Abstract: Tobacco black shank is widely distributed in tobacco growing areas in China and has serious effects on tobacco production and quality.One of the potential control approaches against tobacco black shank is to use antagonistic bacterial.Using the culture plate dilution method,YBM-4 and YJC-4,two bacterial strains were isolated from tobacco rhizosphere soil to have an antagonistic effect on tobacco black shank.Based on comprehensive analysis on morphology,biological traits and physiological and biochemical characteristics,and 16S rDNA gene sequence analysis,YBM-4 was identified as Bacillus subtilis and YJC-4 was identified as Bacillus safensis.The two strains of YJC-4 and YBM-4 had significant antagonism against tobacco black shank.In paired cultures,relative inhibition rate of YJC-4 and YBM-4 reached 85.34% and 72.99%.In the pot experiment,effects of YBM-4 and YJC-4 on tobacco agronomic traits and biomass growth promoting were obvious.The control effects on tobacco black shank of YBM-4,YJC-4 and YBM-4+YJC-4 mixture (1:1 ratio) were 76.37%,80.06% and 81.50%,with the AUDPC lower than that of the control group by 265.01,281.58,and 287.32 respectively.YBM-4 and YJC-4 have a potential in bio-control of tobacco black shank.