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Influence of larval exposure to salinity and cadmium stress on juvenile performance of two marine invertebrates (Capitella sp. I and Crepidula fornicata)
Journal of Experimental Marine Biology and Ecology 264 Ž2001. 101–114 www.elsevier.comrlocaterjembe
Influence of larval exposure to salinity and cadmium stress on juvenile performance of two marine invertebrates žCapitella sp. I and Crepidula fornicata/ Jan A. Pechenik a,) , Tim Gleason b, Dara Daniels a , Denise Champlin b b
a Biology Department, Tufts UniÕersity, Medford, MA 02155, USA U.S. EnÕironmental Protection Agency, Office of Research and DeÕelopment, National Health and EnÕironmental Effects Research Laboratory, Atlantic Ecology DiÕision, 27 Tarzwell DriÕe, Narragansett, RI 02882, USA
Received 9 November 2000; received in revised form 28 April 2001; accepted 30 June 2001
Abstract Delayed metamorphosis and short-term food limitation reduce juvenile or adult fitness in a number of marine invertebrate species. In this study, we tested the ability of pollutant and salinity stress to bring about similar effects on juvenile or adult performance. Larvae of the polychaete Capitella sp. I were exposed to sublethal cadmium stress Žup to 2000 mg ly1 . or salinity stress Ždown to 10‰. for 24 and 48 h at 23 8C. Following exposure, we induced surviving larvae to metamorphose and monitored the subsequent survival, growth, and reproductive output of juveniles reared under control conditions Žno added cadmium, 32‰ salinity.. Similarly, larvae of the gastropod Crepidula fornicata were exposed for 24 and 48 h to cadmium in seawater Žup to a nominal concentration of 20,000 mg ly1 .. Surviving larvae were reared to metamorphic competence in the absence of cadmium, induced to metamorphose, and maintained under control conditions for an additional 5 days to monitor juvenile growth rates and survival. Exposing larvae of Capitella sp. I to low salinity Ž10–12‰. for 48 h generally did not affect adult fecundity, but stressing the larvae for as little as 24 h significantly reduced post-settlement survival and juvenile growth rates Ž P - 0.05.. In contrast, exposing larvae of this species to cadmium for even 48 h had no significant effects on post-settlement survival or fecundity, and no consistent effect on mean juvenile growth rate. Similarly, cadmium exposure did not significantly affect mean juvenile growth rates for C. fornicata, even when larvae were severely stressed Ži.e., when larval mortality
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Corresponding author. Tel.: q1-617-627-3195; fax: q1-617-627-3805. E-mail address: [email protected] ŽJ.A. Pechenik..
0022-0981r01r$ - see front matter q 2001 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 2 - 0 9 8 1 Ž 0 1 . 0 0 3 1 3 - 6
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exceeded 50% during exposures.. We suggest that heavy metal stressors do not act through the same mechanism as the stresses of inadequate food supply, reduced salinity, and delayed metamorphosis. q 2001 Elsevier Science B.V. All rights reserved. Keywords: Capitella; Cadmium; Crepidula; Larvae; Metamorphosis; Salinity stress
1. Introduction It has become increasingly clear in recent years that metamorphosis is not always a fresh beginning for marine invertebrates Žreviewed by Pechenik et al., 1998; Pechenik, 1999.. For example, newly metamorphosed juveniles of Crepidula fornicata grew nearly 70% more slowly if they were starved for 5 days as larvae ŽPechenik et al., 1996b. and grew about 37% more slowly if food concentrations were dramatically reduced for just 3 days during larval development ŽPechenik et al., 1996a.. Similarly, newly settled worms of Capitella sp. I showed dramatically lower survival if larvae had been forced to delay metamorphosis for as little as 72 h ŽPechenik and Cerulli, 1991., and newly metamorphosed barnacles Ž Balanus amphitrite. exhibited markedly reduced rates of tissue growth if cyprids were forced to delay metamorphosis for as little as 3 days ŽPechenik et al., 1993.. Similar findings have been reported for a variety of invertebrate species from several taxonomic groups Žreviewed by Pechenik et al., 1998.. Thus, sublethal stresses experienced by larvae have the potential to dramatically reduce post-metamorphic performance, either directly or through reduced competitive ability or reproductive output. Pechenik et al. Ž1998. predicted that exposing larvae to sublethal pollutant stress would have similar impact on juvenile survival, growth rates, competitive ability, time to reproductive maturity, or fecundity. In this study we examined the consequences on juvenile performance of exposing larvae to either low salinity stress Žthe deposit-feeding polychaete Capitella sp. I. or elevated cadmium Ž Capitella sp. I and the suspension-feeding gastropod C. fornicata.. The larvae of Capitella sp. I emerge into the plankton as non-feeding trochophores that are competent to metamorphose within minutes of being released Že.g., Pechenik and Cerulli, 1991.. In contrast, larvae of C. fornicata are released as feeding veligers that must develop for at least a week before becoming competent to metamorphose Že.g., Pechenik et al., 1996b.. Capitella sp. I is particularly common in areas of fluctuating salinity and high heavy metal concentrations, as at sewage outfalls ŽLevin et al., 1996.. C. fornicata is a common inhabitant of bays and estuaries in many parts of the world ŽBlanchard, 1997., and is therefore exposed to a variety of coastal pollutants. Cadmium is a major component of industrial discharge and is highly toxic to a variety of aquatic animals Že.g., Eisler, 1985; Williams et al., 1986; Au et al., 2001.. 2. Materials and methods 2.1. Exposure of Capitella sp. I to cadmium Adult worms were obtained from Dr. J. Grassle ŽRutgers University. and maintained in our laboratory for multiple generations at about 30‰ salinity on a diet of fine mud
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collected from the Little Sippewissett salt marsh on Cape Cod, Massachusetts, USA, where members of this species occur naturally. The mud was forced through a 1-mm sieve to remove debris and then frozen for at least several days to kill animal residents before use ŽCohen and Pechenik, 1999.. Two experiments ŽCad-1 and Cad-2. were conducted at 23 8C, both using static exposures to toxicant. Approximately 600 larvae released from brood tubes by four females were pooled and subsampled for the first experiment. The larvae were distributed among plastic cups containing 50 ml of either 0.45-mm-filtered seawater or CdCl 2 seawater solution. Seven CdCl 2 concentrations were tested, up to a nominal concentration of 1000 mg ly1 , plus control seawater, with three replicates per treatment and 12 larvae per replicate. Survival and incidence of metamorphosis were assessed after 24 and 48 h; individuals were observed at 100 = using a compound microscope to confirm loss of larval cilia. Metamorphosed individuals were discarded. Remaining larvae were then transferred to cups of seawater with mud to stimulate natural metamorphosis ŽButman et al., 1988; Dubilier, 1988; Pechenik and Cerulli, 1991; Cohen and Pechenik, 1999., and maintained at 20 8C for 2–2.5 weeks to assess juvenile survival and growth, and adult fecundity. At the end of the experiment we retrieved all surviving juveniles and measured the length and width of each individual at 8–20 = using a dissecting microscope equipped with an ocular micrometer. Worms were anesthetized in 0.36 M MgCl 2 before measurements were taken. Worm volumes were calculated assuming that each individual was cylindrical. The second experiment was conducted about 1 week later, subsampling from 310 larvae pooled from two females. The experiment was similar to the first one, except that we added one higher nominal CdCl 2 concentration Ž2000 mg ly1 ., and examined larvae only after 48 h. Again, we tested three replicates at each concentration Žincluding seawater controls., with 12 larvae per replicate. 2.2. Exposure of C. fornicata to cadmium One experiment was conducted, using 360 larvae. Stacks of adults were collected from North Kingston, RI, USA in late August 1998. Larvae were released from one female in the laboratory 4 days later and were fed the naked flagellate Isochrysis galbana Žclone T-ISO. for 24 h before the experiment began on the following day. Larvae were exposed to CdCl 2 in static conditions Ž23 8C. at nine concentrations Žincluding a seawater control., up to a nominal concentration of 20,000 mg ly1 , with four replicates per treatment and 10 larvae per replicate. Nominal and measured cadmium concentrations for this experiment are shown in Table 1. Each replicate
