Project description:Nitrification, the oxidation of ammonia via nitrite to nitrate, has always been considered to be a two-step process catalysed by chemolithoautotrophic microorganisms oxidizing either ammonia or nitrite. No known nitrifier carries out both steps, although complete nitrification should be energetically advantageous. This functional separation has puzzled microbiologists for a century. Here we report on the discovery and cultivation of a completely nitrifying bacterium from the genus Nitrospira, a globally distributed group of nitrite oxidizers. The genome of this chemolithoautotrophic organism encodes the pathways both for ammonia and nitrite oxidation, which are concomitantly activated during growth by ammonia oxidation to nitrate. Genes affiliated with the phylogenetically distinct ammonia monooxygenase and hydroxylamine dehydrogenase genes of Nitrospira are present in many environments and were retrieved on Nitrospira contigs in new metagenomes from engineered systems. These findings fundamentally change our picture of nitrification and point to completely nitrifying Nitrospira as key components of nitrogen-cycling microbial communities.
Project description:Nitrification, the oxidation of ammonia via nitrite to nitrate, has been considered to be a stepwise process mediated by two distinct functional groups of microorganisms. The identification of complete nitrifying Nitrospira challenged not only the paradigm of labor division in nitrification, it also raises fundamental questions regarding the environmental distribution, diversity, and ecological significance of complete nitrifiers compared to canonical nitrifying microorganisms. Recent genomic and physiological surveys identified factors controlling their ecology and niche specialization, which thus potentially regulate abundances and population dynamics of the different nitrifying guilds. This review summarizes the recently obtained insights into metabolic differences of the known nitrifiers and discusses these in light of potential functional adaptation and niche differentiation between canonical and complete nitrifiers.
Project description:Aquaponic systems are sustainable solutions for food production combining fish growth (aquaculture) and production of vegetables (hydroponic) in one recirculating system. In aquaponics, nitrogen-enriched wastewater from fish in the aquaculture serves as fertilizer for the plants in the hydroponics, while the nitrogen-depleted and detoxified water flows back to the aquaculture. To investigate bacterial nitrogen-cycling in such an aquaponic system, measurements of nitrogen species were coupled with time-resolved 16S rRNA gene profiling and the functional capacity of organisms was studied using metagenomics. The aquaponic system was consistently removing ammonia and nitrite below 23 µM and 19 µM, and nitrate to steady-state concentrations of about 0.5 mM. 16S rRNA gene amplicon sequencing of sediments exposed in the pump sump revealed that typical signatures of canonical ammonia-oxidising microorganisms were below detection limit. However, one of the most abundant operational taxonomic units (OTU) was classified as a member of the genus Nitrospira with a relative abundance of 3.8%. For this genus, also genome scaffolds were recovered encoding the only ammonia monooxygenase genes identified in the metagenome. This study indicates that even in highly efficient aquaponic systems, comammox Nitrospira were found to participate in ammonium removal at low steady-state ammonia concentrations.
Project description:Microbes commonly exist in diverse and complex communities where species interact, and their genomic repertoires evolve over time. Our understanding of species interaction and evolution has increased during the last decades, but most studies of evolutionary dynamics are based on single species in isolation or in experimental systems composed of few interacting species. Here, we use the microbial ecosystem found in groundwater-fed sand filter as a model to avoid this limitation. In these open systems, diverse microbial communities experience relatively stable conditions, and the coupling between chemical and biological processes is generally well defined. Metagenomic analysis of 12 sand filters communities revealed systematic co-occurrence of at least five comammox Nitrospira species, likely promoted by low ammonium concentrations. These Nitrospira species showed intrapopulation sequence diversity, although possible clonal expansion was detected in a few abundant local comammox populations. Nitrospira species showed low homologous recombination and strong purifying selection, the latter process being especially strong in genes essential in energy metabolism. Positive selection was detected for genes related to resistance to foreign DNA and phages. We found that, compared to other habitats, groundwater-fed sand filters impose strong purifying selection and low recombination on comammox Nitrospira populations. These results suggest that evolutionary processes are more affected by habitat type than by species identity. Together, this study improves our understanding of species interaction and evolution in complex microbial communities and sheds light on the environmental dependency of evolutionary processes. IMPORTANCE Microbial species interact with each other and their environment (ecological processes) and undergo changes in their genomic repertoire over time (evolutionary processes). How these two classes of processes interact is largely unknown, especially for complex communities, as most studies of microbial evolutionary dynamics consider single species in isolation or a few interacting species in simplified experimental systems. In this study, these limitations are circumvented by examining the microbial communities found in stable and well-described groundwater-fed sand filters. Combining metagenomics and strain-level analyses, we identified the microbial interactions and evolutionary processes affecting comammox Nitrospira, a recently discovered bacterial type capable of performing the whole nitrification process. We found that abundant and co-occurrent Nitrospira populations in groundwater-fed sand filters are characterized by low recombination and strong purifying selection. In addition, by comparing these observations with those obtained from Nitrospira species inhabiting other environments, we revealed that evolutionary processes are more affected by habitat type than by species identity.
