Development and evaluation of the inactivated vaccine against Aeromonas schubertii in hybrid snakehead

Abstract

Aeromonas schubertii is a global pathogen of fish that causes internal nodular diseases and death in severe cases. In the present study, we isolated nine pathogenic strains from diseased hybrid snakehead (Channa argus♂ × Channa maculata♀) which were formally identified as A. schubertii by molecular identification, including 16S rRNA and gyrB gene sequencing. Subsequently, we determined the extracellular enzyme activity of six isolates and compared pathogenicity to investigate potential relationships with geographic characteristics. Two highly pathogenic strains, AS-14 and AS-16, were finally identified as inactivated vaccine candidates. Subsequently, we evaluated different concentrations of the antigen for each vaccine by performing experiments in hybrid snakehead; fish were inoculated with inactivated vaccine via the intraperitoneal route and a booster injection was administered 14 days later. Relative percent survival (RPS) for fish immunized with high doses of the two vaccines (1 × 108 CFU/fish) were 54.4% and 57.4%, respectively; these values were higher than other groups of fish receiving different doses (1 × 106 and 1 × 107 CFU/fish). The bacterial load of tissues harvested from the fish immunized with high doses of the vaccine groups were significantly lower than those of fish immunized with low doses. Titers of specific antibodies against A. schubertii in fish immunized with a high dose of AS-16 vaccine were maintained at high levels when tested six weeks after the booster injection. Furthermore, immune-related genes in the spleen, including MHC I, MHC II, CD4, CD8α and IgM, all exhibited high levels of expression, thus suggesting that hybrid snakehead underwent an adaptive immune response post-vaccination. Collectively, our findings enrich our understanding of the specific causative agents underlying nodular diseases in hybrid snakehead and provide scientific reference guidelines for the development of vaccines against A. schubertii. The development of such vaccines will be conducive to the sustainable development of hybrid snakehead in commercial aquaculture.