Quantitative dietary leucine requirement of juvenile Pacific white shrimp,Litopenaeus vannamei (Boone) reared in low‐salinity water

The experiment was conducted to determine the leucine requirement of juvenile Pacific white shrimp Litopenaeus vannamei (Boone) in low‐salinity water (0.50–1.20 g L^?1). Six diets were formulated to contain 410 g kg^?1 crude protein with fish meal, peanut meal and precoated crystalline amino acids with different concentration of l ‐leucine (16.72, 19.60, 22.06, 24.79, 27.28 and 30.16 g kg^?1 dry diet). Each diet was randomly assigned to triplicate groups of 30 shrimps (0.38 ± 0.002 g), and the feed trial lasted for 8 weeks. The results indicated that the maximum weight gain was observed at 24.95 g kg^?1 dietary leucine group, whereas the diets containing higher leucine concentration conversely reduced the growth performance (P < 0.05). Moreover, the highest body protein content and body protein deposition and the lowest haemolymph AST and ALT activities were also found at 24.95 g kg^?1 dietary leucine group. With the increase in leucine in diets, a dose‐dependent increase was found in body lipid content and haemolymph urea concentration. The polynomial regression calculated using weight gain, feed efficiency and body protein deposition indicated that the optimal dietary leucine requirement for L. vannamei reared in low‐salinity water was 23.73 g kg^?1 leucine of dry diet, correspondingly 57.88 g kg^?1 of dietary protein.     

Protein‐sparing effect of carbohydrate in the diet of white shrimp Litopenaeus vannamei at low salinity

Juvenile Litopenaeus vannamei were fed for 8 weeks with diets containing four ratios of protein to carbohydrate (CBH) at P26 : C30, P30 : C25, P34 : C19 and P38 : C14, respectively, at 3.0 g L^?1 salinity. Shrimp weight gain of P34 : C19 group was the highest and differed from the shrimp fed the P26 : C30 or P30 : C25 diet. Shrimp fed the P26 : C30 diet obtained higher survival than those fed other diets. Shrimp fed the P34 : C19 diet contained the highest body protein and lipid, which were significantly higher than those fed the P38 : C14 diet. Shrimp fed the P30 : C25 diet had the highest haemolymph glucose content, which was significantly higher than those fed the P26 : C30 or P38 : C14 diet. Shrimp muscle glycogen of the P26 : C30 group was the highest. Hepatopancreas B‐cell number of shrimp fed the P26 : C30 diet was lower than those fed other diets, and the R cell number was the highest in the shrimp fed the P30 : C25 diet. This study indicates that the protein‐sparing effect by CBH occurred in the P30 : C25 and P34 : C19 groups because these proteins to CBH ratios can support normal growth. Within the range of basic energy demand, the high dietary CBH to protein ratio can improve L. vannamei survival at low salinity.     

Influence of variation in water temperature on survival,growth and yield of Pacific white shrimp Litopenaeus vannamei in inland ponds for low‐salinity culture

There is considerable interest in the culture of whiteleg shrimp (Litopenaeus vannamei) in inland low‐salinity water in Alabama and other states in the Sunbelt region of the US. However, the growing season is truncated as compared with tropical or subtropical areas where this species is typically cultured, and temperature is thought to be a major factor influencing shrimp production in the US. This study, conducted at Greene Prairie Aquafarm located in west‐central Alabama, considered water temperature patterns on a shrimp farm in different ponds and different years; and sought possible effects of bottom water temperature in ponds on variation in shrimp survival, growth and production. Water temperature at 1.2 m depth in 22 ponds and air temperature were monitored at 1‐hr intervals during the 2012, 2013, 2014 and 2015 growing seasons. Records of stocking rates, survival rates and production were provided by the farm owner. Correlation analysis and linear mixed model analysis of variance were used. Results showed that hourly water temperatures differed among ponds. The range of water temperature in each pond explained 41% of the variance in average final weight of shrimp harvested from each pond. In conclusion, the results suggest that variation in water temperature patterns has considerable influence on shrimp growth and survival in ponds.     