Table 1 Nominal and measured total cadmium concentrations Žmg ly1 . for the C. fornicata study Nominal concentration Measured concentration
0 Žcontrol. -10
156.25 243
312.5 361
625 568
1250 1058
2500 2189
5000 4363
10,000 8294
20,000 16,392
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contained 50 ml of the test solution, to which phytoplankton Žclone T-ISO. was added to attain a concentration of approximately 10 5 cells mly1 . Test solutions were changed after 24 h and the exposures were terminated after 48 h. Because larvae of this species are not competent to metamorphose when released from the parent, surviving larvae were then reared for another 8 days in non-polluted Žcontrol. seawater on a diet of T-ISO with the container and feeding suspension changed every other day, after which they were induced to metamorphose by elevating the Kq concentration of seawater by 20 mM for 6 h ŽPechenik and Heyman, 1987; Pechenik and Gee, 1993.. Mean juvenile growth rates were assessed for individuals from seven of the treatments Ž0, 325, 625, 1250, 2500, 5000 and 10,000 mg ly1 .; no larvae survived the 48 h exposure at the highest concentration tested Ž20,000 mg ly1 .. Within each treatment group at least three juveniles between 860 and 1115 mm shell length were haphazardly selected from each replicate, so that juvenile growth rates were monitored for about 12 individuals per treatment. Shell lengths were measured nondestructively at 32 = . Each juvenile was reared at 25 8C in a separate glass dish with 40 ml of phytoplankton suspension ŽT-ISO, 18 = 10 4 cells mly1 .. Food and water were replaced daily for 5 days and individual shell lengths were remeasured to determine growth. Cadmium concentrations were measured at the start of the experiment following Lussier et al. Ž1999.. Prior to analysis, all aqueous samples were acidified with 1 ml of concentrated nitric acid per ml of sample at room temperature and placed on an orbital shaker table at 40 rpm for 24 h. Metal concentrations were determined using a Model 3410 inductively coupled plasma atomic emission spectrometer ŽICP-AES. ŽApplied Research Laboratories, Valencia, CA. with a detection limit of about 0.01 mg Cd ly1 . We verified instrumental calibrations by analyzing independent reference solutions before and during analysis of unknowns Žone verification for every 10 unknowns.; freedom from matrix interference was verified by analyzing spiked unknowns Žone spiked sample for every 20 unknowns.. Analyses were acceptable if verification standard or spike recovery was between 85% and 115%. Concentrations reported in Table 1 are the means of three replicate measurements Žcorrected for dilution where appropriate.. 2.3. Exposure of Capitella sp. I larÕae to reduced salinity Three experiments were conducted, all at 20 8C and a 12 L:12 D photoperiod. In each experiment, the different salinities were created by diluting seawater Žabout 32‰ salinity. with deionized water. Salinities were confirmed using a refractometer. The first experiment ŽExp S-1. used larvae pooled from four females, with three replicates per treatment and 10 larvae per dish. Five salinities were tested, ranging between 10‰ and 30‰. After exposure to the test salinities for either 24 or 48 h, unmetamorphosed larvae were transferred to full strength seawater Ž45 ml. and about 1 g of mud. As with cadmium exposure, exposure to water of reduced salinity stimulates metamorphosis in a substantial fraction of Capitella larvae ŽPechenik et al., 2001.; this had the effect of substantially reducing sample sizes in some of our treatments. Juveniles were reared for an additional 2–2.5 weeks, with water changed every 5 days and additional aliquots of
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mud added as needed, to promote continuous growth and prevent competition for food among individuals. At the end of the experiment we retrieved all surviving juveniles and measured the length and width of each individual at 8–20 = using a dissecting microscope equipped with an ocular micrometer, as described above.
Fig. 1. Influence of larval cadmium exposure Žas CdCl 2 . on juvenile mortality of Capitella sp. I in two experiments. Larvae were exposed to the concentrations indicated for 24 or 48 h and then transferred to control seawater with sediment for the duration of the study. Each bar represents the mean of three replicates Žq1 S.D.. with 10–12 larvae per replicate.
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Fig. 2. Influence of larval cadmium exposure Žas CdCl 2 . on mean fecundity Žq1 S.D.. of Capitella sp. I in experiment Cad-1. Larvae were exposed to the concentrations indicated for 24 or 48 h and then transferred to control seawater with sediment to induce metamorphosis. Juveniles were reared under those control conditions for the duration of the study. Numbers above each bar indicate the number of females that produced larvae before the end of the study.
The second set of two experiments were similar to the first except that in Exp S-2, we established four replicates per treatment, each with 12 larvae per replicate, but tested only four salinities including the 30‰ control. In Exp S-3, larvae were exposed to only three salinities Ž30‰, 12‰, 10‰.. They were generally stressed either for 24 h Žthree replicates, 12 larvae per replicate. or 48 h Žthree replicates, 12 larvae per replicate. except that larvae were exposed to the lowest salinity only for 24 h. 2.4. Data analysis Data were analyzed by one-way analysis of variance ŽANOVA. followed by Bonferroni Multiple Comparisons tests to determine the source of significant differences among means. Statistical comparisons were made against the 30‰ control data unless otherwise indicated.
Fig. 3. Influence of larval cadmium exposure Žas CdCl 2 . on final mean juvenile body volume in two experiments ŽCad-1, Cad-2. for the polychaete Capitella sp. I. Larvae were exposed to cadmium for 24 or 48 h as indicated and then transferred to control seawater with sediment for the duration of the study. Body volumes were estimated by measuring juvenile length and width and using the formula for the volume of a cylinder. The number above each bar indicates the number of individuals measured for each treatment. ) Indicates means that differ significantly Ž P - 0.05. from the control mean.
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3. Results 3.1. Effect of cadmium on Capitella sp. I No larvae died during the 24 or 48 h exposures to elevated cadmium at concentrations up to 1000 mg ly1 in either experiment; 2–3 larvae from each replicate died within
Fig. 4. Influence of larval cadmium exposure on larval mortality Ža. and juvenile growth rate Žb. for C. fornicata. Larvae were exposed to the indicated cadmium concentrations Žas CdCl 2 . for 48 h and then reared for additional week in control seawater before inducing metamorphosis. Juveniles were cultured in control seawater on a diet of T-ISO. In Ža., each bar represents the mean mortality Žq1 S.D.. of four replicates per treatment and 10 larvae per replicate. In Žb., the error bars indicate 1 S.D. above the mean and the number above each bar indicates the number of juveniles measured for each treatment.
