Abstract: Aiming for the changes of microbial community composition, structure, and species balance in cigar tobacco fermentation, the surface bacteria and fungi of cigar tobacco at different fermentation stages were analyzed by using high-throughput sequencing technology and ratio analysis method of bacterial and fungal diversity index. Results showed that with the fermentation process, the α diversity of bacteria increased at first and then decreased, and peaked in PEF3 (middle stage). The α diversity of fungi was basically stable, yet significantly decreased in PEF4 (late stage). The species balance slightly increased in early and middle stages, while significantly decreased after PEF4. PCoA analysis showed that the bacterial community structure of unfermented and pre fermentation exhibited similar, while significantly separated from that of post fermentation. The dominant genera in fermentation were bacterial Staphylococcus, Pseudomonas, and fungal Streptomyces, Aspergillus, Penicillium, respectively. The key microbial community changes affecting species balance are from Pantoea, Methylobacterium, and Sphingomonas of early stage to Massilia, Agromyces, Rhizobiaceae of middle stage, and finally stabilized with Staphylococcus, Pseudomonas, Trichomonas, Ralstonia and Monogramella. Microorganisms were divided into two modules, and a total of 8 key genera beneficial for improving species balance were selected out. The final structural equation model indicates that in fermentation of cigar tobacco, the main environmental factors affecting species balance are temperature and humidity, which affect species balance by affecting the abundance of key bacterial genera.