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Interactive effect of diet and temperature on the growth of juvenile clams
J. Exp. Mar. Biol. Ecol., 1987, Vol. 113, pp. 23-38 Elsevier
23
JEM 00959
Interactive effect of diet and temperature on the growth of juvenile clams I. L&g,
S. D. Utting and R. W. S. Kilada
Fisheries ~aboruto~, Conwy, Gwynedd, U.K., Directorate of Fisheries Research, Ministry ofAgr<ure, Fisheries and Food
(Received 6 April 1987; revision received 3 July 1987; accepted 14 July 1987) Abstract: Small juveniles (0.3-3 mg live weight) of three clam species, Tapes semidecussata Reeve, T. decussata L., and Mercenariu mercenarfa L., were fed eight single-species algal diets at a range of temperatures. Nutritional value of the algae tested was in the order: Isochrysis aff gulbana (T. ISO) = Skeletonema costatum > Chaetoceros calciirans = Chroomonas salina = Thalassiosira pseudonana > Tetraselmis suecica z Phaeodactylum tricornutum > Chlamydomonas coccoides, when the same weight of these species was given
in the diet to each clam species, Reasons for differences in nutritional value are discussed with reference to the biochemical content of the algae and the relative growth efficiency of the animals. Respiration rate, food cell clearance rate, and growth (as increase in organic weight) of M. mercenariu and T. semidecussatu increased with temperature from 10 to 25 “C. Growth rates decreased at > 25 “C. T. decussata showed only a slightly increased growth response at > 15 “C. Key words: Algal diet; Temperature; Clam; Growth eflkiency
INTRODUCTION
The commercial hatchery culture of bivalve mollusc’s requires a supply of live algal (phytoplankton) food cells of high nutritional value. The food value of a wide range of algae has been assessed for the culture of oysters (e.g., Walne, 1970; Tenore & Dunstan, 1973; Enright et al., 1986). A few selected species are now used routinely, often in mixtures, to provide a diet on which the animals will grow efficiently (Epifanio, 1979; Romberger & Epifanio, 1981; Laing & Millican, 1986). Recent work has focussed on the ident~~ation of particular components of the diet that are essential for healthy growth (Langdon, 1983) and the importance of polyunsaturated fatty acids (PUFA) has been reported (Waldock & Holland, 1984). There is an increasing interest in the hatchery rearing of the commercially valuable clam species, Tapes semidecussata Reeve and Mercenaria mercenaria L., for which there are markets in Asia, North and South America, and Europe (Lucas, 1977; Claus, 198 1; McHugh, 1981; Burrell, 1983), and also T. decussatu L., for which there is demand in Europe (Rodriguez, 1983; Breber, 1985). Depletion of natural and introduced stocks Correspondence address: 1. Laing, Fisheries Laboratory, Directorate of Fisheries Research, Ministry of Agriculture, Fisheries and Food, Benarth Road, Conwy, Gwynedd LL3.2 8UB, U.K. 0022-098 1/87i$O3.50 0 1987 Elsevier Science Publishers B.V. (Biomedical Division)
24
I. LAING &‘-AL.
of these cfams has brought about an increased demand for hatchery-produced young (seed). They are often cultured on algal diets designed for oyster culture, but there is very little evidence to support the assumption that these diets are suitable for clams. Some reports, however, indicate dflerences in nutritional value between certain algal species used to feed clams and oysters (Loosanoff & Davis, 1963; Walne, 1970; Epifanio, 1979) although comparisons can be unreliable as foods were not from the same growth phase (Webb & Chu, 1983). Any differences may play a significant part in juvenile growth performance. Physical aspects of the hatchery environment for clams, especially temperature, have also been adopted from oyster culture. Again, few detailed analyses of the biological effects of variation in temperat~e on clam culture have been made, other than a few ecological studies (Mann & Glomb, 1977; Mill&n & Williams, 1985). Diet and temperature are probably two of the most important factors regulating the growth of clams in hatcheries and this paper presents data on their interactive effects for eight marine microalgae. Three commercially valuable clam species were studied and juveniles were chosen as it is this phase of the hatchery-rearing operation which requires the greater resources in terms of seawater, algal food, and other facilities and so is economically the most important. Organic weight-growth increments of whole juveniles were measured, and growth rates evaluated in terms of nutritional status of the different algae fed. Evidence from previous work with Ostrea edulis L. juveniles (Laing & Mill&n, 1986) suggested that consideration of growth efticiency (Newell, 1983) was necessary to gain a better underst~ding of the biological processes involved in the growth response. Not all of the energy from the algal food cells consumed will be available for growth. Only a proportion of the cells consumed is assimilated and of this assimilated energy a proportion is used in respiration. The effect that efficiency of utilization of the diet had on growth was assessed by comparing the proportion of assimilated energy used in growth and respiration for clams of the same organic weight fed different algae at various temperatures.
MATERIALS
ANDMETHODS
Juveniles of three species of clam, Tapes semidecussata~ T. dec~sata, and ~ercena~a mercenaria, were cultured in the hatchery (Walne, 1974). For the experiments, initial live weights of the animals were 0.26-3.08 mg (T. semidecussata), 0.17-1.25 mg (T. decussata), and 0.24-0.82 mg @I. mercenaria). Experimental treatments were carried out in from six to twelve 50-l recirculation systems (Laing & Millican, 1986) in 12 trials, Diets were added to the water of each system within which two upwelling units (Bayes, 198 1) were placed, each upweller containing 150-200 juveniles. Single-species algal diets tested were the diatoms Skeletonema costatum (Grev.) Cleve, Chaetoceros calcitrans (Paulsen) Takano, P~eodact~lum tricornutum Bohhn, and T~aIassiosjra~seudonana (Hustedt) Hastle et Heimdal(3H), and the flagellates Tetra-
GROWTH
OF JUVENILE
25
CLAMS
selmis suecica (Kylin) Butch., Isochrysis affgulbana Green (T. ISO), Chroomonas sulinu Butch.,
and
exponential
Chlumydomonus coccoides Butch. growth
phase
from cultures
grown
These
foods
in medium
were harvested prepared
in the
in autoclaved
seawater (Walne, 1970) in 3-l borosilicate-glass flat-bottomed boiling flasks. Cultures were continuously illuminated (7 mW . cm - ’ at the culture surface) at 2 1 2 1 “C and bubbled with 2.5 1. min - ’ of air enriched with 1 y0 carbon dioxide, filtered to 0.3 pm. All cultures were operated cultured.
except for C. calcitrans, which was batch-
semicontinuously
TABLE I Experimental treatments: diets fed to three clam species (S = T. semidecussafa, D = T. decussata, M = M. mercenaria) at a range of temperatures, each in a 50-l recirculating seawater system containing duplicate upwelling cylinders. Figures given are number of trials in which the treatment was tested. Algal food
T. IS0
S. costatum C. calcitrans
C. salina T. pseudonana
T. suecica P. tricomutum C. coccoides Unfed
Clam species
S D M S S D M S S D M S S D S S
Temperature
(‘C)
10
12
15
16
20
24
25
28
1
3 2 2
5
2 2 2
2 2 2
5
2 2 2
1
1
4 2 2 3 2 2 1
1
1
3 1
4 3
1 2 1 1 1 1 3 3
5 2
2 2
2 2 5 5
Table I gives the treatments and the number of trials in which each was tested. Each of the above algae was fed to T. semidecussatu at 25 “C and selected diets were fed to all three clam species at various temperatures between 10 and 28 “C. Temperatures were maintained within 1 “C of the nominal values. At > 20 ‘C, this was achieved in recirculating water baths in constant temperature rooms. At Q 15 “C, the 50-l recirculating systems were operated in through-flow water baths at the required temperature. Food was added daily to ensure a residual level of > 25 y0 of the ration fed after 24 h, subject to a minimum ration of 2.2 pg (organic weight) - ml- ’ to stimulate efficient filtration (Urban et al., 1983). Organic weight of the food and the equivalent minimum ration, as cells . ,ul - ‘, are given in Table II. Algal cell concentrations were estimated daily using a “Coulter” counter (Model ZB) immediately before and after
26
I. LAING ETAL. TABLE II
Organic weights of food species, and minimum feeding rations used, equivaient to 2.2 pg.mlexperimental systems. Algal food
T. IS0 S. costatum C. calcitrans C. salina T. pseudonana T. suecica P. tricom~t~m C. coccoides
Organic weight (pg.cell-‘) 20 32 7
130 20 200 20 80
* in 50-I
Minimum ration (cells *pcl-‘) 110 69 314 17 110 11 110 27.5
feeding. From these values the number of algal cells of each species cleared from suspension was calculated and from this and the organic weight of the algal cells (Table II) the weight of food cells cleared each day was determined. All trials were of 3-wk duration. The recirculation systems were emptied, cleaned and refilled twice each week with seawater at a salinity of 30-33x,, filtered to ~2.5pm particle diameter. At ‘f-day intervals, samples of about half the clams in each upwelling tube were taken for respiration-rate (oxygen-consumption), live-weight, dry-weight, and org~ic-wei~t determinations. At the end of the 3-wk experimental period, all the remaining clams were sampled. Thus, three duplicate samples of 35-100 juveniles were taken from each treatment during the course of each trial. Oxygen consumption was determined as the difference between initial and final dissolved oxygen content of filtered seawater in duplicate 500-ml stoppered bottles in which about half of the sample (i.e., 20-50 clams) was incubated for 4 h at the experimental temperature. The seawater used had previously been aerated for 24 h to ensure that it was saturated with oxygen under the prevailing conditions of temperature and barometric pressure. The oxygen tension of the enclosed water did not drop below 80% saturation in the 4-h expe~ment~ period. One bottle without animals acted as a control for each treatment. A dissolved oxygen electrode with integral stirrer and YSI Model 58 oxygen meter was used to determine oxygen content. Mean live weight was determined for each duplicate sample by counting all the juveniles in the sample, drying them on an absorbent cloth to remove surface water, and then weighing them together to within an error of 0.1 mg. The whole juveniles were dried for 48 h at 60 “C, then reweighed to give the mean dry weight. The organic weight was defmed as the AFDW. This was determined on each duplicate sample by grinding all of the whole dry juveniles together to a fine powder in a ball mill. Triplicate subsamples of 50-60 mg were weighed to within an error of 1 pg, ashed at 475 ‘C in a mume furnace and then reweighed. From the final (wk 3) samples,
21
GROWTHOF JUVENILECLAMS
further triplicate subsamples of 20-30 mg of weighed, dried, milled juveniles from the duplicate samples were prepared for analysis of total lipid. The lipid was extracted in chloroform : methanol (2 : 1, v/v) and charred using the method of Marsh & Weinstein (1966), then estimated by reading the absorbance at 375 pm against a range of cholesterol standards and blanks. Total lipid content of the algae was also estimated using this method. RESULTS GROWTH In all treatments, juvenile clams showed an exponential increase in organic weight with time. For example, organic weights of T. ~e~~ec~~~at~ fed T. ISO, C. calcjt~a~, and C. coccoides at 25 oC during a 3-wk trial are shown in Fig. 1. Linear regression lines were fitted in the form: In organic weight = kt + a, 1.2
7
(1)
0 Y=O.O416e
;
'.081
1.0
E _J
il 3 ; s 0 L 0
0.8
Y=0.0416e0~99X
0.6
0.4
3.2 Y=0.0416e0~'6X 0 0
----,I_ 0.5 1.0 Time
1.5
I 2.0
2.5
3.0
(weeks1
Fig. 1. Weekly organic weight growth of T. semidecussafu juveniles fed single-species diets of T. IS0 (O), C. cu~c~~~~~ (X), and C. camides ( + ) at 25 oC in duplicate upwelfmg cylinders in 50-l recirculating seawater systems.