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24–48 h at 2,000 mg ly1 . There was no significant effect Ž P ) 0.10. of either 24 h or 48 h cadmium exposures on post-settlement survival ŽFig. 1a, b. or fecundity ŽFig. 2. at any concentration tested. Even if the control mortality of juveniles in experiment Cad-2 had been as low as the control mortality in Cad-1 ŽFig. 1., the differences in mean survival would still not have been significant Ž p s 0.21..
Fig. 5. The influence of larval exposure to reduced salinity Ža. on the survival and growth Žb. of Capitella sp. I reared under control conditions Ž30 ppt s 30‰.. Larvae were exposed to the indicated salinities for 24 and 48 h and then transferred to control seawater with sediment for the duration of the study. Juvenile mortality and final body volume were determined after 2–2.5 weeks. In Ža., each bar represents the mean mortality Žq1 S.D.. from four replicates with 12 larvae per replicate. In Žb., the number above each bar represents the number of individuals measured from each treatment, and error bars represent 1 S.D. above the mean. ) Indicates means that differ significantly from the control means.
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Experiment
Salinity Ž‰.
Percent juvenile mortality
Final body volume Žmm3 .
Eggs per female
24 h exposure
48 h exposure
24 h
48 h
24 h
48 h
1.5"2.6 Ž19. 2.3"1.3 Ž16. 0.78"0.4 Ž11. 0.82"0.4 Ž17. –
3.1"0.8 Ž13. 2.8"2.2 Ž17. 3.5"1.5 Ž6. 0.5"0.3 Ž5. ) 0.1 Ž1.
84.5"26.1 Ž6. 123.0"23.4 Ž10. – 45.9"29.3 Ž12. ) –
105.4"67.3 Ž7. 137.8"44.6 Ž8. 120.3"66.4 Ž4. 170.0"124.5 Ž2.
S-1
30 20 15 12 10
29.6"23.2 Ž3. 40.7"17.0 Ž3. 56.2"26.5 Ž2. 21.9"29.6 Ž3. –
24.3"6.1 Ž2. 19.8"5.4 Ž3. 33.3 Ž1. 37.5 Ž1. 92.9"10.1 Ž2. )
S-2
30 15 12 10
0"0 Ž2. 9.5"12.7 Ž3. 17.4"15.1 Ž3. 58.4"16.7 Ž3. )
17.3"6.8 Ž3. 21.4"9.3 Ž4. 53.3"41.2 Ž3. ) 90.0"14.1 Ž2. )
3.5"1.6 Ž16. 1.9"1.3 Ž29. ) 1.4"1.1 Ž27. ) 0.5"0.7 Ž14. )
3.1"1.8 Ž19. 1.8"0.9 Ž34. ) 0.6"0.4 Ž12. ) 2.8 Ž1.
138.8"48.6 Ž11. 80.0"47.2 Ž14. ) 129.4"69.7 Ž9. 155.3"47.9 Ž3.
87.3"39.1 Ž9. 87.1"59.3 Ž16. – –
S-3
30 12 10
16.7"11.6 Ž3. 31.8"17.3 Ž4. 35.0"12.9 Ž4.
42.0"32.0 Ž3. 100 Ž1. –
1.8"1.7 Ž25. 0.9"0.9 Ž22. 1.1"1.1 Ž26.
– – –
77.0"47.7 Ž5. 106.0"89.1 Ž2. 112.8"84.3 Ž4.
– – –
)
Indicates means that differ significantly from the control Ž30‰. means.
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Table 2 Influence of exposing larvae of Capitella sp. I to low salinity on post-settlement performance Each entry is the mean"1 S.D. Ž N .. N s number of replicates for juvenile mortality columns, number of individuals for other columns. Each replicate contained 10–12 larvae.
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There was also no consistent effect of larval exposure to cadmium on juvenile growth rate ŽFig. 3.. No significant differences in final mean body volume resulted from 24 h exposures at any cadmium concentration tested ŽFig. 3a.. Following 48 h exposures in this same experiment ŽCad-1., mean juvenile growth was significantly reduced below that of control worms for individuals exposed as larvae to four of the cadmium concentrations. However, growth was not significantly reduced following exposure to the highest concentration tested Ž1000 mg ly1 ., and indeed, mean body volume did not differ significantly for individuals at any concentration between 15.6 and 1000 mg ly1 ŽFig. 3b.; any differences in final mean body size were significant Ž P - 0.05. only relative to the final mean body size of the control individuals. 3.2. Effects of cadmium on C. fornicata Larval mortality exceeded 50% by the end of 48 h at the four highest cadmium concentrations tested ŽFig. 4a.; no larvae survived the full 48 h at the highest concentration tested, 20,000 mg ly1 . Nevertheless, survivors showed comparable mean juvenile growth rates regardless of treatment ŽFig. 4b.. 3.3. Effects of salinity on Capitella sp. I No larvae died during the 24 or 48 h exposures to reduced salinity, although the larvae were pale and sluggish at 12‰ and were pale and completely inactive at 10‰. However, the percentage of juveniles recovered several weeks after adding mud to induce metamorphosis was significantly reduced for individuals subjected as larvae to the lowest salinity Ž10‰. for as little as 24 h in two of the three experiments Že.g., Fig. 5a; Table 2.. In one experiment ŽExp S-2. post-settlement mortality was significantly elevated for larvae exposed to 12‰ salinity for 48 h ŽTable 2.. Significant declines in juvenile growth rate were recorded in two of the three experiments ŽS-1 and S-2. for individuals that had been exposed for either 24 or 48 h to reduced salinity as larvae Že.g., Fig. 5b; Table 2.. In contrast, fecundity was significantly reduced for individuals in some of the 24 h low-salinity exposures, but was not significantly reduced in any experiment following 48 h exposures to reduced salinity ŽTable 2..
