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The present study was conducted to evaluate the supplementation of three autochthonous Bacillus strains (B. subtilis, B. amyloliquefaciens and B. cereus) and a commercial B. amyloliquefaciensin doses of 1 × 1010 CFU/kg on the growth performance, hematology, antioxidant activities, digestive enzyme levels, immune status and disease resistance of Clarias gariepinus. A total of 300 fish (75.23 ± 1.6 g) were randomly divided into 5 groups (each group was subdivided into 2 subgroups, 30 fish/each). The control group was fed basal diet (D0). Diets D1, D2, D3 and D4were supplemented with B. subtilis, B. amyloliquefaciens, B. cereus and a commercial B. amyloliquefaciens, respectively. During the course of the experiment, D3 showed the best body weight, weight gain, specific growth rate and food conversion ratio. The measured hemogram blood parameters had the highest significant increase in D3. WBCs and monocyte counts had no significant differences among the experimental groups. The serum antioxidant and digestive enzymes were the highest in D3 and were the lowest in D0. After 15 d, the non-specific immune parameters were markedly increased in fish fed probiotic-containing diet compared with the control. After 30 d, the highest significant immune parameters were observed in D3; D1 and D2 had no significant differences in serum lysozyme activity, nitric oxide and IgM compared with D0. Myostatin cDNA levels were adversely affected by probiotic supplements compare with the control. The PACAP expression showed the highest significant value in D3 followed by D1and D4then D2. The relative survival percentages of the Aeromonas sobria challenged C. gariepinus were the highest in D3, D2, D4 and then D1. Among the three isolated Bacillus species, dietary supplementation with the B. cereus had the highest performance in C. gariepinus compared with the commercial B. amyloliquefaciens and the control group.Bacillus species were improved growth performance of C. gariepinus.Bacillus species were enhanced digestive enzymes of C. gariepinus.The antioxidant response was improved by Bacillus dietary supplementation.PACAP of C. gariepinus was increased after the supplementation of Bacillus sp.C. gariepinus fed Bacillus supplemented diet had highest post-challenge relative percent survival.