Incidence,Growth, and Inactivation of Listeria monocytogenes in Cooked and Peeled Cold-Water Shrimp

ABSTRACT

Cooked and peeled cold-water shrimp (Pandalus jordani) naturally contaminated with Listeria monocytogenes were obtained from a processor for a series of studies to determine the level of contamination and growth characteristics of this bacterium in the naturally contaminated product. L. monocytogenes was isolated from every 25-g sample of individually quick frozen (IQF) shrimp that was tested. The level of contamination in each composite sample ranged from 5 to 16 colony forming units (CFU) per 25 g. When individual shrimp taken from the 25-g sample portions were tested separately, samples positive for L. monocytogenes ranged from 1 of 12 to 5 of 15 shrimp tested. The project also evaluated the effectiveness of three methods to inactivate the bacterium: ozone, chlorine dioxide, and steam as possible product reconditioning strategies. Ozone and chlorine dioxide were both found to be ineffective reconditioning treatments for shrimp naturally contaminated with L. monocytogenes. Experiments with steam conducted at the laboratory and later at the shrimp processing plant verified that shrimp contaminated with L. monocytogenes could be safely reconditioned by steam pasteurization. Steam was used successfully to pasteurize several thousand pounds of contaminated shrimp in the processing plant. When the naturally contaminated product was packaged in either oxygen-permeable or impermeable films and stored at 5°C and 10°C, the product was deemed spoiled by sensory evaluation after 9 days of storage, at which time the L. monocytogenes population were 3 × 10⁴ CFU per g. By comparison, when an isolate (strain 4311) from naturally contaminated shrimp was inoculated onto the pasteurized shrimp at a concentration of 12 cells /25g, the L. monocytogenes population reached 3.0 × 10⁸ per g after 9 days of storage. The pasteurization process used in this study would not be effective in inactivation of Clostridium botulinum. Ready-to-eat-shrimp must therefore be stored below 3°C or frozen.     

Effects of Fumonisin B1-Containing Feed on the Muscle Proteins and Ice-Storage Life of White Shrimp (Litopenaeus vannamei)

Juvenile (5.5–5.7 g) white leg shrimp were exposed to mycotoxin in one indoor trial by feeding fumonisin B₁ (FB₁) at levels from 0.2 to 2.0 μg g^?1 FB₁ for 30 days. Shrimp growth was affected after exposure to more than 0.6 μg g^?1 FB₁. Soluble muscle protein concentration decreased, and changes in myosin thermodynamic properties were observed in shrimp after 30 days of exposure to FB₁. Marked histological changes in tissue of shrimp fed a diet containing FB₁ at 2.0 μg g^?1 were also observed. Shrimp fed diets containing more than 0.6 μg g^?1 FB₁ showed greater decrease in shear forces after 12 days of ice storage.     

Isolation and identification of Vibrio campbellii as a bacterial pathogen for luminous vibriosis of Litopenaeus vannamei

Outbreak of luminescent disease was reported from Litopenaeus vannamei shrimp farms in Zhangpu County, Southern China during May–July 2011. The clinical signs included fluorescent, less food consumption and high mortality. Bacteria were isolated from the infected shrimps. The pathogen, a luminescent bacterium named VH1 was identified as Vibrio campbellii based on MLSA analysis (16S rDNA, rpoD and toxR). The haemolysin (hly) gene specific in V. campbellii was detected in strain VH1. Pathogenicity test using immersion infection confirmed that strain VH1 was virulent to L. vannamei postlarvae and juveniles, and the LC₅₀ value was 1.55 × 10⁶ CFU mL^?1 and 1.7 × 10⁶ CFU mL^?1 respectively.     

