 73% for larvae from summer eggs: Paschke, 1998). Zoeae 1 of C. granulata may

be less adapted to cope with food stress immediately after hatching than those of marine

crabs. However, zoeae 1 should initially experience an environment with high levels of

food availability, since larvae are released to the estuarine waters (Anger et al., 1994).

The salinity experienced during embryonic development did not affect PNR50. Instead,

variability in PNR50 among broods was correlated to variability in initial larval biomass:

larvae with higher biomass showed a higher tolerance to starvation. Thus, the relationship

between larval survival and salinity or initial larval biomass depended on whether the

larvae were initially fed or deprived of food. Under initial food stress, the metabolism must

be based on initial larval reserves, otherwise it is also fuelled by the reserves accumulated

during the initial feeding period. Under food stress the zoeae 1 of other crabs (e.g., H.


araneus: Anger and Dawirs, 1982; Anger, 1986; C. maenas: Dawirs, 1987) catabolise

lipids and then proteins. However, they suffer higher losses of N respect to C. A similar

pattern of loss of C and N would explain why PNR50 was in C. granulata better correlated

with N than with C.

4.2. Duration of development

There was a significant effect of starvation period and prehatching salinity on duration

of development of zoeae 1. There was a delay in moulting time when larvae were initially

fed for 2 days and later deprived of food. A similar fact has been found in other decapod

larvae (C. maenas, Hyas coarctatus: Anger, 1987b; H. araneus: Anger and Dawirs, 1981).

This delay could be attributed to a low rate of ecdysteroid secretion (Anger and Spindler,

1987). Duration of development was considerably affected by the extent of the initial

starvation period. A similar pattern has been found in other decapod crustaceans (Anger

and Dawirs, 1981; Anger et al., 1981a,b; Dawirs, 1986; Paschke, 1998). In C. granulata

each day of starvation increased the duration of development for more than 1 day. Thus, as

in H. araneus (Anger and Dawirs, 1981), in C. granulata (1) feeding is necessary for

starting physiological processes leading to moulting, and (2) an additional compensation

period is needed to recover the biomass lost during the starvation.

Zoeae 1 that hatched from embryos maintained at low prehatching salinities had a

shorter duration of development than the larvae that hatched from embryos in 32x. The

latter must have reached the exuviation threshold later as a consequence of a lower rate of

accumulation of reserves. The correlation between duration of development and initial

individual larval biomass was significant when (1) the larvae were fed 2 days and then

deprived of food (PRS experiment) or (2) initially deprived of food for at least 1 day (PNR

experiment). Thus, under food stress the amount of accumulated reserves during the

feeding period would not mask the effect of initial larval biomass on duration of

development, so that small differences in initial larval biomass may be important to

explain variability in the rate of development. These differences should be related to lipid

reserves, since in planktotrophic decapod crustacean larvae C content reflects lipid content

(Anger and Harms, 1990; Anger, 2001). Besides, after initial starvation and subsequent

feeding, there was a weak effect of biomass on the sensitivity to an increment of 1 day in

the starvation period (Fig. 6): larvae with higher biomass showed in general smaller

increments in duration of development after a lengthening of the starvation period. A

similar pattern was found for zoeae 1 of the shrimp C. crangon (Paschke, 1998): zoeae 1

from smaller summer eggs were more sensitive to an increment in the starvation period

than those from larger winter eggs. Thus, a lower initial larval biomass seems to increase

negative effects of the starvation period on duration of development synergistically.

In summary, the zoea 1 ofC. granulata resembles that of marine crabs but with somewhat

enhanced ability to tolerate food deprivation after an initial feeding period. Low prehatching

salinities (<12x) affect survival of embryos (Bas and Spivak, 2000), moderately low

salinities (15–20x) have negative effects on larval survival through an increased loss of

mass during embryogenesis (Gimenez and Anger, 2001). However, low prehatching

salinities have a positive effect on larval survival and growth under low posthatching

salinities (Charmantier et al., in press), and short starvation periods (this report). Larval


survival and duration of development under food stress depended on the interaction among

prehatching salinity, initial larval C and N content, and food stress. Further work should the

consider the variability in quality of eggs and larvae within and among estuarine habitats,

and their relationship with salinity and other factors as a relevant topic.

Acknowledgements

This work was part of my dissertation. I very much appreciated the help and suggestions

made by Dr. Klaus Anger during my stay at Biologische Anstalt Helgoland. I also

acknowledge C. Puschel for biomass measurements, K. Riesebeck and G. Torres for their

help in rearing Artemia and in other activities during the course of the experiments. Thanks

are also due to Dr. K. Anger, Dr. O. Defeo, Dr. D Calliari, Lic. G. Torres, Dipl. L. Gutow,

and to anonymous reviewers for their comments on earlier versions of this manuscript.

Financial support was provided by Deutscher Akademischer Austausch Dienst (DAAD) in

Germany and Programa de Desarrollo de las Ciencias Basicas (PEDECIBA) in Uruguay.

[SS]

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Effects of prehatching salinity and initial larval biomass on survival and duration of development in the zoea 1 of the estuarine crab, Chasmagnathus granulata, under nutritional stress