As an indicator and regulator of climate and environmental change, the Tibet Plateau is an important barrier for ecological security. However, despite the importance of soil microbial communities in almost all soil biochemical processes and ecosystem functions, the biogeography of soil microbial communities on the Tibet Plateau is poorly understood, especially at large scales over different ecosystem types. In this study, we collected samples from 64 sampling sites representing different grassland ecosystem types and spanning 2121 km across on the Tibet Plateau. We then used next generation high-throughput sequencing to investigate the soil prokaryote community (i.e. bacteria and archaea) diversity and spatial patterns and to explore their relationship with biotic (e.g. plant functional group diversity and biomass) and abiotic (e.g. aridity index, soil carbon and nitrogen levels) factors. Among the four alpine grassland types (i.e. alpine meadow, alpine steppe, alpine shrub and alpine desert) sampled in this study, alpine meadow had the highest soil microbial biomass and alpine desert had the lowest soil microbial richness and Shannon diversity. The soil microbial diversity in the alpine grassland correlated with plant diversity and climate factors. Soil microbial diversity negatively correlated with the annual average air temperature, but was not correlated with the annual average precipitation, indicating that temperature, rather than precipitation, may be more important in controlling the soil microbial diversity in alpine grassland ecosystems at cold temperatures. Higher air temperature likely led to an intensified aridity under limited precipitation, and thus decreased microbial diversity. As a result, the aridity index combined with temperature and precipitation explained more of the variance in the soil microbial diversity than air temperature or precipitation did individually. Moreover, after separating plant species into four functional groups (grass, forb, legume and sedge), microbial diversity positively correlated with plant functional group diversity, explaining more of the variance in microbial diversity than plant species diversity did. Results of structural equation modeling revealed that the aridity index and annual air temperature affected soil microbial diversity, directly or indirectly, through influencing plant functional group diversity and aboveground biomass; while aboveground biomass changed the soil carbon to nitrogen ratio in the upper soil layers and thus impacted soil microbial diversity. However, in contrast to microbial diversity, soil microbial biomass carbon was not correlated with plant functional group diversity, plant species diversity, or the climate factors annual average air temperature, annual precipitation and aridity index, but were linked to soil nutrient status (e.g. soil dissolved organic carbon, ammonia, available phosphorus, and carbon to nitrogen ratio) and plant biomass of sedges and forbs, demonstrating that microbial biomass and diversity were likely controlled by different factors. In summary, this study investigated the spatial patterns of soil microbial communities across different alpine grassland ecosystem types on the Tibet Plateau and enhanced our understanding of biotic and abiotic factors controlling microbial biomass and diversity, which will be important in predicting microbial changes on the Tibet Plateau under future climate change. Under future warming and wetting scenarios on the Tibet Plateau, it is possible that the aridity index would decrease, leading to increased soil microbial diversity. Results of this study also suggest a focus on the aridity index and plant functional group diversity in future microbial biogeography studies in order to further determine their roles in controlling or mediating soil microbial biomass and diversity.