Genome-Centered Metagenomics Analysis Reveals the Microbial Interactions of a Syntrophic Consortium during Methane Generation in a Decentralized Wastewater Treatment System

The application of anaerobic digestors to decentralized wastewater treatment systems (DWTS) has gained momentum worldwide due to their ease of operation, high efficiency, and ability to recycle wastewater. However, the microbial mechanisms responsible for the high efficiency and ability of DWTS to recycle wastewater are still unclear. In this study, the microbial community structure and function of two different anaerobic bioreactors (a primary sludge digestor, PSD, and anaerobic membrane bioreactor, AnMBR) of a DWTS located in Germany was investigated using 16S rRNA gene amplicon and metagenomic sequencing, respectively. The results showed that the microbial community structure was remarkably different in PSD and AnMBR. Methanobacteriaceae and Syntrophaceae were identified as the families that significantly differed in abundance between these two bioreactors. We also used genome-centered metagenomics to predict the microbial interactions and methane-generating pathway, which yielded 21 near-complete assembled genomes (MAGs) (average completeness of 93.0% and contamination of 2.9%). These MAGs together represented the majority of the microbial community. MAGs affiliated with methanogenic archaea, including Methanobacterium sp., Methanomicrobiales archaea, Methanomassiliicoccales archaea, and Methanosaeta concilii, were recruited, along with other syntrophic bacterial MAGs associated with anaerobic digestion. Key genes encoding enzymes involved in specific carbohydrate-active and methanogenic pathways in MAGs were identified to illustrate the microbial functions and interactions that occur during anaerobic digestion in the wastewater treatment. From the MAG information, it was predicted that bacteria affiliated with Bacteroidetes, Prolixibacteraceae, and Synergistaceae were the key bacteria involved in anaerobic digestion. In the methane production step, Methanobacterium sp. performed hydrogenotrophic methanogenesis, which reduced carbon dioxide to methane with hydrogen as the primary electron donor. Taken together, our findings provide a clear understanding of the methane-generating pathways and highlight the syntrophic interactions that occur during anaerobic digestion in DWTS.