Nauplii of the Calanoid Copepod, Acartia sinjiensis as an Initial Food Organism for Larval Red Snapper, Lutjanus argentimaculatus.

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1 SUISANZOSHOKU (1997-H9) l Nauplii of the Calanoid Copepod, Acartia sinjiensis as an Initial Food Organism for Larval Red Snapper, Lutjanus argentimaculatus. Masanori Doi1), Atsushi OHNO1), Yasuhiko TAKI1), Tanin SINGHAGRAIWAN2), and Hiroshi KOHNO1) 1)Tokyo University of Fisheries, Konan, Minato-ku, Tokyo 108, Japan 2)The Eastern Marine Fisheries Development Center, Ban Phe, Muang District, Rayong 21160, Thailand Abstract Initial feeding traits of larval red snapper Lutjanus argentimaculatus were investigated by supplying nauplii of the calanoid copepod Acartia sinjiensis and rotifers. The gut content of larvae consisted solely of A. sinjiensis nauplii (mean nauplii/larva) on 0-2 days after initial mouth opening (days 0-2). The mean body width of the nauplii ingested was mm on day 0, subsequently increasing with larval age. Feeding on rotifers (mean lorica width, mm) was observed from day 3 (mean 0.92 rotifers/larva), with the number ingested increasing markedly on days 4 and 5. The mouth width of red snapper larvae on day 0 ( mm) was larger than the body width of the nauplii and rotifers but smaller than those of other fish species conventionally reared using only rotifers. This small mouth size may have been initially restrictive, allowing only the seizure of early nauplii at the onset of feeding. The larvae grew well from 2.84 mm in mean total length (TL) on day 0 to 3.38 mm TL on day 5, without significant mortality. Nauplii of A. sinjiensis constitute a promising initial food for early red snapper larvae and very likely also for those of other marine fishes with a small mouth size. Introduction Mass seed production of marine fishes has been successful for those species of which larvae can feed on the rotifer, Brachionus plicatilis from the onset of feeding. Among these are the red seabream, Pagrus major and the flounder, Paralichthys olivaceus in East Asia, the seabass, Lates calcarifer in Southeast Asia, and the turbot, Scophthalmus maximus in Europe. On the other hand, seed production trials, utilizing rotifers as an initial food organism for groupers and snappers in Southeast Asia have not been very successful1-4). This is attributable to failures in ensuring the larvae of a smooth transition from endogenous to exogenous feeding. Their mouth size at the onset of feeding may not be large enough to accept rotifers and the amount of endogenous energy resource remaining at this time is small in these species5-7). A search for food organisms suitable for the larval rearing of such species, considering factors of body size, nutritional quality and Received : July 1, 1996 Key words: Acartia sinjiensis, Lutjanus argentimaculatus, Initial feeding, Larval rearing

2 32 M.Doi et al. (1997) ease of mass cultivation, has been made by many workers, e.g., larval bivalves2,8-10), phytoplankton9,11) screened small Brachionus rotifers9) and a small Thaistrain rotifer, Brachionus sp2,12,13) Moderately successful results have been obtained in the larval rearing of the red snapper, Lutjanus argentimaculatus by supplying unsorted, tank-cultured copepods prior to rotifer feeding14). Because they are the main diets of many marine fish larvae in the wild, copepods have been considered promising candidates for initial larval food15). Among the various coastal copepod species, those of the calanoid genus Acartia seem to have great potential as initial dietary organisms; they occur world wide16) and can be propagated in both indoor17) and outdoor18-20) tanks. Successful results have been obtained in the larval rearing of red seabream, Pagrus major using Acartia tsuensis19). In the present study, early feeding traits of Lutjanus argentimaculatus larvae were investigated by supplying them with