where t = time (wk), a and k are constants. The constant k, which represents the slope of the line, may be considered as a coetTicient of growth rate for comparisons of different treatments. Higher values of k represent faster growth rates. In the example above, k values were 1,08,0.99, and 0.16 for T. ISO, C. calcitrans, and C. coccoides, respectively.
28
I. LAING ET AL.
EFFECT OF DIET
Mean growth-rate coefficients (k values) for the three clam species fed a range of single-species algal diets at 20 “C and for T. semidecussata at 25 “C are shown in Table III. The algae are ranked in order of nutritional value and in each column the growth coefficients for the algal diets are divided by the coefficient for T. ISO, one of the best foods. These calculated values show that the growth rate for each diet, relative to that with a diet of T. ISO, was much the same for all three species of clam, although actual growth rates were lower for T. decussata than for the other species. For Ustrea edulis, a different pattern emerged, with C. cal~itrans and C. salina giving greater growth than T. ISO. Overall, C. calcitrans was more effective in 0. edzdis than in any of the clams. TABLE III
Growth rate coefficient of three species of clams fed eight single-species algal diets at 20 “C. Results are also given for T. semidecussatu at 25 “C and for 0. eduh (Laing & Milliean, 1986) at 20 “C for comparison. The values in parentheses give the growth rates relative to those measured with T. IS0 for the diets. Results are the means of duplicate determinations for all trials in which the treatment was carried out (Table I). Algal food
_~_~ T. IS0 S. ro~t~t~
C. ~~~l~itrans C. T. T. P. C.
salina pseudonana suucica tricornutum coccoides
llnfed
Tapes semidecussata (25 “C)
1.105 1.105 (1.0) 0.990 (0.9) 0.990 (0.9) 0.967 (0.88) 0.898 (0.61) 0.553 (0.50) 0.184 (0.17) 0.175 (0.16)
Tapes semidecussatu 0.829 0.829(1.0)
0.737 (0.89) 0.714 (0.86) 0.668 (0.81) 0.392 (0.47) 0.322 (0.39) 0.222 (0.27)
Mercenaria mercenaria 0.944
0.806 (0.85) 0.714 (0.77)
Tapes decussaza
0.622 0.553 (0.89) 0.530 (0.85) 0.438 (0.70)
Ostrea edulis
0.875 0.852 (0.97) 1.082 (1.23) 0.990(1.13) 0.737 (0.84) 0.438 (0.50)
-
C. coccoides supported very poor growth in T. semidecussata juveniles. Organic weight-growth increments given by this flagellate were only slightly greater than those of unfed clams. EFFECT OF TEMPERATURE
Growth-rate coefficients of T. semidecussata, M. mercenaria, and T. decussata varied with temperature, as shown in Fig. 2, where data are given for diets of T. ISO, C. ccllcitrans and C. coccoides. With a diet of T. ISO, growth rates of T. semidecussata and M. mercenaria increased up to 25 ’ C, and then decreased above this temperature. A4. mercenaria grew slightly better than T. semidecussata below this optimum temperature and slower above it. Growth-rate coefficients of T. decussata were similar to those of the other two clams from 12 to 16 “C. At > 16 “C, the rate of increase was much lower, although it was rn~nt~n~ to 28 “C with a T. IS0 diet.
GROWTH OF JUVENILE CLAMS
10 Temp
15 ('Cl
0,
z
0 0
5
IQ Temp
is
20
25
30
?Cl
Fig. 2. Organic-weight growth-rate coefficients (k) at a range of temperatures for A: T. semidecussatn juveniles fed single-species diets ofT. IS0 (Q), C. culcihans (X), and C. coccoides ( + ); and B: T. semidecussata (Of, M. ~?ce~u~~ (a), and T. ~ec~~u#~ (A) juveniles fed T. ISO. Resufts are means of duplicate determinatians for atI trials in which the treatmentwas can-i& out (Table X).
For 7: semidecussata juveniles, the coefficient of the C. calcitmns diet was SS-90% of that for T. ISU at al1 temperatures. In Table III it can be seen that, with most ofthe other algal species fed to T. ~ern~de~u~s~tff, the growth-rate coefficients reIative to the coeffEents on the T. IS0 diet (figures in brackets) at.20 “C, were similar to those at 25 ’ C, Overall, coefficients at 20 ‘C were z 74 % of the 25 ’ C values for all algae except C. coccaidtq which supported less growth of T. se~~~~sa~ juveniles at 25 “C than at20OC.
I. LANG ETA..
30
ORGANIC RESERVES
The organic weight, as a percentage of the live weight, of T. semidecussata juveniles varied with temperature and diet (Table IV). Juveniles fed T. IS0 at 20 “C had a similar organic content to juveniles fed the same diet at 15 “C (t = 0.23, P > 0.1 for 36 df), but a significantly higher content than juveniles fed T. IS0 at 25 “C (t = 2.39, P < 0.05 for 43 df). Juveniles fed a very poor diet, e.g., C. coccoides, had a significantly lower proportion of organic weight to live weight than juveniles fed T. IS0 at 20 “C (t = 8.05, P < 0.001 for 30 df) and 25 “C (t = 7.35, P < 0.001 for 39 df).
TABLE IV Organic content, as a percentage of the live weight, of 1.2-3-mg-live-weight T. semidecussafa juveniles fed T. IS0 and C. coccoides at a range of temperatures. Results are means ( 2 SD) from weekly samples from all trials in which the treatment was tested (Table I). Temperature (‘C)
Algal food
._~
15
20
T. IS0
8.00 + 1.48
8.05 & 0.66
25 -..7.33 i_ 0.73
C. coccoides
5.58 k 1.72
5.73 * 1.73
5.55 i: 1.16
The effect of temperature and diet on the lipid content of T. semidecussata after 3 wk growth is given in Table V. The percentage lipid content of juveniles fed T. IS0 was similar over the temperature range tested, with the highest mean value at 20 “C. The percentage lipid content in juveniles fed with the diatoms C. calcitrans or S. costatum
TABLE
V
Lipid content (as percentage of AFDW) of 7’.semidecussata juveniles and a range of single-species algal diets fed at different temperatures. Results are means for the final (Wk 3) sample of all trials in which the treatment was tested (Table I). Initial lipid levels of juveniles were 4.3-5.8x AFDW. Algal food
Temperature (“C) .-___ -.