Mycoflora of the 'green water' culture system of tiger shrimp Penaeus monodon Fabricius

This study was conducted to quantify and characterize the mycoflora associated with the ‘green water’ culture system of Penaeus monodon. Samples of water, tilapia gut and mucus, and shrimp hepatopancreas from three shrimp farms were collected during 15, 30, 45 and 60 days of culture (DOC). Results showed that high fungal loads were observed in tilapia gut (total: 117–1352 colony forming unit (CFU) 5 cm hind gut^?1; yeasts: 0–136 CFU 5 cm hind gut^?1) and mucus (total: 12–311 CFU (5 cm²)^?1; yeasts: 0–88 CFU (5 cm²)^?1), while minimal fungal populations were observed in water samples (total: 0–110CFU mL^?1; yeasts: 0–5 CFU ml^?1). Shrimp hepatopancreas harboured a very low number of filamentous fungi (0–27 CFU 0.1 g^?1) and yeasts (0–7CFU 0.1 g^?1) especially at 60 DOC. The filamentous fungal isolates were dominated by Penicillium and Aspergillus species, while the yeast populations were dominated by Rhodotorula and Saccharomyces species. The dominance of these fungi on tilapia mucus and gut and their presence in the rearing water might play an important role in the overall mechanisms involved in the control of luminous Vibrio in the ‘green water’ grow‐out culture of P. monodon.     

Effect of light limitation on the water quality,bacterial counts and performance of Litopenaeus vannamei postlarvae reared with biofloc at low salinity

The aim of this study was to evaluate the effect of light limitation on the water quality, bacterial counts and performance of Litopenaeus vannamei postlarvae reared with biofloc at low salinity (≈9 g L^?1). Two treatments were designed: T₁ = culture with natural sunlight and T₂ = culture in darkness. After 28 days, in both treatments, the final weight of shrimp was over 0.6 g with a specific growth rate over 7.4% d^?1, and a survival rate over 70%. In both treatments, Vibrio sp. concentration presented low values (culture with natural sunlight = 0.1 to 9.9 × 10² CFU mL^?1, culture in darkness = 0.4 to 11.7 × 10² CFU mL^?1) and Bacillus sp. had high values (culture with natural sunlight = 0.7 to 66.0 × 10⁴ CFU mL^?1, culture in darkness = 0.7 to 65.8 × 10⁴ CFU mL^?1). All water quality parameters remained within the ranges suitable for shrimp culture, except for alkalinity during the first stage of the study. Although in some sampling periods some significant differences were found in bacterial counts and water quality parameters, shrimp productive performance under culture with biofloc at low salinity was not affected significantly by light limitation.     

Effects of dietary Lactobicillus plantarum on the growth,carcass quality,and immune response of African catfish (Clarias gariepinus) challenged with Salmonella typhi

An experiment was conducted to investigate the effects of Lactobacillus plantarum on the production of African catfish (Clarias gariepinus). Five experimental diets containing 0, 10³, 10⁵, 10⁷, and 10⁹ CFU of Lactobacillus plantarum g^?1 diet (T₁–T₅ treatments respectively) were fed to African catfish (Clarias gariepinus) (9.20 ± 0.1 g initial body mass), for 70 days. Results indicated significantly lower growth performance in the fish fed diet without probiotics and in those fed diet with the least probiotic level. Treatments with probiotics significantly improved the blood profile and carcass protein but significantly reduced the carcass fat (P < 0.05); these treatments also marginally improved the carcass minerals in comparison with the treatment without probiotic (P < 0.05). Challenging the fish fed the experimental diets with Salmonella typhi showed higher immunity of fish fed the probiotic diets than those fed the nonprobiotic diet. Duncan’s multiple range test showed that the best fish performance was observed with 10³ CFU g^?1 L. plantarum for very parameter measured. However, regression analyses showed the optimum level of the bacteria to be 10^4.13–10⁵ CFU g^?1     

Changes in Biogenic Amines and Bacteria of Tiger-Toothed Croaker (Otolithes ruber) during Ice Storage

Changes in the content of biogenic amines (BA; e.g., putrescine, tyramine, and histamine) and the mesophilic and psychrotrophic bacterial counts of tiger-tooth croaker (Otolithes ruber) were monitored during ice storage for 18 days. BA and bacterial loads gradually increased (p < 0.05) with storage time. On the first day of storage, no amines were detected, and no histamine was detected up to 6 days. The best linear regressions between BA and bacterial counts were found between putrescine and psychrotrophic bacteria. Furthermore, it was found that psychrotrophs were the dominant microorganisms during storage and showed significant (p < 0.05) differences with storage time. The initial concentration of putrescine was 1.27 μg g^?1 and reached 18.8 μg g^?1 when the psychrotrophic bacterial load was over 10⁸ CFU g^?1. The results suggest that the level of putrescine is a suitable indicator of freshness for O. ruber, and the shelf life of O. ruber stored in ice as determined by BA content and microbiological data was 10 days.     