Project description:The recently discovered comammox process encompasses both nitrification steps, the aerobic oxidation of ammonia and nitrite, in a single organism. All known comammox bacteria are affiliated with Nitrospira sublineage II and can be grouped into two distinct clades, referred to as A and B, based on ammonia monooxygenase phylogeny. In this study, we report high-quality draft genomes of two novel comammox Nitrospira from the terrestrial subsurface, representing one clade A and one clade B comammox organism. The two metagenome-assembled genomes were compared with other representatives of Nitrospira sublineage II, including both canonical and comammox Nitrospira. Phylogenomic analyses confirmed the affiliation of the two novel Nitrospira with comammox clades A and B respectively. Based on phylogenetic distance and pairwise average nucleotide identity values, both comammox Nitrospira were classified as novel species. Genomic comparison revealed high conservation of key metabolic features in sublineage II Nitrospira, including respiratory complexes I-V and the machineries for nitrite oxidation and carbon fixation via the reductive tricarboxylic acid cycle. In addition, the presence of the enzymatic repertoire for formate and hydrogen oxidation in the Rifle clades A and B comammox genomes, respectively, suggest a broader distribution of these metabolic features than previously anticipated.
Project description:Nitrospira has been revealed as a high versatile genus. Although previously considered only responsible for the conversion of nitrite to nitrate, now we know that Nitrospira can perform complete ammonia oxidation to nitrate too (comammox). Comammox activity was firstly reported as dominant in extremely limited oxygen environments, where anaerobic ammonia oxidation was also occurring (anammox). To explain the comammox selection, we developed an Individual-based Model able to describe Nitrospira and anammox growth in suspended flocs assembled in a dynamic nitrogen and oxygen-limiting environment. All known and hypothesized nitrogen transformations of Nitrospira were considered: ammonia and nitrite oxidation, comammox, nitrate-reducing ammonia oxidation, and anaerobic nitrite-reducing ammonia oxidation. Through bioenergetics analysis, the growth yield associated to each activity was estimated. The other kinetic parameters necessary to describe growth were calibrated according to the reported literature values. Our modeling results suggest that even extremely low oxygen concentrations (~1.0 µM) allow for a proportional growth of anammox versus Nitrospira similar to the one experimentally observed. The strong oxygen limitation was followed by a limitation of ammonia and nitrite, because anammox, without strong competitors, were able to grow faster than Nitrospira depleting the environment in nitrogen. These substrate limitations created an extremely competitive environment that proved to be decisive in the community assembly of Nitrospira and anammox. Additionally, a diversity of metabolic activities for Nitrospira was observed in all tested conditions, which in turn, explained the transient nitrite accumulation observed in aerobic environments with higher ammonia availability.
Project description:In agricultural soils fertilized with a high amount of ammonium nitrogen, the pH decreases because of the oxidation of ammonia by nitrifiers. Molecular-based analyses have revealed that members of the genus Nitrospira dominate over other nitrifiers in some acidic soils. However, terrestrial Nitrospira are rarely cultivated and little is known about their ecophysiology. In addition, recent studies discovered a single microbe with the potential to oxidize both ammonia and nitrite (complete ammonia oxidizer; comammox) within Nitrospira, which had been previously recognized as a nitrite oxidizer. Despite their broad distribution, there are no enrichment samples of comammox from terrestrial or acidic environments. Here, we report the selective enrichment of both comammox and nitrite-oxidizing Nitrospira from the acidic soil of a heavily fertilized tea field. Long-term enrichment was performed with two individual continuous-feeding bioreactors capable of controlling ammonia or nitrite concentration and pH. We found that excessive ammonium supply was a key factor to enhance the growth of comammox Nitrospira under acidic conditions. Additionally, a low concentration of nitrite was fed to prevent the accumulation of free nitrous acid and inhibition of cell growth under low pH, resulting in the selective enrichment of nitrite-oxidizing Nitrospira. Based on 16S rRNA gene analysis, Nitrospira accounting for only 1.2% in an initial soil increased to approximately 80% of the total microorganisms in both ammonia- and nitrite-fed bioreactors. Furthermore, amoA amplicon sequencing revealed that two phylotypes belonging to comammox clade A were enriched in an ammonia-fed bioreactor. One group was closely related to previously cultivated strains, and the other was classified into a different cluster consisting of only uncultivated representatives. These two groups coexisted in the bioreactor controlled at pH 6.0, but the latter became dominant after the pH decreased to 5.5. Additionally, a physiological experiment revealed that the enrichment sample oxidizes ammonia at pH <4, which is in accordance with the strongly acidic tea field soil; this value is lower than the active pH range of isolated acid-adapted nitrifiers. In conclusion, we successfully enriched multiple phylotypes of comammox and nitrite-oxidizing Nitrospira and revealed that the pH and concentrations of protonated N-compounds were potential niche determinants.