Abstract: In order to systematically understand the main pathogenic fungi in the rhizosphere of tobacco, diseased rhizosphere samples were collected from three tobacco planting areas. Then pathogen isolation and identification were performed in the laboratory, and 64 strains of pathogenic fungi were obtained. According to the morphological characteristics of colonies, mycelium, and spores, as well as the phylogenetic analysis of multiple gene loci, the strains were identified at the species level; and their pathogenicity were tested in leaf, stem, and root of tobacco. The results showed that the isolated pathogens included Fusarium oxysporum, F. sambucinum, F. commune, F. sulawesiense, F. citri, F. ipomoeae, Lasiodiplodia brasiliense, Rhizopus microspores, and Cladosporium cladosporioides. Pathogenicity assays indicated that these pathogens had certain pathogenicity to tobacco. The pathogenicity of F. sambucinum, F. oxysporum, F. commune and L. brasiliense were strong. The pathogenicity of F. sulawesiense, F. citri, F. ipomoeae, R. microspores and C. cladosporioides were weak. The classification and identification results of the pathogenic fungi will provide a foundation for the scientific, reasonable, and effective control of tobacco rhizosphere diseases.