Fatty acids and starvation in larval striped bass (Morone saxatilis)

Comp. Biochem. Physiol. Vol. 77B, No. 4, pp. 785 790, 1984 Printed in Great Britain

0305-0491/84 $3.00+0.00 © 1984 Pergamon Press Ltd

FATTY ACIDS A N D STARVATION IN LARVAL STRIPED BASS (MORONE SAXATILIS) F. DOUGLAS MARTIN, DAVID A. WRIGHT and JAY C. MEANS University of Maryland, Center for Environmental and Estuarine Studies, Chesepeake Biological Laboratory, Solomons, MD 20688-00038, USA (Tel: 301 326-4281) (Received 30 August 1983)

Abstract--1. Growth of 4-day-old striped bass larvae was followed for 14 days under various starvation/feeding regimes. Fully fed larvae were supplied with 1000 Artemia nauplii per liter every day, although in experimental animals feeding was delayed for up to 10 days. All starved control larvae were dead by day 11. 2. Major constituent fatty acids were measured throughout the experimental period in fully fed, fully starved and feeding-delayed animals. The following fatty acids were found to be the principal constituents oflipids of both fed and starved larvae: 14:0, 16:0, 16:1, 18:1, 20:1, 22:1, 18:2, 18:3, 20:5, 22:5 and 22:6. 3. Two days of starvation caused a dramatic fall in total known fatty acid levels by comparison with fed larvae. Fed controls steadily increased in fatty acid content throughout the experimental period reaching a level of 202 pg per larva on day 14. 4. Starved controls showed an increase in fatty acid levels for the first six days, with a rapid decrease after day 6. Treatments having 4 and 5 days of delay in feeding also showed peak concentrations of total fatty acids on day 6, a fall in these levels on day 8, followed by a later recovery. Most component fatty acids were seen to vary with basically the same patterns. 5. Growth rates, as measured by notochord length or dry weight, were significantly inhibited by 5 days delay in feeding, but were apparently able to recover on resumption of feeding. 6. Longer periods of starvation resulted in irreversible growth inhibition.

Inc. (Castro Valley, CA, USA) and feeding level was 1000 nauplii per liter. Aquaria were maintained at room ambient temperature (19.6-21.0°C). The following treatments were established. Starved controls were maintained. Fully fed controls began feeding on day 6 past hatching. Feeding delays of 4, 5, 6 and 7 days were used. Replicate aquaria of each treatment were maintained so that at each sampling period a whole aquarium was drained and 20 larvae were frozen with liquid nitrogen for fatty acid determination, 20 larvae were frozen with liquid nitrogen from RNA:DNA determination and the rest were fixed in buffered 2~ formaldehyde for morphometric and histologic examination. Frozen larvae were maintained at -20°C until analysis could be performed. Groups of larvae were weighed, homogenized, extracted by capillary gas chromatography using the methods described in an earlier paper (Martin et el., 1984).

INTRODUCTION It has been hypothesized that either a lack of food for larvae or a mismatch in larval fish and food organism distributions is a principal cause of poor year-class strength (Hunter, 1976). Our objective was to establish criteria for assessing nutritional state in larval striped bass in order to test this hypothesis. A m o n g assessment methods tested was fatty acid content and composition. Studies of fatty acid (FA) utilization during endogenous feeding have been done for salmon (Takama et el., 1969), brook trout (Atchison, 1975), Pacific sardine (Lasker, 1962) and striped bass (Eldridge et el., 1977, 1982, 1983) and changes in F A composition at or near starvation have been shown in all cases. The use of this type of information to assess nutritional state of larvae has not been attempted previously. The data presented here represent laboratory produced normative data for such an assessment.

RESULTS

METHODS AND MATERIALS

Striped bass larvae of the Potomac River stock were obtained from the Cedarville State Fish Hatchery, Cedarville, Maryland at four days past hatching. Thirty-eight liter capacity aquaria holding 20 liters of water were stocked at 10 larvae per liter. Water used was Solomons well water brought up to 1.5 ppt salinity by addition of Patuxent River water which had been filtered using a sand bed filter rated at 5 ~m. Water was conditioned by the addition of small amounts of Chorella stock culture. All feedings were with Australian strain Artemia nauplii obtained from Artemia, Contribution No. 1506 of the University of Maryland, Center for Environmental and Estuarine Studies. 785

The following fatty acids were found to be the principal constituents of the lipids of both fed and starved larvae: 14:0; 16:0; 18:0; 16:1; 18:1; 20:1; 22:1; 18:2; 18:3; 20:5; 22:5 and 22:6. Figure 1 shows the changes in total known fatty acids as/~g per larva. By day 2 of the experiment it can be seen that there is a large difference in total known fatty acid levels with the fed controls containing 26.5/~g per larvae and starved controls containing only 1.9/~g per larvae. The fed controls show a steady increase in fatty acid levels throughout the experiment reaching a level of 201.9 p g per larva on day 14 of the experiment. The starved controls showed an increase in fatty acid levels for the first six days, with a rapid decrease following day 6. The peak level was 9.3 # g per larva on day 6 and by day 10 levels had dropped to 1.9 #g.

