Soil microbial communities play key roles in biogeochemical cycles and greenhouse gas formation during the decomposition of the released organic matter in the thawing permafrost. The aim of our research was to assess the taxonomic prokaryotic diversity in soil-ecological niches of the Darkhituy-Khaimisan transect during the initial period of soil thawing. We investigated changes in the microbial communities present in the active layer of four sites representing distinct habitats (larch forest, birch forest, meadow steppe and thermokarst lake). We explore the relationship between the biogeochemical differences among habitats and the active layer microbial community via a spatial (across habitats, and with depth through the active layer) community survey using high-throughput Illumina sequencing. Microbial communities showed significant differences between active and frozen layers and across ecosystem types, including a high relative abundance of Alphaproteobacteria, Firmicutes, Crenarchaeota, Bacteroidota and Gemmatimonadota in the active layer and a high relative abundance of Actinobacteriota and Desulfobacterota in the frozen layer. Soil pH, temperature and moisture were the most significant parameters underlying the variations in the microbial community composition. CCA suggested that the differing environmental conditions between the four soil habitats had strong influences on microbial distribution and diversity and further explained the variability of soil microbial community structures.