Dietary administration of Bacillus (B. licheniformis and B. subtilis) and isomaltooligosaccharide influences the intestinal microflora,immunological parameters and resistance against Vibrio alginolyticus in shrimp,Penaeus japonicus (Decapoda: Penaeidae)

The experiment was conducted to investigate the effects of oral administration of probiotics (Bacillus licheniformis and Bacillus subtilis) in combination with prebiotic isomaltooligosaccharide (IMO) on the intestinal microflora and immunological parameters of Penaeus japonicus and its resistence against Vibrio alginolyticus. A basal diet was supplemented with 0% probiotics and prebiotic (control), 10⁸ colony forming unit (CFU) g^?1B. licheniformis and 0.2% IMO (T1), 10⁸ CFU g^?1B. subtilis and 0.2% IMO (T2), 10⁸ CFU g^?1B. licheniformis in combination with 10⁸ CFU g^?1B. subtilis and 0.2% IMO (T3). The results showed that the total bacterial counts significantly increased (P<0.05) and Vibrio counts significantly decreased (P<0.05) in the intestine of shrimp with supplementation of dietary synbiotics. Shrimp fed the diet with both Bacillus probiotics and IMO (T3) produced significantly higher immune parameters (phenoloxidase activity, lysozyme activity and nitric oxide synthase activity and superoxide dismutase activity) than the control group (P<0.05). Significant lower cumulative mortality after challenge with V. alginolyticus was also observed in shrimp fed diet with both Bacillus probiotics and IMO (T3) than that in the control group (P<0.05). The results indicated that the oral administration of probiotics (B. licheniformis and B. subtilis) and prebiotic IMO has positive effects on bacterial flora in P. japonicus gut, and can activate non‐specific immunity of shrimp.     

Biological characteristics of the improved extensive shrimp system in the Mekong delta of Vietnam

Low and unstable shrimp yields of the improved extensive shrimp system has been a tremendous obstacle for economic development in the coastal areas of Southern Vietnam. To investigate the biological characteristics of this system, ponds in the coastal Cai Nuoc district, Mekong delta of Vietnam, were monitored. Results showed that the system was not optimal for shrimp. While chlorophyll a (chl a) (1.51–37.2 μg L^?1), phytoplankton density (6333–974 444 cells L^?1) and zooplankton density (7.1–517.2 ind L^?1) were abundant and comparable to shrimp farms elsewhere, zoobenthic community was very poor (7–1971 ind m^?2). Toxin‐producing cyanobacteria (Oscillatoria limosa, Oscillatoria formosa, Anabaena sp. and Phormidium tenue) were found. Total bacteria and Vibrios were present in large numbers (respectively 1.04 × 10⁵ and 6.64 × 10² CFU mL^?1 in pond water, 6.33 × 10⁵ and 9.47 × 10³ CFU g^?1 in sediment). Presence of toxin‐producing organisms, poor zoobenthic community and abundance of Vibrios all can enhance shrimp susceptibility to diseases. The following measures are recommended to improve the situation: (1) complete testing of seeds for pathogens, (2) not to incorporate fish into shrimp ponds and (3) applying no‐culture breaks and pathogen‐killing chemicals.     

