[Objective] Strain YBT2532 was screened from 400 Bacillus thuringiensis strains against Xanthomonas oryzae pv. oryzae. Its antimicrobial substance was isolated. [Methods] Physical and biochemistry characteristics of the antimicrobial active substance produced by the B. thuringiensis strain YBT-2532 were determined. [Results] It is not sensitive to temperature, protease and pH. After incubation at 70 °C for 1 h, still 75% activity retained. Proteinase K, trypsin and pepsin had no effect on its activity. The active substance was stable in the pH range between 2.0 and 12.0. It can be dissolved in water and methanol, weakly dissolved in ethanol, insoluble in acetone, chloroform and dichloromethane. Purification of the active substance was achieved using gel chromatography, ion exchange chromatography, solid phase extraction and HPLC. The purified active substance was analyzed by HPLC-IT-MS. It is a polar water soluble small molecule of 797.8 Da. [Conclusion] A new antibacterial substance was obtained from an isolate of Bacillus thuringiensis.