T. IS0 S. costatum C. calcitrans C. coccoides P. bicornutum C. salina T. suecica
Unfed
IO-12
15
20
25 ~_
10.13 7.41
20.6 13.3
9.60
9.91 8.21
17.3
9.18
8.87
12.5 12.0 21.3 4.7
6.40
5.24 -
4.07
4.02
4.14
9.72 7.12 7.71
3.19 7.41 9.50 9.44 3.41
GROWTH OF JUVENILE CLAMS
31
decreased as temperature increased. At 25 “C, the percentage of lipid content was greater in juveniles fed a flagellate diet than in juveniles fed a diatom diet. Percentage lipid content of the juveniles was not related to that in the alga fed, or to the growth rate of the animals. The percentage of lipid in juveniles fed C. coccoides and in unfed animals was less than with other diets and decreased with increasing temperature. At 20 and 25 “C lipid content decreased to less than the initial value with these treatments. RESPIRATION
AND GROWTH EFFICIENCY
Respiration, measured as oxygen-consumption diet as well as with temperature (Table VI).
rate, of T. semidecussata, varied with
TABLE VI Respiration rate (~1 oxygen consumed. mg clam- ’ 1h - ‘) of 0.15-0.9 -mg-organic-weight T. semidecussata juveniles fed a range of single-species algal diets at different temperatures. Results are means ( f SD) from weekly samples from all trials in which the treatment was tested (Table I). Temperature (“C)
Algal food
T. IS0 S. cosiafum C. c&nuns C. coccoides
I5
20
25
0.92 + 0.24 1.17 * 0.31
1.33 + 0.26 1.70 2 0.38 2.64 + 0.63
2.03 + 0.36 2.57 + 0.49
0.75 k 0.27
0.85 k 0.29
1.07 & 0.29
Higher temperatures were associated with higher respiration rates. At 20 “C, sizerelated oxygen-consumption rate for juveniles fed C. cafc~tra~ was si~i~c~tly greater than that for juveniles fed S. costatum, which in turn was significantly higher than the rate on the T. IS0 diet (F2,,6 = 42.37, P < 0.001). Respiration rates of juveniles fed C. coccoides were lower than those with the other diets. With diets of T. ISO, C. calcitrans, and S. costatum, algal cell-clearance rates (Fig. 3) showed a pattern similar to that for oxygen-consumption rate. Juveniles fed on diets that gave higher size-related respiration rates consumed a greater weight of algal food cells. Gross-gosh efficiency, K, = G. C- ‘, is the propo~ion of the cleared ration, C, incorporated into organic growth, G. Net-growth efficiency, KZ = G *(G + R) _ ‘, is the proportion of the assimilated ration used for growth, where R is the AFDW equivalent of the energy used in respiration, calculated as 1 mg . 1.2 ml 0, consumed - l (Walne, 1965). Growth efficiencies, K, and K2, for 0.3 mg organic weight T. semidecussata juveniles fed T. IS0 or S. costatum at 15-25 “C are shown in Table VII. The weekly growth increment, G, was calculated from the equation: G = 0.3ek - 0.3,
I. LAING ET AL.
---
0.2
0.4 aqmlc
Organic
I 0.8
0.6 wt.
of
wt.of
--I 1.0
1.2
,uveniles
(mg)
juveniles
(mg)
1.4
Fig. 3. The relationship between algal food cell-clearance rate (C, mg organic weight of food cells cleared. clam ’ wk- ‘) and organic weight of T. semidecussata juveniles fed A: T. IS0 at 25 “C ( q), 20 “C (0). and 15 “C (A); and B: T. IS0 (Q), C. calcitrans (X), and S. cost&urn (V) at 20 “C.
where k is the growth-rate coefficient (from Eqn. 1). Cleared ration values, C, for 0.3-mg-org~ic-weight juveniles were calculated from food cons~ption-clam size relationships (e.g., Fig. 3). Respiratory requirements, R, were calculated from datagiven in Table VI. In Table VII, it can be seen that growth of T. semidec~sa~a juveniles was similar with the two diets, but, at each temperature, cleared ration and respiratory values were higher with S. costatum leading to lower growth efficiencies than with T. ISO. Both diets were utilized most efficiently at 20 “C.
S. S. C C. C. C.
costatum costatum calcitrans calcitrans ~alc~~rans calcitrans
-r. iso S. costatum
T. IS0 T. IS0
15 20 2.5 15 20 25 20 20 20 20
(“Cl
Temperature
T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. decussata M. mercenaria 0. edulis
Bivalve species
0.211 0.395 0.638 0.211 0.395 0.638 0.330 0.220 0.370 0.580
G
0.056 0.091 0.182 0.070 0.116 0.224 0.161 0.403 0.191 0.102
R f R)-‘)
0.790 0.866 0.780 0.750 0.773 0.740 0.672 0.353 0.660 0.850
K,(G(G
0.68 1.30 2.00 0.90 2.18 4.98 5.00 5.50 4.80 4.20
c
0.31 0.30 0.32 0.23 0.18 0.13 0.07 0.04 0.08 0.14
K,(GC_‘)
of diet and temperature on the growth response of four species of juvenile bivalves of 0.3 mg initial organic weight. Weekly growth increment, G, respiration rate, R, and weight of food cells consumed, C, all expressed as mg AFDW. Data for 0. eduh from Laing & Millican (1986).
Diet
Effect
TABLE VII
34
I. LAING ET AL..
Growth efficiencies of T. semidecussuta, T. decussataa,M. mer~ena~a and 0. edulis, all fed C. calcitrans at 20 “C, are also given in Tabie VII. T. semidecussata and M. mercenaria were very similar in their growth response to this diet, giving similar size-specific values of G, R, C, K,, and K2. All four bivalves cleared similar amounts of food from suspension but the diet was used most efficiently by 0. eduh juveniles. They used less energy in respiration and grew more quickly than the three clams. The respiration rate of T. decussata was much higher than with the other two clam species on this diet. Discussion The relative food value of the algal species examined ranked the same irrespective of the clam species used. Although the growth data presented refer mainly to the two Tapes species, results obtained with M. mercenarib by other workers show a similar ranking of these algal diets as that shown here (Table III). Walne (1970) found very high growth rates of A4. mercenaria fed with S. costatum, low growth rates with P. tricornutum and very little growth with C. coccoides. Epifanio (1979) showed that T. suecica promoted moderate growth of M. mercenatia juveniles. Other molluscan species fed with these diets show similar relative growth rates (Peirson, 1983; Laing & Millican, 1986; Enright et al., 1986).
These authors have suggested that, although total lipid content of the diet does not correlate with food value, one of the more important factors dete~ining nutritional value of algae is their PUFA content. In particular, eicosapentaenoic acid, 20 : 5 ~3, and docosahexaenoic acid, 22: 6~3, are considered to be essential: algae that contain relatively high amounts of one or other generally support much better growth than species without either (Langdon & Waldock, 1981; Webb & Chu, 1983). Consequently, T. ISO, which has a high 22 : 6~3 content, and S. costatum, C. calcitrans, and T. pseudonana, which are rich in 20 : 5 03, were of good food value. T. suecica, which contains less 20 : 5 w3 than the diatoms, was of moderate food value (Table III). The high nutritional value of S. costatum was of interest and this species can be cultured successfully on a large scale, both in intensive indoor systems (Laing, 1985) and in outdoor tanks (De Pauw et al., 1983), making it a p~ic~~ly useful alga for commercial clam rearing. P. tri~om~~rn gave poor growth of T. semide~~ssata, even though this alga contains both 20 : 5 03 and 22 : 6 w3 fatty acids (Moreno et al., 1979). This is probably because P. tricornutum is indigestible to many bivalves (Epifanio, 1983). T. decussata performed moderately well with P. tricornutum, showing a higher growth-rate coefficient than T. semidecussata with this diet. C. coccoides was of very little food value. This species is known to lack essential PUFAs (Chuecas & Riley, 1969) but may also be of limited food value due to indigestibility of the cell wall (Webb & Chu, 1983) or a toxicity factor (Taub & Dollar, 1968). The small growth increment observed with this alga, and with unfed animals, may reflect uptake of other organic matter in the seawater (Walne, 1970; Manahan et a/., 1982).
