Protective Effects and Potential Mechanisms of Bacillus subtilis Ya3.1 Against Aeromonas hydrophila Infection in Hybrid Sturgeon (Acipenser schrenckii ♂ × Huso dauricus ♀)

Bacterial disease caused by Aeromonas hydrophila is a major constraint in intensive sturgeon aquaculture. This study examined the intestinal, humoral, and hepatic responses associated with dietary or waterborne pretreatment with the host-derived probiotic Bacillus subtilis Ya3.1 during A. hydrophila infection in hybrid sturgeon (Acipenser schrenckii ♂ × Huso dauricus ♀). Absolute taxon-targeted qPCR, serum innate immune assays, intestinal 16S rRNA gene sequencing, and untargeted liver metabolomics were integrated to evaluate probiotic-associated protective responses. Ya3.1 pretreatment increased intestinal B. subtilis-associated gene copy numbers before challenge and reduced the post-challenge A. hydrophila-associated burden. Among surviving fish at 10 days post-infection, both probiotic-pretreated groups showed significantly higher alkaline phosphatase activity than the infected control group; the dietary pretreatment group also exhibited a significantly higher complement C3 concentration. Microbiota analysis revealed marked community restructuring in untreated challenged fish, characterized by enrichment of Aeromonas and other opportunistic taxa, whereas probiotic-pretreated fish showed reduced challenge-associated compositional displacement and relative enrichment of Bacillus-related taxa. Liver metabolomics further indicated that Ya3.1 pretreatment was associated with altered hepatic metabolic profiles, with glycerophospholipid metabolism emerging as a recurrent candidate pathway. Overall, Ya3.1-associated protection was associated with a lower pathogen burden, reduced microbiota displacement, partial preservation of humoral immune indices, and altered hepatic metabolic responses.