Nutrition of red drum,Sciaenops ocellatus L.: An additional evaluation of the effects of soya‐based diets and supplemental prebiotic

We assessed the effects of a fishmeal (FM)‐ and two soya‐based diets [without or with prebiotic (GroBiotic^® ‐A; GBA) supplementation] on the production performance, non‐specific immunity, blood parameters and gut microbiota diversity of red drum, Sciaenops ocellatus. All diets were formulated to contain 450 g/kg crude protein (CP) and 120 g/kg lipid. A combination of soya bean meal and soya protein concentrate (SP) was used to replace 88% of the FM in the FM‐based diet (REF) originating the first soya‐based diet (SOY). The second soya‐based diet (SOY + GBA) was designed identically to SOY except for the supplementation of GBA (20 g/kg) substituting for wheat flour. After feeding the experimental diets to triplicate groups of 25 (~3.4 g/fish) and 90 (~4.1 g/fish) red drum juveniles for 9 (Trial I) and 16 weeks (Trial II), our results showed that (i) red drum was highly sensitive to the soya‐based diets in the first week of feeding; (ii) replacing 88% of FM with SP reduced the production performance of red drum without affecting intestinal micromorphology; (iii) whole‐blood leucocyte oxidative radical production was unaffected while lower haematocrit and higher plasma lysozyme activity were found in fish fed REF; (iv) plasma osmolality was unaffected while lower glucose and higher lactate levels were observed in fish fed SOY and SOY + GBA; (v) 16S rRNA gene‐based diversity of fish gut microbiota was affected both by SP and by GBA. This study expands the information on red drum responses to soya‐based diets and prebiotic supplementation.     

Development of Digestive Enzymes in California Halibut <Emphasis Type="Italic">Paralichthys californicus</Emphasis> Larvae

California halibut Paralichthys californicus is an important commercial species with high aquaculture potential in Baja California Sur, México. To optimize the feeding process using live prey and/or inert diets, we evaluated alkaline proteases, pepsin, trypsin, chymotrypsin, leucine aminopeptidase, lipase, α-amylase, and acid and alkaline phosphatase activities on starved larvae and larvae fed live prey. Highest activities were observed for alkaline protease, trypsin, chymotrypsin, leucine aminopeptidase, and alkaline phosphatase in feeding larvae than starved larvae on day 4 after hatching. At day 5, a sizeable increase in all enzymatic activities was detected in feeding larvae. Alkaline protease, trypsin, chymotrypsin, and alkaline phosphatase decreases progressively from day 5 until day 18. At day 18, a slight pepsin activity was observed. This was considered an indicator of the start of digestive system maturation. We concluded that total enzymatic equipment for this species is complete between day 18 and 30 after hatching. Based on this evidence, early weaning from live prey to inert feed would be possible at this time.     

Feeding Larval Red Drum on Microparticulate Diets in a Closed Recirculating Water System1

A feeding protocol was developed for red drum larvae based on combining a commercial microparticulate diet (Kyowa Fry Feed) with live prey (rotifers) in a closed, water reuse system. In five trials, growth and survival were measured on larvae reared on a combination of live and microdiet for 1–5 d and then microdiet alone. Results in each trial were compared to control larvae reared on live rotifers Brachionus plicatilis and brine shrimp nauplii Artemia salinas. The most satisfactory combination was feeding live food and microdiet together for the first five days and then completely discontinuing live prey, eliminating the need to feed brine shrimp to the larvae. Growth rates of larvae fed progressively larger sizes of the microdiet were as good as larvae reared on live prey. Both groups metamorphosed to the juvenile stage at less than one month. Survival rates on the five day live food and microdiet combination were a remarkable 60% from egg to the juvenile stage. The successful weaning of red drum to microdiets paves the way to produce a semipurified diet to test nutrient requirements of larval fish.     

