Abstract: In order to research the biocontrol of Bacillus cereus NJSZ-13 strain against pine wilt disease, single factor and orthogonal experimentswere conducted on optimization of medium and culture conditions. Single factor tests showed that the best carbon source in the medium was solublestarch, nitrogen source was soy peptone, and inorganic salt was calcium chloride. Orthogonal test demonstrated that the optimum proportion was:soluble starch 10 g/L, soy peptone 14 g/L, calcium chloride 6 g/L. The experiment resulted that the optimal culture conditions were: fermentationtemperature at 28℃, initial pH of 7.5; inoculum size of 2% and liquid volume of 30%. Compared with the culture conditions before optimization,viable count of strain NJSZ-13 in fermentation broth under the optimized conditions reached 4.10×109 cfu/mL, 4.1 times of that before one.