United States
Environmental Protection
Agency
Environmental Research
Laboratory
Narragansett Rl 02882
Research and Development
EPA-600/S3-81-052 Mar. 1982
Project Summary
Marine Fish Larvae
Growth and Survival
Edward D. Houde and A. Keith Taniguchi
This report describes the research
undertaken to develop and standardize
culture methods for larvae of two
common marine fishes, the spotted
seatrout Cynoscion nebulosus and the
lined sole Achirus lineatus. Culture
methods are explained and the rela-
tionships of survival, growth and yield
to temperatures, food concentration
and egg stocking densities were
determined.
Two different diets, a laboratory-
cultured diet based on the rotifer
Brachionus plicatifis, supplemented in
some experiments with brine shrimp
Anemia salina nauplii, and a net-
collected zooplankton diet were com-
pared. Feeding rates and rations also
were estimated for each species as
functions of age, size, food type and,
in the case of seatrout, temperature.
Despite considerable variability in
survival and growth rates, it is possible
to produce large numbers of larvae at
any stage from hatching to meta-
morphosis for toxicological studies
including hazard assessment and
water quality bioassays. Spotted
seatrout larvae are easier to culture
successfully than a re lined sole, which
might be a factor to consider in choice
of a bioassay test organism.
This Project Summary was devel-
oped by EPA's Environmental Re-
search Laboratory, Narragansett, Rl,
to announce key findings of the
research project that is fully docu-
mented in a separate report of the
same title (see Project Report ordering
information at back).
Introduction
The larval stages of marine organ isms
are particularly sensitive to environ-
mental stresses. Despite more difficulty
in culturing and maintaining these
stages it is useful to evaluate the
potential effects of environmental
contaminants on larval development.
The ability to culture marine fish larvae
for experimental purposes has increased
greatly in the past 10 years. However,
successful rearing techniques have not
been standardized although there is a
general recognition of important factors
that affect both survival and condition of
larvae. The role of food and nutrition in
larval culture is most important.
The purpose of this research was to
develop and evaluate standard method-
ology to culture larvae of two common
marine fishes, so that large numbers of
healthy larvae or juveniles could be
predictably produced for toxicological
research. Two species that are found in
coastal waters of the Gulf of Mexico and
southeastern United States, the spotted
seatrout (Cynoscion nebulosus) and
lined sole (Achirus lineatus). were
selected for this project to investigate
how survival, growth and condition of
the larvae were affected by culture
methods.
Research results reported here fo-
cused on the roles of prey type and prey
availability in predictably producing
metamorphosed spotted seatrout and
lined sole. Feeding rates of the larvae
were determined as a function of age
and in relation to the culture regime. In
addition to aiding design of pollution
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and contaminant-oriented experiments,
results of this study have helped to
document environmental requirements
of larvae of the two species.
Results and Discussion
Both spotted seatrout and lined sole
can be reared on a diet of zooplankton or
lab cultured food including rotifers and
brine shrimp nauplii. Survival rates at
metamorphosis increased as food con-
centrations were raised. Embryo stock-
ing levels had a relatively small effect on
survival, but lowest survival rates
usually were observed at highest
stocking levels.
Spotted Seatrout
Percent survival over all factor
combinations ranged up to 73.4% for
spotted seatrout farvae reared on
zooplankton and to 80.5% for larvae
reared on rotifers (Table 1). On a
zooplankton diet mean survival rate
increased from 15.1 to 73.4% as food
level was raised from 25 to 5000 per
liter. Survival was significantly higher at
28° than at 24° or 32°C.
Survival of seatrout larvae that were
fed rotifers did not differ significantly
from survival of zooplankton-fed larvae.
Mean survival rates over all food
concentrations and stock densities
were 37.9% for zooplankton-fed larvae
and 43.6% for rotifer-fed larvae. Survival
of larvae increased significantly as food
level was raised for both diets.
Growth of seatrout larvae, expressed
as standard length and dry weight,
increased significantly as zooplankton
concentrations and temperatures were
raised (Table 2). Lengths and weights of
larvae decreased as stock density
increased. Growth increased as food
concentration was raised for larvae
reared on the rotifer diet and decreased
as stock density increased, but the
effects were not as clear as for
zooplankton-fed larvae (Table 3). At the
1 000 per liter food concentration mean
dry weights of seatrout larvae were 6.1
times higher on the zooplankton diet
than on the rotifer diet.