Effect of dietary supplementation with Bacillus subtilis on the growth,performance, immune response and antioxidant activities of the shrimp (Litopenaeus vannamei)

The objective of this study was to evaluate the effect of dietary supplementation of a probiotic bacterium, Bacillus subtilis, on the growth, immune response and antioxidant activities of shrimp (Litopenaeus vannamei). Shrimps with an average initial weight of 2.11±0.17 g were randomly assigned to four groups with three replicates. The control group was fed a basal diet, and three treated groups were fed diets supplemented with B. subtilis at doses of 1 × 10⁴, 5 × 10⁴ and 10 × 10⁴ colony‐forming unit (CFU) g^?1 feed respectively. After 40 days of culture, 10 shrimps from each replicate were taken randomly for the determination of immune response and oxidization resistance indices. The results showed that the shrimps fed with B. subtilis at a dose of 1 × 10⁴, 5 × 10⁴ CFU g^?1 feed showed significantly better growth than that of the control diet. The phenoloxidase activities showed a tendency to increase with an increased dose of B. subtilis in diets but there was no significant difference among the three treated groups. In addition, phenoloxidase activities were found to be significantly higher (P<0.05) in the groups treated with B. subtilis than that of the control group. Shrimps treated with 5 × 10⁴ CFU g^?1 feed probiotic bacterium showed the highest lysozyme activity and it was significantly higher (P<0.05) than the other groups. However, there was no significant difference in acid phosphatase and alkaline phosphatase activity across all the groups. The total antioxidant capacity, superoxide dismutase and glutathione peroxidase activities in the probiotic‐treated groups were significantly increased (P<0.05) as compared with the control groups. Both maleic dialdehyde concentration and superoxide anion activities in the probiotic‐treated groups were significantly lower (P<0.05) than those of the control. The probiotic did not affect the nitric oxide synthase and the catalase activity in any of the control and treated groups. These results indicated that the probiotic B. subtilis could significantly promote the growth rate of shrimp by increasing the immune function and antioxidant capacity. The most effective dose of B. subtilis in the diet was 5 × 10⁴ CFU g^?1 feed.     

Pharmacokinetics and tissue residues of marbofloxacin in crucian carp (Carassius auratus) after oral administration

Pharmacokinetics and residue elimination of marbofloxacin (MBF) were studied in crucian carp (Carassius auratus, 250±30 g) kept at two water temperatures of 15 and 25 °C. Marbofloxacin concentrations in plasma and tissues were analysed by means of high‐performance liquid chromatography using an ultraviolet detector. The limits of detection were 0.02 μg mL^?1, 0.02 μg g^?1, 0.025 μg g^?1, 0.02 μg g^?1 and 0.025 μg g^?1 in plasma and muscle, skin, liver and kidney respectively. Fish were administered orally at a single dosage of 10 mg kg^?1 body weight in the PK group. The data were fitted to two‐compartment open models at both temperatures. At 15 °C, the absorption half‐life () and distribution half‐life (t_1/2α) of the drug were 0.36 and 4.48 h respectively. The corresponding values at 25 °C were 0.23 and 0.87 h respectively. The elimination half‐life (t_1/2β) was 50.75 h at 15 °C and 25.05 h at 25 °C. The maximum MBF concentration (C_max) differed little between 15 (6.43 μg mL^?1) and 25 °C (8.36 μg mL^?1). The time to peak concentration was 1.74 h at 15 °C and 0.78 h at 25 °C. The apparent volume of distribution (V_d/F) of MBF was estimated to be 1.36 and 0.87 L kg^?1 at 15 and 25 °C respectively. The area under the concentration–time curve (AUC) was 301.80 μg mL^?1 h at 15 °C and 182.80 μg mL^?1 h at 25 °C. The total clearance of MBF was computed as 0.03 and 0.05 L h^?1 kg^?1 at 15 and 25 °C respectively. After repeated oral administration at a dosage of 10 mg kg^?1 body weight per day for 3 days, the results showed that the elimination half‐lives () of MBF from all tissues at 15 °C were longer than that at 25 °C. Therefore, water temperature is an important factor to be considered when deciding a reasonable withdrawal time.     

