Abstract: In order to improve the flocculation activity of the microbial flocculant which can treat tannery wastewater and was produced by Bacillus fusiformis, the culture conditions of Bacillus fusiformis were optimized. Taking the flocculation rate as an index, the optimum culture temperature, inoculation quantity, pH and culture time of Bacillus fusiformis producing bioflocculant were determined by single factor experiment at 31 ℃, 6%, 5 or 6 and 48 h respectively. Then, the orthogonal test for four factors and three levels (L₉(3⁴)) was used to determine the optimum temperature for the production of bioflocculant by Bacillus fusiformis,which was 33 ℃.And the optimum inoculation concentration was 6%, the optimum pH was 5, and the optimum culture time was 54 h. Compared with the initial culture conditions, the flocculation rate of the microbial flocculants prepared under the optimized culture conditions increased from 46.9% to 62.1%, with an increase of 32.4%. Thus the culture conditions of microbial flocculant produced by Bacillus fusiformis were optimized, which was contribute to the application of the microbial flocculant produced by Bacillus fusiformis to treat tannery wastewater.