An oral delivery system for controlling white spot syndrome virus infection in shrimp using transgenic microalgae

White spot disease (WSD) is a longstanding and serious viral disease of various shrimp species that has caused high mortality rates for many decades. Currently, there is no practical method to control this disease. Therefore, we have explored the development of a novel vaccine-based method to control this disease using transgenic algae. During infection by white spot syndrome virus (WSSV), the interaction between viral envelope proteins and cell surface protein receptors on target cells is the key step of viral entry and replication. Hence, transgenic lines of the green microalga Chlamydomonas reinhardtii harbouring a WSSV VP28 viral envelope protein were created as an oral delivery system for vaccinating shrimp. Two separate transplastomic lines containing wild-type and codon optimized gene sequences for VP28 were evaluated for recombinant protein levels. Only the codon optimized line gave rise to detectable VP28 in western blot analysis, which demonstrated that optimization for chloroplast codon bias improved the efficiency of expression and that the gene design produced a favourable RNA secondary structure with suitable free energy for translation. In addition, bile salt and acid tolerance tests demonstrated that this transgenic Chlamydomonas can tolerate mildly acidic (pH 5.0) conditions and 0.30% bile salts. These features indicated that algal cells are suitable for delivering viral antigens through a shrimp's digestive system. In WSSV infection experiments, the highest survival rate (87%) was recorded in shrimps fed with the codon optimized VP28 line mixed into their feed, indicating that this line could be employed in the control of WSSV spread in shrimp populations. This algal strategy offers a new, efficient, fast and less labour-intensive method for the control of other diseases in aquatic animals through oral delivery

Efficiency of Citronella grass (Cymbopogon winterianus Jowitt) extracts to control Zoothamnium sp. in black tiger shrimp (PL15 stage)

Study on the efficiency of Citronella grass (Cymbopogon winterianus Jowitt) extracts to control Zoothamnium sp. was conducted in the laboratory at Kasetsart University. Comparison between two methods of extractor, water distillation and soxhlet extractor, revealed that soxhlet extractor was about 225 times more efficient than the water distillation, based on the concentration that could give 100% mortality of Zoothamnium sp. in 24 hours (0.008 g/l from soxhlet extractor and 1.8 g/l from water distillation). The LC50 of PL15 shrimp in 24 hours of this extract from water distillation was 21.05 g/l while from soxhlet extractor was 1.88 g/l. Those concentrations, 0.008 g/l from soxhlet extractor and 1.8 g/l from water distillation did not harmful to the PL15 shrimp measured by dissolved oxygen and pH of water

Effects of the dietary supplementation of mixed probiotic spores of Bacillus amyloliquefaciens 54A, and Bacillus pumilus 47B on growth, innate immunity and stress responses of striped catfish (Pangasianodon hypophthalmus)

The study used the mixed probiotics of Bacillus amyloliquefaciens 54A and B. pumilus 47B isolated from striped catfish (Pangasianodon hypophthalmus) intestine aiming to stimulate growth performance, innate immunity, stress tolerance of striped catfish. The average weight gain (AWG), specific growth rate (SGR), and feed conversion ratio (FCR) were analyzed after fish were fed the mixture of probiotics (B. amyloliquefaciens 54A and B. pumilus 47B) at concentrations of 1 108, 3 108, and 5 108 CFU g 1 feed for 90 days. Immunity parameters, survival rate of fish challenged with Edwardsiella ictaluri and ammonia tolerance were also investigated. The amounts of B. amyloliquefaciens and B. pumilus were counted and identified by specific primer pairs of Ba1-F/Ba1-R, and 16-F/Bpu-R to confirm the presence of probiotics in fish intestine. The AWG (476.6 ± 7.81 g fish 1) of fish fed probiotics at 5 108 CFU g 1 was significant higher than the control (390 ± 25.7 g fish 1) after 90 days of feeding, but there was no significant (P > 0.05) effect of probiotics on FCR and SGR. Fish fed diet containing probiotics at 5 108 CFU g 1 also expressed resistance to E. ictaluri infection and higher immune parameters such as phagocytic activity, respiratory bursts, and lysozyme activity than the control. Stress response with ammonia showed significantly lower mortality rate (25%, 20% and 27%) of fish fed probiotics at all three levels of 1, 3 and 5 108 CFU g 1 than the fish fed control diet (75%). The study also demonstrated that the probiotics survived in the intestine of striped catfish after 90 days of feeding. Therefore, the dietary supplementation of a mixture of B. amyloliquefaciens and B. pumilus at 5 108 CFU g 1 can be used to improve the health and growth rate of striped catfish