GROWTH
When diets of good to moderate the effect of an increase in temperature size-related
algal cell-clearance
OF JUVENILE
CLAMS
35
food value were fed to T. semidecussata juveniles, from 15 to 25 ‘C was to give an increase in clam
rate, with the result that more energy was available
and
the animals grew faster. Newell (1980) suggested four potential mechanisms by which bivalves may respond to an increase in environmental temperature such as this. These are: (1) an increase in food consumption, C, to offset metabolic losses, R; (2) an increase in absorption efficiency, and therefore the amount of assimilated energy, A, available; (3) suppression of energy losses through respiration, R; (4) a combination of I, 2, and 3. Here, T. semidecussata juveniles are shown to have followed Strategy 1. There was no evidence of increased absorption efficiency (Strategy 2) or suppression of energy losses (Strategy 3) as temperature increased. K, remained constant or decreased slightly with increasing temperature and oxygen consumption continued to increase curvilinearly from 15 to 25 “C (Table VII). The increased energy expenditure in respiration as temperature increased used a greater proportion of the assimilated energy at 25 “C than at 20 “C, shown by lower K2 values (Table VII). This was associated with a lower rate of organic growth in relation to shell deposition at higher temperatures (Table IV), an effect also noticed in T. semidecussata and M. mercenaria juveniles grown outdoors (Mann & Glomb, 1977; Millican & Williams, 1985). T. decussata appears to have a lower optimum temperature for growth than :r. semidecussata and M. mercenaria (Fig. 2). At 20 ‘C, lower growth rates of T. decussata juveniles were associated with higher oxygen-consumption rates, compared with the two other clam species (Table VII). When food was absent or the cells were of low nutritional value, e.g., C. coccoides, the clams suppressed their oxygen consumption resulting in the conservation of energy and the maintenance of some growth. At 20 “C, the oxygen consumption of l-mg-organic-weight animals was 0.85 ~1. h- ’ with a diet of C. coccoides, compared with 1.33 ~1. h - ’ with T. IS0 (Table VI). This corresponds to Strategy 3 of Newell (1980). At 25 “C, oxygen consumption of clams fed C. coccoides increased to 1.07 ~1. h _ ’ and with a limited amount of food energy available the increased maintenance requirement reduced the capacity for organic growth (Fig. 2A). At > 25 “C, growth of M. mercenaria and T. semidecussata juveniles decreased (Fig. 2). The effect of high temperature on the growth response of bivalves is discussed by Winter (1978). A decline in growth rate is usually associated with a marked decrease in filtration rate at these temperatures. All of the alga species tested are viable in the temperature range of lo-25 “C. At > 25 “C, lower growth rates of clams may be associated with temperature stress of the algal food cells, which tend to settle out of suspension (Ukeles, 1961). At all temperatures, the proportion of the cleared algal ration utilized in growth was low, shown by low observed value of K, in Table VII. Newell (1980) suggests that this is due to an “exploitative strategy”, whereby in conditions of abundant food resources, as in these trials, the clams maintain a maximum energetic gain from the environment by processing the food cells at a high rate, resulting in a high respiratory energy expenditure.
36
I. LAING
ET AL.
Growth efficiency of clams was modified by diet as well as temperature. This was most clearly illustrated by the comparison of the growth response of 0.3-mg-organicweight T, semidec~ssuta juveniles to single-species diets of T. ISO, S. ~~~t~tu~, and C. caZcitrm.~ at 20 “C (Table VII). Respiration utilized proportionally more of the assimilated energy when T. semidecussata juveniles were fed the diatoms S. costatum and C. calcitrans compared with the flagellate T. ISO. Even with this higher respiration rate, juveniles fed S. cost&urn were still able to grow at the same rate as those fed T. IS0 because the lower efficiency of utilization of the diet was compensated for by an increased ration. Although a higher ration of C. calcitrans cells was cleared than of S. costatum, similar amounts of each were assimilated (G + R = 0.5 mg AFDW at 20 ‘C). About 20% more of this assimilated energy was used in respiration, and organic weight-growth increment was 16% lower with a diet of C. caZcitransthan with S. costatum. Utilization efficiency of a diet of C. calcitrans was much higher by 0. edulis juveniles than by T. semidecussatu juveniles. More of the cleared ration was available for growth and respiration, and the proportion of this assimilated energy used in respiration was lower (Table VII). The energetic responses of the clams to diet and temperature affected the lipid reserves of the animals. The higher apparent energy requirement for the utiIization of diatom diets was associated with a smaller proportion of lipid in the organic reserves, particularly at higher temperatures, compared with juveniles fed flagellate diets of moderate to good food value. Mill&n & Williams (1985) found that M. mercenaria and T. semidecussata reared in a lagoon fertilized to encourage blooms of diatom species contained proportion~ly less lipid than animals in the sea. Unfed clams and those fed C. coccoides showed a loss in percentage lipid content at 20 “C and especially at 25 “C when the animals were presumably utilizing reserves to meet increased metabolic requirements. For the commercial culture of these clam species, a balance needs to be maintained between growth rate, growth efficiency, and juvenile quality, in terms of organic and lipid reserves, for any diet-temperature combination. The results in this paper show that certain algal diets promoted good growth in all three clam species, with T. IS0 and S. costatum being especially suitable. C. calcitrans, C. salina, T. pseudonana, and T. suecica are also useful diet species for clam rearing. Although maximum organic growth of T. semidec~sata and M. mercenarib occurred at 2.5 ‘C, a culture temperature of 20 OC may be more appropriate, as this produces animals of a higher quality which utilize the diet fed more effkiently. However, as the clams are growing more slowly at 20 “C than at 25 ‘C, culture facilities are tied up for a greater time. Most efficient growth of T. decussata juveniles occurs at 15-20 “C. The nature of the growth response described gives some insight into the metabolic requ~ements and strategies of these clams for maint~ning m~imum growth with variation in food supply and temperature in their natural environment.
GROWTH OF JUVENILE CLAMS
37
REFERENCES BAYES, J.C., 1981. Forced upwelling nurseries for oysters and clams using im~unded
water systems. Proceedings of the International Workshop on Nursery Culturing of Bivalve Molluscs at Ghent, Belgium. Spec. Publ. Eur. Maricult. Sot., Vol. 7, pp. 73-83. BREBER,P., 1985. On-growing of the carpet-shell clam (T. decussata L.): two years experience in Venice Lagoon. Aquaculture, Vol. 44, pp. 51-56. BURRELL,JR., V.C., 1983. Molluscan aquaculture in the United States: a brief overview. J. lY[d. Mancult. Sot., Vol. 14, pp. 164-169. CHUECZAS, I. & J. P. RILEY, 1969. Component fatty acids of the total lipids of some marine phytoplankton. J. Mar. Biol. Assoc. U.K., Vol. 49, pp. 97-l 16. CLAUS, C., 1981. Trends in nursery rearing ofbivalve molluscs. Proceedings ofthe International Workshop on Nursery Culturing of Bivalve Mol1uscs at Ghent, Belgium. Spec. Publ. Eur. Muricult. SIX., Vol. 7, pp. 21-33. DE PAUW, N., J. VERBOVEN& C. CLAUS, 1983. Large-scale microalgae production for nursery rearing of marine bivalves. Aquacult. Eng., Vol. 2, pp. 27-47. ENRIGHT,CT., C. F. NEWKIRK,J. S. CRAIGIE & J.D. CASTELL,1986. Evaluation of ph~oplankton as diets for juvenile Ostrea edulis L. J. Eyr. Mar. BZoZ.E&of., Vol. 96, pp. l-13. EPIFANIO,C. E., 1979. Growth in bivalve molluscs: nutritional effects oftwo or more species of algae in diets fed to the American oyster, Crussostrea virgimca (Gmelin) and the hard clam, Mercenaria mercenariu (L.). Aquaculture, Vol. 18, pp. l-12. EPIFANIO,C. E., 1983. Phytoplankton and yeast as foods for juvenile bivalves: a review of research at the University of Delaware. in, Proceed&gs of the Second international Conference on Aquaculture Nutrition. Biochem~ea~ and physiolo~~al Qpprouches to shel~~h nut~tion, edited by G.D. Pruder, C. Langdon & D. Conklin, Louisiana State University, Baton Rouge, Louisiana, pp. 292-304. LAING, I.,1985. Factors affecting the large-scale production of four species of commercially important marine algae Aquaculture, Vol. 44, pp. 161-166. LAING, I.& P.F. MILLICAN,1986. Relative growth and growth efficiency of Ostrea edulis L. spat fed various algal diets. Aquaculture, Vol. 54, pp. 245-262. LANGDON,C. J., 1983. New techniques and their application to studies of bivalve nutrition. In, Proceedings
of the Second
Interna~~onaIConference on Aquiculture Nutn.tion. Biochemical and phys~o~o~.~~~ ~pproa&hes to shel~~shnutrition, edited by G. D. Pruder, C. Langdon & D. Conklin, Louisiana State University, Baton
Rouge, Louisiana, pp. 305-320. LANGDON, C.J.& M. J.WALDOCK, 1981. The effect of algal and artificial diets on the growth and fatty acid composition of Crassostrea gigus spat. J. Mar. Biol. Ass. U.K., Vol. 61, pp. 431-448. LOOSANOFF,V. 1.. & H.C. DAVIS, 1963. Rearing of bivalve molluscs. In, Advances in marine biology, Vol. 1,
edited by F. S. Russell, Academic Press, London, pp. 1-136. LVCAS, A., 1977. Carpet shell farming: tradition and new means. Peche Murk, Vol. 1193, pp. 475-478. MANAHAN,D.T., S. H. WRIGHT,G.C. STEPHENS& M.A. RICE, 1982. Transport ofdissolved amino acids by the mussel, Mytilus eduhs: demonstration of net uptake from natural sea water. Science, Vol. 215, pp. 1253-1255. MANN, R. & S. GLOMB, 1977. The effect of temperature on growth and ammonia excretion of the Manila clam, Tapes japonica. Estuarine Coastal Mar. Sci., Vol. 6, pp. 335-339. MARSH,J. B. & D. B. WEINSTEIN,1966. Notes on methodology- simple charring method for determination of lipids. J. Lipid Ref., Vol. 7, pp, 574-576. MCHUGH, J.L., 1981. Recent advances in hard clam mariculture. f. She@ Res., Vol. 1, pp. 51-55. MILKAN, P. F. & D. R. WILLIAMS, 1985.The seasonal variations in the levels of meat content, lipid and carbohydrate in Mercenaria mercenaria L. and Tapes semidecussata Reeve grown in fertilised and unfertilised water. ICES CM. 1985/K:51, 9 pp. MORENO,V. J., J. E. A. DE MORENO& R. R. BRENNER,1979. Biosynthesis of unsaturated fatty acids in the diatom Phaeodactylum tricornutum. Lipids, Vol. 14, pp. 15-19. NEWEI.L, R.C., 1980. The maintenance of energy balance in marine invertebrates exposed to changes in environmental temperature. in, Animals and en~ironmen?uZ~fness, Vol. I, edited by R. Gilles, Pergamon Press, Oxford, pp. 561-582. NEWELL,R. I. E., 1983. Molluscan bioenergetics - a synopsis. In, Proceedings of rhe Second hternafiod Conference on Aquaculture Nutrition. Biochemical and physiological approaches to sheZ!hsh nutrition, edited