tank-propagated Acartia sinjiensis nauplii and rotifers, with the object of examining the availability of A. sinjiensis nauplii as an initial food organism for tropical marine fish larvae having a small mouth. Materials and Methods The study was conducted at the Eastern Marine Fisheries Development Center (EMDEC), Rayong Province, Thailand. Eggs of the red snapper, Lutjanus argentimaculatus were derived from hormone-induced spawning at EMDEC, the inducement and spawning procedures having been described by Singhagraiwan and Doi14) The 2.5m3 circular tanks (2 m in diameter and ca. 0.8 m in water depth) were used in the larval rearing experiment. The tanks were filled with sand-filtered sea water (salinity 32 ppt), and supplied with the green algae, Nannochloropsis sp. at a density of 60 ~ 104 cells/ml and a chemical fertilizer (composition: 78.1% KNO3, 7.8% NaH2PO4, 3.9% Na2SiO3, 2.4% FeC12 and 7.8 % Nat EDTA) at 91 g/m3 on May 5, At night of the same day, copepodids of Acartia sinjiensis was collected from a 2.5 ha earthen pond at EMDEC using a light trap21), and introduced to the tanks as brooders for nauplii propagation. The stocking density of the copepodids was 28 individuals/l. After the introduction of the copepodids, the tank water was found to have been contaminated by the rotifer, Brachionus plicatilis cf. rotundiformis22), due probably to the initial rotifer-contamination of the Nannochloropsis cultures. The study was continued however, including in its parameters the electivity of red snapper larvae toward A. sinjiensis and Brachionus sp. Spawning of L.argentimaculatus took place from 01:00 to 02:00 on May 6.About 8 hours after spawning, 5,000 fertilized eggs were transferred into each rearing tank. The tank water was gently aerated, remaining unchanged during the experiment from May 6 to May 13. On May 10, 40 ~ 104 rotifer individuals were added to each tank. During the experiment, the water temperature in the tanks at 10:00 ranged from 28.8 Žt to 30.2 Ž (mean 29.3 Ž). Of the two tanks, one was used solely for monitoring larval survival, all other data being taken from the other tank. Sampling of food organisms in the tank water was conducted daily at 10:00, by collecting a 5 l water sample using a water column sampler. Sampling of red snapper larvae, made by gentle use of a scoop net, was undertaken more than once a day from May 8, the day of initial mouth opening of the larvae. Larval samples were preserved in 5 % formalin. In each sample, 11 to 20 individuals were used for examination of growth and gut contents, with mouth width measurements taken from an additional 10 larvae. Larval survival was estimated at 21:00 on May 6, 8, 10 and 12 by the water column method. The body width of A. sinjiensis was obtained by conversion from the body length measurement, using the length-width relationship determined from 120 individuals in the rearing tank. Likewise, the lorica width of rotifers was determined from the length-width relationship established from 100 individuals. The degree of fullness of the gut of red snapper larvae was described by the following gut fullness index: 0, food absent; 1, food less than 10 % of total gut capacity; 2, food 10-50% of total capacity; 3, food 50-80% of total capacity; 4, food % of total capacity, the gut having no empty spaces; 5, gut overfull, determined from the smooth, stretched gut surface. The electivity

