Immuno-stimulatory effect and toxicology studies of salt pan bacteria as probiotics to combat shrimp diseases in aquaculture

Abstract

The shrimp aquaculture industry has experienced serious economic losses due to diseases caused by Vibrio species. The application of antibiotics to combat diseases has led to environmental hazards, antibiotic-resistance in pathogens and accumulation of antibiotics in tissues. This study explores the use of probiotics as an alternative to antibiotics. A probiotic consortium SFSK4 (comprising salt pan bacteria Bacillus licheniformis TSK71, Bacillus amyloliquefaciens SK27, Bacillus subtilis SK07, Pseudomonas sp. ABSK55) was used as a water additive during shrimp culture. It significantly increased shrimp (Litopenaeus vannamei) immunity i.e. total hemocyte count, phagocytosis, total plasma protein, respiratory burst and bactericidal activity as compared to the control. It also stimulated the phenoloxidase activity by two-fold. Proteomic analysis revealed the differential expression of 50 immune proteins (39 up-regulated and 11 down-regulated) in SFSK4 treated shrimps. Four major immune modulation proteins viz. Caspase2, GTPase activating protein, Hemocyanin and Glucan pattern-recognition lipoprotein involved in cell mediated immune response were identified in SFSK4 treated shrimp hemolymph. SFSK4 decreased shrimp mortality by more than 50% against pathogens. Toxicology studies revealed that administration of the highest dose of probiotic (10¹² CFU/mL) showed no adverse effect on shrimp survival (LC₅₀ analysis) and neither exhibited cytotoxicity. Genotoxicity study confirmed that the probiotic did not cause DNA damage in shrimps. The findings suggest that the probiotic SFSK4 is an eco-friendly water additive to enhance shrimp immunity against diseases in aquaculture, which could help curtail environmental hazards as an effective alternative to antibiotics.

Keywords: Comet assay; Cytotoxicity; Innate immunity; Litopenaeus vannamei; Proteomic.