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Studies have found Ca. Nitrospira nitrosa-like bacteria to be the principal or sole comammox bacteria in nitrogen removal systems for wastewater treatment. In contrast, multiple populations of strict ammonia and nitrite oxidizers co-exist in similar systems. This apparent lack of diversity is surprising and could impact the feasibility of leveraging comammox bacteria for nitrogen removal. We used full-length 16S rRNA gene sequencing and genome-resolved metagenomics to compare population-level (i.e., species) diversity of comammox bacteria with that of strict nitrifiers in full-scale wastewater treatment systems and assess whether these observations were consistent or diverged at the strain-level. Full-length 16S rRNA gene sequencing indicated that while Nitrosomonas-like bacteria exhibited higher population-level diversity, the effective microdiversity of most Nitrospira-like bacteria were comparatively higher except for one Nitrospira Lineage II population. Comammox bacterial metagenome assembled genomes (MAGs) were associated with Ca. Nitrospira nitrosa. The average amino acid identity between comammox bacterial MAGs (93%) across systems was significantly higher than that of the Nitrosomonas-like ammonia oxidizers (73%) and the Nitrospira-like nitrite oxidizer MAGs (75), suggesting that the same comammox population was detected in all systems. Comammox bacteria and some ammonia oxidizers MAGs were significantly less microdiverse than most ammonia and nitrite oxidizers. Interestingly, strain-resolved analysis also indicates that different nitrogen removal systems harbor different comammox bacterial strains within the Ca. Nitrospira nitrosa cluster. These results suggest that comammox bacteria associated with Ca. Nitrospira nitrosa have low species- and strain-level diversity in nitrogen removal systems and may thus harbor specific adaptations to the wastewater ecosystem.

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