3 Acartia nauplii, initial food for fish larvae 33 of larvae toward different food organisms was calculated using IVLEV's equation23) for food electivity. In this paper, days are numbered from the time of initial mouth opening of the red snapper larvae, which occurred on May 8 (=day 0). Results Size and propagation of food organisms The organisms occurring in the experimental tanks throughout the experimental period comprised mainly the copepod, Acartia sinjiensis and the rotifer, Brachionus sp. which together accounted for about 98 % of the total number of individuals. Other organisms found in the tanks were nauplii of both Oithona sp. and Balanus sp. stage-specific means fitted well to the regression W= 0.510L (r=0.993). Because the linear increase in body size was closely correlated with developmental stage, without any overlapping of body lengths between the stages, identification of naupliar stages could be determined from body length, in addition to appendage morphology, although slight overlaps in body width were seen between more advanced stages. The lorica length and width of the rotifers were }0.020 and }0.016 mm (mean }SD, n =100),respectively, the relationship being expressed as W=0.712L (r=0.758) (Fig.l).Rotifer body width was greater than that of A. sinjiensis of the same body length. By day 0, adult A. sinjiensis had propagated in the tank. The abundance of nauplii was 347 individuals/l on day 0 and 486 individuals/l on day 1(Fig.2).The The mean body length and width of A. sinjiensis nauplii increased as the naupliar stage advanced, from ~ mm (n=20) at stage 1 (N-1) to ~0.123 mm (n=20) at N-6 (Fig.l).The Fig.2.Changes in abundance (individuals/l) of Acartia sinjiensis nauplii (solid circles) and Brachionus sp. (open triangles) larval rearing tank. in the 2.5 m3 Fig.1.Relationships between body length and width of Acartia sinjiensis nauplii (solid circles) and lorica length and width of Brachionus sp. (open circles, n =100) used in the study. Solid circles and bars indicate stage specific means and ranges of the nauplii (n=20 for each stage, N-1-N-6). Some of the points for Brachionus sp. represent multiple measurements. density then decreased gradually to 11 individuals/l on day 5. Of the different developmental stages occurring in the tank, nauplii at N-2 were most abundant on day 0, accounting for 47 % of the total number of nauplii, followed by N-3 (16 %) and N-1 (14 %). The abundance of copepodids was 73 individuals/l on day 2, thereafter remaining steady at individuals/l till day 5. The abundance of rotifers increased from 69 individuals/l on day 0 to 114 individuals/l on day 2 (Fig.2). Following the additional supply of rotifers on day 2 (after the monitoring of rotifer abundance

4 34 M.Doi et al. (1997) Fig.3.Changes in total length (A) and mouth width (B) of larval Lutjanus argentimaculatus reared in the 2.5 m3 tank. Means and ranges are shown. for the day), the abundance increased to 557 individuals/l on day 4, but subsequently declined to 189 individuals/i on day 5. increase in width during days 1 and 2, after which it increased linearly, reaching }0.041 mm (n=10) on day 4. Survival and growth of red snapper larvae The hatching and survival rates from hatching (day -2) to the onset of feeding (day 0) were low (about 40 % and 50 %, respectively), with about 1, 000 larvae being estimated as surviving until day 0 in each tank. However, the number of larvae per tank was calculated as 800-1, 200 on days 2 and 4, indicating high survival rates, nearly 100 %, after the onset of feeding. At 08 :00 on day 0, larvae measured 2.84 }0.06 mm Feeding of larvae The feeding incidence rate (percentage of larvae TL (mean }SD, n=20; Fig. 3A), growing to 2.90 } 0.01 mm TL (n=11) by 17:00 on the same day. Larval length almost did not increase during days 1 (2.90 } 0.01 mm TL, n=20, at 07:00) and 2 (2.95 }0.01 mm TL, n=11, at 10 : 00), but subsequently accelerated, attaining 3.38 }0.09 mm TL (n=12) on day 5. By 03:00 on day 0, the mouth had opened in about 50 % of the larvae. The mouth width, measured at 08:00 on the same day, was }0.010 mm (mean } SD, n =10; Fig. 3B), increasing to }0.008 mm (n=10) at 14:00 on the same day and }0.017 mm (n=10) at 07:00 on day 1. Mouth opening did not Fig. 4. Feeding incidence (% of larvae with food in gut; solid circles) and mean index of gut fullness (see, text; open circles) in larval Lutjanus argentimaculatus.