Delivery of HUFA, probionts and biomedicine through bioencapsulated Artemia as a means to enhance the growth and survival and reduce the pathogenesity in shrimp Penaeus monodon postlarvae

One of the major problems in the shrimp culture industry is the difficulty in producing high-quality shrimp larvae. In larviculture, quality feeds containing a high content of highly unsaturated fatty acids (HUFA) and ingredients that stimulate stress and disease resistance are essential to produce healthy shrimp larvae. In the present study, Penaeus monodon postlarvae (PL15) were fed for 25 days on an unenriched Artemia diet (control; A) or on a diet of Artemia enriched with either HUFA-rich liver oil of the trash fish Odonus niger (B), probionts [Lactobacillus acidophilus (C1) or yeast-Saccharomyces cerevisiae (C2)] or biomedicinal herbal products (D) that have anti-stress, growth-promoting and anti-microbial characteristics. P. monodon postlarvae fed unenriched Artemia exhibited the lowest weight gain (227.9 ± 8.30 mg) and specific growth rate (9.95 ± 0.05%), while those fed the HUFA-enriched Artemia (B) exhibited the highest weight gain and specific growth rate (362.34 ± 12.56 mg and 11.77 ± 0.08%, respectively). At the end of the 25-day rearing experiment, the shrimp postlarvae (PL40) were subjected to a salinity stress study. At both low and high (0 and 50‰) salinities, the group fed the control diet (A) experienced the highest cumulative mortality indices (CMI) 935.7 ± 2.1 and 1270.7 ± 3.1, respectively. Those fed diet D showed the lowest stress-induced mortality, and CMI were reduced by 31.1 and 32.3% under conditions of low and high salinity stress, respectively. A 10-day disease challenge test was conducted with the P. monodon postlarvae (PL40–PL50) by inoculating the shrimp with the pathogen Vibrio harveyi at the rate of 10⁵–10⁷ CFU/ml in all rearing tanks. P. monodon postlarvae fed probiont-encapsulated Artemia diets (C1 and C2) exhibited the highest survival (94.3 and 82.3%, respectively) and lowest pathogen load (V. harveyi) in hepatopancreas (5.2 × 10² ± 9.0 × 10 and 4.6 × 10² ± 9.0 × 10 CFU g⁻¹, respectively) and muscle (2.0 × 10² ± 6 × 10 and 1.7 × 10² ± 8.6 × 10 CFU g⁻¹, respectively) tissues. The shrimp that were fed the unenriched Artemia (Control; A) showed the lowest survival (26.33%) and highest bacterial load in the hepatopancreas (1.0 × 10⁵ ± 5 × 10³ CFU g⁻¹) and muscle (3.6 × 10⁴ ± 6 × 10² CFU g⁻¹). The shrimp fed the herbal product (D)-enriched Artemia also exhibited enhanced survival and reduced V. harveyi load in the tissues tested compared to the control diet (A) group. The results are discussed in terms of developing a quality larval feed to produce healthy shrimp larvae.     

Occurrence of Listonella anguillarum in seed production environments of Japanese flounder Paralichthys olivaceus (Temminck et Schlegel)

The present study was undertaken to investigate the distribution of Listonella anguillarum in the rearing water, fish and diets (rotifers) of Japanese flounder (Paralichthys olivaceus). A total of 793 isolates were obtained from the seed production environment of Japanese flounder and 175 out of them were identified as L. anguillarum by biochemical characterization, polymerase chain reaction (PCR) detection for VAH1 haemolysin gene and phylogenetic analysis of 16S ribosomal deoxyribonucleic acids (rDNA) sequences. These results strongly suggested that L. anguillarum is rapidly and accurately identified by the combination of incubation on thiosulphate–citrate–bile salt–sucrose agar at 35°C overnight and PCR detection for the VAH1 haemolysin gene. All flounder specimens and all rotifer samples harboured L. anguillarum at high densities of 6.9 × 10³–6.3 × 10⁵ colony forming units (CFU) g^?1 and 1.5 × 10⁴–2.3 × 10⁶ CFU g^?1, respectively, while as low as 5.0 × 10⁰–2.0 × 10¹ CFU mL^?1 of L. aguillarum were detected in only two of 11 seawater samples, even though no vibriosis occurred in larval and juvenile flounder of tanks. This fact strongly suggests that L. anguillarum is an inhabitant in the seed production environments of Japanese flounder.     

