ABSTRACT The recently discovered comammox bacteria have the potential to completely oxidize ammonia to nitrate. These microorganisms are part of the Nitrospira genus and are present in a variety of environments, including Biological Nutrient Removal (BNR) systems. However, the physiological traits within and between comammox- and nitrite oxidizing bacteria (NOB)-like Nitrospira species have not been analyzed in these ecosystems. In this study, we identified Nitrospira strains dominating the nitrifying community of a sequencing batch reactor (SBR) performing BNR under micro-aerobic conditions. We recovered metagenomes-derived draft genomes from two Nitrospira strains: (1) Nitrospira sp. UW-LDO-01, a comammox-like organism classified as Candidatus Nitrospira nitrosa, and (2) Nitrospira sp. UW-LDO-02, a nitrite oxidizing strain belonging to the Nitrospira defluvii species. A comparative genomic analysis of these strains with other Nitrospira-like genomes identified genomic differences in Ca. Nitrospira nitrosa mainly attributed to each strains’ niche adaptation. Traits associated with energy metabolism also differentiate comammox from NOB-like genomes. We also identified several transcriptionally regulated adaptive traits, including stress tolerance, biofilm formation and micro-aerobic metabolism, which might explain survival of Nitrospira under multiple environmental conditions. Overall, our analysis expanded our understanding of the genetic functional features of Ca. Nitrospira nitrosa, and identified genomic traits that further illuminate the phylogenetic diversity and metabolic plasticity of the Nitrospira genus.