F. DOUGLAS MARTIN et al.

786 1000

100

"o 10

100

>,

/i

kl_

2

i'°1

'.~~:~ ..'"..,,

ii

i

].

,,. \~, y'X~.x.

i~S2./

,-

o= v

2

I,--

1 2

4

6

8

10 12

2

14

Fig. 1. C h a n g e s in total k n o w n f a t t y acids (/xg p e r l a r v a ) in

fed larvae, starved larvae and larvae which have had feeding delayed for between 4 and 7 days; Larvae fully fed at a daily rate of 1000 Artemia nauplii per 1, larvae starved throughout experimental period; ..... feeding delayed 4 days; . . . . feeding delayed 5 days; - - . - - feeding delayed 6 days; - - x feeding delayed 7 days.

All starved control larvae were dead by day 11. Treatments having 4 and 5 days of delay in feeding showed peak levels of fatty acids on day 6, similar to the starved controls. Like the starved controls, day 8 fatty acid levels were greatly reduced. However following that there was a large increase in levels with day 14 levels being 12.1 and 73.2/tg per larva for feeding delays of 4 and 5 days respectively. Feeding delays of 6 and 7 days showed lower levels of fatty acids on day 8. Larvae for the 6 day feeding delay treatment showed no increase in fatty acid levels by day 12 (1. l / l g per larva) and all were dead by day 13. Larvae from the 7 day feeding delay treatment survived to day 14 but had only recovered to fatty acid levels of 4 . 2 # g per larva. Figure 2 shows total known fatty acid content as percent of dry weight. On this basis, the fatty acid content is seen to slowly decline in fed controls while the starved controls show a peak level at day 6 followed by a rapid decline. All treatments are well below the fed controls for all dates except for the 4-day delay larvae on day 6 and the 5-day delay larvae on day 14. Figure 3 shows the changes in dry weight throughout the experiment. It can be seen that larvae from treatments with feeding delays of 4 or 5 days lag behind the fed controls but have growth rates which are comparable. Larvae from treatments with delays of 6 or 7 days have reduced average growth rates and lag far behind the other treatments in size. Figure 4 shows the pattern of variation in composition of the major constituent fatty acids as percent dry weight. It shows that most components vary with basically the same patterns. Figure 5 shows fatty acid composition of striped bass eggs (obtained from the Potomac River), fed controls on days 2 and 14, starved controls on days 6 and 10 and the Artemia nauplii used for feeding.

4

6

8

10 12 14

Days of Experiment

Days of Experiment

Fig. 2. Changes in total known fatty acids expressed as percent dry wt in fed larvae, starved larvae and larvae which have had feeding delayed for between 4 and 7 days; - larvae fully fed at a daily rate of 1000 Artemia nauplii per 1; - - - larvae starved throughout experimental period days; ..... feeding delayed 4 days; . . . . feeding delayed 5 days; - - . - - feeding delayed 6 days; - - x feeding delayed 7 days. For the saturated and monoenoic fatty acids, the composition of the fed larvae comes to resemble that of their diet, Artemia nauplii, while the profile for 18:3, 22:5 and 22:6 is intermediate between the egg profile and that of Artemia, there is the probability that at least some of the 18:3 is being converted to 22:6. The starved controls show evidence of differential utilization of saturated fatty acids and conservation of 22:5 and 22:6.

10

...../i vE

,,,'"i /

~g

CY"

......

ab

.01 2

4

6

8

10

12 14

Days of Experiment

Fig. 3. Changes in larval dry weight in fed larvae, starved larvae and larvae which have had feeding delayed for between 4 and 7 days, - Larvae fully fed at a daily rate of 1000 Artemia nauplii per 1; - - - larvae starved throughout experimental period; ..... feeding delayed 4 days; . . . . feeding delayed 5 days; . - - feeding delayed 6 days; - - x I feeding delayed 7 days.