38
I. LAING ETAL.
by G.D. Pruder, C. Langdon & D. Conklin, Louisiana State University, Baton Rouge, Louisiana, pp. 252-27 1. PEIRSON,W. M., 1983. Utilisation of eight algal species by the bay scallop Argapecten irradiuns concentricus (Say.). J. Exp. Mar. Biol. Ecol., Vol. 68, pp. l-11. RODRIGUEZ,A., 1983. Culture of Venerupts decussatu L. in the intertidal zone. Proceedings of First Seminar on Marine Sciences, Vigo. Seminar& de Estudos Galegas a Coruna, Spain, pp. 527-540. ROMBERGER,H.P. & C.E. EPIFANIO, 1981. Comparative effects of diets consisting of one or two algal species upon assimilation efftciencies and growth of juvenile oysters, Crassostrea virgimca (Gmelin). Aquaculture, Vol. 25, pp. 11-81. TAUB, F. B. & A.M. DOLLAR, 1968. The nutritional inadequacy of Chlorella and Chlamydomonas SPQ.as food for Daphnia puiex. Limnol. Oceanogr., Vol. 13, pp. 607-617. TENORE,K. R. & W. M. DUNSTAN,1973. Comparison of rates of feeding and biodeposition of the American oyster, Crassostrea vir~n~ca Gmefin, fed different species of ph~oplankton. J. Exp. Mar. Biol. Ecol., vol. 12, QQ.19-26. UKELES, R., 1961. The effect of temperature on the growth and survival of several marine algae species. Biol. Bull., Vol. 120, pp. 255-264. URBAN,E. R., C. D. PRUNER& C. J. LANGDON,1983. Effect of ration on growth and growth efficiency of juveniles of Crassostrea virgtnica (Gmelin). J. .She@ Res., Vol. 3, pp. 5 l-57. WALDOCK,M.J. & D.L. HOLLAND, 1984. Fatty acid metabolism in young oysters, Crassostrea gigus: polyunsaturated fatty acids. Lipids, Vol. 19, QQ.332-336. WALNE, P.R., 1965. Observations on the influence of food supply and temperature on the feeding and growth of the larvae of Ostrea edulti I.. Fish. Invest. (London) Ser. 2, Vol. 24, pp. l-45. WALNE,P.R., 1970. Studies on the food value of nineteen genera of algae to juvenile bivalves of the genera Ostrea, Crassostreu, Mercenaria and Mytilus. Fish Invest. (London) Ser. 2, Vol. 26, pp. l-62. WALNE,P.R., 1974. Culture of bivalve molluscs. 50 years experience at Conwy. Fishing News (Books), West Byfleet, 173 QQ. WEBB, K. L. & F. L. CHU, 1983. Ph~oplankto~ as a food source for bivalve larvae. In, Proceedings of the Second International Conference on Aquaculture Nutrition. 3iochem~a~ and physioio~.cai approaches to .~he~~f~hnutrjr~on, edited by G. D. Pruder, C. Langdon & D. Conklin, Louisiana State University, Baton
Rouge, Louisiana, pp. 272-291. WINTER,J. E., 1978. A review ofknowledge of suspension feeding in Iamellibranchiate bivalves, with special reference to artificial aquaculture systems. Aquacuhure, Vol. 13, pp. l-33.
23
JEM 00959
Interactive effect of diet and temperature on the growth of juvenile clams I. L&g,
S. D. Utting and R. W. S. Kilada
Fisheries ~aboruto~, Conwy, Gwynedd, U.K., Directorate of Fisheries Research, Ministry ofAgr<ure, Fisheries and Food
(Received 6 April 1987; revision received 3 July 1987; accepted 14 July 1987) Abstract: Small juveniles (0.3-3 mg live weight) of three clam species, Tapes semidecussata Reeve, T. decussata L., and Mercenariu mercenarfa L., were fed eight single-species algal diets at a range of temperatures. Nutritional value of the algae tested was in the order: Isochrysis aff gulbana (T. ISO) = Skeletonema costatum > Chaetoceros calciirans = Chroomonas salina = Thalassiosira pseudonana > Tetraselmis suecica z Phaeodactylum tricornutum > Chlamydomonas coccoides, when the same weight of these species was given
in the diet to each clam species, Reasons for differences in nutritional value are discussed with reference to the biochemical content of the algae and the relative growth efficiency of the animals. Respiration rate, food cell clearance rate, and growth (as increase in organic weight) of M. mercenariu and T. semidecussatu increased with temperature from 10 to 25 “C. Growth rates decreased at > 25 “C. T. decussata showed only a slightly increased growth response at > 15 “C. Key words: Algal diet; Temperature; Clam; Growth eflkiency
INTRODUCTION
The commercial hatchery culture of bivalve mollusc’s requires a supply of live algal (phytoplankton) food cells of high nutritional value. The food value of a wide range of algae has been assessed for the culture of oysters (e.g., Walne, 1970; Tenore & Dunstan, 1973; Enright et al., 1986). A few selected species are now used routinely, often in mixtures, to provide a diet on which the animals will grow efficiently (Epifanio, 1979; Romberger & Epifanio, 1981; Laing & Millican, 1986). Recent work has focussed on the ident~~ation of particular components of the diet that are essential for healthy growth (Langdon, 1983) and the importance of polyunsaturated fatty acids (PUFA) has been reported (Waldock & Holland, 1984). There is an increasing interest in the hatchery rearing of the commercially valuable clam species, Tapes semidecussata Reeve and Mercenaria mercenaria L., for which there are markets in Asia, North and South America, and Europe (Lucas, 1977; Claus, 198 1; McHugh, 1981; Burrell, 1983), and also T. decussatu L., for which there is demand in Europe (Rodriguez, 1983; Breber, 1985). Depletion of natural and introduced stocks Correspondence address: 1. Laing, Fisheries Laboratory, Directorate of Fisheries Research, Ministry of Agriculture, Fisheries and Food, Benarth Road, Conwy, Gwynedd LL3.2 8UB, U.K. 0022-098 1/87i$O3.50 0 1987 Elsevier Science Publishers B.V. (Biomedical Division)
24
I. LAING &‘-AL.
of these cfams has brought about an increased demand for hatchery-produced young (seed). They are often cultured on algal diets designed for oyster culture, but there is very little evidence to support the assumption that these diets are suitable for clams. Some reports, however, indicate dflerences in nutritional value between certain algal species used to feed clams and oysters (Loosanoff & Davis, 1963; Walne, 1970; Epifanio, 1979) although comparisons can be unreliable as foods were not from the same growth phase (Webb & Chu, 1983). Any differences may play a significant part in juvenile growth performance. Physical aspects of the hatchery environment for clams, especially temperature, have also been adopted from oyster culture. Again, few detailed analyses of the biological effects of variation in temperat~e on clam culture have been made, other than a few ecological studies (Mann & Glomb, 1977; Mill&n & Williams, 1985). Diet and temperature are probably two of the most important factors regulating the growth of clams in hatcheries and this paper presents data on their interactive effects for eight marine microalgae. Three commercially valuable clam species were studied and juveniles were chosen as it is this phase of the hatchery-rearing operation which requires the greater resources in terms of seawater, algal food, and other facilities and so is economically the most important. Organic weight-growth increments of whole juveniles were measured, and growth rates evaluated in terms of nutritional status of the different algae fed. Evidence from previous work with Ostrea edulis L. juveniles (Laing & Mill&n, 1986) suggested that consideration of growth efticiency (Newell, 1983) was necessary to gain a better underst~ding of the biological processes involved in the growth response. Not all of the energy from the algal food cells consumed will be available for growth. Only a proportion of the cells consumed is assimilated and of this assimilated energy a proportion is used in respiration. The effect that efficiency of utilization of the diet had on growth was assessed by comparing the proportion of assimilated energy used in growth and respiration for clams of the same organic weight fed different algae at various temperatures.
