cDNA cloning and expression of a novel group IB phospholipase A2 in the intestine of the red sea bream,Pagrus (Chrysophrys) major

A cDNA encoding a novel phospholipase A₂ (PLA₂), which was named IN PLA₂, was cloned from the intestine of the red sea bream. The amino acid sequence of IN PLA₂ showed 49–75% homology with those of red sea bream group IB sPLA₂, hepatopancreas DE-1 and DE-2 PLA₂, and gill G-3 PLA₂. IN PLA₂ consists of a prepropeptide of 24 amino acid residues, followed by a mature protein. IN PLA₂ contains 14 cysteines, and includes Cys11, the calcium binding loop and the pancreatic loop that are commonly conserved in group IB sPLA₂ enzymes. In addition, IN PLA₂ is a cationic protein with a pI of 8.52. Therefore, IN PLA₂ was identified as a novel group IB sPLA₂ isoform in red sea bream. IN PLA₂ mRNA was found by northern blot analysis to be expressed mainly in the pyloric caeca and the intestine, and was detected in the goblet cells of the intestine by in situ hybridization. The expression level of IN PLA₂ mRNA was elevated by intravenous injection of lipopolysaccharide—the outer-membrane component of Gram-negative bacteria. These results suggest that IN PLA₂ is secreted from the goblet cells of the intestine in response to stimulus such as bacterial infection, and that it contributes to antimicrobial defense in addition to the digestion of dietary phospholipids in the gastrointestinal tract.     

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.     

Purification and characterization of phospholipase A2 from the pyloric caeca of red sea bream, Pagrus major

A phospholipase A₂ was purified 55,000-fold in a yield of 10% from the lipid-free extract of powder of the pyloric caeca of red sea bream to near homogeneity by sequential column chromatography on S-sepharose fast flow, butyl-cellulofine, Asahipak ES-502C cation-exchange HPLC, TSK gel G3000SW gel-filtration HPLC, and Asahipak ODP-50 reversed-phase HPLC. The final preparation showed a single band with the apparent molecular mass of 14 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and an estimated specific activity was 717 µmol min^-1 mg^-1 protein. The purified enzyme had a pH optimum in the range of pH 8.0–9.0 and required the presence of both 8 mM of Ca²⁺ and from 2 to 10 mM of sodium deoxycholate for its maximal activity, using 2 mM of phosphatidylcholine as a substrate. The purified enzyme preferentially hydrolyzed the 2-acyl ester bonds of both phosphatidylglycerol and phosphatidylcholine in the presence of sodium deoxycholate, followed in order by phosphatidylethanolamine and phosphatidyl-serine. In contrast to porcine pancreatic PLA₂, pyloric caeca PLA₂ hydrolyzed mixed-micellar phosphatidylcholine substrate effectively, regardless of the kinds of bile salts used. These results indicate that Ca²⁺-dependent low molecular mass PLA₂, so called secretory PLA₂, occurs in the pyloric caeca of red sea beam.     

Experimental infection of Atlantic halibut, Hippoglossus hippoglossus L., yolk-sac larvae with infectious pancreatic necrosis virus: detection of virus by immunohistochemistry and in situ hybridization

Three different concentrations (10⁷, 10⁵ and 10³ TCID₅₀ ml^-1) of infectious pancreatic necrosis virus (IPNV) serotype Sp isolated from Atlantic halibut, Hippoglossus hippoglossus L., were used to bath-challenge Atlantic halibut yolk-sac larvae. The larvae challenged with 10⁷ TCID₅₀ ml^-1 suffered significantly higher cumulative mortality than the other challenged groups and the control group, and affected individuals displayed necrosis of the intestine, liver and kidney. In larvae from the groups challenged with 10⁷ and 10⁵ TCID₅₀ ml^-1, IPNV was detected by immunohistochemistry and in situ RNA/DNA hybridization in the intestine, liver and kidney. In addition, some individuals stained IPNV-positive in the heart and eye/brain region. Detection by in situ hybridization did not appear to be more sensitive than immunohistochemistry. However, background staining was virtually absent in comparison with immunohistochemistry, and the staining seemed to be more distinctly localized to the cytoplasm of infected cells. The results show that farmed halibut yolk-sac larvae can be infected by IPNV immediately after hatching, with resulting high mortality. As the larvae are not immunologically mature at this stage of development, vaccination is not recommended.     