Fatty acids in striped bass

787

100 ----14:0

16:0 - - - - 1 6 : 1

Fed . . . . 18:1 --x--18:2 - - * - - 1 8 : 3 -~-22:1

Feeding Delayed 4 days

22:5 ~ 2 2 : 6

.::...'

--

total

Starved 10

,. \,i

./

/

.

.

\

./.

-/

\

.~.. .".. .~" \ '..." .

/."

'\ /\.

-

./

..'.

'-..." '.. \

: A

\.

" "\ ,-/'\.

/ ./:x \\,

'

: : .'- ;

i .//:./:~".:~. .x

/

i..r~ '. \

/'.

. ..\.

./' ...)'~

\

"o

/ " <

"..\.\

••

\\,,7

.1 100

LL

f Feeding Delayed

Feeding Delayed

Feeding Delayed

5 Days

6 Days

7 Days

t--

10

I

ii!i

//

:~ i ~.:~ \ ~:~ .1

:':~, ,~

L_

,.~ ~

6

8

i";

t /

\

."i

",.

.,."! ~/ .>-~ ,,.,.7

• \ -',

\. /

i

\

I. 10 12

14

2

4

6

8

10 12 14

D a y s of E x p e r i m e n t

Fig. 4. Changes in total and major constituent fatty acids (expressed as percentage dry weight) in fed and starved larvae and larvae which had feeding delayed for 4-7 days.

DISCUSSION

The fatty acid composition of the eggs found in this study was similar to that reported by Eldridge et al. (1983) as was that of the starving larvae. The increase

in known fatty acids up to day 6 followed by the rapid decrease of levels in starved and delayed feeding larvae deserves further attention. The treatments with delays of 4 or 5 days show the same peak on day 6 followed by a large decrease on day 8 despite the fact

788

F. DOUGLASMARTINet al. 45,

I starved 10 days starved 6 days eggs fed 2 days fed 14 days Artemia

40

35

•~

3o

u_

25

8 "s

20

c c3

0.

15

10

0

14:0

16:0

16:1

18:1

20:1

22:1

18:2

18:3

22:5

22:6

Fig. 5, Constituent fatty acids (expressed as percentage of total known fatty acids) in larvae starved or fed for varying periods, eggs and Artemia nauplii (used as food in these experiments).

that they had been feeding for 2 and 1 days respectively on day 6. This suggests a massive shift in enzyme systems involved in lipid metabolism. This may result from completion of catabolism of the lipoprotein of the yolk material, as striped bass larvae begin feeding while still containing considerable amounts of yolk. No delayed feeding larva showed evidence of fatty acid accumulation before day 10 indicating that utilization rate was equal to dietary intake rate for up to 4 days after these enzyme systems were turned on. Eldridge et al. (1977) and Rogers and Westin (1979) report that there is no "point-of-no-return" for starving striped bass larvae in that, even late in the starvation sequence, the surviving larvae will feed if offered food and will survive. We feel that while this is correct, larvae which have delayed feeding for 6 or more days have passed a point where they have almost no chance to survive. Planktonic larvae have higher mortality rates than do the demersal juveniles (Dey, 1981) so that an increase in time spent in the planktonic stage (or inversely a lower growth rate) will eliminate a disproportionately large proportion of such larvae (Houde, 1983). Figure 6 shows growth rates for larvae from an unpublished pilot study. This pilot study differed from the study presented here in that the larvae were held in 31 containers and Houde (personal communication) has shown that striped bass larvae held in containers of less than 101 capacity have reduced growth rates. Despite this it can be seen that up to 5 days of delay in feeding cause no large change in growth rate, while delays of 7 days or more cause large decrease in growth. We

believe that this large change is related to the shift over in lipid metabolism which occurs on day 6. Our findings that fatty acid composition reflects levels in diets is consistent with patterns observed in larvae and young of other species such as channel catfish (Yingst and Stickney, 1979), Atlantic silverside (Simpson et al., 1979), gourami (Rahn et al., 1977), turbot (Cowey et al., 1976) and rainbow trout (Lee et al., 1967; Castledine and Buckley, 1980; Leger, 1981). Other sources of shifts in fatty composition of young fish have been selective consumption (or retention) of particular fatty acids (Rahn et al., 1977; Yingst and Stickney, 1979; Simpson et al., 1979) or synthesis (de novo or otherwise) of particular fatty acid (Atchinson, 1975; Cowey et al., 1976; Yingst and Stickney, 1979). Takama et al. (1969) and Simpson et al. (1979) found no evidence of selective utilization or retention, however Lee et al. (1967) and Castledine and Thomas (1980) found that o)3 PUFAs are selectively included into the phospholipid portion of the lipid pool. Eeger (1981) states that because phospholipids are primarily included in membranes they are retained far into starvation. This would give the appearance of differential retention. Our data would indicate that selective consumption, especially of saturated fatty acids, occurs as well as conversion from 18:3 to 22:6 probably occurring in striped bass larvae. The retention of 22:6 even under starvation conditions and the apparent conversion of 18:3 to 22:6 in larvae fed A r t e m i a having no 22:6 indicate that the 033 group of fatty acids are probably essential