MATERIALS
ANDMETHODS
Juveniles of three species of clam, Tapes semidecussata~ T. dec~sata, and ~ercena~a mercenaria, were cultured in the hatchery (Walne, 1974). For the experiments, initial live weights of the animals were 0.26-3.08 mg (T. semidecussata), 0.17-1.25 mg (T. decussata), and 0.24-0.82 mg @I. mercenaria). Experimental treatments were carried out in from six to twelve 50-l recirculation systems (Laing & Millican, 1986) in 12 trials, Diets were added to the water of each system within which two upwelling units (Bayes, 198 1) were placed, each upweller containing 150-200 juveniles. Single-species algal diets tested were the diatoms Skeletonema costatum (Grev.) Cleve, Chaetoceros calcitrans (Paulsen) Takano, P~eodact~lum tricornutum Bohhn, and T~aIassiosjra~seudonana (Hustedt) Hastle et Heimdal(3H), and the flagellates Tetra-
GROWTH
OF JUVENILE
25
CLAMS
selmis suecica (Kylin) Butch., Isochrysis affgulbana Green (T. ISO), Chroomonas sulinu Butch.,
and
exponential
Chlumydomonus coccoides Butch. growth
phase
from cultures
grown
These
foods
in medium
were harvested prepared
in the
in autoclaved
seawater (Walne, 1970) in 3-l borosilicate-glass flat-bottomed boiling flasks. Cultures were continuously illuminated (7 mW . cm - ’ at the culture surface) at 2 1 2 1 “C and bubbled with 2.5 1. min - ’ of air enriched with 1 y0 carbon dioxide, filtered to 0.3 pm. All cultures were operated cultured.
except for C. calcitrans, which was batch-
semicontinuously
TABLE I Experimental treatments: diets fed to three clam species (S = T. semidecussafa, D = T. decussata, M = M. mercenaria) at a range of temperatures, each in a 50-l recirculating seawater system containing duplicate upwelling cylinders. Figures given are number of trials in which the treatment was tested. Algal food
T. IS0
S. costatum C. calcitrans
C. salina T. pseudonana
T. suecica P. tricomutum C. coccoides Unfed
Clam species
S D M S S D M S S D M S S D S S
Temperature
(‘C)
10
12
15
16
20
24
25
28
1
3 2 2
5
2 2 2
2 2 2
5
2 2 2
1
1
4 2 2 3 2 2 1
1
1
3 1
4 3
1 2 1 1 1 1 3 3
5 2
2 2
2 2 5 5
Table I gives the treatments and the number of trials in which each was tested. Each of the above algae was fed to T. semidecussatu at 25 “C and selected diets were fed to all three clam species at various temperatures between 10 and 28 “C. Temperatures were maintained within 1 “C of the nominal values. At > 20 ‘C, this was achieved in recirculating water baths in constant temperature rooms. At Q 15 “C, the 50-l recirculating systems were operated in through-flow water baths at the required temperature. Food was added daily to ensure a residual level of > 25 y0 of the ration fed after 24 h, subject to a minimum ration of 2.2 pg (organic weight) - ml- ’ to stimulate efficient filtration (Urban et al., 1983). Organic weight of the food and the equivalent minimum ration, as cells . ,ul - ‘, are given in Table II. Algal cell concentrations were estimated daily using a “Coulter” counter (Model ZB) immediately before and after
26
I. LAING ETAL. TABLE II
Organic weights of food species, and minimum feeding rations used, equivaient to 2.2 pg.mlexperimental systems. Algal food
T. IS0 S. costatum C. calcitrans C. salina T. pseudonana T. suecica P. tricom~t~m C. coccoides
Organic weight (pg.cell-‘) 20 32 7
130 20 200 20 80
* in 50-I
Minimum ration (cells *pcl-‘) 110 69 314 17 110 11 110 27.5
feeding. From these values the number of algal cells of each species cleared from suspension was calculated and from this and the organic weight of the algal cells (Table II) the weight of food cells cleared each day was determined. All trials were of 3-wk duration. The recirculation systems were emptied, cleaned and refilled twice each week with seawater at a salinity of 30-33x,, filtered to ~2.5pm particle diameter. At ‘f-day intervals, samples of about half the clams in each upwelling tube were taken for respiration-rate (oxygen-consumption), live-weight, dry-weight, and org~ic-wei~t determinations. At the end of the 3-wk experimental period, all the remaining clams were sampled. Thus, three duplicate samples of 35-100 juveniles were taken from each treatment during the course of each trial. Oxygen consumption was determined as the difference between initial and final dissolved oxygen content of filtered seawater in duplicate 500-ml stoppered bottles in which about half of the sample (i.e., 20-50 clams) was incubated for 4 h at the experimental temperature. The seawater used had previously been aerated for 24 h to ensure that it was saturated with oxygen under the prevailing conditions of temperature and barometric pressure. The oxygen tension of the enclosed water did not drop below 80% saturation in the 4-h expe~ment~ period. One bottle without animals acted as a control for each treatment. A dissolved oxygen electrode with integral stirrer and YSI Model 58 oxygen meter was used to determine oxygen content. Mean live weight was determined for each duplicate sample by counting all the juveniles in the sample, drying them on an absorbent cloth to remove surface water, and then weighing them together to within an error of 0.1 mg. The whole juveniles were dried for 48 h at 60 “C, then reweighed to give the mean dry weight. The organic weight was defmed as the AFDW. This was determined on each duplicate sample by grinding all of the whole dry juveniles together to a fine powder in a ball mill. Triplicate subsamples of 50-60 mg were weighed to within an error of 1 pg, ashed at 475 ‘C in a mume furnace and then reweighed. From the final (wk 3) samples,
21
GROWTHOF JUVENILECLAMS
further triplicate subsamples of 20-30 mg of weighed, dried, milled juveniles from the duplicate samples were prepared for analysis of total lipid. The lipid was extracted in chloroform : methanol (2 : 1, v/v) and charred using the method of Marsh & Weinstein (1966), then estimated by reading the absorbance at 375 pm against a range of cholesterol standards and blanks. Total lipid content of the algae was also estimated using this method. RESULTS GROWTH In all treatments, juvenile clams showed an exponential increase in organic weight with time. For example, organic weights of T. ~e~~ec~~~at~ fed T. ISO, C. calcjt~a~, and C. coccoides at 25 oC during a 3-wk trial are shown in Fig. 1. Linear regression lines were fitted in the form: In organic weight = kt + a, 1.2
7
(1)
0 Y=O.O416e
;
'.081
1.0
E _J
il 3 ; s 0 L 0
0.8
Y=0.0416e0~99X
0.6
0.4
3.2 Y=0.0416e0~'6X 0 0
----,I_ 0.5 1.0 Time
1.5
I 2.0
2.5
3.0
(weeks1
Fig. 1. Weekly organic weight growth of T. semidecussafu juveniles fed single-species diets of T. IS0 (O), C. cu~c~~~~~ (X), and C. camides ( + ) at 25 oC in duplicate upwelfmg cylinders in 50-l recirculating seawater systems.
where t = time (wk), a and k are constants. The constant k, which represents the slope of the line, may be considered as a coetTicient of growth rate for comparisons of different treatments. Higher values of k represent faster growth rates. In the example above, k values were 1,08,0.99, and 0.16 for T. ISO, C. calcitrans, and C. coccoides, respectively.
28
I. LAING ET AL.
EFFECT OF DIET
Mean growth-rate coefficients (k values) for the three clam species fed a range of single-species algal diets at 20 “C and for T. semidecussata at 25 “C are shown in Table III. The algae are ranked in order of nutritional value and in each column the growth coefficients for the algal diets are divided by the coefficient for T. ISO, one of the best foods. These calculated values show that the growth rate for each diet, relative to that with a diet of T. ISO, was much the same for all three species of clam, although actual growth rates were lower for T. decussata than for the other species. For Ustrea edulis, a different pattern emerged, with C. cal~itrans and C. salina giving greater growth than T. ISO. Overall, C. calcitrans was more effective in 0. edzdis than in any of the clams. TABLE III
Growth rate coefficient of three species of clams fed eight single-species algal diets at 20 “C. Results are also given for T. semidecussatu at 25 “C and for 0. eduh (Laing & Milliean, 1986) at 20 “C for comparison. The values in parentheses give the growth rates relative to those measured with T. IS0 for the diets. Results are the means of duplicate determinations for all trials in which the treatment was carried out (Table I). Algal food
_~_~ T. IS0 S. ro~t~t~
C. ~~~l~itrans C. T. T. P. C.
salina pseudonana suucica tricornutum coccoides
llnfed
Tapes semidecussata (25 “C)
1.105 1.105 (1.0) 0.990 (0.9) 0.990 (0.9) 0.967 (0.88) 0.898 (0.61) 0.553 (0.50) 0.184 (0.17) 0.175 (0.16)
Tapes semidecussatu 0.829 0.829(1.0)
0.737 (0.89) 0.714 (0.86) 0.668 (0.81) 0.392 (0.47) 0.322 (0.39) 0.222 (0.27)
Mercenaria mercenaria 0.944
0.806 (0.85) 0.714 (0.77)
Tapes decussaza
0.622 0.553 (0.89) 0.530 (0.85) 0.438 (0.70)
Ostrea edulis
0.875 0.852 (0.97) 1.082 (1.23) 0.990(1.13) 0.737 (0.84) 0.438 (0.50)
-
C. coccoides supported very poor growth in T. semidecussata juveniles. Organic weight-growth increments given by this flagellate were only slightly greater than those of unfed clams. EFFECT OF TEMPERATURE
Growth-rate coefficients of T. semidecussata, M. mercenaria, and T. decussata varied with temperature, as shown in Fig. 2, where data are given for diets of T. ISO, C. ccllcitrans and C. coccoides. With a diet of T. ISO, growth rates of T. semidecussata and M. mercenaria increased up to 25 ’ C, and then decreased above this temperature. A4. mercenaria grew slightly better than T. semidecussata below this optimum temperature and slower above it. Growth-rate coefficients of T. decussata were similar to those of the other two clams from 12 to 16 “C. At > 16 “C, the rate of increase was much lower, although it was rn~nt~n~ to 28 “C with a T. IS0 diet.
