Expanded Diversity and Metabolic Versatility of Marine Nitrite-Oxidizing Bacteria Revealed by Cultivation- and Genomics-Based Approaches

Abstract

Nitrite-oxidizing bacteria (NOB) are ubiquitous and abundant microor- ganisms that play key roles in global nitrogen and carbon biogeochemical cycling. Despite recent advances in understanding NOB physiology and taxonomy, currently very few cultured NOB or representative NOB genome sequences from marine envi-ronments exist. In this study, we employed enrichment culturing and genomic ap-proaches to shed light on the phylogeny and metabolic capacity of marine NOB. We successfully enriched two marine NOB (designated MSP and DJ) and obtained a high-quality metagenome-assembled genome (MAG) from each organism. The maxi-mum nitrite oxidation rates of the MSP and DJ enrichment cultures were 13.8 and 30.0 µM nitrite per day, respectively, with these optimum rates occurring at 0.1 mM and 0.3 mM nitrite, respectively. Each enrichment culture exhibited a different toler-ance to various nitrite and salt concentrations. Based on phylogenomic position and overall genome relatedness indices, both NOB MAGs were proposed as novel taxa within the Nitrospinota and Nitrospirota phyla. Functional predictions indicated that both NOB MAGs shared many highly conserved metabolic features with other NOB. Both NOB MAGs encoded proteins for hydrogen and organic compound metabolism and defense mechanisms for oxidative stress. Additionally, these organisms may have the genetic potential to produce cobalamin (an essential enzyme cofactor that is limiting in many environments) and, thus, may play an important role in recycling cobalamin in marine sediment. Overall, this study appreciably expands our under-standing of the Nitrospinota and Nitrospirota phyla and suggests that these NOB play important biogeochemical roles in marine habitats. IMPORTANCE Nitrification is a key process in the biogeochemical and global nitro-gen cycle. Nitrite-oxidizing bacteria (NOB) perform the second step of aerobic nitrifi-cation (converting nitrite to nitrate), which is critical for transferring nitrogen to other organisms for assimilation or energy. Despite their ecological importance, there are few cultured or genomic representatives from marine systems. Here, we obtained two NOB (designated MSP and DJ) enriched from marine sediments and estimated the physiological and genomic traits of these marine microbes. Both NOB enrichment cultures exhibit distinct responses to various nitrite and salt concentra-tions. Genomic analyses suggest that these NOB are metabolically flexible (similar to other previously described NOB) yet also have individual genomic differences that likely support distinct niche distribution. In conclusion, this study provides more in-sights into the ecological roles of NOB in marine environments.