Nitrosomonas supralitoralis sp. nov., an ammonia-oxidizing bacterium from beach sand in a supralittoral zone.

A betaproteobacterial chemolithotrophic ammonia-oxidizing bacterium designated APG5T was isolated from supralittoral sand of the Edmonds City Beach, WA, USA. Growth was observed at 10-35 °C (optimum, 30 °C), pH 5-9 (optimum, pH 8) and ammonia concentrations as high as 100 mM (optimum, 1-30 mM NH4Cl). The strain grows optimally in a freshwater medium but tolerates up to 400 mM NaCl. It is most closely related to 'Nitrosomonas ureae' (96.7% 16S rRNA and 92.4% amoA sequence identity). The 3.75-Mbp of AGP5T draft genome contained a single rRNA operon and all necessary tRNA genes and has the lowest G+C content (43.5%) when compared to the previously reported genomes of reference strains in cluster 6 Nitrosomonas. Based on an average nucleotide identity of 82% with its closest relative ('N. ureae' Nm10T) and the suggested species boundary of 95-96%, a new species Nitrosomonas supralitoralis sp. nov. is proposed. The type strain of Nitrosomonas supralitoralis is APG5T (= NCIMB 14870T = ATCC TSD-116T).

High Synteny and Sequence Identity between Genomes of Nitrosococcus oceani Strains Isolated from Different Oceanic Gyres Reveals Genome Economization and Autochthonous Clonal Evolution.

The ammonia-oxidizing obligate aerobic chemolithoautotrophic gammaproteobacterium, Nitrosococcus oceani, is omnipresent in the world's oceans and as such important to the global nitrogen cycle. We generated and compared high quality draft genome sequences of N. oceani strains isolated from the Northeast (AFC27) and Southeast (AFC132) Pacific Ocean and the coastal waters near Barbados at the interface between the Caribbean Sea and the North Atlantic Ocean (C-27) with the recently published Draft Genome Sequence of N. oceani Strain NS58 (West Pacific Ocean) and the complete genome sequence of N. oceani C-107, the type strain (ATCC 19707) isolated from the open North Atlantic, with the goal to identify indicators for the evolutionary origin of the species. The genomes of strains C-107, NS58, C-27, and AFC27 were highly conserved in content and synteny, and these four genomes contained one nearly sequence-identical plasmid. The genome of strain AFC132 revealed the presence of genetic inventory unknown from other marine ammonia-oxidizing bacteria such as genes encoding NiFe-hydrogenase and a non-ribosomal peptide synthetase (NRPS)-like siderophore biosynthesis module. Comparative genome analysis in context with the literature suggests that AFC132 represents a metabolically more diverse ancestral lineage to the other strains with C-107 and NS58 potentially being the youngest. The results suggest that the N. oceani species evolved by genome economization characterized by the loss of genes encoding catabolic diversity while acquiring a higher redundancy in inventory dedicated to nitrogen catabolism, both of which could have been facilitated by their rich complements of CRISPR/Cas and Restriction Modification systems.

A Physiological and Genomic Comparison of Nitrosomonas Cluster 6a and 7 Ammonia-Oxidizing Bacteria.

Ammonia-oxidizing bacteria (AOB) within the genus Nitrosomonas perform the first step in nitrification, ammonia oxidation, and are found in diverse aquatic and terrestrial environments. Nitrosomonas AOB were grouped into six defined clusters, which correlate with physiological characteristics that contribute to adaptations to a variety of abiotic environmental factors. A fundamental physiological trait differentiating Nitrosomonas AOB is the adaptation to either low (cluster 6a) or high (cluster 7) ammonium concentrations. Here, we present physiological growth studies and genome analysis of Nitrosomonas cluster 6a and 7 AOB. Cluster 6a AOB displayed maximum growth rates at ≤ 1 mM ammonium, while cluster 7 AOB had maximum growth rates at ≥ 5 mM ammonium. In addition, cluster 7 AOB were more tolerant of high initial ammonium and nitrite concentrations than cluster 6a AOB. Cluster 6a AOB were completely inhibited by an initial nitrite concentration of 5 mM. Genomic comparisons were used to link genomic traits to observed physiological adaptations. Cluster 7 AOB encode a suite of genes related to nitrogen oxide detoxification and multiple terminal oxidases, which are absent in cluster 6a AOB. Cluster 6a AOB possess two distinct forms of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and select species encode genes for hydrogen or urea utilization. Several, but not all, cluster 6a AOB can utilize urea as a source of ammonium. Hence, although Nitrosomonas cluster 6a and 7 AOB have the capacity to fulfill the same functional role in microbial communities, i.e., ammonia oxidation, differentiating species-specific and cluster-conserved adaptations is crucial in understanding how AOB community succession can affect overall ecosystem function.