GROWTH OF JUVENILE CLAMS
10 Temp
15 ('Cl
0,
z
0 0
5
IQ Temp
is
20
25
30
?Cl
Fig. 2. Organic-weight growth-rate coefficients (k) at a range of temperatures for A: T. semidecussatn juveniles fed single-species diets ofT. IS0 (Q), C. culcihans (X), and C. coccoides ( + ); and B: T. semidecussata (Of, M. ~?ce~u~~ (a), and T. ~ec~~u#~ (A) juveniles fed T. ISO. Resufts are means of duplicate determinatians for atI trials in which the treatmentwas can-i& out (Table X).
For 7: semidecussata juveniles, the coefficient of the C. calcitmns diet was SS-90% of that for T. ISU at al1 temperatures. In Table III it can be seen that, with most ofthe other algal species fed to T. ~ern~de~u~s~tff, the growth-rate coefficients reIative to the coeffEents on the T. IS0 diet (figures in brackets) at.20 “C, were similar to those at 25 ’ C, Overall, coefficients at 20 ‘C were z 74 % of the 25 ’ C values for all algae except C. coccaidtq which supported less growth of T. se~~~~sa~ juveniles at 25 “C than at20OC.
I. LANG ETA..
30
ORGANIC RESERVES
The organic weight, as a percentage of the live weight, of T. semidecussata juveniles varied with temperature and diet (Table IV). Juveniles fed T. IS0 at 20 “C had a similar organic content to juveniles fed the same diet at 15 “C (t = 0.23, P > 0.1 for 36 df), but a significantly higher content than juveniles fed T. IS0 at 25 “C (t = 2.39, P < 0.05 for 43 df). Juveniles fed a very poor diet, e.g., C. coccoides, had a significantly lower proportion of organic weight to live weight than juveniles fed T. IS0 at 20 “C (t = 8.05, P < 0.001 for 30 df) and 25 “C (t = 7.35, P < 0.001 for 39 df).
TABLE IV Organic content, as a percentage of the live weight, of 1.2-3-mg-live-weight T. semidecussafa juveniles fed T. IS0 and C. coccoides at a range of temperatures. Results are means ( 2 SD) from weekly samples from all trials in which the treatment was tested (Table I). Temperature (‘C)
Algal food
._~
15
20
T. IS0
8.00 + 1.48
8.05 & 0.66
25 -..7.33 i_ 0.73
C. coccoides
5.58 k 1.72
5.73 * 1.73
5.55 i: 1.16
The effect of temperature and diet on the lipid content of T. semidecussata after 3 wk growth is given in Table V. The percentage lipid content of juveniles fed T. IS0 was similar over the temperature range tested, with the highest mean value at 20 “C. The percentage lipid content in juveniles fed with the diatoms C. calcitrans or S. costatum
TABLE
V
Lipid content (as percentage of AFDW) of 7’.semidecussata juveniles and a range of single-species algal diets fed at different temperatures. Results are means for the final (Wk 3) sample of all trials in which the treatment was tested (Table I). Initial lipid levels of juveniles were 4.3-5.8x AFDW. Algal food
Temperature (“C) .-___ -.
T. IS0 S. costatum C. calcitrans C. coccoides P. bicornutum C. salina T. suecica
Unfed
IO-12
15
20
25 ~_
10.13 7.41
20.6 13.3
9.60
9.91 8.21
17.3
9.18
8.87
12.5 12.0 21.3 4.7
6.40
5.24 -
4.07
4.02
4.14
9.72 7.12 7.71
3.19 7.41 9.50 9.44 3.41
GROWTH OF JUVENILE CLAMS
31
decreased as temperature increased. At 25 “C, the percentage of lipid content was greater in juveniles fed a flagellate diet than in juveniles fed a diatom diet. Percentage lipid content of the juveniles was not related to that in the alga fed, or to the growth rate of the animals. The percentage of lipid in juveniles fed C. coccoides and in unfed animals was less than with other diets and decreased with increasing temperature. At 20 and 25 “C lipid content decreased to less than the initial value with these treatments. RESPIRATION
AND GROWTH EFFICIENCY
Respiration, measured as oxygen-consumption diet as well as with temperature (Table VI).
rate, of T. semidecussata, varied with
TABLE VI Respiration rate (~1 oxygen consumed. mg clam- ’ 1h - ‘) of 0.15-0.9 -mg-organic-weight T. semidecussata juveniles fed a range of single-species algal diets at different temperatures. Results are means ( f SD) from weekly samples from all trials in which the treatment was tested (Table I). Temperature (“C)
Algal food
T. IS0 S. cosiafum C. c&nuns C. coccoides
I5
20
25
0.92 + 0.24 1.17 * 0.31
1.33 + 0.26 1.70 2 0.38 2.64 + 0.63
2.03 + 0.36 2.57 + 0.49
0.75 k 0.27
0.85 k 0.29
1.07 & 0.29
Higher temperatures were associated with higher respiration rates. At 20 “C, sizerelated oxygen-consumption rate for juveniles fed C. cafc~tra~ was si~i~c~tly greater than that for juveniles fed S. costatum, which in turn was significantly higher than the rate on the T. IS0 diet (F2,,6 = 42.37, P < 0.001). Respiration rates of juveniles fed C. coccoides were lower than those with the other diets. With diets of T. ISO, C. calcitrans, and S. costatum, algal cell-clearance rates (Fig. 3) showed a pattern similar to that for oxygen-consumption rate. Juveniles fed on diets that gave higher size-related respiration rates consumed a greater weight of algal food cells. Gross-gosh efficiency, K, = G. C- ‘, is the propo~ion of the cleared ration, C, incorporated into organic growth, G. Net-growth efficiency, KZ = G *(G + R) _ ‘, is the proportion of the assimilated ration used for growth, where R is the AFDW equivalent of the energy used in respiration, calculated as 1 mg . 1.2 ml 0, consumed - l (Walne, 1965). Growth efficiencies, K, and K2, for 0.3 mg organic weight T. semidecussata juveniles fed T. IS0 or S. costatum at 15-25 “C are shown in Table VII. The weekly growth increment, G, was calculated from the equation: G = 0.3ek - 0.3,
I. LAING ET AL.
---
0.2
0.4 aqmlc
Organic
I 0.8
0.6 wt.
of
wt.of
--I 1.0
1.2
,uveniles
(mg)
juveniles
(mg)
1.4
Fig. 3. The relationship between algal food cell-clearance rate (C, mg organic weight of food cells cleared. clam ’ wk- ‘) and organic weight of T. semidecussata juveniles fed A: T. IS0 at 25 “C ( q), 20 “C (0). and 15 “C (A); and B: T. IS0 (Q), C. calcitrans (X), and S. cost&urn (V) at 20 “C.
where k is the growth-rate coefficient (from Eqn. 1). Cleared ration values, C, for 0.3-mg-org~ic-weight juveniles were calculated from food cons~ption-clam size relationships (e.g., Fig. 3). Respiratory requirements, R, were calculated from datagiven in Table VI. In Table VII, it can be seen that growth of T. semidec~sa~a juveniles was similar with the two diets, but, at each temperature, cleared ration and respiratory values were higher with S. costatum leading to lower growth efficiencies than with T. ISO. Both diets were utilized most efficiently at 20 “C.
S. S. C C. C. C.
costatum costatum calcitrans calcitrans ~alc~~rans calcitrans
-r. iso S. costatum
T. IS0 T. IS0
15 20 2.5 15 20 25 20 20 20 20
(“Cl
Temperature
T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. semidecussata T. decussata M. mercenaria 0. edulis
Bivalve species
0.211 0.395 0.638 0.211 0.395 0.638 0.330 0.220 0.370 0.580
G
0.056 0.091 0.182 0.070 0.116 0.224 0.161 0.403 0.191 0.102
R f R)-‘)
0.790 0.866 0.780 0.750 0.773 0.740 0.672 0.353 0.660 0.850
K,(G(G
0.68 1.30 2.00 0.90 2.18 4.98 5.00 5.50 4.80 4.20
c
0.31 0.30 0.32 0.23 0.18 0.13 0.07 0.04 0.08 0.14
K,(GC_‘)
of diet and temperature on the growth response of four species of juvenile bivalves of 0.3 mg initial organic weight. Weekly growth increment, G, respiration rate, R, and weight of food cells consumed, C, all expressed as mg AFDW. Data for 0. eduh from Laing & Millican (1986).