Fatty acids in striped bass

789

fed controls

9.0

/ / ~ 4 12day daysdelay delay 5 days delay 3 days delay days delay

8.0

E

7 days delay

7.0

a~

....1

o

6.0

9 days delay Z

5.0

,.starved controls

2

4

6

8

10

12

14

Days of Experiment

Fig. 6. Growth of striped bass larvae as determined by notochord length in fed and starved specimens, and larvae subjected to feeding delays of betwen 1 and 9 days. Lines were fitted to parabolic formulae derived from the raw data.

fatty acids for striped bass as they are for almost all fishes investigated (Castell et al., 1971a,b,c; Yu and Sinnhuber, 1975, 1979; Owen et al., 1975; Fujii et al., 1976; Cowey and Sargent, 1977; Farkas et al., 1977, 1980; Leger et al., 1979; Scott and Middleton, 1979; Watanabe, 1982). The ability to convert 18:3 to 22:6 has been correlated to living in freshwater habitats (Watanabe, 1982) or to omnivory (Owen et al., 1975). It would be interesting to see whether striped bass retain the ability to elongate and desaturate 18:3 as adults, since as adults they are nearly complete piscivores after living in marine environments and as piscivores they should obtain sufficient levels of c~3 P U F A s without the need for conversion of 18:3 to 22:6. Acknowledgement--This research was supported by the U.S. Fish and Wildlife Service through contact 14-160009-81-031 to the University of Maryland Chesapeake Biological Laboratory. REFERENCES

Atchison G. J. (1975) Fatty acid levels in developing brook trout (Salvelinusfontinalis) eggs and fry. J. Fish. Res. Bd Can. 32, 2513-2515. Castell J. D., Lee D. J. and Sinnhuber R. O. (1972) Essential fatty acids in the diet of rainbow trout (Salmo gairdneri): Lipid metabolism and fatty acid composition. J. Nutr. 102, 93-100. Castell J. D., Sinnhuber R. O., Wales J. H. and Lee D. J. (1972) Essential fatty acids in the diet of rainbow trout (Salmo gairdneri): Growth, feed conversion and some gross deficiency symptoms. J. Nutr. 102, 77-86.

Castell J. D., Sinnhuber R. O., Lee D. J. and Wales J. H. (1972) Essential fatty acids in the diet of rainbow trout (Salmo gairdneri): Physiological symptoms of EFA deficiency. J. Nutr. 102, 87-92. Castledine A. J. and Buckley J. T. (1980) Distribution and mobility of 3 fatty acids in rainbow trout fed varying levels and types of dietary lipid. J. Nutr. 110, 675-685. Cowey C. B., Owen J. M., Adron J. W. and Middleton C. (1976) Studies on the nutrition of marine flatfish. The effects of different dietary fatty acids on the growth and fatty acid composition of turbot (Scophthalrnus maximus). Br. J. Nutr. 36, 479-486. Cowey C. B. and Sargent J. R. (1977) Lipid nutrition in fish. Comp. Biochem. Physiol. 57, 269 273. Eldridge M. B., Joseph J. D., Taberski K. M. and Seaborn G. (1983) Lipid and fatty acid composite of the endogenous energy sources of striped bass (Morone saxatilis) eggs. Lipids 18, 510-513. Eldridge M. B., King D. J., Eng D. and Bowers M. J. (1977) Role of the oil globule in survival and growth of striped bass (Morone saxatilis) larvae. Proceedings of the 5th Annual Conference of the Western Association of State Game and Fish Commissioners, pp. 303-313. Eldridge M. B., Whipple J. A. and Bowers M. J. (1982) Bioenergetics and growth of striped bass, Morone saxatilis, embryos and larvae. Fish. Bull. 80, 461-474. Farkas T., Csengeri I., Majoros F. and Olah J. (1977) Metabolism of fatty acids in fish. I. Development of essential fatty acid deficiency in the carp, Cyprinus carpio Linn. Aquaculture 11, 147-157. Farkas T., Csengeri I., Majoros F. and Olfi.h J. (1980) Metabolism of fatty acids in fish. Ili. Combined effect of environmental temperature and diet on formation and deposition of fatty acids in the carp, Cyprinus carpio Linnaeus 1758. Aquaculture 20, 29-40.

