[Background] Tobacco aging is a process involving the interaction of many factors. [Objective] In order to investigate the relationship between the structure, function and chemical composition of culturable microflora on the surface of aging tobacco leaves. [Methods] The microflora of tobacco leaves stored in Guiyang, Tanchang and Maotai storerooms were isolated at different aging time respectively. The function of bacteria and fungi were analyzed by annotating to the FAPROTAX and FUNGuild databases respectively. At the same time, combining with the main chemical components of tobacco samples, the correlation analysis of culturable microflora was carried out. [Results] The results showed that 189 strains of dominant bacteria and 229 strains of dominant fungi were isolated from 243 tobacco leaf samples. Altogether, Bacillus was the dominant population in bacteria, whereas Aspergillus and Penicillium were accounted for dominant fungal populations. With the extension of aging time, the proportion of dominant populations and dominant functional groups were decreased gradually, and in the other side, the main chemical substance was correlated with the change of microbial communities significantly. [Conclusion] It is demonstrate that the structural changes of microbial functional groups have a contribution to accelerate tobacco aging process. In turn, the changes of major chemical components during the aging process could affect the composition and function of microbial communities.