Experimental Studies of Feeding in Larval Red Drum

The red drum ( Sciaenops ocellatus ) is the only marine fish species in commercial culture in the US. Red drum can be induced to spawn on demand at any time of the year and the young grown to marketable size in indoor raceways or in ponds. Although many eggs are produced (up to 1 million per spawning pair), grow-out through the larval stage remains a bottleneck in commercial production. Most larvae for commercial use are grown in outdoor ponds where natural blooms of phytoplankton and zooplankton are induced, but pond production is limited seasonally and geographically since red drum larvae require sea water and are intolerant of temperatures below 20 C.
It is possible to raise larvae indoors in tanks year-round by controlling temperature and salinity and by culturing live food, but not in sufficient number to supply commercial fish growers. Experimental studies are being carried out to modify the feeding and rearing systems to improve production of post-larvae. Of utmost importance is the development of a complete diet independent of live foods. Results of studies of highly unsaturated fatty acid requirements, digestive system development and activity levels of digestive enzymes are being incorporated into the development of an improved diet for red drum larvae.     

Involvement of trypsin and chymotrypsin activities in Atlantic cod (Gadus morhua) embryogenesis

Proteases play a key role in yolk formation and degradation during embryogenesis of marine fish. This study presents the first clear data on the involvement of trypsin and chymotrypsin activities in the embryogenesis of Atlantic cod (Gadus morhua). Both enzyme activities were shown to be present in unfertilized eggs followed by a significant decline (P < 0.01) in trypsin activity during the first 4 days post fertilization (dpf). Thereafter, the trypsin- and chymotrypsin activities increased to a maximum around day 9 pf. A decline in trypsin and chymotrypsin activities was observed from day 10 pf with minimal activity just prior to first feeding (day 15 pf). Western blot analysis, using polyclonal antibodies raised to Atlantic cod trypsins I and Y, mostly coincided with the trypsin activity profile. The novel trypsin Y was previously shown to have both trypsin- and chymotrypsin-like activities. Thus, some of the chymotrypsin activity observed in the samples may be originated from trypsin Y. The low trypsin and chymotrypsin activities just prior to first feeding (13–15 dpf) may indicate insufficient digestive function as trypsin has been shown to be a suitable short-term indicator reflecting the nutritional quality of marine fish larvae.     

Effect of Three Probiotics Administered Through Live Feed on Digestive Enzyme Activity in California Halibut,Paralichthys californicus,Larvae

The objective of this study was to evaluate the effect of probiotics as digestive system maturation promoters in larvae of the California halibut, Paralichthys californicus, during ontogeny. Three commercial probiotics: Levucell^®, Bactocell^®, and live yeast, were supplied at a concentration of 50 mg/L, using enriched rotifers and Artemia nauplii as vectors. Total and specific activities of trypsin, leucine‐aminopeptidase (LAP), lipase, chymotrypsin, and pepsin were assessed throughout the development. Growth in terms of length was not significantly different between treatments. Total pepsin and chymotrypsin activity were significantly higher at the end of the experiment, 46‐d posthatch (dph) in larvae supplied with Bactocell. However, no significant effect of the probiotic was detected for trypsin, lipase, and LAP. On the basis of the results, California halibut larvae seemed to have adequate alkaline proteolytic activity early in development, whereas acid proteolytic activity was not detected until the complete formation of a functional stomach. The feeding protocol and probiotics evaluated were unable to improve the morphological development of digestive system or the digestive enzyme activity early on. Nevertheless, the increase in activity of pepsin and chymotrypsin observed at 46 dph suggests a potential use of these probiotics once the metamorphosis is completed, and thus warrants further evaluation.     

Effects of the Prebiotics GroBiotic®‐A and Inulin on the Intestinal Microbiota of Red Drum,Sciaenops ocellatus

