Abstract: Bacillus subtilis strain HMB19198 is a promising biocontrol agent for tomato gray mold. Fengycin, a kind of lipopeptide, plays an important role in strain HMB19198 suppressing tomato gray mold. To improve the fermentation level of strain HMB19198, the basic medium benefiting for the sporulation of strain HMB19198 was selected from eight kinds of industrial mediums. The carbon and nitrogen sources suitable for sporulation of strain HMB19198 were screened by a single factor test. Results showed that soluble corn starch and molasses as carbon sources, peanut cake powder, and peptone as nitrogen sources were conducive to the sporulation of strain HMB19198. The Plackett-Burman analysis and response surface analysis were used to optimize the concentrations of soluble corn starch, molasses, peanut cake powder, peptone, K₂HPO₄•3H₂O, and MgSO₄•7H₂O in the medium. The optimized medium included 67.0 g/L soluble corn starch, 14.1 g/L molasses, 41.6 g/L peanut cake powder, 10 g/L peptone, 9.2 g/L K₂HPO₄•3H₂O and 1.5 g/L MgSO₄•7H₂O. In the optimized medium, the spore concentration of strain HMB19198 reached 6.92 × 10⁹ spore/mL. Compared with the basic medium, using the optimized medium, the fermentation period of the strain was shortened by 40%, the spore concentration in the fermentation broth was increased by 107%, and the fengycin concentration was increased by 39.4%.