Supplementation of sodium chloride in diets to improve the meat quality of Pacific white shrimp,Litopenaeus vannamei,reared in low‐salinity water

To investigate the effect of dietary sodium chloride (NaCl) on meat quality of white shrimp (Litopenaeus vannamei) reared in low‐salinity (2 g L^?1) water, shrimp were distributed into four groups (treatments T‐1, T‐2, T‐3 and control) with three replicates. All shrimps were completely randomised stocked into 12 tanks at an initial density of 40 shrimps per tank. Diets for the control, T‐1, T‐2 and T‐3 groups consisted of the basal diet supplemented with 0 g kg^?1, 10 g kg^?1, 20 g kg^?1 and 40 g kg^?1 of NaCl respectively. After 50 days, shrimps in T‐3 showed significantly better (P < 0.05) moisture, crude protein and ash than those of the control and T‐1. Higher muscle Na content was observed (P < 0.05) in T‐3 than that of the control. Significant increases (P < 0.05) in contents of inosinic acid, total free amino acid (TFAA) and essential free amino acid (EFAA) were also found in T‐3. Texture assays showed significant differences (P < 0.05) in hardness, adhesiveness and springiness between group T‐3 as compared with those of T‐1 and control. It indicated that dietary supplementation of NaCl appeared to be a promising practice to improve meat quality of white shrimp reared in low‐salinity waters.     

White spot syndrome virus epizootic in cultured Pacific white shrimp Litopenaeus vannamei (Boone) in Taiwan

White spot syndrome virus (WSSV) has caused significant losses in shrimp farms worldwide. Between 2004 and 2006, Pacific white shrimp Litopenaeus vannamei (Boone) were collected from 220 farms in Taiwan to determine the prevalence and impact of WSSV infection on the shrimp farm industry. Polymerase chain reaction (PCR) analysis detected WSSV in shrimp from 26% of farms. Juvenile shrimp farms had the highest infection levels (38%; 19/50 farms) and brooder shrimp farms had the lowest (5%; one of 20 farms). The average extent of infection at each farm was as follows for WSSV‐positive farms: post‐larvae farms, 71%; juvenile farms, 61%; subadult farms, 62%; adult farms, 49%; and brooder farms, 40%. Characteristic white spots, hypertrophied nuclei and basophilic viral inclusion bodies were found in the epithelia of gills and tail fans, appendages, cephalothorax and hepatopancreas, and virions of WSSV were observed. Of shrimp that had WSSV lesions, 100% had lesions on the cephalothorax, 96% in gills and tail fans, 91% on appendages and 17% in the hepatopancreas. WSSV was also detected in copepoda and crustaceans from the shrimp farms. Sequence comparison using the pms146 gene fragment of WSSV showed that isolates from the farms had 99.7–100% nucleotide sequence identity with four strains in the GenBank database – China ( AF332093 ), Taiwan ( AF440570 and U50923 ) and Thailand ( AF369029 ). This is the first broad study of WSSV infection in L. vannamei in Taiwan.     

Histological alterations in gills of shrimp Litopenaeus vannamei in low‐salinity waters under different stocking densities: Potential relationship with nitrogen compounds

Two experimental modules with different stocking densities (M1 = 70 and M2 = 120 shrimp /m²) were examined weekly over a culture cycle in tanks with low‐salinity water (1.9 g/L) and zero water exchange. Results showed survival rates of 87.7 and 11.9% in M1 and M2, respectively. Water temperature, pH, dissolved oxygen, electrical conductivity and chlorophyll a were not significantly (p > .05) different between modules. In contrast, the concentrations of nitrogen compounds were significantly (p < .05) different between modules, except nitrite‐N (M2 were 2.31 ± 1.38 mg/L N‐TAN, 0.18 ± 0.49 mg/L N‐NO₂^? and 6.83 ± 6.52 mg/L N‐NO₃^?; in M1: 0.97 ± 0.73 mg/L N‐TAN, 0.05 ± 0.21 mg/L N‐NO₂^? and 0.63 ± 0.70 mg/L N‐NO₃^?). When waters of both modules reached higher levels of ammonia and nitrite, histological alterations were observed in gills. The histological alterations index (HAI) was higher in M2 (5‐112) than in M1 (2‐22).     

