Enrichment and Physiological Characterization of a Cold-Adapted Nitrite-Oxidizing Nitrotoga sp. from an Eelgrass Sediment

Abstract

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 M NO 2 Ϫ and 54 h, respectively. The nitrite oxidation rate of AM1 was stimulated at concentrations of Ͻ30 mM NH 4 Cl but completely inhibited at 50 mM NH 4 Cl. 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.IMPORTANCE Nitrite-oxidizing bacteria (NOB) are key players in the second step of nitrification, which is an important process of the nitrogen cycle. Recent studies have suggested that the organisms of the novel NOB genus Nitrotoga were widely distributed and played a functional role in natural and engineered ecosystems. However, only a few Nitrotoga enrichments have been obtained, and little is known about their ecology and physiology. In this study, we successfully enriched a Nitrotoga sp. from sand in a shallow coastal marine ecosystem and undertook a physiological characterization. The laboratory experiments showed that the Nitrotoga enrichment culture could adapt not only to low temperature but also to relatively high concentrations of free ammonia. The determination of as-yet-unknown unique characteristics of Nitrotoga contributes to the improvement of our insights into the microbiology of nitrification.KEYWORDS Nitrospira, Nitrotoga, ammonia, coastal sand, cultivation, enrichment, microbial communities, nitrification, nitrite-oxidizing bacteria, physiology I n nitrification, ammonia is oxidized into nitrate via nitrite by phylogenetically different chemolithoautotrophic microorganisms, and this is an integral part of the global nitrogen cycle. The reactions are catalyzed by ammonia-oxidizing bacteria (AOB),

Genomic profiling of four cultivated Candidatus Nitrotoga spp. predicts broad metabolic potential and environmental distribution

“…If Ca. Nitrotoga are indeed motile, they may migrate towards areas of greater reducing potential, and likely lose external motile structures when growing in biofilms or in culture, as the presence of flagella or pili have not been noted in previous studies [2,13,[33][34][35].…”

“…A Ca. Nitrotoga enrichment culture from coastal sands grew faster when ammonium was added to the culture medium [35], likely due to the reduced need for assimilatory nitrite reduction.…”

“…All previously reported Ca. Nitrotoga cultures were enriched at low temperatures (4-17°C) [2,[33][34][35]. Sampling for the Ca.…”

Longterm Monitoring of Nitrification and Nitrifying Communities during Biofilter Activation of Two Marine Recirculation Aquaculture Systems (RAS)

“…Only recently was a non-marine Nitrotoga enriched from eelgrass sediment growing best at 16°C, with a 16S rRNA gene similarity of 99.6 % to Cand. Nitrotoga arctica and 99.3 % to the sequences in this study [60]. Thus, the frequent detection of this nitrite oxidizer especially in the relatively warm biofi lter of RAS 2 was quite surprising and should encourage future research to consider the use of Nitrotoga-specifi c primers and probes when analyzing nitrifying communities in marine environments.…”

Unexpected versatility in the metabolism and ecophysiology of globally relevant nitrite-oxidizingNitrotogabacteria

“…Exopolysaccharide (EPS) synthesis is also likely in Nitrotoga, as an extensive array of PEP-CTERM exopolysaccharide sorting enzymes (Haft et al, 2006), polysaccharide export pumps, and saccharide modifiers (i.e., UDP-glucose dehydrogenase, polyprenyl glycosylphosphotransferase, D-alanyl-lipoteichoic acid acyltransferase) were present in all genomes. EPS production has been observed in Nitrotoga via microscopy (Ishii et al, 2017), and may explain the formation of microcolonies in WWTPs (Lücker et al, 2015).…”