Two separate feeding trials examined the effects of dietary supplementation of the prebiotics GroBiotic^®‐A and inulin on growth performance and gastrointestinal tract microbiota of the red drum, Sciaenops ocellatus. In the first feeding trial, fish meal‐based diets without prebiotics or supplemented with either GroBiotic^®‐A or inulin at 1% of dry weight were fed to triplicate groups of juvenile red drum (initial weight of 2.6 g) in 110‐L aquaria operated as a brackish water (7 ppt) recirculating system for 8 wk. In the second feeding trial, soybean meal/fish meal‐based diets supplemented with either GroBiotic^®‐A or inulin at 1% of dry weight were fed to triplicate groups of red drum (initial weight of 15.8 g) in 110‐L aquaria operated as either a common recirculating water system or closed system with individual biofilters (independent aquaria) for 6 wk. Supplementation of the prebiotics in either feeding trial did not alter weight gain, feed efficiency ratio, or protein efficiency ratio of red drum fed the various diets. In the second feeding trial, the culture system significantly affected weight gain, feed efficiency ratio, and protein efficiency ratio although there were no effects of dietary treatments on fish performance or whole‐body protein, lipid, moisture, or ash. Denaturing gradient gel electrophoresis (DGGE) analysis of the gastrointestinal tract microbial community showed no effect of the dietary prebiotics as the microbial community appeared to be dominated by a single organism with very low diversity when compared with other livestock and fish species. DGGE of the microbial community in the biofilters of the independent aquariums showed a diverse microbial community that was not affected by the dietary prebiotics.     

Changes in digestive enzyme activity during initial ontogeny of bay snook Petenia splendida

Several samples of P. splendida larvae were obtained from eggs until day 60 after hatching (dah) to determine acid and alkaline proteases, trypsin, chymotrypsin, leucine aminopeptidase, α-amylase, lipase, and acid and alkaline phosphatase activities using biochemical techniques. Additionally, SDS–PAGE alkaline protease zymogram and PAGE acid protease zymogram were carried out to identify active isoforms during larviculture. Alkaline protease and chymotrypsin were present at the moment of hatching, increased gradually reaching the maximum values at 35 dah. Trypsin and leucine aminopeptidase activities were low from hatching, increasing gradually as larvae grew. Alkaline protease zymogram showed four zymogens, which appears at different days, remaining present until the end of the larviculture (95.2 kDa at 11 dah, 26.4 kDa at 9 dah, 21.4 kDa at 3 dah, and 23.3 kDa at hatching). Pepsin activity was present at day 7 after hatching and increased progressively until the end of the larviculture. Acid protease zymogram only showed one zymogen (0.65 rf), which appear at 6 dah. Lipase was high at the time of hatching and increased until 15 dah, after which decreased gradually. Amylase was high from the beginning and until 15 dah and then decreased rapidly to almost nothing onward. Alkaline and acid phosphatases presented a high activity at the egg stage, fell slightly during the first feeding and increased again from 20 to 30 dah. Results obtained in this study show that larvae can be fed artificial diets starting on day 10 after hatching.     

Effects of dietary animal proteins on growth and body composition of the red drum (Sciaenops ocellatus)

Two experiments were conducted to determine the nutritional value of various dietary proteins for juvenile red drum. In the first 8-week feeding trial, diets containing similar quantities of lipid, carbohydrate, available energy and ash with 35% crude protein from either lyophilized whole-body croaker (Micropogon undulatus), striated beef muscle, red drum processing waste or commercially processed menhaden (Brevoortia tyrannus) fish meal were fed to juvenile red drum in brackish (6 ppt) water along with a control diet containing lyophilized muscle of red drum. The control diet produced significantly (P<0.05) greater weight gain (WG), feed efficiency (FE), and protein efficiency ratio (PER) values than all other diets; intermediate responses were observed for fish fed diets containing protein from red drum waste and whole-body croaker, while diets containing striated beef muscle and menhaden fish meal yielded the lowest values. Some differences in tissue indices and body composition of red drum including hepatosomatic index, whole-body ash and lipid, as well as liver lipid and glycogen were induced by the various diets. In the second 8-week feeding trial, the control diet containing red drum muscle was compared with similar diets containing protein from whole-body croaker and menhaden fish meal. Again the control diet produced the greatest WG, FE, and PER values followed by whole-body croaker and then menhaden fish meal. Effects of the dietary proteins on tissue indices and body composition were limited. The excellent protein quality and low-temperature processing of lyophilized red drum muscle resulted in superior performance of red drum relative to the other evaluated protein products, and lyophilized whole-body croaker provided better performance than commercially processed menhaden fish meal.     