Evolutionary History of Copper Membrane Monooxygenases.

Copper membrane monooxygenases (CuMMOs) oxidize ammonia, methane and some short-chain alkanes and alkenes. They are encoded by three genes, usually in an operon of xmoCAB. We aligned xmo operons from 66 microbial genomes, including members of the Alpha-, Beta-, and Gamma-proteobacteria, Verrucomicrobia, Actinobacteria, Thaumarchaeota and the candidate phylum NC10. Phylogenetic and compositional analyses were used to reconstruct the evolutionary history of the enzyme and detect potential lateral gene transfer (LGT) events. The phylogenetic analyses showed at least 10 clusters corresponding to a combination of substrate specificity and bacterial taxonomy, but with no overriding structure based on either function or taxonomy alone. Adaptation of the enzyme to preferentially oxidize either ammonia or methane has occurred more than once. Individual phylogenies of all three genes, xmoA, xmoB and xmoC, closely matched, indicating that this operon evolved or was consistently transferred as a unit, with the possible exception of the methane monooxygenase operons in Verrucomicrobia, where the pmoB gene has a distinct phylogeny from pmoA and pmoC. Compositional analyses indicated that some clusters of xmoCAB operons (for example, the pmoCAB in gammaproteobacterial methanotrophs and the amoCAB in betaproteobacterial nitrifiers) were compositionally very different from their genomes, possibly indicating recent lateral transfer of these operons. The combined phylogenetic and compositional analyses support the hypothesis that an ancestor of the nitrifying bacterium Nitrosococcus was the donor of methane monooxygenase (pMMO) to both the alphaproteobacterial and gammaproteobacterial methanotrophs, but that before this event the gammaproteobacterial methanotrophs originally possessed another CuMMO (Pxm), which has since been lost in many species.

Erratum: Addendum: Comparative Genomic Analysis of the Class Epsilonproteobacteria and Proposed Reclassification to Epsilonbacteraeota (phyl. nov.).

[This corrects the article on p. 682 in vol. 8, PMID: 28484436.].

Draft Genome Sequences of Two Gammaproteobacterial Methanotrophs Isolated from Rice Ecosystems.

The genomes of the aerobic methanotrophs "Methyloterricola oryzae" strain 73aT and Methylomagnum ishizawai strain 175 were sequenced. Both strains were isolated from rice plants. Methyloterricola oryzae strain 73aT represents the first isolate of rice paddy cluster I, and strain 175 is the second representative of the recently described genus Methylomagnum.

Comparative Genomic Analysis of the Class Epsilonproteobacteria and Proposed Reclassification to Epsilonbacteraeota (phyl. nov.).

The Epsilonproteobacteria is the fifth validly described class of the phylum Proteobacteria, known primarily for clinical relevance and for chemolithotrophy in various terrestrial and marine environments, including deep-sea hydrothermal vents. As 16S rRNA gene repositories have expanded and protein marker analysis become more common, the phylogenetic placement of this class has become less certain. A number of recent analyses of the bacterial tree of life using both 16S rRNA and concatenated marker gene analyses have failed to recover the Epsilonproteobacteria as monophyletic with all other classes of Proteobacteria. In order to address this issue, we investigated the phylogenetic placement of this class in the bacterial domain using 16S and 23S rRNA genes, as well as 120 single-copy marker proteins. Single- and concatenated-marker trees were created using a data set of 4,170 bacterial representatives, including 98 Epsilonproteobacteria. Phylogenies were inferred under a variety of tree building methods, with sequential jackknifing of outgroup phyla to ensure robustness of phylogenetic affiliations under differing combinations of bacterial genomes. Based on the assessment of nearly 300 phylogenetic tree topologies, we conclude that the continued inclusion of Epsilonproteobacteria within the Proteobacteria is not warranted, and that this group should be reassigned to a novel phylum for which we propose the name Epsilonbacteraeota (phyl. nov.). We further recommend the reclassification of the order Desulfurellales (Deltaproteobacteria) to a novel class within this phylum and a number of subordinate changes to ensure consistency with the genome-based phylogeny. Phylogenomic analysis of 658 genomes belonging to the newly proposed Epsilonbacteraeota suggests that the ancestor of this phylum was an autotrophic, motile, thermophilic chemolithotroph that likely assimilated nitrogen from ammonium taken up from the environment or generated from environmental nitrate and nitrite by employing a variety of functional redox modules. The emergence of chemoorganoheterotrophic lifestyles in several Epsilonbacteraeota families is the result of multiple independent losses of various ancestral chemolithoautotrophic pathways. Our proposed reclassification of this group resolves an important anomaly in bacterial systematics and ensures that the taxonomy of Proteobacteria remains robust, specifically as genome-based taxonomies become more common.