Effect of feeding Lactobacillus-based probiotics on the gut microflora, growth and survival of postlarvae of Macrobrachium rosenbergii (de Man)

The present study was conducted to study probiotics treatment in the post‐larval diet of Macrobrachium rosenbergii. Three hundred postlarvae (average weight, 114–118 mg±0.11) were divided in five experimental groups each with four replicates. The experiment was conducted for 60 days. Experimental diets were identical in all the aspects except for variation in the probiotics strain. T1 and T2 groups were fed Lactobacillus acidophilus (140×10¹¹ CFU 100 g^?1) and L. sporogenes (24×10⁷ CFU 100 g^?1) respectively. The T3 group was fed L. sporogenes bioencapsulated in Artemia nauplii. T4 was the control group (without probiotic) whereas T5 was fed Artemia along with control diet. The bacteriological study indicated that the gut microflora of postlarvae are devoid of lactic acid bacteria. The probiotic strains were found to have inhibitory effects against the gram‐negative bacterial flora present in the gut. Growth of the probiotic fed groups was significantly higher (P<0.05) than the control group. Significantly higher growth (P<0.05), per cent weight gain (132.5%), specific growth rate (1.41%), feed efficiency ratio (FER) (0.45), protein efficiency ratio (1.29) and protein gain (161.6%) were recorded in T3 group fed Artemia bioencapsulated L. sporogenes over the control group (P<0.05). Although insignificant (P>0.05), growth‐promoting effects of L. sporogenes were found to be higher than L. acidophillus. Survival of the postlarvae was not affected by probiotics in the diet.     

Characterization of a novel marine origin aerobic nitrifying–denitrifying bacterium isolated from shrimp culture ponds

A novel marine origin Bacillus subtilis strain H1 isolated from a shrimp culture pond effectively removed NH₄⁺‐N, ‐N and ‐N, with a maximum ammonium, nitrite and nitrate removal rate of 2.35 mg NH₄⁺‐N L^?1 hr^?1 per OD, 9.64 mg ‐N L^?1 hr^?1 per OD and 0.75 mg ‐N L^?1 hr^?1 respectively. The gas chromatography–isotope ratio mass spectrometry results indicated that N₂O was emitted when ¹⁵NH₄Cl, Na¹⁵NO₂ or Na¹⁵NO₃ was used. Additionally, N₂ was also produced when Na¹⁵NO₂ was used. Single‐factor experiments suggested that the optimal conditions for NH₄⁺‐N and ‐N removal were glucose as a carbon source, C/N 15, initial pH 7.5, 30 g/L NaCl, 28°C and a shaking speed of 160 rpm. Orthogonal tests showed that the optimal conditions for NH₄⁺‐N removal were C/N 15, pH 9, 10 g/L NaCl and shaking speed 160 rpm when ammonium chloride was used as the substrate. The optimal conditions for ‐N removal were C/N 10, pH 6, 10 g/L NaCl and a shaking speed of 160 rpm when sodium nitrite was used as the substrate. In summary, B. subtilis strain H1 had highly efficient aerobic nitrifying–denitrifying ability and high adaptability, suggesting that it is potentially valuable to marine aquaculture.     

Effects of Lactobacillus pentosus on the growth performance,digestive enzyme and disease resistance of white shrimp,Litopenaeus vannamei (Boone, 1931)