A laboratory cultured diet of rotifers-
Artemia was similar to the zooplankton
diet in supporting survival of spotted
seatrout, Table 2.
Lined Sole
Survival rates, presented as 90%
confidence intervals, ranged up to
63.2% over all factor combinations for
larvae fed zooplankton and to 55.4% for
larvae fed rotifers, (Table 3). Survival
increased significantly as food levels
were raised for both zooplankton and
rotifer foods. There was some evidence
that survival was higher on rotifers at
the lowest food levels and on zooplankton
at the highest food level. At food levels
of 1000 per liter or higher, either
zooplankton or rotifers could be used to
rear large numbers of lined sole to
metamorphosis.
Lined sole grew relatively fast on
zooplankton compared to rotifers. Over
all factor combinations, mean standard
lengths at 16 days were 3.93 mm on the
zooplankton diet and only 3.40 mm on
the rotifer diet. The difference in mean
weights at 16 days was even more
striking; survivors averaged 209-pg on
the zooplankton diet but only 1 23-jug on
the rotifer diet. Mean standard lengths
and dry weights were significantly
higher at the 1000 per liter food levels
compared to the 50 and 100 per liter
levels for both diets.
Thirteen to 25 day old lined sole had
equally good survival and slightly better
growth on the lab-reared diet of rotifers-
Artemia compared to zooplankton fed
larvae, Table 4.
Length-Weight Relationships
Spotted Seatrout
The exponent in power functions that
described the length-weight relationship
for spotted seatrout larvae reared at
three different temperatures, over three
food levels (25, 100, 1000 food items
Table 1. Spotted Seairout . Ninety Percent Confidence Intervals about the Means for Percent Survival. Standard Length.
Dry Weight Per Larva and Total Yields of Larvae at 12 Days After Hatching Reared at 28°C on Either a Zooplankton
or Rotifer Diet. Confidence Limits are Placed on Means Over All Factor Combinations at the Designated Factor Level.
.90 Confidence Limits
Factor
and Level
1. Zooplankton Food 28°
Food Concentration
25
WO
WOO
5000
Stock Density (Food Concentration -
0.5
5.0
25.0
II. Rotifer Food 28°
Food Concentration
25
WO
WOO
5000
Stock Density {Food Concentration -
0.5
5.0
25.0
Percent
Survival
6.7 to 15.1
18.7 to 33.6
49. 3 to 53.1
65.7 to 7 3. 4
WOO /I)
44.5 to 51. 7
37. 2 to 52.0
15.4 to 29.4
10.9 to 35.5
36.3 to 44.3
31. 8 to 53.6
56.9 to 80.5
WOO /I)
21. 3 to 49.7
47. 9 to 53.3
39.9 to 50. 1
Standard
Length (mm)
3. 13 to 4.75
2.71 to 5.85
4.31 to7.51
5.75 to 8.81
7.23 to 10.37
5. 52 to 8.48
5. 18 to 6.88
2.33 to 3. 57
2.68 to 4. 38
3.42 to 4. 1O
3. 50 to 4. 42
3.88 to 5.20
3.7 8 to 4. 50
3.26 to 3.86
Larval
Dry Weight fag)
30.7to 341.7
O.Oto 509.7
0.0 to 1285.4
205.7 to 221 4.0
11 59.6 to 3067.4
697.9 to 1628.7
453.0 to 938.2
0.0 to 11 9.7
0.0 to 233.1
77.2 to 122.8
44. 1 to 224. 7
87.3(0372.1
96.1 to 197.7
65.1 to 11 5.1
Total Yield fmg!
O.Oto 5.2
6.2 to 11.1
65. 6 to 74.0
166.2 to 179.2
5.3 to 12.5
25.5 to 33.1
33.3to41.7
O.Oto 8.9
2.0 to 10.0
O.Oto 13.1
15.2 to 22.0
O.Oto 6.9
7.0 to 12.4
18.2 to 24.6
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Egg Source
Plankton
Plankton
Hormone
Hormone
Plankton
Plankton
Hormone
Hormone
Food Type
(1000/1)
Rotifer-Anc.miB
Rotifer-Anemia
Rotifer-Anemia
Rotifer-Anemia
Zooplankton
Zooplankton
Zooplankton
Zooplankton
Percent Survival
IStock Density 0.5/1)
657
27.1
20.0
62.3
60.0
27.4
37.7
643
Mean Standard
Length (mm)
/••- 1 S.E.)
