Enrichment and physiological characterization of a coldadapted nitrite-oxidizing Nitrotoga sp. from an eelgrass sediment

Nitrite-oxidizing bacteria (NOB) are responsible for the second step of nitrification in natural and engineered ecosystems. The recently discovered genus Nitrotoga belongs to the Betaproteobacteria and potentially has high environmental importance. Although environmental clones affiliated with Nitrotoga are widely distributed, the limited number of cultivated Nitrotoga spp. results in a poor understanding of their ecophysiological features. In this study, we successfully enriched the nonmarine cold-adapted Nitrotoga sp. strain AM1 from coastal sand in an eelgrass zone and investigated its physiological characteristics. Multistep-enrichment approaches led to an increase in the abundance of AM1 to approximately 80% of the total bacterial population. AM1 was the only detectable NOB in the bacterial community. The 16S rRNA gene sequence of AM1 was 99.6% identical to that of "Candidatus Nitrotoga arctica," which was enriched from permafrost-affected soil. The highest nitrogen oxidation rate of AM1 was observed at 16°C. The half-saturation constant (K_m) and the generation time were determined to be 25 μM NO₂ ^- and 54 h, respectively. The nitrite oxidation rate of AM1 was stimulated at concentrations of <30 mM NH₄Cl but completely inhibited at 50 mM NH₄Cl. AM1 can grow well under specific environmental conditions, such as low temperature and in the presence of a relatively high concentration of free ammonia. These results help improve our comprehension of the functional importance of Nitrotoga.