Experimental infection of Atlantic halibut,Hippoglossus hippoglossus L., yolk-sac larvae with infectious pancreatic necrosis virus: detection of virus by immunohistochemistry and in situ hybridization*

Three different concentrations (10⁷, 10⁵ and 10³ TCID₅₀ ml^?1) of infectious pancreatic necrosis virus (IPNV) serotype Sp isolated from Atlantic halibut, Hippoglossus hippoglossus L., were used to bath-challenge Atlantic halibut yolk-sac larvae. The larvae challenged with 10⁷ TCID₅₀ ml^?1 suffered significantly higher cumulative mortality than the other challenged groups and the control group, and affected individuals displayed necrosis of the intestine, liver and kidney. In larvae from the groups challenged with 10⁷ and 10⁵ TCID₅₀ ml^?1, IPNV was detected by immunohistochemistry and in situ RNA/DNA hybridization in the intestine, liver and kidney. In addition, some individuals stained IPNV-positive in the heart and eye/ brain region. Detection by in situ hybridization did not appear to be more sensitive than immunohistochemistry. However, background staining was virtually absent in comparison with immunohistochemistry, and the staining seemed to be more distinctly localized to the cytoplasm of infected cells. The results show that farmed halibut yolk-sac larvae can be infected by IPNV immediately after hatching, with resulting high mortality. As the larvae are not immunologically mature at this stage of development, vaccination is not recommended.     

Morphological changes of digestive structures in starved tench Tinca tinca (L.) juveniles

The aim of the study was to evaluate the effect of short-term starvation on the morphology of the digestive system of juvenile tench Tinca tinca (L.). The morphological changes in digestive structures of juveniles, fed 70 days starter or Chironomidae larvae, were examined after 1, 12, 24 and 48 h of starvation at 28 °C. The morphological changes in the organisation of the digestive structures were as follows: (1) Intestine – proteolysis of the intestinal mucosa, especially of enterocytes in apical part of mucosal folds; (2) Liver – progressive reduction of lipid vacuoles and glycogen granules, and, in effect, volume reduction of hepatocytes cytoplasm; (3) Pancreas – progressive degeneration of exocrine pancreatic cells, parallel with decrease of zymogen activity. The fish fed natural food prior to starvation seem to be more resistant to starvation than those fed starter.     

Purification and characterization of phospholipase A2 isoforms from the hepatopancreas of red sea bream, Pagrus major1

Two phospholipase A₂ (PLA₂) isoforms, tentatively denoted as DE-1 and DE-2 PLA₂s, were purified from the hepatopancreas of red sea bream (Pagrus major) to near homogeneity by sequential column chromatography on S-Sepharose fast flow, DEAE-Sepharose fast flow and butyl-Cellulofine, and by ion-exchange, gel-filtration and reversed-phase HPLC. The purified DE-1 and DE-2 PLA₂s both showed a single band with the apparent molecular mass of approx. 13.5 kDa by SDS-polyacrylamide gel electrophoresis, and were found to be both related to group I PLA₂ based on the N-terminal amino acid sequences. DE-1 PLA₂ had a pH optimum in the alkaline region at around pH 10 and required approximately 10 mM of Ca²⁺ and 4-10 mM of sodium deoxycholate for its maximal activity, using 2 mM of phosphatidylcholine and phosphatidylethanolamine as substrates. DE-2 PLA₂ also had a pH optimum in the alkaline region at around pH 8-9 and required >10 mM of Ca²⁺ and approximately 6 mM of sodium deoxycholate for its maximum activity with 2 mM of phosphatidylcholine as a substrate; its enzymatic activity towards phosphatidylethanolamine was greatly inhibited by the addition of sodium deoxycholate. The results demonstrate that red sea bream hepatopancreas contains two enzymatically distinct group I PLA₂ isoforms.     