4. Discussion Short-term larval experiences such as delayed metamorphosis and nutritional stress have been shown to affect several aspects of juvenile performance for a variety of species in a variety of taxonomic groups Žreviewed by Pechenik et al., 1998.. Similarly, exposing larvae to sublethal low-salinity stress Že.g., 10–12‰. for only 24–48 h significantly reduced post-settlement survival and mean juvenile growth rates for Capitella sp. I Žthis study. and reduced mean juvenile growth rates for both C. fornicata and C. plana ŽJ. Jarrett, personal communication.. Remarkably, however, we could
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detect no significant post-metamorphic response to cadmium exposure in the present experiments for either of the two species tested. For Capitella sp. I., the highest concentration tested Ž2000 mg ly1 . was more than 2.5 times higher than the reported LD-50 for this species ŽReish et al., 1976. and several larvae died at this concentration in each replicate before the exposure ended. For C. fornicata, more than 50% of the larvae died during the 48 h exposures at concentrations of 2500 mg ly1 or higher. Thus, the larvae of both species clearly perceived the stress. Nevertheless, individuals surviving even the most toxic treatments performed as well after metamorphosis as control individuals. Exposing larvae of the bryozoan Bugula neritina to copper Ž100 mg ly1 . for 6 h reduced the survival of colonies transplanted to the field Žat one of the two sites included in the study. but did not affect any measure of reproductive potential ŽNg and Keough, in press.. Colonies transplanted to the field also showed slightly reduced growth rates, but only at one of the sites and only six weeks after being transplanted. Similarly, exposing bryozoan larvae ŽWatersipora subtorquata. to produced water Ža toxic byproduct of oil drilling. did not substantially affect the ability of juveniles to grow or compete for space ŽA. Boxhall and P.T. Raimondi, personal communication., even though delaying metamorphosis for less than 24 h dramatically reduces the growth rate and competitive ability of other bryozoan species Že.g., Wendt, 1996, 1998.. Why do some stressful larval experiences carry over so dramatically into postmetamorphic life whereas others do not? Answering that question will require a better understanding of the mechanisms through which the non-pollutant stresses reduce juvenile or adult performance. Delayed metamorphosis, direct nutritional stress, and reduced salinity may affect juvenile performance by interfering with the development of feeding structures Že.g., Wendt, 1996., or the development of adult tissues involved in nutrient assimilation, or, in the case of elevated juvenile mortality, by reducing nutritional reserves below those needed for successfully completing metamorphosis. Although larvae of C. fornicata may be more sensitive to cadmium later in development than they were in these experiments, it appears that sublethal cadmium exposure does not affect the same processes that are affected by delayed metamorphosis, salinity stress, or nutritive stress. Cadmium generally acts as a broad-spectrum calcium channel blocker Že.g., Colwell and Levine, 1999.. Exposing larvae to sublethal concentrations of toxic hydrocarbons or some other pollutants may exert subtle carry-over effects even though cadmium exposure does not. Heintz et al. Ž2000., for example, report that exposing embryos of the pink salmon Ž Oncorhynchus gorbuscha. to sublethal concentrations of crude oil hydrocarbons caused a 15% decline in survival over the next 2 years, possibly through an effect on juvenile growth rate. It will be interesting to look for carry-over effects in Capitella sp. I, C. fornicata, and other invertebrates in response to other pollutants.
Acknowledgements We thank Jerry Jarrett and an anonymous reviewer for their comments on a draft of the manuscript. [SS]
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References Au, D.W.T., Lee, C.Y., Chan, K.L., Wu, R.S.S., 2001. Reproductive impairment of sea urchins upon chronic exposure to cadmium Part I: Effects on gamete quality. Environ. Pollut. 111, 1–9. Blanchard, M., 1997. Spread of the slipper limpet Crepidula fornicata ŽL. 1758. in Europe. Current state and consequences. Sci. Mar. 61 ŽSuppl 2., 109–118. Butman, C.A., Grassle, J.P., Webb, C.M., 1988. Substrate choices made by marine larvae settling in still water and in a flume flow. Nature 333, 771–773. Cohen, R.A., Pechenik, J.A., 1999. Relationship between sediment organic content, metamorphosis, and postlarval performance in the deposit-feeding polychaete Capitella sp. I. J. Exp. Mar. Biol. Ecol. 240, 1–18. Colwell, C.S., Levine, M.S., 1999. Metabotropic glutamate receptor modulation of excitotoxicity in the neostriatum: role of calcium channels. Brain Res. 833, 234–241. Dubilier, N., 1988. H 2 S—A settlement cue or a toxic substance of Capitella sp. I larvae? Biol. Bull. 174, 30–38. Eisler, R., 1985. Cadmium hazards to fish, wildlife, and invertebrates: a synoptic review. U. S. Fish Wildl. Serv. Biol. Rep. No. 2, 46 pp. Heintz, R.A., Rice, S.D., Wertheimer, A.C., Bradshaw, R.F., Thrower, F.P., Joyce, J.E., Short, J.W., 2000. Delayed effects on growth and marine survival of pink salmon Oncorhynchus gorbuscha after exposure to crude oil during embryonic development. Mar. Ecol. Progr. Ser. 208, 205–216. Levin, L., Caswell, H., Bridges, T., DiBacco, C., Dabrera, D., Plaia, G., 1996. Demographic responses of estuarine polychaetes to pollutants: life table response experiments. Ecol. Appl. 6, 1295–1313. Lussier, S.M., Boothman, W.S., Poucher, S., Champlin, D., Helmstetten, A., 1999. Comparison of dissolved and total metals concentrations from acute tests with saltwater organisms. Environ. Toxicol. Chem. 18, 889–898. Ng, T.Y.T., Keough, M.J., in press. Effects of short-term larval exposure to copper on the performance of the bryozoan Bugula neritina in laboratory and field-transplant studies. J. Exp. Mar. Biol. Ecol. Pechenik, J.A., 1999. On the advantages and disadvantages of larval stages in benthic marine invertebrate life cycles. Mar. Ecol. Progr. Ser. 177, 269–297. Pechenik, J.A., Cerulli, T.R., 1991. Influence of delayed metamorphosis on survival, growth, and reproduction of the marine polychaete Capitella sp. I. J. Exp. Mar. Biol. Ecol. 151, 17–27. Pechenik, J.A., Gee, C.C., 1993. Onset of metamorphic competence in larvae of the gastropod Crepidula fornicata ŽL.., judged by a natural and an artificial cue. J. Exp. Mar. Biol. Ecol. 167, 59–72. Pechenik, J.A., Heyman, W.D., 1987. Using KCl to determine size at competence for larvae of the marine gastropod Crepidula fornicata ŽL... J. Exp. Mar. Biol. Ecol. 112, 27–38. Pechenik, J.A., Rittschof, D., Schmidt, A.R., 1993. Influence of delayed metamorphosis on survival and growth of juvenile barnacles Balanus amphitrite. Mar. Biol. 115, 287–294. Pechenik, J.A., Estrella, M.S., Hammer, K., 1996a. Food limitation stimulates metamorphosis of competent larvae and alters postmetamorphic growth rate in the marine prosobranch gastropod Crepidula fornicata. Mar. Biol. 127, 267–275. Pechenik, J.A., Hammer, K., Weise, C., 1996b. The effect of starvation on acquisition of competence and post-metamorphic performance in the marine prosobranch gastropod Crepidula fornicata ŽL... J. Exp. Mar. Biol. Ecol. 199, 137–152. Pechenik, J.A., Wendt, D.E., Jarrett, J.N., 1998. Metamorphosis is not a new beginning. BioScience 48, 901–910. Pechenik, J.A., Berard, R., Daniels, D., Gleason, T.R., Champlin, D., 2001. Influence of lowered salinity and elevated cadmium on the survival and metamorphosis of trochophores in Capitella sp. I. Invertebr. Biol. 120, 142–148. Reish, D.J., Martin, J.M., Piltz, F.M., Word, J.Q., 1976. The effect of heavy metals on laboratory populations of two polychaetes with comparisons to the water quality conditions and standards in southern California marine waters. Water Res. 10, 299–302. Wendt, D.E., 1996. Effect of larval swimming duration on success of metamorphosis and size of the ancestrular lophophore in Bugula neritina ŽBryozoa.. Biol. Bull. 191, 224–233.
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Wendt, D.E., 1998. Effect of larval swimming duration on growth and reproduction of Bugula neritina ŽBryozoa. under field conditions. Biol. Bull. 195, 126–135. Williams, K.A., Green, D.W.J., Pascoe, D., Gower, D.E., 1986. The acute toxicity of cadmium to different larval stages of Chironomus riparius ŽDiptera: Chironomidae. and its ecological significance for pollution regulation. Oecologia 70, 362–366.
