Bacillus fermented complete feed: a green alternative to improve disease resistance, growth performance and hepato-intestinal health of Nile tilapia

Due to the prominent antibacterial activity and favorable fermentation characteristics, Bacillus strains are widely utilized in the preparation of fermented complete feed in aquafeed industry. However, synthetic microbial consortia are preferentially applied in industrial-scale production than single Bacillus strains alone. To date, the evaluation and comparison of the probiotic effects of single strains or synthetic microbial consortia fermented complete feed for fish are still insufficient. In this study, a Bacillus strain (D7-HSJ9) with inhibiting activity against Streptococcus iniae was isolated, Three probiotic combinations were used to ferment complete fish feed: Bacillus sp. D7-HSJ9 alone (RKL), D7-HSJ9 + Lactobacillus rhamnosus (BL), and D7-HSJ9 + L. rhamnosus and Saccharomyces cerevisiae (BLS). The antibacterial activity of the fermented feed was evaluated, and the RKL group exhibited the strongest antibacterial activity. An in vivo challenge trial in Nile tilapia ( Oreochromis niloticus ) also showed that the RKL group achieved the highest relative protection rate (42.86%) following S. iniae challenge. Moreover, an 8-week feeding trial showed that RKL group exhibited the best growth promotion (higher WGR/SGR, lower FCR), improved intestinal villus width and digestive enzyme activity, reduced hepatic steatosis, and reshaped gut microbiota (decreased pathogen Plesiomonas , increased probiotics Bacillus/Pediococcus ). Transcriptomic analysis revealed RKL group upregulated immune-related genes ( E3-TRIM39, NLRC3, CTL10, Mx, Ho1 ) and downregulated immunosuppressive genes ( LTA4, Plc2, angptl3, VTCN-1, PD1 ). This is the first report of Bacillus spizizenii fermenting complete fish feed, highlighting its superiority over mixed-strain fermentation and potential as a green alternative to antibiotics in aquaculture.