Investigation of the underlying microbial community and active acetate oxidation pathway in a full-scale commercial dry anaerobic digestion system

Anaerobic digestion (AD) is a well-established organic waste treatment biotechnology that enables resource recovery in the form of biomethane and nutrient-rich biosolids. AD systems treating municipal organic waste are prone to reduced biomethane yields and process upsets due to ammonia inhibition within their underlying microbial community. Microbial communities that are resilient to high ammonia conditions within AD are therefore of interest, given their ability to maintain stable and efficient AD reactor operation. Such microbial communities are generally characterized by the presence of a microbial guild performing syntrophic acetate oxidation (SAO) in partnership with hydrogenotrophic methanogens. So far, few members of the SAO guild have been identified and characterized. The objective of this study was to characterize the microbial community present within a high ammonia AD system and identify the taxa that were performing SAO. In this study, a commercial-scale biogas facility with a dry mesophilic AD system treating municipal organic wastes was monitored over the course of a year. Chemical monitoring of the AD system revealed high in situ ammonia concentrations, while a microbial community profile obtained via 16S rRNA gene amplicon sequencing suggested SAO as a dominant means of acetate utilization. In situ activity profiling was performed in batch microcosms with biomass sampled from the AD reactor using DNA stable isotope probing (SIP) amended with universally labelled ¹³C-acetate. DNA extracts recovered from the SIP incubations revealed a 5% shift in total community DNA ¹³C content. Following a consensus-based approach of high-resolution and quantitative SIP data analysis, 13 bacterial amplicon sequencing variants (ASVs) belonging to 8 genera were determined as isotopically enriched. Linking short-read 16S rRNA ASVs to long-read metagenomic assemblies allowed for accurate phylogenetic placement of acetate-incorporating taxa, as well as for an examination of potential metabolic pathways used for acetate oxidation by the active members. The results of this study provide new insights that can help guide process operations and optimization of the study AD system, as well as contribute to the emerging body of knowledge regarding SAO and microbial communities underpinning dry AD systems with high ammonia levels.Applied Science, Faculty ofCivil Engineering, Department ofGraduat