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Background  A solid-state anaerobic digestion method is used to produce biogas from various solid wastes in China but the efficiency of methane production requires constant improvement. The diversity and abundance of relevant microorganisms play important roles in methanogenesis of biomass. The next-generation high-throughput pyrosequencing platform (Roche/454 GS FLX Titanium) provides a powerful tool for the discovery of novel microbes within the biogas-generating microbial communities.    Results  To improve the power of our metagenomic analysis, we first evaluated five different protocols for extracting total DNA from biogas-producing mesophilic solid-state fermentation materials and then chose two high-quality protocols for a full-scale analysis. The characterization of both sequencing reads and assembled contigs revealed that the most prevalent microbes of the fermentation materials are derived from Clostridiales (Firmicutes), which contribute to degrading both protein and cellulose. Other important bacterial species for decomposing fat and carbohydrate are Bacilli, Gammaproteobacteria, and Bacteroidetes (belonging to Firmicutes, Proteobacteria, and Bacteroidetes, respectively). The dominant bacterial species are from six genera:  Clostridium ,  Aminobacterium ,  Psychrobacter ,  Anaerococcus ,  Syntrophomonas , and  Bacteroides . Among them, abundant  Psychrobacter  species, which produce low temperature-adaptive lipases, and  Anaerococcus  species, which have weak fermentation capabilities, were identified for the first time in biogas fermentation. Archaea, represented by genera  Methanosarcina ,  Methanosaeta  and  Methanoculleus  of Euryarchaeota, constitute only a small fraction of the entire microbial community. The most abundant archaeal species include  Methanosarcina barkeri fusaro ,  Methanoculleus marisnigri JR1 , and  Methanosaeta theromphila , and all are involved in both acetotrophic and hydrogenotrophic methanogenesis.    Conclusions  The identification of new bacterial genera and species involved in biogas production provides insights into novel designs of solid-state fermentation under mesophilic or low-temperature conditions.

A pyrosequencing-based metagenomic study of methane-producing microbial community in solid-state biogas reactor