5 Acartia nauplii, initial food for fish larvae 35 with food in the gut) was 0 % (n = 20) at 08 :00 on day 0 (5 hours after mouth opening in about 50 % of larvae). The rate then increased abruptly to 87.5 % (n =16) by 11:00 and 90.9% (n=11) by 17:00 on the same day (Fig. 4). The feeding incidence rate was 100 % at 10:00 on day 1 and thereafter (n=11-20). The mean gut fullness index was 2.50 (n=16) 11:00 and 2.95 (n=11) at 17:00 on day 0, and 2.81 (n=20) at 07:00 and 3.75 (n=12) at 10:00 on day 1 (Fig.4). The index maintained a level greater than 3.3 in all observations made at 10 : 00 on days 2-5 (n= 11-12). Throughout the experimental period, the gut contents comprised almost entirely of A. sinjiensis nauplii and rotifers. Occasional other items found were eggs of the above two species. Copepodids of A. sinjiensis were not found in the gut contents until the end of the experiment (day 5). At 11:00 and 17:00 on day 0, the gut contents of larvae consisted solely of A. sinjiensis nauplii (mean 2.1 (n=16) and 3.4 (n=11) individuals/larva, respectively) (Fig. 5). The number of nauplii per gut at increased to 7.2 on day 2 (n ii), subsequently decreasing gradually to 1.7 on day 5 (n=12). Rotifers were first observed in the larval gut on day Fig. 5. Average numbers (individuals/larva) of Acartia sinjiensis nauplii (solid circles) and Brachionus sp. (open triangles) observed in gut of larval Lutjanus argentimaculatus. Fig. 6. Frequency distribution (individuals/larva) of body width of Acartia sinjiensis nauplii (open areas) and Brachionus sp. (shaded areas) ingested by larval Lutjanus argentimaculatus.

6 36 M. Doi et al. (1997) Table 1. Ivlev's food electivity index toward Acartia sinjiensis nauplii and Brachionus sp. in Lutjanus argentimaculatus larvae 1 Numbers in parentheses indicate number of larval specimens examined 2 N ot available.. 3 (mean 0.92 individuals/larva, n=12) (Fig. 5). Subsequently, the number of rotifers ingested increased exponentially, reaching 3.75 (n=12) and 8.92 individuals/larva (n=12) on days 4 and 5, respectively. The body size of A. sinjiensis nauplii in the larval gut increased with larval growth (Fig. 6). The mean body width was mm (range mm, n- 70) on day 0, thereafter increasing to mm (range mm, n=79) on day 1, mm (range mm, n=79) on day 2, mm (range mm, n=75) on day 3, mm (range mm, n=44) on day 4 and mm (range mm, n=19) on day 5, respectively. Based on the linear relationship between body size and naupliar stage and the size distribution patterns shown in Fig. 1, the stage of the nauplii in the gut was considered to be mostly N-1 on day 0, N-2 on days 1 and 2, N-2 to N-4 on day 3, and N-2 to N-3 on days 4 and 5. Changes in the food electivity index indicated that the above advances in the size and stage of nauplii ingested were due to changes in larval preference toward larger and hence more advanced nauplii (Table 1). On day 0, the index was highest for N-l, 0.59 at 11:00 and 0.55 at 17:00. The index for N-1 became negative on day 1, being at 07:00 and at 10:00, and remained so on days 3-5, from to In contrast, electivity to N-2 shifted from essentially neutral on day 0 (-0.02 at 11:00 and 0.01 at 17:00), to positive, on days 1-3 and 0.73 and 0.87 on days 4 and 5, respectively, The mean lorica width of rotifers ingested was mm (range mm, n=11) on day 3 and mm (range mm, n = 152) on days 4 and 5 (Fig. 6). Although there were no statistical differences between the mean lorica width ingested on days 3-5 (ANOVA, p > 0.05), the mode changed from 0.10 mm on day 3 to 0.12 mm on days 4 and 5 (Fig. 6). The mode of lorica width on days 4 and 5, 0.12 mm, was equivalent to the mode for rotifers in the tank water, indicating that the larvae on day 3 tended to feed on relatively small sized rotifers. The food electivity index for rotifers was -1.0 (no feeding) on days 0-2, on day 3, on day 4 and 0.15 on day 5 (Table 1). Although the index had gradual positive increase, it was much lower than that for A, sinjiensis nauplii, indicating that the larvae fed selectively on the latter, rather than on rotifers, throughout the experimental period. Preference of food organisms Discussion The increases in size of the food organisms (Acartia sinjiensis nauplii and rotifers) ingested by the red snapper larvae (Fig. 6) and in mouth width of the latter (Fig. 3B) strongly suggested that the size of food organisms in relation to mouth size was the primary factor in the selection of food organisms by the larvae, as in the seabream, Archosargus rhomboidalis24), turbot, Scophthalmus muximus25), gilt-head seabream, Sparus aurata26) and many other marine fish larvae15,27)