The use of Pseudoalteromonas sp. F15 in larviculture of the Yesso scallop,Patinopecten yessoensis

A 49‐day rearing trial was conducted to investigate the effects of Pseudoalteromonas sp. F15 on growth, survival, digestive and immune‐related enzyme activities in larval and juvenile Yesso scallop, Patinopecten yessoensis. Larvae (Day 2 post‐fertilization) (d2 PF)) were provided with a diet only of microalgae (control) and supplemented with F15 at final concentrations of 1×10⁴ and 1×10⁶ cells/mL, respectively. Results indicated that the specific growth rates and survival rates of umbo larvae (d13 PF), crawling pediveliger larvae (d28 PF) and juveniles (d51 PF) fed the F15 supplementation diets at 10⁴ and/or 10⁶ cells/mL were significantly higher than those fed the diet of microalgae. Larvae and juveniles fed 10⁴ and/or 10⁶ cells/mL F15 supplementation diets showed a significant increase in pepsin, amylase and cellulase activities compared with the control group. The lysozyme, superoxide dismutase and catalase activities of larvae and juveniles fed the F15 supplementation diets at 10⁴ and/or 10⁶ cells/mL significantly increased compared to those fed the diet of microalgae. In addition, F15 protected juveniles against challenge with Vibrio splendidus. These data suggest that F15 can be useful in scallop larviculture.     

Changes in Na+,K+-ATPase activity and gill chloride cell morphology in the grouper Epinephelus coioides larvae and juveniles in response to salinity and temperature

The activity of the enzyme Na⁺,K⁺-ATPase and morphological changes of gill chloride cells in grouper, Epinephelus coioides larvae and juveniles were determined 6–48 h after abrupt transfer from ambient rearing conditions (30–32 ppt, 26.5–30 °C) to different salinity (8, 18, 32, 40 ppt) and temperature (25, 30 °C) combinations. Na⁺,K⁺-ATPase activity in day 20 larvae did not change at salinities 8–32 ppt. Activity decreased significantly (P <0.01) after exposure to 40 ppt at 25–30 °C, which was accompanied by an increase (P <0.05) in density and fractional area of chloride cells. Enzyme activity in 40 ppt did not reach a stable level and larvae failed to recover from an osmotic imbalance that produced a low survival at 25 °C and death of all larvae at 30 °C. Enzyme activity and chloride cell morphology in day 40 groupers did not change in 8–40 ppt at 25 °C and 8–32 ppt at 30 °C. A significant decrease and a subsequent increase in Na⁺,K⁺-ATPase activity in 40 ppt at 30 °C was associated with the increase in chloride cell density resulting in an increased fractional area but a decreased cell size. Enzyme activity and chloride cells of day 60 grouper were unaffected by abrupt transfer to test salinities and temperatures. These results demonstrate that grouper larvae and juveniles are efficient osmoregulators over a wide range of salinities. Salinity adaptation showed an ontogenetic shift as the larvae grew and reached the juvenile stage. This development of tolerance limits may reflect their response to actual conditions existing in the natural environment.     