The Effects of Enriching Live Foods With Highly Unsaturated Fatty Acids on the Growth and Fatty Acid Composition of Larval Red Drum Sciaenops ocellatus

Three experiments were carried out to test the effects of enrichment of live food (rotifers) with varying levels of n-3 highly unsaturated fatty acids (HUFA) on the growth rate and fatty acid composition of red drum larvae. Additionally, the fatty acid compositions of red drum eggs and day-1 larvae were compared. The enrichment techniques were successful in that the levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were elevated in the rotifers fed the enrichment diet. Red drum larvae fed the control rotifers produced the highest growth rate of the three experiments. Larvae fed rotifers with no HUFA supplement (NHUFA) had a significantly lower growth rate than the controls for that experiment. The fatty acid compositions of the eggs and day-1 larvae did not vary significantly and contained high levels of 16:0, 16:1 n-7 and DHA (22:6 n-3). Based on these data, the lack of DHA in the diet significantly reduced the growth rates of larval red drum. The 10-day-old red drum larvae had similar fatty acid profiles at the end of the experiments regardless of the diet they were fed, indicating that dietary inputs have little effect on the fatty acid composition of larvae during the first ten days of growth. Red drum larvae appear to have the ability, though limited, to bioconvert EPA to DHA since there was a significant increase in the levels of DHA from day 1 to day 10 in the NHUFA larvae. However, the efficiency of this bioconversion is not sufficient for optimal growth and supplemental DHA at least to the level found in the control rotifers (0.3–0.4mg/100mg tissue) is necessary to maximize growth. The exact role of EPA could not be determined from this study due to the inability to produce an EPA-free rotifer.     

Applying the ontogeny of digestive enzyme activity to guide early weaning of pigfish,Orthopristis chrysoptera (L.)

Weaning marine fish larvae from live prey to a dry microdiet is an important step towards optimizing the commercial production, but early weaning is constrained by the lack of sufficient digestive enzymes at first feeding. This study quantified the activity of five digestive enzymes throughout the larval period of pigfish (Orthopristis chrysoptera [L.]) to assess ontogenetic changes in digestive abilities, and then trials were conducted that determined the optimal time for weaning. The activity of all digestive enzymes was low or undetectable at first feeding (3 days post hatching, dph; 2.5 mm standard length, SL). A substantial increase in activity occurred at 5.7 mm SL (17 dph), 6.9 mm SL (21 dph), 7.7 mm SL (23 dph), 8.4 mm SL (25 dph) and 11.2 mm SL (30 dph) for bile salt‐dependent lipase, trypsin, chymotrypsin, amylase and acid protease respectively. During the weaning experiment, larvae were co‐fed live prey and microdiet beginning 15 dph (4.8 mm SL). Live prey was withdrawn from the diet at 24, 28, 32 or 36 dph, with the control receiving live prey and microdiet throughout (to 43 dph). There were no significant differences in mean final SL among treatments, but survival was significantly lower when larvae were weaned at 24 dph compared to 32–43 dph. Based on the digestive enzyme activity and survival, weaning larval pigfish at 32 dph (11.7 mm SL) when reared at 24°C is recommended.     

Effects of the timing of initial feeding on growth and survival of loach (<Emphasis Type="Italic">Misgurnus anguillicaudatus</Emphasis>) larvae