11.97 ± 0.19
8.56 ± 0.18
8.94 ± 034
7.72 ± 0.09
15.49 ± 022
1482 ± 033
15.51 ±0.17
11.69 ±0.11
Means
Table2. Spotted Seairout . Percent Survival and Mean Length of Larvae Reared per liter), differed significantly. The
to 16 Days After Hatching at 28°C on Either a Rot/fer-Anem\a or exponent at 28°C, for either rotifer or
Zooplankton Diet. Two Egg Sources. Plankton-Collected or Hormone- zooplankton diets, was significantly
Induced, also Were Compared. higher (Analysis of Covariance) than
those at 24° or 32°C. The equations are:
• zooplankton food 24°—W =
1.071L34'
• zooplankton food 28°—W= 0.51 L39'
• rotifer food 28°—W = 0.50L393
• zooplankton food 32°—W = 0.83L362
There was no evidence that food
concentrations had a significant effect
on the length-weight relationships. The
combined length-weight relationship
for both of the food types, all temperature
and food concentrations was: W =
0.75L366.
Length-weight relationships, in theory,
can be used as an index of larval
conditions. High values of the exponent
in the power function relationships
indicate that larvae are relatively "fat"
and presumably better conditioned. We
did not find effects of food concentration
on length-weight relationships of
spotted seatrout. This result indicates
that the length-weight relationship may
not be useful to judge effects of diet on
larval condition for this species. Tem-
~~~ perature did seem to have an easily
detected effect of length-weight rela-
tionships for spotted seatrout. The
Lined Sole . Ninety Percent Confidence Intervals About the Means for Percent Survival. Standard Length, Dry
Weight Per Larva and Total Yields of Larvae at 12 Days After Hatching Reared at 28°C on Either a Zooplankton or
Rotifer Diet. Confidence Limits are Placed on Means Over All Factor Combinations at the Designated Factor Level.
.90 Confidence Limits
Percent Survival
Plankton eggs
Hormone eggs
Rotifer-Anemia diet
Zooplankton diet
Standard Length
Plankton eggs
Hormone eggs
Rotifer-Artem\a diet
Zooplankton diet
43.6%
46.1%
43.9%
45.7%
12.71 mm
10.97 mm
9.30 mm
14.38 mm
Table 3.
Percent
Survival
Standard
Length (mm)
Larval
Dry Weight
Total Yield (mg)
I. Zooplankton Food 28°
Food Concentration fStock Density 0.5/1)
50 0.0 to 2.0
100 2.2 to 12.5
WOO 30.8 to 63.2
Stock Density (Food Concentration - 1000/1)
0.5 2.6 to 60.8
2.0 0.0 to 30.0
8.0 0.8 to 38.8
16.0 0.0to21.7
II. Rotifer Food 28°
Food Concentration (Stock Density - 0.5/1)
3.31 to3.33
3.32 to 3.78
3.97 to 4.49
3 50 to 4.62
3.12 to 4.52
3.31 to 4.19
3.51 to3.81
56.6 to 97.4
87.8to 161.0
196 3 to 313.1
99.1 to 372.6
34.5 to 249.3
93.1 to 229.5
90.2 to 249.5
0.0 to <0.1
<0.1 to 1.8
5.3 to 26.2
<0.1 to 4.6
<0.1 to 3.1
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Table 4. Lined Sole . Percent Survival and Mean Length of Larvae Reared
to 25 Days After Hatching at 28°C on Either a Rotifer-Anemia or
Zooplankton Diet.
Food Type
(500/1)
Rotifer-Anemia
/?of//er-Artemia
/?or//er-Artemia
Zooplankton
Zooplankton
Zooplankton
Percent Survival
(Stock Density, 0.5/1)
54.3
75.7
40.0
51.4
48.6
70.0
Mean Standard
Length (mm)
(± 1 S.E.)
7.72 ±0.11
7.01 ± 0.07
6.84 ± 0.15
677 ±0.14
6.29 ±011
6.38 ± 0.08
Means
Percent Survival
/?or//er-Artemia diet
Zooplankton diet
Standard Length
Rotifer-Anemia diet
Zooplankton diet
56.7%
56.7%
7.19 mm
6.48 mm
"fattest" larvae, regardless of diet type,
were reared at 28°C. Larvae at lower
and higher temperatures apparently
were less well conditioned.