Investigation of the cytotoxicity of apidaecin on intestinal epithelial cells of tilapia (Oreochromis niloticus) in vitro

Antimicrobial peptides (AMPs) are a diverse group of small polypeptides that can inhibit the growth of microbes; the investigation of their cytotoxicity is critical before they can be considered for use. The purpose of the present study was to examine the cytotoxicity of apidaecin at different concentrations (0.0, 10.0, 20.0, and 30.0 μg/ml) on tilapia, Oreochromis niloticus, primary intestinal epithelial cells in vitro through cells morphology observation, methylthiazolyldiphenyl-tetrazolium bromide (MTT) conversion assay, and cellular viability assay. The lactate dehydrogenase (LDH) activity, cytosolic free Ca²⁺ concentration, and phospholipase A₂ (PLA₂) activity of primary cells treated with/without apidaecin were also determined. No significant differences (P > 0.05) in MTT assay and viability were found between the treated and the control cells. LDH activities were slightly increased when treated with apidaecin for 24 h compared with the control (19.97 ± 1.24 U/l), but not significantly (P > 0.05). When treated with 20.0 μg/ml apidaecin for 24 h, the concentrations of cytosolic free Ca²⁺ was the highest among all the treatments (133.65 ± 9.65 nmol/l), but not the difference was not significant when compared with the control (P > 0.05). Similar results were found for the activity of cytosolic PLA₂. The current results indicate that apidaecin did not show significant cytotoxicity to primary epithelial cells within 30.0 μg/ml and that it can be used as a new candidate for infection control in fish culture.     

Ontogeny of the digestive tract and enzymes in rock bream Oplegnathus fasciatus (Temminck et Schlegel 1844) larvae

Histological development of the digestive tract and specific activities of three digestive enzymes (trypsin, alkaline phosphatase, and pepsin) were studied in rock bream Oplegnathus fasciatus from hatching to 50 days after hatching (DAH). At hatching, the digestive tract appeared as an undifferentiated straight tube and differentiated into the buccopharynx, esophagus, stomach, intestine, and rectum at mouth opening by 3 DAH. The taste bud and mandibular teeth were present in the buccopharyx at 8 DAH. The goblet cells appeared in the esophagus at 8 DAH and in the buccopharyx at 9 DAH. The stomach anlage was formed at 2 DAH and developed into cardia, fundus, and pylorus at 14 DAH. The gastric glands were visible at 16 DAH, and the pepsin was firstly detected on 22 DAH. At 2 DAH, the intestinal valve appeared and divided the intestine into anterior intestine (AI) and posterior intestine (PI). The rectum was differentiated from the PI at 3 DAH. The supranuclear vacuoles were visible in the rectum by 6 DAH, and the lipid inclusions were present in the AI at 8 DAH. The alkaline phosphatase was detected at 1 DAH, and the increase in its activity indicated the maturation of the intestine after 40 DAH. The hepatocytes and pancreatic cells were differentiated from the blast cells at 2 DAH, and the acidophilic zymogen granules in the exocrine pancreas were observed simultaneously. The trypsin was detected by 1 DAH and increased to the maximum at 19 DAH, followed by a decrease as the stomach became functional.     

FOOD AND FEEDING STRATEGIES IN CULTURE OF LARVAL AND EARLY JUVENILE PURPLE-HINGE ROCK SCALLOPS,Hinnites multirugosus (GALE)

The purple-hinge rock scallop is receiving increasing interest as a candidate for commercial mariculture. Hatchery culture of juveniles is prerequisite since natural recruitment of juveniles is variable and not dependable as a source of stock. Success in carrying larvae through metamorphosis has been limited. Critical to larval culture is a sound understanding of optimal food and feeding strategies. Growth and survival were examined in early larvae fed six algal diets at four separate concentrations. The data clearly illustrate the superiority of both Isochrysis galbana and a Tahitian (T) strain of Isochrysis. Feed concentrations of 1 × 10⁵ cells/ml were found to be optimal for these species of Isochrysis. An I. galbana diet supported 80% survival through metamorphosis. The other algae examined included Mono-chrysis lutheri, Rhodomonas sp., Carteria pallida, and a 1:1:1 mixture of Rhodomonas, Monochrysis, and Isochrysis. All these diets supported 5–6% survival, and the growth was only slightly better than starvation controls. A similar study was done with early juveniles (1 mm dia). The diets included: I. galbana, M. lutheri, Thalassiosira pseudonana (3H), T-Isochrysis sp., and Phaedactylum tricornutum. As with the larvae, superior growth and survival occurred with diets of both species of Isochrysis. Cell concentrations between 1 × 10⁵ and 2 × 10⁵ cells/ml were optimal. The other diets supported minimal growth. In the feeding runs of both larvae and early juveniles, cell densities in excess of 5 × 10⁵ cells/ml were detrimental.     

