Denitrifying anaerobic methane oxidizing (DAMO) microorganisms were enriched from paddy field soils using continuous-flow and batch cultures fed with nitrate or nitrite as a sole electron acceptor. After several months of cultivation, the continuous-flow cultures using nitrite showed remarkable simultaneous methane oxidation and nitrite reduction and DAMO bacteria belonging to phylum NC10 were enriched. A maximum volumetric nitrite consumption rate of 70.4±3.4 mg-N·L−1·day−1 was achieved with very short hydraulic retention time of 2.1 hour. In the culture, about 68% of total microbial cells were bacteria and no archaeal cells were detected by fluorescence in situ hybridization. In the nitrate-fed continuous-flow cultures, 58% of total microbial cells were bacteria while archaeal cells accounted for 7% of total cell numbers. Phylogenetic analysis of pmoA gene sequence showed that enriched DAMO bacteria in the continuous-flow cultivation had over 98% sequence similarity to DAMO bacteria in the inoculum. In contrast, for batch culture, the enriched pmoA gene sequences had 89–91% sequence similarity to DAMO bacteria in the inoculum. These results indicate that electron acceptor and cultivation method strongly affect the microbial community structures of DAMO consortia.
Anaerobic treatment of sewage has many advantages; however, the effluent contains high levels of dissolved methane. In this study, we investigated the use of a closed-type downflow hanging sponge (DHS) reactor for application of the denitrifying anaerobic methane oxidation (DAMO) reaction for nitrogen and dissolved methane removal. When using nitrate, the DAMO reaction achieved a denitrification rate of 84.4 g N m day, which is close to that required for practical application of denitrification to anaerobic sewage treatment. The microbial community that developed in the DHS was investigated using16S rRNA, and novel species of DAMO bacteria affiliated with Group b of NC10 phylum were enriched. This contrasted with the results of previous studies in which the Candidatus Methylomirabilis oxyfera affiliated with Group a was enriched. The results obtained herein suggest that a post-treatment system for anaerobically treated sewage using a closed-type DHS reactor may become practical in the near future.
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