Influence of larval exposure to salinity and cadmium stress on juvenile performance of two marine invertebrates žCapitella sp. I and Crepidula fornicata/ Jan A. Pechenik a,) , Tim Gleason b, Dara Daniels a , Denise Champlin b b
a Biology Department, Tufts UniÕersity, Medford, MA 02155, USA U.S. EnÕironmental Protection Agency, Office of Research and DeÕelopment, National Health and EnÕironmental Effects Research Laboratory, Atlantic Ecology DiÕision, 27 Tarzwell DriÕe, Narragansett, RI 02882, USA
Received 9 November 2000; received in revised form 28 April 2001; accepted 30 June 2001
Abstract Delayed metamorphosis and short-term food limitation reduce juvenile or adult fitness in a number of marine invertebrate species. In this study, we tested the ability of pollutant and salinity stress to bring about similar effects on juvenile or adult performance. Larvae of the polychaete Capitella sp. I were exposed to sublethal cadmium stress Žup to 2000 mg ly1 . or salinity stress Ždown to 10‰. for 24 and 48 h at 23 8C. Following exposure, we induced surviving larvae to metamorphose and monitored the subsequent survival, growth, and reproductive output of juveniles reared under control conditions Žno added cadmium, 32‰ salinity.. Similarly, larvae of the gastropod Crepidula fornicata were exposed for 24 and 48 h to cadmium in seawater Žup to a nominal concentration of 20,000 mg ly1 .. Surviving larvae were reared to metamorphic competence in the absence of cadmium, induced to metamorphose, and maintained under control conditions for an additional 5 days to monitor juvenile growth rates and survival. Exposing larvae of Capitella sp. I to low salinity Ž10–12‰. for 48 h generally did not affect adult fecundity, but stressing the larvae for as little as 24 h significantly reduced post-settlement survival and juvenile growth rates Ž P - 0.05.. In contrast, exposing larvae of this species to cadmium for even 48 h had no significant effects on post-settlement survival or fecundity, and no consistent effect on mean juvenile growth rate. Similarly, cadmium exposure did not significantly affect mean juvenile growth rates for C. fornicata, even when larvae were severely stressed Ži.e., when larval mortality
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Corresponding author. Tel.: q1-617-627-3195; fax: q1-617-627-3805. E-mail address: [email protected] ŽJ.A. Pechenik..
0022-0981r01r$ - see front matter q 2001 Elsevier Science B.V. All rights reserved. PII: S 0 0 2 2 - 0 9 8 1 Ž 0 1 . 0 0 3 1 3 - 6
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exceeded 50% during exposures.. We suggest that heavy metal stressors do not act through the same mechanism as the stresses of inadequate food supply, reduced salinity, and delayed metamorphosis. q 2001 Elsevier Science B.V. All rights reserved. Keywords: Capitella; Cadmium; Crepidula; Larvae; Metamorphosis; Salinity stress
1. Introduction It has become increasingly clear in recent years that metamorphosis is not always a fresh beginning for marine invertebrates Žreviewed by Pechenik et al., 1998; Pechenik, 1999.. For example, newly metamorphosed juveniles of Crepidula fornicata grew nearly 70% more slowly if they were starved for 5 days as larvae ŽPechenik et al., 1996b. and grew about 37% more slowly if food concentrations were dramatically reduced for just 3 days during larval development ŽPechenik et al., 1996a.. Similarly, newly settled worms of Capitella sp. I showed dramatically lower survival if larvae had been forced to delay metamorphosis for as little as 72 h ŽPechenik and Cerulli, 1991., and newly metamorphosed barnacles Ž Balanus amphitrite. exhibited markedly reduced rates of tissue growth if cyprids were forced to delay metamorphosis for as little as 3 days ŽPechenik et al., 1993.. Similar findings have been reported for a variety of invertebrate species from several taxonomic groups Žreviewed by Pechenik et al., 1998.. Thus, sublethal stresses experienced by larvae have the potential to dramatically reduce post-metamorphic performance, either directly or through reduced competitive ability or reproductive output. Pechenik et al. Ž1998. predicted that exposing larvae to sublethal pollutant stress would have similar impact on juvenile survival, growth rates, competitive ability, time to reproductive maturity, or fecundity. In this study we examined the consequences on juvenile performance of exposing larvae to either low salinity stress Žthe deposit-feeding polychaete Capitella sp. I. or elevated cadmium Ž Capitella sp. I and the suspension-feeding gastropod C. fornicata.. The larvae of Capitella sp. I emerge into the plankton as non-feeding trochophores that are competent to metamorphose within minutes of being released Že.g., Pechenik and Cerulli, 1991.. In contrast, larvae of C. fornicata are released as feeding veligers that must develop for at least a week before becoming competent to metamorphose Že.g., Pechenik et al., 1996b.. Capitella sp. I is particularly common in areas of fluctuating salinity and high heavy metal concentrations, as at sewage outfalls ŽLevin et al., 1996.. C. fornicata is a common inhabitant of bays and estuaries in many parts of the world ŽBlanchard, 1997., and is therefore exposed to a variety of coastal pollutants. Cadmium is a major component of industrial discharge and is highly toxic to a variety of aquatic animals Že.g., Eisler, 1985; Williams et al., 1986; Au et al., 2001.. 2. Materials and methods 2.1. Exposure of Capitella sp. I to cadmium Adult worms were obtained from Dr. J. Grassle ŽRutgers University. and maintained in our laboratory for multiple generations at about 30‰ salinity on a diet of fine mud
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collected from the Little Sippewissett salt marsh on Cape Cod, Massachusetts, USA, where members of this species occur naturally. The mud was forced through a 1-mm sieve to remove debris and then frozen for at least several days to kill animal residents before use ŽCohen and Pechenik, 1999.. Two experiments ŽCad-1 and Cad-2. were conducted at 23 8C, both using static exposures to toxicant. Approximately 600 larvae released from brood tubes by four females were pooled and subsampled for the first experiment. The larvae were distributed among plastic cups containing 50 ml of either 0.45-mm-filtered seawater or CdCl 2 seawater solution. Seven CdCl 2 concentrations were tested, up to a nominal concentration of 1000 mg ly1 , plus control seawater, with three replicates per treatment and 12 larvae per replicate. Survival and incidence of metamorphosis were assessed after 24 and 48 h; individuals were observed at 100 = using a compound microscope to confirm loss of larval cilia. Metamorphosed individuals were discarded. Remaining larvae were then transferred to cups of seawater with mud to stimulate natural metamorphosis ŽButman et al., 1988; Dubilier, 1988; Pechenik and Cerulli, 1991; Cohen and Pechenik, 1999., and maintained at 20 8C for 2–2.5 weeks to assess juvenile survival and growth, and adult fecundity. At the end of the experiment we retrieved all surviving juveniles and measured the length and width of each individual at 8–20 = using a dissecting microscope equipped with an ocular micrometer. Worms were anesthetized in 0.36 M MgCl 2 before measurements were taken. Worm volumes were calculated assuming that each individual was cylindrical. The second experiment was conducted about 1 week later, subsampling from 310 larvae pooled from two females. The experiment was similar to the first one, except that we added one higher nominal CdCl 2 concentration Ž2000 mg ly1 ., and examined larvae only after 48 h. Again, we tested three replicates at each concentration Žincluding seawater controls., with 12 larvae per replicate. 2.2. Exposure of C. fornicata to cadmium One experiment was conducted, using 360 larvae. Stacks of adults were collected from North Kingston, RI, USA in late August 1998. Larvae were released from one female in the laboratory 4 days later and were fed the naked flagellate Isochrysis galbana Žclone T-ISO. for 24 h before the experiment began on the following day. Larvae were exposed to CdCl 2 in static conditions Ž23 8C. at nine concentrations Žincluding a seawater control., up to a nominal concentration of 20,000 mg ly1 , with four replicates per treatment and 10 larvae per replicate. Nominal and measured cadmium concentrations for this experiment are shown in Table 1. Each replicate