The effects of delayed first feeding on growth and survival and starvation on the point-of-no-return of loach Misgurnus anguillicaudatus larvae were studied by evaluating morphometric characteristics under controlled conditions. Larvae began to feed exogenously at 3 days after hatching (DAH) and the PNR occurred between 9 and 10 DAH at 23 ± 1.0°C. The experimental design included a conventional feeding regime with initial feeding from 3 DAH as a control, delayed first feeding for 4, 5 and 6 DAH. Morphometric characteristics (head depth, body depth, eye diameter, mouth diameter, musculature height, total length and yolk sac volume) were evaluated under different initial feeding time (3, 4, 5 and 6 days after hatching). Loach larvae initiated first feeding at 4, 5 and 6 days after hatching achieved comparatively lesser growth performance in all morphometric characteristics than that of 3 days at the end of the experiment. By day 6, significant differences were observed between 3 and 6 days initial feeding larvae for all morphometric characteristics except eye diameter and mouth diameter. Similarly, significant differences were noticed between 3 and 5 days initial feeding. However, there were no significant differences in head depth, body depth, eye diameter, mouth diameter, and total length between 3 and 4 days initial feeding until 12 DAH. After 15 days rearing, significant differences in all morphometric characteristics appeared between 3 and 4 days initial feeding and followed to the end of the experiment. It was also observed that the yolk absorption in loach larvae was completed by 6 days irrespective of the differences in the initial feeding. The yolk volume of 4 and 5 DAH larvae initiated first feeding at 3 days (0.0125 ± 0.0015; 0.0077 ± 0.0009 mm³) had significant differences compared with yolk volume of larvae initiated first feeding at 4 days (0.0081 ± 0.0011; 0.0039 ± 0.0004 mm³), 5 days (0.0079 ± 0.0010; 0.0017 ± 0.0002 mm³) and 6 days (0.0082 ± 0.0011; 0.0016 ± 0.0001 mm³). Survival rates of four treatments were estimated daily for 30 days and significant differences were observed between the treatments at the end of the experiment. The final survival rate was higher when the loach larvae initiated feeding at 3 days (75.9%) when compared with 4 days (31.8%), 5 days (14.5%) and 6 days (6.4%). The present study suggests that the first feeding of loach larvae should be initiated at 3 DAH for achieving better growth and survival or else bad growth performance will engender if the first feeding is delayed.     

Growth and survival of grouper <Emphasis Type="Italic">Epinephelus</Emphasis><Emphasis Type="Italic">coioides</Emphasis> (Hamilton) larvae fed free-living nematode <Emphasis Type="Italic">Panagrellus redivivus</Emphasis> at first feeding

The free-living nematode, Panagrellus redivivus, was tested as live food for grouper Epinephelus coioides larvae during the first feeding stage. A series of experiments were conducted to determine the acceptability of the free-living nematodes in grouper larvae at first feeding, the optimum nematode density and the response of the larvae to nutritionally enriched nematode. All experiments were conducted in 200-L conical tanks filled with 150-L filtered seawater and stocked at 15 larvae L⁻¹. Duration of feeding experiments was up to day 21 (experiment 1) and 14 days (experiment 2 and 3). Brachionus plicatilis and Artemia (experiment 1) and Brachionus plicatilis alone (experiment 2 & 3) was used as the control treatment. Observations indicated that the grouper larvae readily fed on free-living nematodes as early as 3 days posthatching, the start of exogenous feeding. Optimum feeding density for the larvae was 75 nematodes ml⁻¹. The enrichment of cod liver oil or sunflower oil influenced the total lipids and n-3 highly unsaturated fatty acids of P. redivivus, which in turn influenced those of the grouper larvae, however, growth and survival of the larvae were not affected (P > 0.05). The results from this investigation showed that the nematode, P. redivivus, can be used as first live food for grouper larvae from the onset of exogenous feeding until they could feed on Artemia nauplii.     

饲料维生素E含量对美国红鱼生长和非特异性免疫的影响

采用单因子实验设计方法,进行了饲料中添加维生素E对美国红鱼生长（存活率、特定生长率和饲料效率）和非特异性免疫相关指标（血清中溶菌酶活性、血清总补体活性和超氧化物岐化酶（SOD）活性）影响的研究,设计了5个不同维生素E水平（0．1、25．3、51．0、102．8和203．2mg／kg）的等氮等能饲料,对美国红鱼进行为期56d的生长实验,每个水平3个重复,每个养殖单元放养初始体重为151g左右的美国红鱼20尾。饲养实验在海水网箱（1．5m×1．0m×1．0m）中进行。结果表明,随着饲料维生素E含量增加,美国红鱼饲料效率、特定生长率和成活率显著提高,美国红鱼最佳生长性能的饲料维生素E添加量为102．8mg／kg;美国红鱼血清溶菌酶和血清总补体活性随着维生素E的添加量的增加而显著升高,并且维生素E的添加量达到最高值102．8和25．3mg／kg,随后变化不明显;美国红鱼血清SOD活性随维生素E添加量的增加而显著升高,并且含量在102．8mg／kg达到最高值,随后显著下降。综上所述,美国红鱼维生素E的适宜添加量为102．8mg／kg。     

