Abstract:[Objective] Two-stage thermophilic and mesophilic sludge anaerobic digestion process, which combines the processes of thermophilic digestion and mesophilic digestion, is a potentially important strategy for the development of sludge anaerobic digestion. Thus, it is necessary to investigate and compare the microbial community compositions in thermophilic and mesophilic anaerobic digestion systems. [Methods] Based on high-throughput sequencing and GeoChip techniques, we checked the 16S rRNA gene sequences of bacteria and archaea, ITS sequences of fungi and virus and virulence genes in anaerobic digestion systems, then compared the taxonomic and functional compositions between thermophilic and mesophilic microbial community. [Results] The taxonomic compositions of bacteria and archaea and the functional compositions of virus and virulence differed significantly between thermophilic and mesophilic anaerobic digestion systems, while the fungal taxonomic compositions were similar and the sequence reads number were relatively low. Methanogenus and unclassified bacteria and archaea were more abundant in mesophilic condition, while acid-producing and thermophilic microbes were more abundant in thermophilic condition. The relative abundance of genes associated with virus and virulence decreased after thermophilic digestion. In addition, microbial community compositions were significantly correlated with COD, TS and VS. [Conclusion] Microbial community compositions were significantly different between thermophilic and mesophilic anaerobic digestion, which explained functional distinction between the two ecosystems. The succession of microbial community was associated with influent parameters, revealing an important sensitivity.