Effects of aqueous Na/K and dietary K on growth and physiological characters of the Pacific white shrimp,Litopenaeus vannamei,reared in low‐salt well water

The synchronous effects of aqueous Na/K and dietary potassium (K⁺) on growth and physiological characters was studied on the Pacific white shrimp (Litopenaeus vannamei) reared in low‐salinity well water (4 ppt) for 8 weeks with initial weight of 0.28 ± 0.01 g. Three practical diets were formulated with supplement of 0, 0.3%, 0.6% K⁺ which contained 1.29 g/100 g, 1.60 g/100 g, 1.93 g/100 g K⁺ respectively. The supplement of K⁺ to the low‐salinity well water was 10, 20, 40 mg L^?1 which formed Na:K ratios of 42:1, 33:1, 23:1 respectively. Results showed that when the aqueous Na:K ratio was 42 and dietary K⁺ was 1.93 g/100 g K⁺, the WGR and PER of L. vannamei were the highest and the FCR was lower than that of others (P < 0.01). Supplement of K⁺ into well water and diets did not showed significant effects on haemolymph ammonia‐N, uric acid, urea content (P > 0.05), but had a extremely significant effect on arginase activity and Cl^? concentration (P < 0.01). Moreover, similar results were observed in alkaline phosphatase (ALP), bacteriolytic activity (LSZ) and respiratory burst activity (O₂^?) (P < 0.05). These results suggested that aqueous Na/K in the low‐salt well water and dietary K had significant synergistic effect on the growth, osmoregulation and immunity of L. vannamei. Concluded from the growth performance, nitrogen metabolism, osmoregulation and immunity, as the Na/K in the low‐salinity well water descended from 42 to 23, the requirement of dietary K⁺ was also decreased.     

Genetic variation of white spot syndrome virus (WSSV) in Pacific white shrimp Litopenaeus vannamei (Boone 1931) culture of Thailand

White spot syndrome virus (WSSV) is highly pathogenic to penaeid shrimp and has caused significant economic losses in the shrimp farming industry in Thailand. Genotyping analysis was done in 124 WSSV isolates from cultured Pacific white shrimp Litopenaeus vannamei. These samples were obtained during 2007–2014 from eight provinces in Thailand. We investigated five variable loci in the virus genome: deletions in two variable regions, VR14/15 and VR23/24, and three variable number tandem repeats (VNTR) located in open reading frame (ORF) 75, 125 and 94. WSSV genotype was characterized as (X_14/15, X_23/24) (N₇₅‐N₁₂₅‐N₉₄) where X is the number of base pair deletion in the variable region and N is the number of repeat units (RUs) in a specific ORF. The deletion pattern in VR14/15 and VR23/24 regions characterized three WSSV genotypes. The most prevalent genotype was (5950_14/15, 10971_23/24), and it was found in all studied areas. At least 33 genotypes of WSSV were analysed based on 3 VNTR loci, indicating that the VNTRs of WSSV genome are highly variable. From 124 WSSV samples, two samples presented the characteristic of all five variable loci similar to WSSV collected during 2010 in Saudi Arabia (5950_14/15, 10971_23/24) (3₇₅‐6₁₂₅‐7₉₄). Many different WSSV genotypes shown in this study as compared to previously reported genotypes in Thailand suggests current status of disease epidemiology, as well as probable movements of WSSV between countries.     

Growth,body composition,ammonia tolerance and hepatopancreas histology of white shrimp Litopenaeus vannamei fed diets containing different carbohydrate sources at low salinity

