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By catalyzing the second step of nitrification\u2014the oxidation of nitrite to nitrate to generate energy for growth\u2014NOB activity controls the fate of nitrite (NO\n                      <jats:sub>2<\/jats:sub>\n                      <jats:sup>-<\/jats:sup>\n                      ) in aerobic environments. Despite thriving in diverse environments, including soils, freshwater, marine ecosystems, subsurface habitats, and water treatment systems, organisms capable of nitrite oxidation are confined to\n                      <jats:italic toggle=\"yes\">Nitrobacter, Nitrospira, Nitrospina, Nitrotoga,<\/jats:italic>\n                      and a few other specific lineages. The genus\n                      <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                      , recognized for its facultative heterotrophic metabolism, is often associated with high-nitrogen environments. Here, we report the physiological characterization of a novel strain,\n                      <jats:italic toggle=\"yes\">Nitrobacter vulgaris<\/jats:italic>\n                      strain MLSD-S22, isolated from a nitrate- and heavy-metal-contaminated subsurface. Growth inhibition experiments revealed that strain MLSD-S22 and the\n                      <jats:italic toggle=\"yes\">N. vulgaris<\/jats:italic>\n                      type strain Z exhibited similar sensitivities to nitrite and nitrate, with nitrite being the most inhibitory. Microrespirometry demonstrated that the two\n                      <jats:italic toggle=\"yes\">N. vulgaris<\/jats:italic>\n                      strains and\n                      <jats:italic toggle=\"yes\">Nitrobacter winogradskyi<\/jats:italic>\n                      Nb-255 possessed higher affinities for nitrite and oxygen than previously reported for\n                      <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                      , suggesting potential to compete in low-substrate environments. Long-read DNA sequencing provided a complete genome for strain MLSD-S22, revealing two plasmids and an intact nitrous oxide (N\n                      <jats:sub>2<\/jats:sub>\n                      O) reduction operon\u2014an unexpected feature for\n                      <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                      . While N\n                      <jats:sub>2<\/jats:sub>\n                      O reduction activity was not observed under the tested conditions, this discovery raises questions about the contribution of\n                      <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                      NOB to the N\n                      <jats:sub>2<\/jats:sub>\n                      O sink. These findings broaden the physiological and genomic diversity of\n                      <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                      , offering new insights into their adaptation strategies and providing a framework for future evaluation of their potential roles in nitrogen loss.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                        species play a key role in nitrogen cycling; however, their physiological adaptations and genomic diversity remain largely underexplored. This study characterizes\n                        <jats:italic toggle=\"yes\">Nitrobacter vulgaris<\/jats:italic>\n                        strain MLSD-S22, isolated from a nitrate- and heavy-metal-contaminated subsurface, revealing unexpected genomic and metabolic traits. Higher nitrite and oxygen affinities compared to previous reports indicate that some\n                        <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                        are more competitive in low-substrate environments than previously believed. The presence of a complete nitrous oxide (N\n                        <jats:sub>2<\/jats:sub>\n                        O) reduction operon highlights the genomic potential related to N\n                        <jats:sub>2<\/jats:sub>\n                        O reduction within this genus; however, we did not observe N\n                        <jats:sub>2<\/jats:sub>\n                        O reduction under the conditions tested. By expanding\n                        <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                        ecophysiology, this work provides a foundation for future research on the metabolic flexibility of\n                        <jats:italic toggle=\"yes\">Nitrobacter<\/jats:italic>\n                        and their contributions to nitrogen transformations in diverse environments.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/aem.00130-26","type":"journal-article","created":{"date-parts":[[2026,3,24]],"date-time":"2026-03-24T13:00:16Z","timestamp":1774357216000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Comparative physiological and genomic characterization of a novel\n                    <i>Nitrobacter vulgaris<\/i>\n                    strain from a nitrate-contaminated subsurface"],"prefix":"10.1128","volume":"92","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0596-6416","authenticated-orcid":true,"given":"Zachary","family":"Flinkstrom","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/00cvxb145","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Civil and Environmental Engineering, University of Washington","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kristopher A.","family":"Hunt","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/00cvxb145","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Civil and Environmental Engineering, University of Washington","place":["Seattle, USA"]},{"id":[{"id":"https:\/\/ror.org\/011fdmv87","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute for Systems Biology","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Britt","family":"Abrahamson","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/00cvxb145","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Civil and Environmental Engineering, University of Washington","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pierce","family":"Harvell","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02aqsxs83","id-type":"ROR","asserted-by":"publisher"}],"name":"School of Biological Sciences, Institute for Environmental Genomics, University of Oklahoma","place":["Norman, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiangpeng","family":"Li","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02aqsxs83","id-type":"ROR","asserted-by":"publisher"}],"name":"School of Biological Sciences, Institute for Environmental Genomics, University of Oklahoma","place":["Norman, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James","family":"Wilson","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/011fdmv87","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute for Systems Biology","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zachary S.","family":"Cooper","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/011fdmv87","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute for Systems Biology","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacob J.","family":"Valenzuela","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/011fdmv87","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute for Systems Biology","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9157-5974","authenticated-orcid":true,"given":"Nitin S.","family":"Baliga","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/011fdmv87","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute for Systems Biology","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David A.","family":"Stahl","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/00cvxb145","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Civil and Environmental Engineering, University of Washington","place":["Seattle, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0895-0126","authenticated-orcid":true,"given":"Wei","family":"Qin","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/02aqsxs83","id-type":"ROR","asserted-by":"publisher"}],"name":"School of Biological Sciences, Institute for Environmental Genomics, University of Oklahoma","place":["Norman, USA"]},{"id":[{"id":"https:\/\/ror.org\/047426m28","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Microbiology, School of Molecular and Cellular Biology, Carl R. 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