7 Acartia nauplii, initial food for fish larvae 37 Nevertheless, the red snapper larvae showed a strong preference toward advanced A. sinjiensis nauplii (N-4 and N-5) rather than rotifers on days 3 and 4 (Table 1), although their body sizes were nearly equal (Fig. 1). Factors involved in such electivity are considered to include locomotory patterns of the prey28,29), body color and external morphology. Clarification however necessitates further detailed observations. In the present study, the larvae had started to feed on rotifers from day 3, at a time when the density of available A. sinjiensis nauplii had declined, but that of rotifers had increased (due to the additional supply). In case when the density of nauplii had been maintained at a higher level, the larvae might have fed them on more often, if not solely, on and after day 3. Availability of Acartia for initial larval feeding In our four previous attempts to rear red snapper larvae using rotifers, high mortalities occurred during the first few days, not only before but also after the onset of larval feeding, resulting in the nearly complete extinction of the larvae by around day 3 (= 5 days after hatching). Bonlipatanonl) also reported high mortalities occurring in red snapper larvae fed with rotifers, the rate of larvae with rotifers in the gut being % on day 1. All larvae died by day 13 in nine of his 28 rearing experiment replicates. On the other hand, in the present study in which red snapper larvae were provided with both A. sinjiensis and rotifers, the larvae showed high survivorship after the commencement of feeding, preying solely upon A. sinjiensis nauplii during the first three days, shifting gradually from smaller, earlier nauplii to larger, more advanced ones. The larvae started feeding at the latest some eight hours after their initial mouth opening. All individuals examined had significant numbers of nauplii in the gut on and after day 1 (Figs. 4 and 5), although the density of nauplii in the rearing tank (347 individuals / l on day 0 and 486 individuals / l on day 1) was lower than that applied30) in the seed production (5,000-20,000 individuals/l). of rotifers generally of marine fishes Considering the size of the nauplii (mean body width mm for N-1-N-3) and rotifers (mean lorica width mm), the larvae may have been able to seize early nauplii, but not advanced ones or rotifers, at the onset of feeding. Although the mouth width of the larvae was larger than the width of the nauplii and rotifers, the gape of the former seemed to have not been large enough for capturing advanced nauplii and rotifers, a situation similar to that suggested for early grouper larvae31,32) In fact, the mouth width of the red snapper larvae at the onset of feeding observed in this study ( mm) was small compared with other species for which larval rearing is conducted using only rotifers (e. g., mm in seabass, Lates calcarifer33); mm in milkfish, Chanos chanos34)), even considering the possible effect of formalin-preservation on the mouth size measurement. Although the present rearing experiment was on a small and preliminary scale, the findings demonstrated that the early larvae of red snapper, and most probably those of other species with a relatively small mouth size at the time of initial mouth opening, such as grouper, Epinephelus fuscoguttatus rabbitf ish, Siganus guttatus (0.193 mm5)) and (0.168 mm35)), can be reared successfully if early stage nauplii of A. sinjiensis or other Acartia species are provided for the first few days prior to rotifer feeding. Acknowledgments We wish to thank Dr. G. Hardy, Thames, New Zealand, for his critical review of the manuscript. This study was supported partly by the Japan International Cooperation Agency. References 1) Bonlipatanon, P. (1988): Study of red snapper (Lutjanus argentimaculatus Forsskð l) spawning in captivity, in greport of Thailand and Japan Joint Coastal Aquaculture Research Project, No. 3 h, National Institute of Coastal Aquaculture, Thailand, and Japan International Cooperation Agency, pp ) Doi, M., M. N. Munir, N. L. Nik Razali and T. Zulkifli (1991): Artificial propagation of the grouper, Epinephelus suillus at the marine finfish hatchery in Tg. Demong, Terengganu, Malaysia. Kertas Penembangan Perikanan Bil., 167, Department of Fisheries, Malaysia, 41 pp.