The objective of this study was to evaluate the probiotic properties of lactic acid bacteria (LAB) strains isolated from digestive tract of white shrimp Litopenaeus vannamei. Eighteen LAB colonies were isolated and one bacterium was found capable of producing three extracellular enzymes (protease, cellulose and lipase) simultaneously and exhibited antagonistic activity against shrimp pathogens (Vibrio vulnificus, V. rotiferianus and V. campbellii). The putative probiotic strain AS13 was identified as Lactobacillus pentosus based on 16S rRNA sequencing. The L. vannamei were fed diet containing 0 (control), 10⁶, 10⁷ and 10⁸ CFU g^?1 bacterial cells of AS13 for 28 days. The results showed that supplementation of L. pentosus significantly improved the growth performance and feed utilization in the treated groups over the control. Similarly, digestive enzyme activities were elevated in the intestines of treated groups. Moreover, feeding of supplemented diets containing AS13 significantly reduced the mortality rate caused by pathogenic Vibrio species (V. vulnificus, V. rotiferianus and V. campbellii). Our results indicated L. pentosus AS13 addition at 10⁷ CFU g^?1 can effectively enhance the growth performance, feed utilization, digestive enzymes and disease resistance of L. vannamei in the laboratory condition.     

Stocking strategies for production of Litopenaeus vannamei (Boone) in amended freshwater in inland ponds

The performance of the Pacific white shrimp Litopenaeus vannamei (Boone) under various stocking strategies was evaluated in earthen ponds filled with freshwater amended with major ions. Six 0.1‐ha earthen ponds located in Pine Bluff, AR, USA, were filled with freshwater in 2003 and 2004, and potassium magnesium sulphate added to provide 50 mg K⁺ L^?1 and stock salt added to provide 0.5 g L^?1 salinity. In 2003, three ponds either were stocked with PL₁₅ shrimp (39 PL m^?2) for 125 days of grow out or with PL₂₅ shrimp for 55 days (23 PL m^?2) followed by a 65‐day (28 PL m^?2) grow‐out period. In 2004, ponds were stocked with 7, 13 or 30 PL₁₅ m^?2 for 134 days of grow out. Salinity averaged 0.7 g L^?1 during both years, and concentration of SO₄^?2, K⁺, Ca²⁺ and Mg²⁺ was higher, and Na⁺ and Cl^? was lower in amended pond water than in seawater at 0.7 g L^?1 salinity. Potassium concentration in amended water was 52–61% of the target concentration. Shrimp yields ranged from 3449 kg ha^?1 in 2003 to 4966 kg ha^?1 in 2004 in ponds stocked with 30–39 PL₁₅ m^?2 for a 125–134‐day culture period. At harvest, mean individual weight ranged from 17.1 to 19.3 g shrimp^?1. In ponds stocked with PL₂₅ shrimp, yields averaged 988 and 2462 kg ha^?1 for the 1st and 2nd grow‐out periods respectively. Gross shrimp yield in 2004 increased linearly from 1379–4966 kg ha^?1 with increased stocking rate. These experiments demonstrated that L. vannamei can be grown successfully in freshwater supplemented with major ions to a final salinity of 0.7 g L^?1.     

Effect of dietary supplementation of inulin and W. cibaria on haemato‐immunological parameters of hybrid surubim (Pseudoplatystoma sp)

The dietary supplementation of prebiotics, probiotics and symbiotic in hybrid surubins (a Pseudoplatystoma corruscans and P. fasciatum cross) was evaluated for the effects on their autochthonous intestinal microbiota and on haematological and immunological parameters. A total of 160 fish were divided into four treatment groups with four replicates each. The treatment groups were fed with the following diets for 15 days: control diet without supplementation; 0.5% inulin (prebiotic) supplementation; Weissella cibaria (CPQBA 001‐10 DRM 02) (7.87 ± 0.2 log CFU g^?1) supplementation; or 0.5% inulin and W. cibaria supplementation (symbiotic group). The midgut intestines of the fish with the symbiotic diet supplementation had higher concentrations of lactic acid bacteria (7.07 ± 1.11 log CFU g^?1) and low levels of Vibrio spp (1.90 ± 0.60 log CFU g^?1) and Pseudomonas spp (2.23 ± 1.48 log CFU g^?1). In addition, increased erythrocytes and reduced circulating neutrophils were observed in this group. No differences in blood glucose, serum protein or lysozyme levels were detected between treatment groups. However, a higher concentration of total immunoglobulin was observed in fish fed with the probiotic and symbiotic diets. The addition of 0.5% inulin (prebiotic) thus W. cibaria (probiotic) to the diet of Pseudoplatystoma hybrid surubins reduce the number of pathogenic bacteria and stimulate the beneficial intestinal microbiota and may possibly alter their immune defence system.     

