Grazing and mowing are two dominant management regimes in the Inner Mongolia grassland, both of which have a profound impact on grassland ecosystem, and most of the previous studies have primarily focused on the effects on plant and soil. Soil microbial communities are the most abundant and diverse groups of organisms on earth and are responsible for numerous key ecosystem processes. They play an important role in the circulation of materials and nutrients between plants and soil ecosystems. However, their response to these management regimes has not been comprehensively explored in Stipa grandis grassland. In this study, two management regimes of grazing and mowing were set up in a typical S. grandis area, and enclosure was used as the control. Based on high-throughput sequencing technology, the changes of soil microbial composition and diversity under different management regimes was examined, Redundancy analysis was used to analysis soil physical and chemical characteristics influencing on it. The results showed that there was no significant difference in soil bacterial α diversity index under different management regimes, but mowing significantly increased soil fungal Observed_species, Chao1 and ACE index. Amplicon sequencing revealed Proteobacteria and Actinobacteria were the dominant bacterial phylum under different grassland management regimes, and the dominant phyla of soil fungal communities were Ascomycota and Mortierellomycota. Grazing significantly increased the relative abundance of Proteobacteria, Verrucomicrobia and Gemmatimonadetes of the bacterial community, and mowing significantly increased the relative abundance of Basidiomycota of fungal community. In addition, grazing and mowing significantly reduced the relative abundance of Firmicutes. Redundancy analysis showed that soil bacteria were mainly affected by nitrate nitrogen and ammonium nitrogen, which were positively correlated with Acidobacteria and negatively correlated with Proteobacteria. Soil fungi were mainly affected by available phosphorus and nitrate nitrogen. In conclusion, changes in grassland usage methods altered the composition and diversity of soil microflora through soil physicochemical factors, and soil ammonium nitrogen, nitrate nitrogen and available phosphorus were the main driving factors on microbial community formation and evolution. This study revealed the structural characteristics and diversity of soil microbial communities in the S. grandis steppe under different management regimes and their relationship with soil physicochemical properties, which may provide a theoretical basis for the rational utilization of S. grandis steppe.