浙江自然海区逃逸美国红鱼食性研究

通过对浙江自然海区逃逸美国红鱼胃含物的分析,从食料生物组成、食性类型、摄食强度及营养级4个方面对浙江自然海区逃逸美国红鱼的食性进行了探讨。结果表明,美国红鱼属游泳动物食性。主要摄食鱼类、长尾类、短尾类和头足类,兼食口足类和多毛类。其食料生物组成和食性类型与原栖息地有较大差异。逃逸美国红鱼的摄食率为80.28%,最高饱满度系数达60.93‰,平均10.26‰。根据计算,其饵料的生境宽度为4.09,营养级为3.43级。浙江自然海区美国红鱼是一种凶猛的中级肉食性鱼类。     

Histological evaluation of the elimination of Artemia nauplii from larval rearing protocols on the digestive system ontogeny of shi drum (Umbrina cirrosa L.)

The influence of the absence of Artemia nauplii from larval diet protocols on growth and digestive system ontogeny was studied using histological techniques in the shi drum (Umbrina cirrosa). One group of larvae was reared using the standard intensive rearing protocol, which offers a combination of enriched rotifers (Brachionus plicatilis), Artemia spp. nauplii and artificial diet (Std-group). Another group was reared using the same protocol, but without the offering of Artemia nauplii (group No-Artemia). The ontogenesis of the digestive system from hatching to metamorphosis was a very rapid process, and there were no differences between the two feeding regimes in the temporal appearance of the various components of the digestive system. The first organised presence of the hepatic and pancreatic tissue appeared at 2–3 d after hatching (dah), suggesting that these organs function from a very early developmental stage. In the No-Artemia larvae between 13 and 29 dah there was a reduction in the height of enterocytes in the intestinal mucosa, a progressive flattening of the primary intestinal folds in the anterior and posterior intestine and a decrease in lipid stores in the liver, suggesting a period of relative starvation. However, by the end of the study at 41 dah, there were no significant differences in body length, intestinal morphology or liver lipid stores between larvae reared under the two feeding regimes. The study suggests that the diet may influence the maturation and/or function, but not the ontogeny of the digestive system. Furthermore, the rapid differentiation of the digestive system in shi drum and the prompt recovery of the No-Artemia larvae from the symptoms of starvation by 29 dah, indicate a plasticity during ontogenesis and the ability of larvae to adapt to artificial diets at very early developmental stages.     

Phenotypic plasticity in gene expression and physiological response in red drum <Emphasis Type="Italic">Sciaenops ocellatus</Emphasis> exposed to a long-term freshwater environment

Red drum (Sciaenops ocellatus) is a euryhaline fish commonly found in the Gulf of Mexico and along the Atlantic coast of North America. Because of high commercial demand and its euryhaline characteristics, aquaculture of this species has diversified from marine to low-salinity aquaculture systems. In recent years, interest in the feasibility of producing red drum in inland freshwater systems has grown and this prompted us to investigate its osmoregulatory capacity after rearing for 8 months in a freshwater aquaculture system. We compared the activities of several genes and enzymes involved in the osmoregulatory process in freshwater-acclimatized (FW) and seawater (SW) red drum. The gene expression profiles were variable: the expression of genes encoding Na⁺/K⁺-ATPase (NKA) and the cystic fibrosis transmembrane regulator (CFTR) was slightly higher in SW than FW fish, while phosphoenolpyruvate carboxykinase (PEPCK) and the glucocorticoid receptor messenger RNA (mRNA) levels were higher in FW red drum. The total plasma K concentration was 60.3% lower, and gill NKA activity was 63.5% lower in FW than in SW fish. PEPCK activity was twofold higher in FW than in SW red drum. Similarly, liver glycogen was 60% higher in FW fish. In summary, both gene expression and the enzyme activity data support the phenotypic plasticity of red drum and suggest that the limited capacity for ion homeostasis observed, in particular the low plasma K concentration, was due to the composition of freshwater and does not necessarily reflect a physiological inability to osmoregulate.     