Table 1 Nominal and measured total cadmium concentrations Žmg ly1 . for the C. fornicata study Nominal concentration Measured concentration
0 Žcontrol. -10
156.25 243
312.5 361
625 568
1250 1058
2500 2189
5000 4363
10,000 8294
20,000 16,392
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contained 50 ml of the test solution, to which phytoplankton Žclone T-ISO. was added to attain a concentration of approximately 10 5 cells mly1 . Test solutions were changed after 24 h and the exposures were terminated after 48 h. Because larvae of this species are not competent to metamorphose when released from the parent, surviving larvae were then reared for another 8 days in non-polluted Žcontrol. seawater on a diet of T-ISO with the container and feeding suspension changed every other day, after which they were induced to metamorphose by elevating the Kq concentration of seawater by 20 mM for 6 h ŽPechenik and Heyman, 1987; Pechenik and Gee, 1993.. Mean juvenile growth rates were assessed for individuals from seven of the treatments Ž0, 325, 625, 1250, 2500, 5000 and 10,000 mg ly1 .; no larvae survived the 48 h exposure at the highest concentration tested Ž20,000 mg ly1 .. Within each treatment group at least three juveniles between 860 and 1115 mm shell length were haphazardly selected from each replicate, so that juvenile growth rates were monitored for about 12 individuals per treatment. Shell lengths were measured nondestructively at 32 = . Each juvenile was reared at 25 8C in a separate glass dish with 40 ml of phytoplankton suspension ŽT-ISO, 18 = 10 4 cells mly1 .. Food and water were replaced daily for 5 days and individual shell lengths were remeasured to determine growth. Cadmium concentrations were measured at the start of the experiment following Lussier et al. Ž1999.. Prior to analysis, all aqueous samples were acidified with 1 ml of concentrated nitric acid per ml of sample at room temperature and placed on an orbital shaker table at 40 rpm for 24 h. Metal concentrations were determined using a Model 3410 inductively coupled plasma atomic emission spectrometer ŽICP-AES. ŽApplied Research Laboratories, Valencia, CA. with a detection limit of about 0.01 mg Cd ly1 . We verified instrumental calibrations by analyzing independent reference solutions before and during analysis of unknowns Žone verification for every 10 unknowns.; freedom from matrix interference was verified by analyzing spiked unknowns Žone spiked sample for every 20 unknowns.. Analyses were acceptable if verification standard or spike recovery was between 85% and 115%. Concentrations reported in Table 1 are the means of three replicate measurements Žcorrected for dilution where appropriate.. 2.3. Exposure of Capitella sp. I larÕae to reduced salinity Three experiments were conducted, all at 20 8C and a 12 L:12 D photoperiod. In each experiment, the different salinities were created by diluting seawater Žabout 32‰ salinity. with deionized water. Salinities were confirmed using a refractometer. The first experiment ŽExp S-1. used larvae pooled from four females, with three replicates per treatment and 10 larvae per dish. Five salinities were tested, ranging between 10‰ and 30‰. After exposure to the test salinities for either 24 or 48 h, unmetamorphosed larvae were transferred to full strength seawater Ž45 ml. and about 1 g of mud. As with cadmium exposure, exposure to water of reduced salinity stimulates metamorphosis in a substantial fraction of Capitella larvae ŽPechenik et al., 2001.; this had the effect of substantially reducing sample sizes in some of our treatments. Juveniles were reared for an additional 2–2.5 weeks, with water changed every 5 days and additional aliquots of
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mud added as needed, to promote continuous growth and prevent competition for food among individuals. At the end of the experiment we retrieved all surviving juveniles and measured the length and width of each individual at 8–20 = using a dissecting microscope equipped with an ocular micrometer, as described above.
Fig. 1. Influence of larval cadmium exposure Žas CdCl 2 . on juvenile mortality of Capitella sp. I in two experiments. Larvae were exposed to the concentrations indicated for 24 or 48 h and then transferred to control seawater with sediment for the duration of the study. Each bar represents the mean of three replicates Žq1 S.D.. with 10–12 larvae per replicate.
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Fig. 2. Influence of larval cadmium exposure Žas CdCl 2 . on mean fecundity Žq1 S.D.. of Capitella sp. I in experiment Cad-1. Larvae were exposed to the concentrations indicated for 24 or 48 h and then transferred to control seawater with sediment to induce metamorphosis. Juveniles were reared under those control conditions for the duration of the study. Numbers above each bar indicate the number of females that produced larvae before the end of the study.
The second set of two experiments were similar to the first except that in Exp S-2, we established four replicates per treatment, each with 12 larvae per replicate, but tested only four salinities including the 30‰ control. In Exp S-3, larvae were exposed to only three salinities Ž30‰, 12‰, 10‰.. They were generally stressed either for 24 h Žthree replicates, 12 larvae per replicate. or 48 h Žthree replicates, 12 larvae per replicate. except that larvae were exposed to the lowest salinity only for 24 h. 2.4. Data analysis Data were analyzed by one-way analysis of variance ŽANOVA. followed by Bonferroni Multiple Comparisons tests to determine the source of significant differences among means. Statistical comparisons were made against the 30‰ control data unless otherwise indicated.
Fig. 3. Influence of larval cadmium exposure Žas CdCl 2 . on final mean juvenile body volume in two experiments ŽCad-1, Cad-2. for the polychaete Capitella sp. I. Larvae were exposed to cadmium for 24 or 48 h as indicated and then transferred to control seawater with sediment for the duration of the study. Body volumes were estimated by measuring juvenile length and width and using the formula for the volume of a cylinder. The number above each bar indicates the number of individuals measured for each treatment. ) Indicates means that differ significantly Ž P - 0.05. from the control mean.
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3. Results 3.1. Effect of cadmium on Capitella sp. I No larvae died during the 24 or 48 h exposures to elevated cadmium at concentrations up to 1000 mg ly1 in either experiment; 2–3 larvae from each replicate died within
Fig. 4. Influence of larval cadmium exposure on larval mortality Ža. and juvenile growth rate Žb. for C. fornicata. Larvae were exposed to the indicated cadmium concentrations Žas CdCl 2 . for 48 h and then reared for additional week in control seawater before inducing metamorphosis. Juveniles were cultured in control seawater on a diet of T-ISO. In Ža., each bar represents the mean mortality Žq1 S.D.. of four replicates per treatment and 10 larvae per replicate. In Žb., the error bars indicate 1 S.D. above the mean and the number above each bar indicates the number of juveniles measured for each treatment.