The effect of stocking density on growth,survival and agonistic behaviour of African catfish

A high rate of sibling cannibalism is one of the principal obstacles in the rearing of larvae and juveniles of the African catfish Clarias gariepinus. This paper examines the underlying behavioural components of agonistic behaviour under culture conditions. Ten day old catfish larvae were stocked at 30 larvae l^–1 in three tank designs with different surface areas and equal volumes. Stocking densities were 1.2, 0.6 and 0.3 fish cm^–2 bottom surface. Growth did not differ between treatments. The highest mortality over 30 days was recorded at the medium density (0.6 larvae l^–1). Fish stocked at this density showed the highest rate of aggression, while there was no difference in aggression between the highest and the lowest stocking densities. The results indicate that stocking density should be at least as high as 1.2 larvae cm^–2 bottom surface area to obtain high production at best survival rates.     

Rearing of pike-perch larvae using formulated diets – first success with starter feeds

The present study evaluated the performance of two commercial diets: AgloNorse (AN) and BioKyowa (BK), and two experimental, formulated diets based on casein (C) or casein plus casein hydrolysates (CH) in rearing of pike‐perch larvae (Sander lucioperca L.). All fish were 5 day old and control group was fed live Artemia nauplii. Fish were sampled periodically for histological comparison of morphological changes in the digestive tracts. Survival of fish fed Artemia nauplii, BK and AN was similar: 54.4%, 50.8% and 52.4%, respectively, while the fish fed formulated diets C and CH showed considerably lower survival: 28.4% and 21.6% respectively. After 5 weeks of rearing, the average body mass of fish ranged from 212±32 mg in Artemia fed group to 53.8±6.8 mg in C diet fed group. A considerable vacuolization of supranuclear zone in enterocytes of posterior intestine was observed in the larvae fed commercial diets. No anomalies in liver development were found. Hepatocytes of fish fed BK diet showed larger glycogen storage areas, compared with those occupied by lipids. The highest zymogen accumulation of pancreatic cytoplasm was observed in fish fed Artemia. In fish fed C and CH diets, anomalies in digestive system development were indicated by lower and less numerous intestinal folds, smaller hepatocytes, retarded development of gastric glands, and in CH group – also local fatty degeneration of liver.     

Effect of environmental calcium levels on calcium uptake in tilapia larvae Oreochromis mossambicus

Effects of environmental calcium concentrations on the survival, growth, body calcium content and calcium uptake kinetics in developing tilapia (Oreochromis mossambicus) larvae were studied. Fertilized eggs were incubated in high- and low-calcium artificial freshwater (0.88–0.96 mmol l^–1 vs. 0.02–0.03 mmol l^–1 CaCl₂ or CaSO₄) until 3 days after hatching. Tilapia larvae showed similar hatching rates and wet weights in either high- or low-calcium medium, indicating neither the development nor the growth in tilapia larvae was affected by the environmental calcium levels. The body calcium content in low-calcium groups was about 90–95% that of high-calcium groups, No matter what calcium source was used (CaCl₂ or CaSO₄), acclimation to low calcium medium caused a stimulation of calcium uptake (measured in 0.2 mmol l^–1 calcium),i.e., 1.2–1.3 fold higher than that of high calcium groups. This enhanced calcium uptake capacity was characterized by a 50% decrease in K_m and a 25% increase in J_max. Effect of different calcium salts on calcium influx was significant only in low calcium level,i.e., calcium influx in low-CaCl₂ group higher than that in low-CaSO₄ group. These results suggest that tilapia larvae are able to modulate their calcium uptake mechanism to maintain normal body calcium content and growth in environments with different levels of calcium.     