790

F. DOUGLAS MARTIN et al.

Fujii M., Nakayama H. and Yone Y. (1976) Effect of ~o3 fatty acids on growth, feed efficiency and fatty acid composition of red sea bream (Chrysophrys major). Rept. Fish. Res. Lab., Kyushu University. 3, 65-86. Houde E. D. (1983) The larval stages. [A review of R. Lasker (ed.). 1981. Marine fish larvae. Morphology, ecology and relation to fisheries. Washington Sea Grant Program, Univ. Washington Press, Seattle. 131 pp.] Envir. Biol. Fish. 9, 77-79. Lasker R. (1962) Efficiency and rate of yolk utilization by developing embryos and larvae of the Pacific sardine Sardinops caerulea (Girard). J. Fish. Res. Bd Canada 19, 867-875. Lee D J., Roehm J. N., Yu T. C. and Sinnhuber R. O. (1967) Effect of ~03 fatty acids on the growth rate of rainbow trout, Salmo gairdneri. J. Nutr. 92, 93-98. Leger C. (1981) Eff~t d'un jeune prolonge sur la composition en lipides et en acides gras de la truite arc-en-ciel, Salmo gairdneri. Aquaculture 25, 195-206. Leger C., Gatesoupe F.-J., Metailler R., Luquet P. and Fremont L. (1979) Effect of dietary fatty acids differing by chain lengths and 09 series on the growth and lipid composition of turbot Scophthalnus maximus L. Comp. Biochem. Physiol. 64B, 345-350. Martin F. D., Wright D. A., Means J. C. and SetzlerHamilton, E. (1984) The importance of food supply to survival of larval striped bass in the Potomac River estuary. Trans. Am. Fish. Soc. (in press). Owen J. M., Adron J. W., Middleton C. and Cowey C. B. (1975) Elongation and desaturation of dietary fatty acids in turbot, Scophthalnus maximus L., and rainbow trout, Salmo gairdnerii Rich. Lipids 10, 528-531. Rahn C. H., Sand D. M. and Schlenk H. (1977) Metabolism of oleic, linoleic and linoleic acids in gourami (Tricho-

gaster cosby) fry and mature females. Comp. Biochem. Physiol. 58, 17-20. Rogers B. A. and Westin D. T. (1979) The combined effect of temperature and delayed initial feeding of the survival and growth of larval striped bass Morone saxatilis (Walbaum). Advances in Marine Environmental Research U.S. E.P.A. (Edited by Jacoff F. S.), pp. 234-250. Environmental Research Laboratory EPA-600/9-79-035. Scott A. P. and Middleton C. (1979) Unicellular algae as a food for turbot (Seophthalnus maximus L.) larvae--The importance of dietary long-chain polyunsaturated fatty acids. Aquaculture 18, 227-240. Simpson K. L., Richardson L. M. and Schauer P. S. (1979) Evaluation of various diets on the lipid and protein composition of early life stages of the Atlantic silverside. In Advances in Marine Environmental Research U.S. E.P.A. (Edited by Jacoff F. S.), pp. 214-233. Environmental Research Laboratory EPA-600/9-79-035. Takama K., Zama K. and Igarashi H. (1969) Changes in the lipids during the development of salmon eggs. Bull. of Faculty of Fisheries. Hokkaido University. 20, 118-126. Watanabe T. (1982) Lipid nutrition in fish. Comp. Biochem. Physiol. 73B, 3-15. Yingst W. L. III and Stickney R. R. (1979) Effect of dietary lipids on fatty acid composition of channel catfish fry. Trans. Am. Fish. Soc. 108, 62~625. Yu T. C. and Sinnhuber R. O. (1975) Effect of dietary linolenic and linoleic acids upon growth and lipid metabolism of rainbow trout (Salmo gairdneri). Lipids 10, 63-66. Yu T. C. and Sinnhuber R. O. (1979) Effect ofo~3 and ~o6 fatty acids on growth and feed conversion efficiency of coho salmon (Oncorhynchus kisuch). Aquaculture 16, 31-38.