Nitrite oxidation in geothermal springs: evidence of an upper temperature limit for thermophilic nitrite-oxidizing bacteria of 60 - 65 °C

Despite its importance in the nitrogen cycle, little is known about nitrite oxidation at high temperatures. In order to bridge this knowledge gap, enrichment cultures were inoculated with sediment slurries from a variety of geothermal springs. While attempts failed to enrich nitrite-oxidizing bacteria (NOB) from >10 hot spring samples at temperatures ≥61 °C, NOB were successfully enriched from the outflow of 7 hot springs located in U.S. Great Basin, southwestern China, and Armenia at temperatures ≤57.9 °C. The stoichiometric conversion of nitrite to nitrate, chlorate sensitivity, and the loss of activity after autoclaving all confirmed biological nitrite oxidation in these enrichments. Regardless of their origin, all successful enrichments contained organisms with high 16S rRNA gene sequence identity (≥97%) with Nitrospira calida. In addition, Armenian enrichments also contained organisms with high 16S rRNA gene sequence identity to Nitrospira moscoviensis. Physiological properties of all enrichments were similar, with a temperature optimum of 45 50 °C and an upper temperature limit between 60 °C and 65 °C. The highest rates of NOB activity occurred with initial NO2 concentrations of 0.5 0.75 mM; however, lower initial nitrite concentrations resulted in shorter lag times. The results presented here suggest a possible upper temperature limit of 60 – 65 °C for 7 chemolithotrophic nitrite oxidation and demonstrate the wide geographic range of Nitrospira spp. in geothermal environments.