Amyloid associated with neoplasia in two captive tricolour sharkminnows Balantiocheilus melanopterus Bleeker

Amyloid associated with pancreatic adenocarcinoma was discovered in two captive adult tricolour sharkminnows Balantiocheilus melanopterus Bleeker found dead in a freshwater display. Enlarged abdomens expanded by bloody ascitic fluid and grossly visible masses of abnormal tissue were present surrounding sections of the stomach and intestine. Histologically, the masses were composed of areas of well‐organized exocrine pancreatic acini interspersed with cords of poorly differentiated, spindle‐shaped cells that compressed and effaced normal parenchyma. These cells possessed small numbers of cytoplasmic zymogen granules; the exocrine nature of these cells was confirmed using transmission electron microscopy (TEM). Fibrovascular connective tissue of the hepatopancreas and mesenteries was expanded by lightly eosinophilic, hyaline, homogeneous acellular material. Similar material greatly expanded the tunica media of large blood vessels in the hepatopancreas. After staining with Congo red or thioflavin T, this material exhibited red–green dichroism under polarized light or bright green fluorescence under ultraviolet light (255 nm), respectively. The non‐branching fibrils, of indeterminate length, had an approximate diameter of 10–20 nm using TEM. Although exocrine pancreatic neoplasia is relatively common in fish, the presence of amyloid is not. To our current knowledge, the latter has not yet been described in association with a neoplastic lesion in fish.     

Formation of Biogenic Amines and Associated Biochemical and Microbial Attributes of Whole Sutchi Catfish (Pangasianodon hypophthalmus) during Iced Storage

Sutchi catfish (Pangasianodon hypophthalmus) is a highly preferred farmed species that is produced in huge quantities. Meat from ice-stored (4 ± 2°C) whole Sutchi catfish was evaluated for formation of biogenic amines, such as putrescine, cadaverine, histamine (HIM), agmatine, tyramine, spermine, and spermidine, and compared with biochemical, microbial, and sensory attributes during 22 days. Analysis of content of biogenic amines in the meat was carried out by liquid chromatography with tandem mass spectrometry using a without derivatization method. Three amines, namely, tyramine, spermidine, and spermine, were only present on 0^th day of storage. Presence of cadaverine was noticed from 9^th day onwards, where the aerobic plate count (APC) reached 4.85 log CFU/g. Putrescine was detected on the 22^nd day of storage, where the APC crossed 7 log CFU/g. HIM was detected in lower quantities from 3^rd day onwards. A shelf life of 15 days was determined based on sensory and microbiological evaluation. Although the samples were biochemically acceptable throughout the storage period, APC exceeded the limit on day 19, and the gradual increase of H₂S-producing bacteria, Brochothrix thermosphacta, Aeromonas, and Enterobacteriaceae, was observed during storage.     

不同月龄缢蛏新品种数量性状的相关与通径分析

为了检测缢蛏新品种"申浙1号"不同月龄数量性状间的相关性。随机选取4、9、11月龄缢蛏个体60个以上,测量其壳长(SL)、壳高(SH)、壳宽(SW)、活体质量(W),采用相关与通径分析方法分析壳形态性状对活体质量的作用效果。结果显示,各数量性状之间的相关关系均达到极显著水平,其中与4、11月龄质量相关系数最大的是壳长,分别为0.911、0.631,与9月龄活体质量相关系数最大的是壳高,为0.653。通径分析结果显示,对4、9、11月龄活体质量的直接影响最大的分别是壳长(0.562)、壳宽(0.433)、壳高(0.394),决定系数与以上通径分析结果的变化趋势一致。利用逐步回归的方法分别建立4、9、11月龄缢蛏活体质量(W)的最优回归方程:W4=–0.756+0.039SL+0.112SW,R2=0.849;W9=–10.622+0.095SL+0.404SH+0.644SW,R2=0.631;W11=–23.772+0.207SL+0.805SH+0.587SW,R2=0.909。研究结果为缢蛏人工选育过程中的亲贝选择和产量预测等提供了基础数据。     