8 38 M. Doi et al. (1997) 3) Lim, L. C. (1993): Larviculture of the greasy grouper Epinephelus tauvina F. and the brownmarbled grouper E. fuscoguttatus F. in Singaphore. J. World Aquaculture Soc., 24(2), ) Emata, A. C., B. Eullaran, and T. U. Bagarinao (1994): Induced spawning and early life description of the mangrove red snapper, Lutjanus argentimaculatus. Aquaculture, 121, ) Kohno, H., S. Diani, P. Sunyoto, B. Slamet and P. T. Imanto (1990) : Early developmental events associated with changeover of nutrient sources in the grouper, Epinephelus fuscoguttatus, larvae. Bull. Pen. Perikanan (Indonesia), Spec. Ed., No. 1, ) Doi, M., H. Kohno, A. Ohno and Y. Taki (1994): Development of mixed-feeding stage larvae of red snapper. Lutjanus argentimaculatus. Suisanzoshoku, 42(3), ) Kohno, H., A. Ohno and Y. Taki (1994): Why is grouper larval rearing difficult? : a comparison of the biological natures of early larvae of four tropical marine fish species. in gthe Third Asian Fisheries Forum h (ed. by L. M. Chou, A. D. Munro, T. J. Lam, T. W. Chen, L. K. K. Cheong, J. K. Ding, H. W. Khoo, V. P. E. Phang, K. F. Shin and C. H. Tan), Asian Fisheries Society, Manila, Philippines, pp ) Lim, L. C., L. Cheong, H. B. Lee and H. H. Heng (1985): Induced breeding studies of the John's snapper Lutjanus johnii (Block), in Singapore. Singapore J. Pri. Ind., 13(2), ) Hamamoto, S., M. Tochino and K. Yokogawa (1986): Effects of small size foods and illumination in breeding the larvae of red spotted grouper, Epinephelus akaara (Temminck et Schlegel). Bull. Kagawa Pref. Fish. Exp. Stn., 2, (in Japanese with English abstract) 10) Lin, K. J., C. L. Yen, T. S. Huang, C. Y. Liu and C. L. Chen (1986): Experiments on the artificial propagation of black spotted grouper, Epinephelus salmonoides (Lacepede)-II, experiment of fry nursing of Epinephelus salmonoides (Lacepede) and its morphological study. Annual Collected Rep., The Penghu Branch (Penghu Fishery Laboratory), Taiwan Fisheries Research Institute, 5, (in Chinese with English abstract) 11) Kayano, Y. and T. Oda (1987): Rearing experiment of red spotted grouper, Epinephelus akaara larvae with diatoms. Bull. Okayama Pref. Fish. Exp. Stn., 2, (in Japanese) 12) Fukunaga, K., K. Nogami, Y. Yoshida, K. Hamazaki and K. Maruyama (1990): Recent increase of Epinephelus akaara seed production amount and its problems at the Tamano Branch Station of the Japan Sea-Farming Association. Saibai Giken, 19 (1), (in Japanese) 13) Kayano, Y. and S. Yamamoto (1991): Optimal quantities of the small typed rotifer Brachionus plicatilis taken on first feeding by larvae of the red spotted grouper Epinephelus akaara. Bull. Okayama Pref. Fish. Exp. Stn., 6, (in Japanese) 14) Singhagraiwan, T. and M. Doi (1993): Induced spawning and larval rearing of the red snapper, Lutjanus argentimaculatus at the Eastern Marine Fisheries Res. Bull., 4, Development Center. Thai. Mar. Fish. 15) Hunter, J. R. (1981) : Feeding ecology and predation of marine fish larvae, in gmarine Fish Larvae h (ed. by R. I. Lasker), Univ. Washington Press, Seattle, pp ) Ueda, H. and J. Hiromi (1987): The Acartia plumosa T. Scott species group (Copepoda, Calanoida) with a description of A. tropica n, sp. Crustacean, 53, ) Stð²ttrup, J. G., K. Richardson, E. Kirkegaard and N. J. Pihl (1986): The cultivation of Acartia tonsa Dana for use as a live food source for marine fish larvae. Aquaculture, 52, ) Ohno, A. and Y. Okamura (1988): Propagation of the calanoid copepod, Acartia tsuensis, in outdoor tanks. Aquaculture, 70, ) Ohno, A. (1992): Fundamental study on the extensive seed production of the red seabream, Pagrus major. Special Research Report No. 2, Japan Sea Farming Association, 110 pp. (in Japanese with English abstract) 20) Doi, M., S. Singhagraiwan, T. Singhagraiwan, M. Kitade and A. Ohno (1994): Culture of the calanoid copepod, Acartia sinjiensis, in outdoor tanks in the tropics. Thai Mar. Fish. Res. Bull., 5,