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24–48 h at 2,000 mg ly1 . There was no significant effect Ž P ) 0.10. of either 24 h or 48 h cadmium exposures on post-settlement survival ŽFig. 1a, b. or fecundity ŽFig. 2. at any concentration tested. Even if the control mortality of juveniles in experiment Cad-2 had been as low as the control mortality in Cad-1 ŽFig. 1., the differences in mean survival would still not have been significant Ž p s 0.21..
Fig. 5. The influence of larval exposure to reduced salinity Ža. on the survival and growth Žb. of Capitella sp. I reared under control conditions Ž30 ppt s 30‰.. Larvae were exposed to the indicated salinities for 24 and 48 h and then transferred to control seawater with sediment for the duration of the study. Juvenile mortality and final body volume were determined after 2–2.5 weeks. In Ža., each bar represents the mean mortality Žq1 S.D.. from four replicates with 12 larvae per replicate. In Žb., the number above each bar represents the number of individuals measured from each treatment, and error bars represent 1 S.D. above the mean. ) Indicates means that differ significantly from the control means.
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Experiment
Salinity Ž‰.
Percent juvenile mortality
Final body volume Žmm3 .
Eggs per female
24 h exposure
48 h exposure
24 h
48 h
24 h
48 h
1.5"2.6 Ž19. 2.3"1.3 Ž16. 0.78"0.4 Ž11. 0.82"0.4 Ž17. –
3.1"0.8 Ž13. 2.8"2.2 Ž17. 3.5"1.5 Ž6. 0.5"0.3 Ž5. ) 0.1 Ž1.
84.5"26.1 Ž6. 123.0"23.4 Ž10. – 45.9"29.3 Ž12. ) –
105.4"67.3 Ž7. 137.8"44.6 Ž8. 120.3"66.4 Ž4. 170.0"124.5 Ž2.
S-1
30 20 15 12 10
29.6"23.2 Ž3. 40.7"17.0 Ž3. 56.2"26.5 Ž2. 21.9"29.6 Ž3. –
24.3"6.1 Ž2. 19.8"5.4 Ž3. 33.3 Ž1. 37.5 Ž1. 92.9"10.1 Ž2. )
S-2
30 15 12 10
0"0 Ž2. 9.5"12.7 Ž3. 17.4"15.1 Ž3. 58.4"16.7 Ž3. )
17.3"6.8 Ž3. 21.4"9.3 Ž4. 53.3"41.2 Ž3. ) 90.0"14.1 Ž2. )
3.5"1.6 Ž16. 1.9"1.3 Ž29. ) 1.4"1.1 Ž27. ) 0.5"0.7 Ž14. )
3.1"1.8 Ž19. 1.8"0.9 Ž34. ) 0.6"0.4 Ž12. ) 2.8 Ž1.
138.8"48.6 Ž11. 80.0"47.2 Ž14. ) 129.4"69.7 Ž9. 155.3"47.9 Ž3.
87.3"39.1 Ž9. 87.1"59.3 Ž16. – –
S-3
30 12 10
16.7"11.6 Ž3. 31.8"17.3 Ž4. 35.0"12.9 Ž4.
42.0"32.0 Ž3. 100 Ž1. –
1.8"1.7 Ž25. 0.9"0.9 Ž22. 1.1"1.1 Ž26.
– – –
77.0"47.7 Ž5. 106.0"89.1 Ž2. 112.8"84.3 Ž4.
– – –
)
Indicates means that differ significantly from the control Ž30‰. means.
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Table 2 Influence of exposing larvae of Capitella sp. I to low salinity on post-settlement performance Each entry is the mean"1 S.D. Ž N .. N s number of replicates for juvenile mortality columns, number of individuals for other columns. Each replicate contained 10–12 larvae.
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There was also no consistent effect of larval exposure to cadmium on juvenile growth rate ŽFig. 3.. No significant differences in final mean body volume resulted from 24 h exposures at any cadmium concentration tested ŽFig. 3a.. Following 48 h exposures in this same experiment ŽCad-1., mean juvenile growth was significantly reduced below that of control worms for individuals exposed as larvae to four of the cadmium concentrations. However, growth was not significantly reduced following exposure to the highest concentration tested Ž1000 mg ly1 ., and indeed, mean body volume did not differ significantly for individuals at any concentration between 15.6 and 1000 mg ly1 ŽFig. 3b.; any differences in final mean body size were significant Ž P - 0.05. only relative to the final mean body size of the control individuals. 3.2. Effects of cadmium on C. fornicata Larval mortality exceeded 50% by the end of 48 h at the four highest cadmium concentrations tested ŽFig. 4a.; no larvae survived the full 48 h at the highest concentration tested, 20,000 mg ly1 . Nevertheless, survivors showed comparable mean juvenile growth rates regardless of treatment ŽFig. 4b.. 3.3. Effects of salinity on Capitella sp. I No larvae died during the 24 or 48 h exposures to reduced salinity, although the larvae were pale and sluggish at 12‰ and were pale and completely inactive at 10‰. However, the percentage of juveniles recovered several weeks after adding mud to induce metamorphosis was significantly reduced for individuals subjected as larvae to the lowest salinity Ž10‰. for as little as 24 h in two of the three experiments Že.g., Fig. 5a; Table 2.. In one experiment ŽExp S-2. post-settlement mortality was significantly elevated for larvae exposed to 12‰ salinity for 48 h ŽTable 2.. Significant declines in juvenile growth rate were recorded in two of the three experiments ŽS-1 and S-2. for individuals that had been exposed for either 24 or 48 h to reduced salinity as larvae Že.g., Fig. 5b; Table 2.. In contrast, fecundity was significantly reduced for individuals in some of the 24 h low-salinity exposures, but was not significantly reduced in any experiment following 48 h exposures to reduced salinity ŽTable 2..
