Influence of Season, Occupancy Pattern, and Technology on Structure and Composition of Nitrifying and Denitrifying Bacterial Communities in Advanced Nitrogen-Removal Onsite Wastewater Treatment Systems

Advanced onsite wastewater treatment systems (OWTS) use biological nitrogen removal (BNR) to mitigate the threat that N-rich wastewater poses to coastal waterbodies and groundwater. These systems lower the N concentration of effluent via sequential microbial nitrification and denitrification. We used high-throughput sequencing to evaluate the structure and composition of nitrifying and denitrifying bacterial communities in advanced N-removal OWTS, targeting the genes encoding ammonia monooxygenase (<i>amoA</i>) and nitrous oxide reductase (<i>nosZ</i>) present in effluent from 44 advanced systems. We used QIIME2 and the phyloseq package in R to examine differences in taxonomy and alpha and beta diversity as a function of advanced OWTS technology, occupancy pattern (seasonal vs. year-round use), and season (June vs. September). Richness and Shannon’s diversity index for <i>amoA</i> were significantly influenced by season, whereas technology influenced <i>nosZ</i> diversity significantly. Season also had a strong influence on differences in beta diversity among <i>amoA</i> communities, and had less influence on <i>nosZ</i> communities, whereas technology had a stronger influence on <i>nosZ</i> communities. <i>Nitrosospira</i> and <i>Nitrosomonas</i> were the main genera of nitrifiers in advanced N-removal OWTS, and the predominant genera of denitrifiers included <i>Zoogloea</i>, <i>Thauera</i>, and <i>Acidovorax</i>. Differences in taxonomy for each gene generally mirrored those observed in diversity patterns, highlighting the possible importance of season and technology in shaping communities of <i>amoA</i> and <i>nosZ</i>, respectively. Knowledge gained from this study may be useful in understanding the connections between microbial communities and OWTS performance and may help manage systems in a way that maximizes N removal.