Lined Sole
The only length-weight relationships
that were obtained for lined sole were
those at 28°C for both the zooplankton
and rotifer diet, when food concentra-
tions were 1000 per liter. There were no
significant differences in the relation-
ships:
• zooplankton food—W = 0.60L403
• rotifer food—W = 0.63L405
Thus, there appeared to be no differences
in condition of lined sole when reared at
high food concentration for either type
of food. The length-weight relationship
from pooled zooplankton and rotifer-fed
lined sole larvae was W = 0.60L406.
Conclusions
Both spotted seatrout and lined sole
can be routinely reared from hatching
through metamorphosis on either a
zooplankton or rotifer diet. Survival
rates exceeding 50% from hatching to
metamorphosis can be predictably
obtained. Survival did not differ between
the diets, but growth and yield were
significantly higher on the zooplankton
diet when equal food concentrations
were compared. Survival, growth and
yield can be predicted with reasonable
confidence as a function of food
concentration, but with less confidence
as a function of stock density. At
moderately high food concentrations,
the degree of variability in survival and
growth responses was similar on either
the zooplankton or rotifer diet.
Spotted seatrout larvae survived
better at 28° than at 24 or 32°C, but
individual growth was best at 32°C.
Growth and survival was increased as
food levels were raised. Despite poorer
survival and growth of individuals at
high stock densities, total yields of both
seatrout and lined sole increased as
stocking density was raised.
A laboratory-cultured diet of rotifers
and Anemia salina nauplii was adequate
to grow spotted seatrout and lined sole
well into the juvenile stage with equal
survival rates compared to fish fed
zooplankton. Growth of spotted seatrout
was relatively poor on rotifers-/4rfem/a
compared to zooplankton, but lined sole
growth did not differ between diets. The
Artemia component of the lab-cultured
diet was responsible for the good
growth of lined sole on that diet.
Eggs of spotted seatrout from plankton
collections or from hormone-induced
spawning gave equal survival and
growth results in larval rearing experi-
ments.
Growth of spotted seatrout larvae
was faster than that of line sole larvae at
equal food concentrations. Specific
growth rates of spotted seatrout fed
zooplankton at 28°C ranged from 44.9
to 69.4% per day over the range of food
concentrations that was fed, while lined
sole specific growth rates ranged from
20.3 to 38.5% per day. Seatrout larvae
growth rates were among the fastest
reported for fish larvae, the maximum
measured rate being 76.5% per day at
32°C and 1000 zooplankters per liter.
Feeding rates and rations were
determined as functions of age and size
of larvae as well as of food concen-
trations, and in the case of spotted
seatrout the effect of temperature also
was documented. Under the same
conditions seatrout larvae consumed
more food per unit time than did lined
sole. Consumption of rotifers by 2-6
day-old seatrout larvae was higher than
zooplankton consumption, but subse-
quently zooplankton was consumed at
higher rates, as larval predatory ability
increased. Both seatrout and lined sole
larvae could consume much more than
100% of their body weight perday when
food concentrations were high.
Length-weight relationships were
determined as functions of food con-
centration, food type (zooplankton or
rotifers) and temperature for spotted
seatrout larvae. Only temperature had a
significant effect on the relationship;
the fattest larvae were reared at 28°C.
No significant difference was found in
the length-weight relationship for lined
sole reared at 28° on zooplankton or
rotifers at a 1000 per liter food concen-
tration.
A routine culture procedure to provide
spotted seatrout or lined sole larvae for
aquatic toxicology research could be
based upon 28°C rearing temperatures,
1000 per liter food concentrations
(either zooplankton or rotifers) and a 5.0
per liter egg stocking density. Growth
rates and survival rates of both species
are higher at food levels exceeding
1000 per liter, but such levelsare unlike
any found in nature. If the goal of the
toxicological study were to provide a
large amount of assay material (i.e.,
yield), then egg stocking densities
higher than 5.0 per liter could be used.
4
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Edward D. Houde and A. Keith Taniguchi are with the Rosenstiel School of
Marine and Atmospheric Science, University of Miami. Miami, FL 33J49.
Allan D. Beck is the EPA Project Officer (see below).
The complete report, entitled "Marine Fish Larvae Growth and Survival,"
(Order No. PB 82-1OJ 395; Cost: $9.50. subject to change! will be available
only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA 22161
Telephone: 703-487-4650
The EPA Protect Officer can be contacted at:
Environmental Research Laboratory
US. Environmental Protection Agency
South Ferry Road
Narragansett, Rl 02882
. S. GOVERNMENT PRINTING OFFICE: 1982/559-092/3378
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United States
Environmental Protection
Agency
Center for Environmental Research
Information
Cincinnati OH 45268
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