Abstract: One hundred and sixty endophytic bacteria strains were isolated from health tobacco plants in Guiyang county of Hunan province in disease severely infested plots. Thirty two, in particular three endophytic bacteria strains showed relatively strong antagonistic actions against Ralstonia solanacearum. Culturing these three bacteria and Ralstonia solanacearum together, the results showed that three endophytic bacteria H-1, H-9, and H-11 had the best antagonistic effects, and bacteriostasis diameters were 3.0, 3.0, and 4.5 mm, respectively. With regard to morphologic, physiological, and biochemical characterics, strain H-1, H-9, and H-11 were identified. The results showed that H-1 was Bacillus subtilis. H-9 and H-11 were Bacillus brevis. We also identified that they were all endophytic bacteria in tobacco.