Mortality,hematology, and histopathology of common snook Centropomus undecimalis (Perciformes: Centropomidae) exposed to acute toxicity of ammonia

This study aimed to evaluate the tolerance of common snook Centropomus undecimalis larvae and juveniles exposed to acute concentrations of un-ionized ammonia for 96 h at 35g L^?1 salinity, after 24 h starvation. For that, 10 larvae (20.85 ± 1.46 mm) of 47 days post hatch (DPH) per experimental unit (1.5 L) were exposed to 0.00 ± 0.00, 0.65 ± 0.04, 1.29 ± 0.09, 2.59 ± 0.18, 3.88 ± 0.27, 5.17 ± 0.34, and 6.47 ± 0.43 mg L^?1 NH₃, in triplicates, at 26.72 ± 0.08°C, dissolved oxygen at 5.72 ± 0.10 mg L^?1 and pH 8.45 ± 0.06. During this period, no mortalities were observed. Another trial was performed with five juveniles (20.35 ± 6.10 g, 13.90 ± 1.75 cm) per experimental unit (60 L) exposed to 0.00 ± 0.00, 2.26 ± 0.07, 2.68 ± 0.11, 3.20 ± 0.13, 3.68 ± 0.17, and 4.27 ± 0.16 mg L^?1 NH₃, in triplicates, at 21.90 ± 0.76°C, dissolved oxygen at 6.27 ± 0.21 mg L^?1 and pH at 8.38 ± 0.04. Fish mortality increased as ammonia concentrations increased at each day, and LC₅₀ 96 h was 3.52 mg L^?1 NH₃. Larvae were less sensitive than juveniles, demonstrating that the environmental toxicity of ammonia to common snook is influenced by age. Sublethal exposition to ammonia caused histological damages in gills of common snook juveniles and variation on glucose levels, hematocrit, and red blood cells number, showing negative effects on fish homeostasis. Moreover, compared to other species, the common snook has great resistance to ammonia.     

Enhancing growth and resistance to Vibrio alginolyticus disease in catarina scallop (Argopecten ventricosus) with Bacillus and Lactobacillus probiotic strains during early development

In bivalve aquaculture, selecting suitable probiotic treatments can be crucial for improving hatchery‐rearing of larvae and juveniles. We assessed the potential of five bacterial strains, previously selected in vitro, to improve survival, growth and resistance of catarina scallop Argopecten ventricosus during early and late larval and juvenile developmental stages, as well as during exposure to the pathogen Vibrio alginolyticus. Hatchery‐reared larvae and juveniles were treated with eight treatments of single or combined strains of Bacillus and Lactobacillus at 1 × 10⁶ CFU mL^?1 every 48 h for 9 days (larvae) and 21 days (juveniles). Compared with the control, significantly higher survival and growth in size and weight of early veliger larvae occurred with the antibiotic and the RL5 (Lactobacillus graminis) treatments. Significantly enhanced settlement of pediveliger larvae occurred with a different probiotic strain, the mix of Lactobacillus and Bacillus (MIX‐LB), while higher survival and growth of early juveniles occurred with C3 (Lactobacillus plantarum). The mix of Bacillus (MIX‐B) significantly increased survival of juveniles from V. alginolyticus after 120‐h infection, consistent with maximum activity of superoxide dismutase enzyme. In contrast, all untreated and infected scallops died by 96 h. The three Bacillus strains performed poorly when used as single treatments and when given to early developing larvae. Our results indicate that the action mechanism of probiotic strains is stage specific and strain specific, generating different responses by the host, including improved survival and growth (likely from better nutrient assimilation) and higher resistance against pathogens (possibly from strengthening the immune system).