The shift toward plant-based feeds in aquaculture necessitates a search for probiotics to improve the digestibility of such feeds and fish growth. This study characterizes four Bacillus strains for their probiotic potential in carp (Cyprinus carpio). Strains isolated from river sediments underwent in vitro screening for proteolytic/amylolytic activity, antibiotic sensitivity, and biofilm formation. Whole-genome sequencing was performed and functional annotation identified probiotic-related genes. All strains were identified as Bacillus velezensis and harbored genes providing thermal, osmotic, and oxidative stress resistance, alongside enzymes and bioactive metabolite synthesis pathways. Two non-antagonistic strain pairs were solid-phase cultivated on soybeans and incorporated into carp feed (0.1% w/w) for a 76-day trial, and growth metrics and gene expression were analyzed in juvenile fish. Group 1 (strains MT14 + MT42) exhibited a 40.75% higher biomass increase than the control group while Group 2 (strains MT141 + MT142) exhibited a 56.62% higher biomass growth. No significant gene expression changes occurred in juveniles, though MT141 + MT142 transiently modulated fry expression profiles. Therefore, these Bacillus velezensis strains synergistically enhance growth in carp that are fed plant-rich diets, highlighting their viability as aquaculture probiotics.

The potential probiotic properties of three Bacillus strains were studied. A probiotic supplement for the African catfish Clarias gariepinus was produced via the solid-state fermentation protocol and incorporated into the fish feed for a period of seven weeks. Since the 36th day of the experiment, all experimental groups had a statistically significant increase in their weight gain than the control group. The maximum weight gain observed in fish fed the probiotic-supplemented feed was 29.16% higher than that of the control group, and the maximum feed conversion rate improvement was 24%. Cell-free extracts from these strains showed antioxidant (11.55–27.40%) and DNA-protective (45.33–61.83%) activity in a series of in vitro biosensor tests. Further investigation into the antimutagenic activity of the strains revealed that two of them reduced the level of induced mutagenesis in an Escherichia coli model (by 33.58% and 54.35%, respectively). We also assessed the impact of probiotic strains on the expression of several key genes in the host (C. gariepinus), including hsp70, cxc, tnfα, il1β, and lysC. More than a 10-fold increase in expression rates was observed for hsp70 in gonads and liver; for cxc in muscles and gonads; for tnfα in brain, gills, and liver; for il1β in the brain, gills, gonads, and liver; and for lysC in gills, gonads, liver, and muscles. This study provides evidence that probiotics exhibiting antioxidant and antimutagenic properties can provide significant benefits in vivo within aquaculture systems. The molecular effects of these probiotics appear to be complex and tissue-specific, with both upregulation and downregulation of immune system genes observed. Nevertheless, at the organismal level, the impact was unequivocally positive in terms of aquaculture objectives, manifested as enhanced body weight gain in the fish. Consequently, these Bacillus strains warrant serious consideration as potential probiotics for this species.