9 Acartia nauplii, initial food for fish larvae 39 21) Doi, M. and T. Singhagraiwan (1993): Biology and culture of the red snapper, Lutjanus argentimaculatus. The research project of fishery resource development in the Kingdom of Thailand, The Eastern Marine Fisheries Development Center, Thailand, and the Japan International Agency, 51 pp. Cooperation 22) Furusawa, M. (1990): L-type and S-type of rotifers. in grotifers as initial food organisms h (ed. by K. Fukusho and K. Hirayama), Koseisha Koseikaku, Tokyo, pp (in Japanese) 23) Ivlev, U. S. (1961): Experimental ecology of the feeding of fishes (translated from the Russian by D. Scott). New Haven, Yale Univ. Press, 302 pp. 24) Stepien, W. P. (1976): Feeding of laboratoryreared larvae of the sea bream Archosargus rhomboidalis (Sparidae). Mar. Biol., 38, ) Kuhlmann, D., G. Quantz and U. Witt (1981): Rearing of turbot larvae L.) on cultured food organisms (Scophthalmus maximus and postmetamorphosis growth on natural and artificial food. Aquaculture, 23, ) Polo, A., M. Yufera and E. Pascual (1992): Feeding and growth of gilt-head seabream (Spares aurata L.) larvae in relation to the size of the rotifer strain used as food. Aquaculture, 103, ) Shirota, A. (1970): Studies on the mouth size of fish larvae. Bull. Japan. Soc. Sci. Fish., 36(4), (in Japanese with English abstract) 28) Van der Meeren, T. (1991): Selective feeding and prediction of food consumption in turbot larvae (Scophthalmus maximus L.) reared on the rotifer Brachionus plicatilis and natural zooplankton. Aquaculture, 93, ) Buskey, E. J., C. Coulter and S. Strom (1993): Locomotory patterns of microzooplankton: potential effects on food selectivity of larval fish. Bull. Mar. Sci., 53(1): ) Anonymous (1986) : Technical manual for seed production of seabass. National Institute of Coastal Aquaculture, Songkhla, Thailand, 49 pp. 31) Maneewong, S., P. Akkayanont, J. Pongmaneerat and M. Iizawa (1986): Larval rearing and development of grouper, Epinephelus malabaricus (Bloch and Schneider). in greport of Thailand and Japan Joint Coastal Aquaculture Research Project, No. 2 h, Japan International Cooperation Agency, pp ) Kayano, Y. (1988): Development of mouth parts and feeding in the larval and juvenile stages of red spotted grouper, Epinephelus akaara. Bull. Okayama Pref. Fish. Exp. Stn., 3, (in Japanese) 33) Kohno, H., S. Hara and Y. Taki (1986): Early larval development of the seabass Lates calcarifer with emphasis on the transition of energy sources. Nippon Suisan Gakkaishi, 52(10), ) Kohno, H., M. Duray, A. Gallego and Y. Taki (1990): Survival of larval milkfish, Chanos chanos, during changeover from endogenous to exogenous energy sources. in gthe Second Asian Fisheries Forum h(ed. by R. Hirano and I. Hanyu), Asian Fish. Soc., Manila, Philippines, pp ) Kohno, H., S. Hara, M. Duray and A. Gallego (1988): Transition from endogenous to exogenous nutrition sources in larval rabbitfish Siganus guttatus. Nippon Suisan Gakkaishi, 54(7),

10 40 M. Doi et al. (1997)