Juvenile Litopenaeus vannamei farmed at 3.0 psu were fed five diets containing glucose, sucrose, wheat starch, corn starch or potato starch as the carbohydrate (CBH) source. Shrimp were fed for 50 days to explore the effect of dietary CBH source on growth, body composition and ammonia tolerance. The specific growth rate of body length of shrimp fed glucose was the highest and significantly higher than those fed potato starch. The survival rate of shrimp fed glucose was 89.44%, and it was the highest and significantly higher than those fed wheat starch. Whole shrimp body crude protein and lipid of the corn starch group were 140.2 g kg^?1 and 10.1 g kg^?1 respectively. And they were significantly higher than those fed wheat starch. Shrimp fed potato starch had higher hepatopancreas and muscle glycogen. Shrimp fed sucrose had higher glucose‐6‐phosphate dehydrogenase and lower pyruvate kinase activities (P < 0.05). Besides, shrimp fed starch produced more B cells in hepatopancreas tubules than those fed glucose or sucrose. Shrimp fed different sources of CBH differed in the number of R cells. After 96‐h of ammonia nitrogen challenge, the survival rate of the treatments from high to low in turn was glucose, wheat starch, corn starch, sucrose and potato starch, and no significant differences were observed among all treatments. Based on shrimp growth and the economic problems of practical production, we recommend wheat starch as CBH source in practical diets for L. vannamei farmed at low salinities.     

Effects of water hyacinth (Eichhornia crassipes) leaf protein concentrate as soybean protein replacement in white shrimp Litopenaeus vannamei (Boone) postlarvae diet

The potential of water hyacinth leaf protein concentrate (WHLPC) as an alternative protein source to soybean meal in formulated diets for the white shrimp Litopenaeus vannamei (Boone) postlarvae, was assessed. Isonitrogenous and isolipidic diets were formulated containing 40% protein and 8% lipid. WHLPC replaced 0%, 25%, 50% and 75% of soybean meal protein in the formulated diets. Twenty postlarval Litopenaeus vannamei weighing 5.0 ± 0.00032 mg were randomly assigned to twelve 30‐L aquarium tanks and were fed experimental diets for 70 days. Results show that the overall biological performance of shrimp in terms of specific growth rate (SGR), protein efficiency ratio (PER), feed efficiency (FE) and survival in treatment receiving the 25% WHLPC replacement is comparable to that of the control diet. Replacement levels beyond 25% exhibited significant depression in terms of growth performance of the shrimp. Collectively, these findings indicate that WHLPC can replace 25% of soybean meal protein in the formulated diets of L. vannamei.     

Response of facilitative glucose transporter 1 to salinity stress and dietary carbohydrate nutrition in white shrimp Litopenaeus vannamei

Facilitative glucose transporter 1 (GLUT1) is a transporter protein for glucose transport via the plasma membrane of the cells to provide energy through carbohydrate metabolism. GLUT1 cDNA from Litopenaeus vannamei was obtained and analysed in this study. Full‐length GLUT1 cDNA is 2062 bp long and contained a 1506‐bp ORF encoding a 502 amino acid protein, a 270‐bp 5′UTR and a 284‐bp 3′UTR. When shrimp were under acute low salinity stress, the expression in hepatopancreas, muscle, gill and eyestalk was all up‐regulated at 12 h (P < 0.05) and 96 h (P < 0.05), while the expression in the four tissues was all down‐regulated at 6 h (P < 0.05) and 48 h (P < 0.05) . The expression in the muscle of shrimp at water salinity of 3 was lower than that at water salinity of 30 independent of dietary carbohydrate levels, while expression in hepatopancreas, gill and eyestalk was up‐regulated at 200 and 300 g kg^?1 carbohydrate levels. The expression in all tissues fed glucose was up‐regulated when compared to the expression in shrimp held at a water salinity of 30. This study suggests that GLUT1 is a conserved protein in L. vannamei, and changes in expression due to environmental salinity and dietary carbohydrate level and source.     

Effects of dietary administration of Macsumsuk® on growth and stress to low salinity and low dissolved oxygen in whiteleg shrimp,Litopenaeus vannamei (Boone, 1931) juveniles