4. Discussion Short-term larval experiences such as delayed metamorphosis and nutritional stress have been shown to affect several aspects of juvenile performance for a variety of species in a variety of taxonomic groups Žreviewed by Pechenik et al., 1998.. Similarly, exposing larvae to sublethal low-salinity stress Že.g., 10–12‰. for only 24–48 h significantly reduced post-settlement survival and mean juvenile growth rates for Capitella sp. I Žthis study. and reduced mean juvenile growth rates for both C. fornicata and C. plana ŽJ. Jarrett, personal communication.. Remarkably, however, we could
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detect no significant post-metamorphic response to cadmium exposure in the present experiments for either of the two species tested. For Capitella sp. I., the highest concentration tested Ž2000 mg ly1 . was more than 2.5 times higher than the reported LD-50 for this species ŽReish et al., 1976. and several larvae died at this concentration in each replicate before the exposure ended. For C. fornicata, more than 50% of the larvae died during the 48 h exposures at concentrations of 2500 mg ly1 or higher. Thus, the larvae of both species clearly perceived the stress. Nevertheless, individuals surviving even the most toxic treatments performed as well after metamorphosis as control individuals. Exposing larvae of the bryozoan Bugula neritina to copper Ž100 mg ly1 . for 6 h reduced the survival of colonies transplanted to the field Žat one of the two sites included in the study. but did not affect any measure of reproductive potential ŽNg and Keough, in press.. Colonies transplanted to the field also showed slightly reduced growth rates, but only at one of the sites and only six weeks after being transplanted. Similarly, exposing bryozoan larvae ŽWatersipora subtorquata. to produced water Ža toxic byproduct of oil drilling. did not substantially affect the ability of juveniles to grow or compete for space ŽA. Boxhall and P.T. Raimondi, personal communication., even though delaying metamorphosis for less than 24 h dramatically reduces the growth rate and competitive ability of other bryozoan species Že.g., Wendt, 1996, 1998.. Why do some stressful larval experiences carry over so dramatically into postmetamorphic life whereas others do not? Answering that question will require a better understanding of the mechanisms through which the non-pollutant stresses reduce juvenile or adult performance. Delayed metamorphosis, direct nutritional stress, and reduced salinity may affect juvenile performance by interfering with the development of feeding structures Že.g., Wendt, 1996., or the development of adult tissues involved in nutrient assimilation, or, in the case of elevated juvenile mortality, by reducing nutritional reserves below those needed for successfully completing metamorphosis. Although larvae of C. fornicata may be more sensitive to cadmium later in development than they were in these experiments, it appears that sublethal cadmium exposure does not affect the same processes that are affected by delayed metamorphosis, salinity stress, or nutritive stress. Cadmium generally acts as a broad-spectrum calcium channel blocker Že.g., Colwell and Levine, 1999.. Exposing larvae to sublethal concentrations of toxic hydrocarbons or some other pollutants may exert subtle carry-over effects even though cadmium exposure does not. Heintz et al. Ž2000., for example, report that exposing embryos of the pink salmon Ž Oncorhynchus gorbuscha. to sublethal concentrations of crude oil hydrocarbons caused a 15% decline in survival over the next 2 years, possibly through an effect on juvenile growth rate. It will be interesting to look for carry-over effects in Capitella sp. I, C. fornicata, and other invertebrates in response to other pollutants.
Acknowledgements We thank Jerry Jarrett and an anonymous reviewer for their comments on a draft of the manuscript. [SS]
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References Au, D.W.T., Lee, C.Y., Chan, K.L., Wu, R.S.S., 2001. Reproductive impairment of sea urchins upon chronic exposure to cadmium Part I: Effects on gamete quality. Environ. Pollut. 111, 1–9. Blanchard, M., 1997. Spread of the slipper limpet Crepidula fornicata ŽL. 1758. in Europe. Current state and consequences. Sci. Mar. 61 ŽSuppl 2., 109–118. Butman, C.A., Grassle, J.P., Webb, C.M., 1988. Substrate choices made by marine larvae settling in still water and in a flume flow. Nature 333, 771–773. Cohen, R.A., Pechenik, J.A., 1999. Relationship between sediment organic content, metamorphosis, and postlarval performance in the deposit-feeding polychaete Capitella sp. I. J. Exp. Mar. Biol. Ecol. 240, 1–18. Colwell, C.S., Levine, M.S., 1999. Metabotropic glutamate receptor modulation of excitotoxicity in the neostriatum: role of calcium channels. Brain Res. 833, 234–241. Dubilier, N., 1988. H 2 S—A settlement cue or a toxic substance of Capitella sp. I larvae? Biol. Bull. 174, 30–38. Eisler, R., 1985. Cadmium hazards to fish, wildlife, and invertebrates: a synoptic review. U. S. Fish Wildl. Serv. Biol. Rep. No. 2, 46 pp. Heintz, R.A., Rice, S.D., Wertheimer, A.C., Bradshaw, R.F., Thrower, F.P., Joyce, J.E., Short, J.W., 2000. Delayed effects on growth and marine survival of pink salmon Oncorhynchus gorbuscha after exposure to crude oil during embryonic development. Mar. Ecol. Progr. Ser. 208, 205–216. Levin, L., Caswell, H., Bridges, T., DiBacco, C., Dabrera, D., Plaia, G., 1996. Demographic responses of estuarine polychaetes to pollutants: life table response experiments. Ecol. Appl. 6, 1295–1313. Lussier, S.M., Boothman, W.S., Poucher, S., Champlin, D., Helmstetten, A., 1999. Comparison of dissolved and total metals concentrations from acute tests with saltwater organisms. Environ. Toxicol. Chem. 18, 889–898. Ng, T.Y.T., Keough, M.J., in press. Effects of short-term larval exposure to copper on the performance of the bryozoan Bugula neritina in laboratory and field-transplant studies. J. Exp. Mar. Biol. Ecol. Pechenik, J.A., 1999. On the advantages and disadvantages of larval stages in benthic marine invertebrate life cycles. Mar. Ecol. Progr. Ser. 177, 269–297. Pechenik, J.A., Cerulli, T.R., 1991. Influence of delayed metamorphosis on survival, growth, and reproduction of the marine polychaete Capitella sp. I. J. Exp. Mar. Biol. Ecol. 151, 17–27. Pechenik, J.A., Gee, C.C., 1993. Onset of metamorphic competence in larvae of the gastropod Crepidula fornicata ŽL.., judged by a natural and an artificial cue. J. Exp. Mar. Biol. Ecol. 167, 59–72. Pechenik, J.A., Heyman, W.D., 1987. Using KCl to determine size at competence for larvae of the marine gastropod Crepidula fornicata ŽL... J. Exp. Mar. Biol. Ecol. 112, 27–38. Pechenik, J.A., Rittschof, D., Schmidt, A.R., 1993. Influence of delayed metamorphosis on survival and growth of juvenile barnacles Balanus amphitrite. Mar. Biol. 115, 287–294. Pechenik, J.A., Estrella, M.S., Hammer, K., 1996a. Food limitation stimulates metamorphosis of competent larvae and alters postmetamorphic growth rate in the marine prosobranch gastropod Crepidula fornicata. Mar. Biol. 127, 267–275. Pechenik, J.A., Hammer, K., Weise, C., 1996b. The effect of starvation on acquisition of competence and post-metamorphic performance in the marine prosobranch gastropod Crepidula fornicata ŽL... J. Exp. Mar. Biol. Ecol. 199, 137–152. Pechenik, J.A., Wendt, D.E., Jarrett, J.N., 1998. Metamorphosis is not a new beginning. BioScience 48, 901–910. Pechenik, J.A., Berard, R., Daniels, D., Gleason, T.R., Champlin, D., 2001. Influence of lowered salinity and elevated cadmium on the survival and metamorphosis of trochophores in Capitella sp. I. Invertebr. Biol. 120, 142–148. Reish, D.J., Martin, J.M., Piltz, F.M., Word, J.Q., 1976. The effect of heavy metals on laboratory populations of two polychaetes with comparisons to the water quality conditions and standards in southern California marine waters. Water Res. 10, 299–302. Wendt, D.E., 1996. Effect of larval swimming duration on success of metamorphosis and size of the ancestrular lophophore in Bugula neritina ŽBryozoa.. Biol. Bull. 191, 224–233.
114
J.A. Pechenik et al.r J. Exp. Mar. Biol. Ecol. 264 (2001) 101–114
Wendt, D.E., 1998. Effect of larval swimming duration on growth and reproduction of Bugula neritina ŽBryozoa. under field conditions. Biol. Bull. 195, 126–135. Williams, K.A., Green, D.W.J., Pascoe, D., Gower, D.E., 1986. The acute toxicity of cadmium to different larval stages of Chironomus riparius ŽDiptera: Chironomidae. and its ecological significance for pollution regulation. Oecologia 70, 362–366.