Diet
Effect
TABLE VII
34
I. LAING ET AL..
Growth efficiencies of T. semidecussuta, T. decussataa,M. mer~ena~a and 0. edulis, all fed C. calcitrans at 20 “C, are also given in Tabie VII. T. semidecussata and M. mercenaria were very similar in their growth response to this diet, giving similar size-specific values of G, R, C, K,, and K2. All four bivalves cleared similar amounts of food from suspension but the diet was used most efficiently by 0. eduh juveniles. They used less energy in respiration and grew more quickly than the three clams. The respiration rate of T. decussata was much higher than with the other two clam species on this diet. Discussion The relative food value of the algal species examined ranked the same irrespective of the clam species used. Although the growth data presented refer mainly to the two Tapes species, results obtained with M. mercenarib by other workers show a similar ranking of these algal diets as that shown here (Table III). Walne (1970) found very high growth rates of A4. mercenaria fed with S. costatum, low growth rates with P. tricornutum and very little growth with C. coccoides. Epifanio (1979) showed that T. suecica promoted moderate growth of M. mercenatia juveniles. Other molluscan species fed with these diets show similar relative growth rates (Peirson, 1983; Laing & Millican, 1986; Enright et al., 1986).
These authors have suggested that, although total lipid content of the diet does not correlate with food value, one of the more important factors dete~ining nutritional value of algae is their PUFA content. In particular, eicosapentaenoic acid, 20 : 5 ~3, and docosahexaenoic acid, 22: 6~3, are considered to be essential: algae that contain relatively high amounts of one or other generally support much better growth than species without either (Langdon & Waldock, 1981; Webb & Chu, 1983). Consequently, T. ISO, which has a high 22 : 6~3 content, and S. costatum, C. calcitrans, and T. pseudonana, which are rich in 20 : 5 03, were of good food value. T. suecica, which contains less 20 : 5 w3 than the diatoms, was of moderate food value (Table III). The high nutritional value of S. costatum was of interest and this species can be cultured successfully on a large scale, both in intensive indoor systems (Laing, 1985) and in outdoor tanks (De Pauw et al., 1983), making it a p~ic~~ly useful alga for commercial clam rearing. P. tri~om~~rn gave poor growth of T. semide~~ssata, even though this alga contains both 20 : 5 03 and 22 : 6 w3 fatty acids (Moreno et al., 1979). This is probably because P. tricornutum is indigestible to many bivalves (Epifanio, 1983). T. decussata performed moderately well with P. tricornutum, showing a higher growth-rate coefficient than T. semidecussata with this diet. C. coccoides was of very little food value. This species is known to lack essential PUFAs (Chuecas & Riley, 1969) but may also be of limited food value due to indigestibility of the cell wall (Webb & Chu, 1983) or a toxicity factor (Taub & Dollar, 1968). The small growth increment observed with this alga, and with unfed animals, may reflect uptake of other organic matter in the seawater (Walne, 1970; Manahan et a/., 1982).
GROWTH
When diets of good to moderate the effect of an increase in temperature size-related
algal cell-clearance
OF JUVENILE
CLAMS
35
food value were fed to T. semidecussata juveniles, from 15 to 25 ‘C was to give an increase in clam
rate, with the result that more energy was available
and
the animals grew faster. Newell (1980) suggested four potential mechanisms by which bivalves may respond to an increase in environmental temperature such as this. These are: (1) an increase in food consumption, C, to offset metabolic losses, R; (2) an increase in absorption efficiency, and therefore the amount of assimilated energy, A, available; (3) suppression of energy losses through respiration, R; (4) a combination of I, 2, and 3. Here, T. semidecussata juveniles are shown to have followed Strategy 1. There was no evidence of increased absorption efficiency (Strategy 2) or suppression of energy losses (Strategy 3) as temperature increased. K, remained constant or decreased slightly with increasing temperature and oxygen consumption continued to increase curvilinearly from 15 to 25 “C (Table VII). The increased energy expenditure in respiration as temperature increased used a greater proportion of the assimilated energy at 25 “C than at 20 “C, shown by lower K2 values (Table VII). This was associated with a lower rate of organic growth in relation to shell deposition at higher temperatures (Table IV), an effect also noticed in T. semidecussata and M. mercenaria juveniles grown outdoors (Mann & Glomb, 1977; Millican & Williams, 1985). T. decussata appears to have a lower optimum temperature for growth than :r. semidecussata and M. mercenaria (Fig. 2). At 20 ‘C, lower growth rates of T. decussata juveniles were associated with higher oxygen-consumption rates, compared with the two other clam species (Table VII). When food was absent or the cells were of low nutritional value, e.g., C. coccoides, the clams suppressed their oxygen consumption resulting in the conservation of energy and the maintenance of some growth. At 20 “C, the oxygen consumption of l-mg-organic-weight animals was 0.85 ~1. h- ’ with a diet of C. coccoides, compared with 1.33 ~1. h - ’ with T. IS0 (Table VI). This corresponds to Strategy 3 of Newell (1980). At 25 “C, oxygen consumption of clams fed C. coccoides increased to 1.07 ~1. h _ ’ and with a limited amount of food energy available the increased maintenance requirement reduced the capacity for organic growth (Fig. 2A). At > 25 “C, growth of M. mercenaria and T. semidecussata juveniles decreased (Fig. 2). The effect of high temperature on the growth response of bivalves is discussed by Winter (1978). A decline in growth rate is usually associated with a marked decrease in filtration rate at these temperatures. All of the alga species tested are viable in the temperature range of lo-25 “C. At > 25 “C, lower growth rates of clams may be associated with temperature stress of the algal food cells, which tend to settle out of suspension (Ukeles, 1961). At all temperatures, the proportion of the cleared algal ration utilized in growth was low, shown by low observed value of K, in Table VII. Newell (1980) suggests that this is due to an “exploitative strategy”, whereby in conditions of abundant food resources, as in these trials, the clams maintain a maximum energetic gain from the environment by processing the food cells at a high rate, resulting in a high respiratory energy expenditure.
36
I. LAING
ET AL.
Growth efficiency of clams was modified by diet as well as temperature. This was most clearly illustrated by the comparison of the growth response of 0.3-mg-organicweight T, semidec~ssuta juveniles to single-species diets of T. ISO, S. ~~~t~tu~, and C. caZcitrm.~ at 20 “C (Table VII). Respiration utilized proportionally more of the assimilated energy when T. semidecussata juveniles were fed the diatoms S. costatum and C. calcitrans compared with the flagellate T. ISO. Even with this higher respiration rate, juveniles fed S. cost&urn were still able to grow at the same rate as those fed T. IS0 because the lower efficiency of utilization of the diet was compensated for by an increased ration. Although a higher ration of C. calcitrans cells was cleared than of S. costatum, similar amounts of each were assimilated (G + R = 0.5 mg AFDW at 20 ‘C). About 20% more of this assimilated energy was used in respiration, and organic weight-growth increment was 16% lower with a diet of C. caZcitransthan with S. costatum. Utilization efficiency of a diet of C. calcitrans was much higher by 0. edulis juveniles than by T. semidecussatu juveniles. More of the cleared ration was available for growth and respiration, and the proportion of this assimilated energy used in respiration was lower (Table VII). The energetic responses of the clams to diet and temperature affected the lipid reserves of the animals. The higher apparent energy requirement for the utiIization of diatom diets was associated with a smaller proportion of lipid in the organic reserves, particularly at higher temperatures, compared with juveniles fed flagellate diets of moderate to good food value. Mill&n & Williams (1985) found that M. mercenaria and T. semidecussata reared in a lagoon fertilized to encourage blooms of diatom species contained proportion~ly less lipid than animals in the sea. Unfed clams and those fed C. coccoides showed a loss in percentage lipid content at 20 “C and especially at 25 “C when the animals were presumably utilizing reserves to meet increased metabolic requirements. For the commercial culture of these clam species, a balance needs to be maintained between growth rate, growth efficiency, and juvenile quality, in terms of organic and lipid reserves, for any diet-temperature combination. The results in this paper show that certain algal diets promoted good growth in all three clam species, with T. IS0 and S. costatum being especially suitable. C. calcitrans, C. salina, T. pseudonana, and T. suecica are also useful diet species for clam rearing. Although maximum organic growth of T. semidec~sata and M. mercenarib occurred at 2.5 ‘C, a culture temperature of 20 OC may be more appropriate, as this produces animals of a higher quality which utilize the diet fed more effkiently. However, as the clams are growing more slowly at 20 “C than at 25 ‘C, culture facilities are tied up for a greater time. Most efficient growth of T. decussata juveniles occurs at 15-20 “C. The nature of the growth response described gives some insight into the metabolic requ~ements and strategies of these clams for maint~ning m~imum growth with variation in food supply and temperature in their natural environment.
GROWTH OF JUVENILE CLAMS
37
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