An 8‐week study was conducted to explore the results of Macsumsuk^® as a feed additive on the stress tolerance and growth of Litopenaeus vannamei in 15 culture tanks of 36 L each. Three hundred shrimp averaging 0.1 ± 0.01 g were fed with five isonitrogenous (48.38 ± 0.38% CP) diets (in triplicate groups) containing kaolinite (Macsumsuk^®) at 0%, 0.3%, 0.6%, 1.2% and 2.4%, namely Mk₀, Mk_0.3, Mk_0.6, Mk_1.2 and Mk_2.4. Specific growth rate (SGR) and weight gain (WG) of shrimp fed diets Mk_1.2 and Mk_2.4 were significantly better than those of shrimp fed diet Mk₀ (p < .05). However, SGR and WG of shrimp fed diets Mk_0.6, Mk_1.2 and Mk_2.4 were not significantly different. Protein efficiency ratio (PER) and feed efficiency (FE) of shrimp fed diets Mk_1.2 and Mk_2.4 were significantly better than those of shrimp fed diets Mk₀, Mk₀.₃ and Mk_0.6. Furthermore, the survival of shrimp fed diet Mk_2.4 was significantly lower than that of shrimp fed diet Mk_0.6 (p < .05). Cumulative mortality of shrimp fed diet Mk_1.2 was significantly lower than that of shrimp fed diet Mk₀ at 1–1.5 hr post‐stress to low dissolved oxygen (from 6.1 mg/L to 2.9 mg/L) and 4–5 hr post‐stress to low salinity (from 32‰ to 1‰) (p < .05). The optimum dietary Macsumsuk^® level for juvenile L. vannamei was determined as 1.97% by the polynomial regression analysis of weight gain.     

Artificial substrates in zero‐water‐exchange culture system regulate the rearing performance of Pacific white shrimp Litopenaeus vannamei (Boone, 1931) under the winter indoor condition

Effects of artificial substrates in zero‐water‐exchange culture system on the rearing performance of Litopenaeus vannamei under winter indoor condition were investigated in this study. Growth, survival, feed conversion rate (FCR), production rate of L. vannamei and water quality were compared between artificial substrate‐treated group (AST) and control group (without artificial substrates presented in the rearing environment). Artificial substrates can significantly improve the water quality, the ammonia and nitrite‐N concentrations in the AST group were significantly lower than in the control group (P < 0.05), and the total heterotrophic bacteria and Vibrio spp. were also significantly lower in the AST group (P < 0.05). The survival, growth and production rate of L. vannamei in the AST group were significantly higher than in the control group (P < 0.05). Significantly lower FCR was observed in the AST group (P < 0.05). Results from this study indicate that the utilization of artificial substrates in the indoor shrimp culture system could effectively control the water quality, improve the survival and growth of shrimp and significantly reduce the FCR. This study provides a guideline for employing artificial substrates in rearing of shrimp in the zero‐water‐exchange culture system under lower temperature, which could be applicable to other similar species.     

Proteome modifications of Pacific white shrimp (Litopenaeus vannamei) muscle under biofloc system

The objective of the study was to examine the effects of biofloc technology on the muscle proteome of Litopenaeus vannamei. Two biofloc treatments and one control were compared: biofloc‐based tanks under zero‐water exchange fed with 150 g/kg crude protein (BF15), or with 250 g/kg crude protein (BF25) diets, and clear water tanks with 50% of daily water exchange stocked with shrimp fed with similar amount of a 250 g/kg crude protein diet, referred to as control. The shrimp (5.28 ± 0.42 g) were divided into the 300‐L fibreglass tanks (water volume of 200 L) at a density of 35 shrimp per tank and were cultured for 35 days. The biofloc groups displayed better growth and survival compared to the control. The muscle tissue from the control and BF25 groups was subjected to proteomic analysis. Lactate dehydrogenase, enolase, arginine kinase, mitochondrial ATP synthase subunit alpha, mitochondrial ATPase inhibitor factor 1 precursor, serpin 3 and myeloid differentiation factor 88 had an increased abundance in the BF25 group, while myosin heavy chain type 1 and myosin heavy chain type 2 showed a decreased abundance. The results indicate that biofloc technology could alter the expression of proteins involved in structure, metabolism and immune status of cultured shrimp.     