Effects of dietary probiotic on the growth performance,non‐specific immunity and disease resistance of cobia,Rachycentron canadum

The present study was performed to investigate the effects of a commercially available probiotic product (compound probiotic) containing Bacillus subtilis 7.0 × 10⁹ CFU g^?1, Bacillus licheniformis 3.0 × 10⁹ CFU g^?1, Lactobacillus spp. 5.0 × 10⁸ CFU g^?1 and Arthrobacter spp. 1.0 × 10⁸ CFU g^?1 on the growth performance, non‐specific immunity and protection against Vibrio harveyi infection in cobia (Rachycentron canadum). Fish were fed diets containing six graded levels of compound probiotic (0.0, 1.0, 2.0, 3.0, 4.0 and 5.0 g kg^?1) for 8 weeks. The results showed that the survival rate ranged from 81.1% to 84.4% with no significant difference among dietary treatments (P > 0.05) after feeding experiment. Dietary compound probiotic significantly increased the specific growth rate (SGR), serum lysozyme, alternative complement pathway (ACP) activity, phagocytosis percentage (PP) and respiratory burst activity of head‐kidney macrophages of cobia. Moreover, feeding of supplemented diets containing compound probiotic resulted in significantly lower mortality against the pathogens Vibrio harveyi compared with the control group. To elevate the growth and immune resistance ability of cobia, an optimal dose of dietary compound probiotic administration determined by second‐order polynomial regression analysis was 3.3 g kg^?1, on the basis of the SGR and mortality after challenge with V. harveyi.     

Water quality,Vibrio density and growth of Pacific white shrimp Litopenaeus vannamei (Boone) in an integrated biofloc system with red seaweed Gracilaria birdiae (Greville)

An indoor trial was conducted for 42 days to evaluate water quality, Vibrio density and growth of Litopenaeus vannamei in an integrated biofloc system (IBS) with Gracilaria birdiae. Four treatments were used, each in triplicate: Control (monoculture shrimp); IBS 2.5 (L. vannamei and 2.5 kg wet weight seaweed m^?3); IBS 5.0 (L. vannamei and 5.0 kg wet weight seaweed m^?3) and IBS 7.5 (L. vannamei and 7.5 wet weight seaweed m^?3). Shrimp individuals (0.34 ± 0.01 g) were stocked at a density of 500 shrimp m^?3. No water exchange was carried out during the experimental period. Molasses was added once a day as an organic carbon source to maintain the C:N ratio at 12:1. The IBS significantly decreased (P < 0.05) dissolved inorganic nitrogen (DIN) ranging from 19% to 34% (3.12–3.83 mg L^?1), NO₃‐N ranging from 19% to 38% (2.40–3.16 mg L^?1), Vibrio density ranging from by 8–83% (0.40–2.20 log 10³ colony‐forming units mL^?1), and FCR ranging from by 20–30% (1.20–1.37), as compared to the control (4.73 mg L^?1, 3.93 mg L^?1, 2.40 log 10³ colony‐forming units mL^?1, and 1.74 respectively). Moreover, the IBS significantly increased (P < 0.05) crude protein in whole body shrimp, ranging from 8% to 13% (13.2–13.7% wet weight basis); as well as final weight, ranging from 25% to 32% (3.90–4.12 g), weekly growth ranging from 25% to 34% (0.59–0.63 g), and shrimp yield by 22–39% (1.72–1.96 kg m^?3), as compared to control (12.1% wet weight basis, 3.12 g, 0.47 g, and 1.41 kg m^?3 respectively). It can thus be concluded that cultivating Gracilaria birdiae in an IBS with L. vannamei can contribute to DIN and NO₃‐N removal, lower Vibrio density, increased crude protein in whole body shrimp, higher growth and yield parameters in shrimp culture.