Dose/frequency: A critical factor in the administration of glucan as immunostimulant to Indian white shrimp Fenneropenaeus indicus

The immunostimulatory effect of an alkali insoluble glucan extracted from marine yeast isolate Candida sake S165 was tested in Fenneropenaeus indicus. Post larvae (PL) of F. indicus, fed glucan incorporated diet at varying concentrations (0.05, 0.1, 0.2, 0.3, 0.4 g glucan/100 g feed) for 21 days were challenged orally with white spot syndrome virus (WSSV). Maximum survival was observed in PL fed the 0.2% glucan incorporated diet. Subsequently the feed incorporated with 0.2% glucan was fed to F. indicus post larvae at different feeding intervals, i.e. daily, once every two days, once every five days, once every seven days and once every ten days. After 40 days, the prawns were challenged orally with WSSV and post challenge survival was recorded. Shrimp feed containing 0.2% glucan when administered once every seven days gave maximum survival. This was supported by haematological data obtained from adult F. indicus, i.e. total haemocyte count, phenoloxidase activity and nitroblue tetrazolium reduction (NBT). The present observation confirms the importance of dose and frequency of administration of immunostimulants in shrimp health management

Primary hemocyte culture of Penaeus monodon as an in vitro model for white spot syndrome virus titration, viral and immune related gene expression and cytotoxicity assays

Immortal cell lines have not yet been reported from Penaeus monodon, which delimits the prospects of investigating the associated viral pathogens especially white spot syndrome virus (WSSV). In this context, a method of developing primary hemocyte culture from this crustacean has been standardized by employing modified double strength Leibovitz-15 (L-15) growth medium supplemented with 2% glucose, MEM vitamins (1 ), tryptose phosphate broth (2.95 g l 1), 20% FBS, N-phenylthiourea (0.2 mM), 0.06 lgml 1 chloramphenicol, 100 lgml 1 streptomycin and 100 IU ml 1 penicillin and hemolymph drawn from shrimp grown under a bio-secured recirculating aquaculture system (RAS). In this medium the hemocytes remained viable up to 8 days. 5-Bromo-20-deoxyuridine (BrdU) labeling assay revealed its incorporation in 22 ± 7% of cells at 24 h. Susceptibility of the cells to WSSV was confirmed by immunofluoresence assay using a monoclonal antibody against 28 kDa envelope protein of WSSV. A convenient method for determining virus titer as MTT50/ml was standardized employing the primary hemocyte culture. Expression of viral genes and cellular immune genes were also investigated. The cell culture could be demonstrated for determining toxicity of a management chemical (benzalkonium chloride) by determining its IC50. The primary hemocyte culture could serve as a model for WSSV titration and viral and cellular immune related gene expression and also for investigations on cytotoxicity of aquaculture drugs and chemicals