Differences in uptake and killing of pathogenic and non‐pathogenic bacteria by haemocyte subpopulations of penaeid shrimp,Litopenaeus vannamei, (Boone)

Phagocytosis is an important function of both invertebrate and vertebrate blood cells. In this study, the phagocytic activity of haemocyte subpopulations of penaeid shrimp, Litopenaeus vannamei, (Boone), against pathogenic and non‐pathogenic particles was investigated in vitro. The haemocytes of penaeid shrimp were firstly separated by centrifugation on a continuous density gradient of iodixanol into four fractions with five subpopulations (sub), of which sub 1 (hyalinocytes) and sub 4 (semi‐granulocytes) have the main function in phagocytosis of both pathogenic and non‐pathogenic bacteria as well as fluorescent polystyrene beads. It was found that these haemocyte subpopulations engulfed virulent Vibrio campbellii and Vibrio harveyi at a higher rate than non‐virulent Escherichia coli and polystyrene beads. When these bacteria were mixed with shrimp haemocyte subpopulations and incubated for 180 min, the percentage of viable intracellular V. campbellii (25.5 ± 6.0%) recovered was significantly higher than the percentage recovered from V. harveyi (13.5 ± 1.1%). No viable intracellular E. coli was observed in this study. In contrast to V. harveyi and E. coli, V. campbellii containing endosomes did not acidify in time. Incubation of haemocyte subpopulations with the most virulent V. campbellii strain resulted in a significant drop in haemocyte viability (41.4 ± 6.3% in sub 1 and 30.2 ± 15.1% in sub 4) after 180 min post‐inoculation in comparison with the less virulent V. harveyi (84.1 ± 5.6% in sub 1 and 83.4 ± 4.1% in sub 4) and non‐virulent E. coli (92.7 ± 2.8% in sub 1 and 92.3 ± 5.6% in sub 4) and polystyrene beads (91.9 ± 1.6% in sub 1 and 84.4 ± 3.4% in sub 4). These findings may be a valuable tool for monitoring shrimp health and immunological studies.     

Effects of Vibrio harveyi on plasma clotting protein (coagulogen) of white shrimp Litopenaeus vannamei

Blood clotting exhibits various important functions, including the prevention of body fluid loss and invasion of pathogens in shrimp. The effects of pathogenic Vibrio harveyi on plasma of white shrimp (Litopenaeus vannamei) in vitro and in vivo were investigated in this study. The clotting protein (coagulogen) in plasma of white shrimp pre‐incubated with extracellular products (ECP) of V. harveyi was found apparently decreased and fast‐migrated in crossed immunoelectrophoresis (CIE) gels. In addition, the coagulogen had been degraded to many low molecular‐weight protein bands in plasma pre‐incubated with ECP on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) gels. When pre‐challenged with bacterial cells and ECP of V. harveyi, the white shrimp began to die at about 30 and 16 h respectively. Moreover, plasma coagulogen was decreased more obvious in shrimp challenged with ECP than that with bacterial cells as visualized in CIE gels, and total plasma protein in both group of shrimp were all decreased. Haemolymph withdrawn from moribund shrimp pre‐challenged with V. harveyi or its ECP was observed unclottable. However, the addition of clotting factors (transglutaminase and/or Ca²⁺) to these unclottable plasma could apparently promote their re‐clotting ability as jelly‐like solid observed in microtubes. The recovery of clotting ability of plasma from moribund shrimp was due to the reformation of coagulogen (200 kDa) after adding the two clotting factors as shown on CIE and SDS‐PAGE gels. The present results suggest that the infection of V. harveyi in white shrimp may not only degrade coagulogen but also influence the presence of transglutaminase and Ca²⁺ ion.     

Effects of low salinity exposure on immunological,physiological and growth performance in Litopenaeus vannamei

Growth, immunological and physiological parameters of white shrimp Litopenaeus vannamei reared at different salinity levels (1, 10, 15, 25 and 35 g/L) at stocking density of 214 shrimp/m³ were examined at 1, 30 and 63 days. Results showed that the total haemocyte count (THC) of shrimp decreased with time at all salinity levels, indicating a potential reduction in the resistance of shrimp against pathogens, since a low value of THC indicates a perturbation of the immune system. Glucose and protein values observed in the haemolymph throughout the study indicate that shrimp adapted well to low salinities (1, 10 and 15 g/L). Although of those shrimp reared at 10 g/L only 83.3% survived, at this salinity, shrimp depicted a higher glucose concentration in haemolymph at the beginning and end of the study.