AbstractDue to the emergence of antibiotic-resistant pathogens, probiotics in aquaculture are used for the prevention of infectious microbial diseases and substitute for antibiotics and chemotherapeutics. In this study, we evaluated the effect of probiotic Bacillus spp. isolated from African nightcrawler (Eudrilus eugeniae) on the growth, feed utilization, and disease resistance of Nile tilapia (Oreochromis niloticus). Four probiotic strains of Bacillus spp. (ANSCI9, BFAR9, RM3, and RM10) were individually incorporated in the commercial diet (control) at 108 CFU g-1 of feed. The experimental fish were fed at 5% of their body weight for 30 days, and subjected to a 14-day Aeromonas hydrophila challenge test afterward. The results showed the probiotic-treated groups have higher (P<0.05) average body weight (ABW) (4.51 ± 0.34 g) than the control (3.89 ± 0.17 g). The BFAR9 (2.73 ± 0.26 g) and RM10 (3.15 ± 0.30 g) showed higher (P<0.05) absolute growth (AG) than the control (2.20 ± 0.16 g). Furthermore, RM10 had higher (P<0.05) specific growth rate (SGR) (1.60 ± 0.10 % day-1) and relative growth rate (RGR) (181.39 ± 18.16 %) than the control (SGR=1.29 ± 0.07 % day-1; RGR=129.84 ± 9.77 %). Consequently, RM10 had significantly lower (P<0.05) feed conversion ratio (FCR) (1.99 ± 0.13) than the control (2.60 ± 0.16). The challenge test revealed that the probiotic-treated groups have higher (P<0.05) survival (81.25 ± 9.57 %) than the control (55.00 ± 19.15 %). These results revealed that the probiotic Bacillus spp. isolated from E. eugeniae improved the growth, feed utilization, and the disease resistance of Nile tilapia.