Dietary Lysine Requirement of Juvenile Red Drum Sciaenops ocellatus

The dietary lysine requirement of juvenile red drum Sciaenops ocellatus was reevaluated in two separate experiments. In the first experiment, lyophilized red drum muscle was combined with an L-form crystalline amino acid premix to yield diets containing approximately 35% protein on a dry-matter basis. Gradations (0.25%) of L-lysine HCl were added to the basal diet containing approximately 1.0% lysine. Each diet was fed to triplicate groups of juvenile red drum initially weighing 6–7 g for 8 weeks. Based on growth and feed efficiency data, the lysine requirement (±SE) was determined to be 1.55% (±0.079%) of dry diet or 4.430% of dietary protein. In a second experiment, the lysine requirement of red drum was investigated with diets containing 35% intact protein from zein and red drum muscle. Each of the zein-based diets containing incremental levels of lysine was fed to triplicate groups of juvenile red drum initially weighing 1–2 g for 8 weeks. A crystalline amino acid test diet serving as a control in this experiment significantly outperformed the zein-based diets, and palatability of those diets was questionable due to an obvious change in feeding behavior of the fish. Due to the inferior performance of the fish fed the zein-based diets, the authors were not successful in determining a requirement with these diets. Thus, a lysine requirement of 1.55% of dry diet (4.43% of dietary protein) as determined in the first experiment is recommended for juvenile red drum.     

Organogenesis of exocrine pancreas in sharpsnout sea bream (<Emphasis Type="Italic">Diplodus puntazzo</Emphasis>) larvae: characterization of trypsin expression

The ontogeny and differentiation stages of digestive systems related with trypsin expression in larvae of sharpsnout sea bream, Diplodus puntazzo, were investigated from hatching to 40 DAH (days after hatching), and total lengths and weights of larvae were determined. Histologic and enzymatic techniques were used to explain the functional development of the pancreas including trypsin activity. The pancreas was identified as a compact structure located in the region slightly posterior to the liver. At 3 DAH, first anus and then mouth opened. Incipient pancreas secretion polyhedral cells could be first observed as zymogen granules. During larval metamorphosis, the pancreas became diffuse, spreading throughout the mesentery in proximity to the stomach, the anterior intestine and the pyloric caeca. The specific activity of trypsin (42.54 ± 6.8 mU/mg protein⁻¹) was found as early as after hatching at larvae size of 2.87 ± 0.34 mm at 0 DAH. Activity further increased until 10 DAH, especially after exogenous feeding. The highest trypsin activity was detected at 25 DAH as 119.26 ± 11.6 mU/mg protein⁻¹. It is concluded that exocrine pancreas organogenesis is the main critical step in the development of digestive system that results in zymogen granules accumulation and increased trypsin activity.     

First feeding parameters of the milletseed butterflyfish Chaetodon miliaris

First feeding performance of the milletseed butterflyfish (Chaetodon miliaris) was evaluated using three experiments examining prey type, microalgal cell density and prey density. To ensure no size bias existed between trials, morphometric measurements, fertilization and hatch rates were recorded for each trial. The proportion of larvae feeding and feeding intensity were measured in each experiment. To evaluate prey type, larvae were offered small size Brachionus plicatilis and Parvocalanus crassirostris nauplii in clear water and water greened with Tisochrysis lutea (Tiso). In trial 1, green water and P. crassirostris nauplii had a significantly higher proportion of larvae feeding and feeding intensity. In trial 2 no differences were detected between P. crassirostris treatments. Microalgae experiments evaluated six treatments ranging from 0 cells/ml to 8 × 10⁵ cells/ml. Feeding response increased as microalgae was added to the environment, but no statistical differences were discerned between treatments besides the highest cell density (8 × 10⁵ cells/ml) and lowest cell density in both trials (0 cells/ml). To determine if prey density had an effect on first feeding response P. crassirostris nauplii were added to tanks at 1 to 15 nauplii/ml. No significant differences in the feeding response were found between prey densities of 1–15 P. crassirostris nauplii/ml in either trial. This study revealed that C. miliaris larvae had the highest first feeding response, measured by the proportion of larvae feeding and feeding intensity per larvae, with the prey item P. crassirostris nauplii in green water and prey densities between 1 and 15 nauplii/ml.