Abstract: In 2014, 35 suspected tobacco bacterial wilt disease samples were collected from Luoshan, Queshan and Suiping counties of southern areas in Henan Province and 26 bacterial strains were isolated. The sequence similarity between the 26 strains and the Ralstonia solanacearum in NCBI database was more than 98 percent. Typical symptoms of bacterial wilt were observed when the isolated strains were used to inoculate tobacco and Ralstonia solanacearum colonies were obtained on TTC medium from the infected tissues. All of the above indicated that the isolated strains were pathogen of tobacco bacterial wilt disease. This study showed that tobacco bacterial wilt occurred in southern areas of Henan Province.