Complete Genome Sequence of Nitrosomonas cryotolerans ATCC 49181, a Phylogenetically Distinct Ammonia-Oxidizing Bacterium Isolated from Arctic Waters.

Nitrosomonas cryotolerans ATCC 49181 is a cold-tolerant marine ammonia-oxidizing bacterium isolated from seawater collected in the Gulf of Alaska. The high-quality complete genome contains a 2.87-Mbp chromosome and a 56.6-kbp plasmid. Chemolithoautotrophic modules encoding ammonia oxidation and CO2 fixation were identified.

Environmental Conditions Outweigh Geographical Contiguity in Determining the Similarity of nifH-Harboring Microbial Communities in Sediments of Two Disconnected Marginal Seas.

Ecological evidence suggests that heterotrophic diazotrophs fueled by organic carbon respiration in sediments play an important role in marine nitrogen fixation. However, fundamental knowledge about the identities, abundance, diversity, biogeography, and controlling environmental factors of nitrogen-fixing communities in open ocean sediments is still elusive. Surprisingly, little is known also about nitrogen-fixing communities in sediments of the more research-accessible marginal seas. Here we report on an investigation of the environmental geochemistry and putative diazotrophic microbiota in the sediments of Bohai Sea, an eutrophic marginal sea of the western Pacific Ocean. Diverse and abundant nifH gene sequences were identified and sulfate-reducing bacteria (SRB) were found to be the dominant putative nitrogen-fixing microbes. Community statistical analyses suggested bottom water temperature, bottom water chlorophyll a content (or the covarying turbidity) and sediment porewater Eh (or the covarying pH) as the most significant environmental factors controlling the structure and spatial distribution of the putative diazotrophic communities, while sediment Hg content, sulfide content, and porewater [Formula: see text]-Si content were identified as the key environmental factors correlated positively with the nifH gene abundance in Bohai Sea sediments. Comparative analyses between the Bohai Sea and the northern South China Sea (nSCS) identified a significant composition difference of the putative diazotrophic communities in sediments between the shallow-water (estuarine and nearshore) and deep-water (offshore and deep-sea) environments, and sediment porewater dissolved oxygen content, water depth and in situ temperature as the key environmental factors tentatively controlling the species composition, community structure, and spatial distribution of the marginal sea sediment nifH-harboring microbiota. This confirms the ecophysiological specialization and niche differentiation between the shallow-water and deep-water sediment diazotrophic communities and suggests that the in situ physical and geochemical conditions play a more important role than geographical contiguity in determining the community similarity of the diazotrophic microbiota in marginal sea sediments.

Complete genome of Nitrosospira briensis C-128, an ammonia-oxidizing bacterium from agricultural soil.

Nitrosospira briensis C-128 is an ammonia-oxidizing bacterium isolated from an acid agricultural soil. N. briensis C-128 was sequenced with PacBio RS technologies at the DOE-Joint Genome Institute through their Community Science Program (2010). The high-quality finished genome contains one chromosome of 3.21 Mb and no plasmids. We identified 3073 gene models, 3018 of which are protein coding. The two-way average nucleotide identity between the chromosomes of Nitrosospira multiformis ATCC 25196 and Nitrosospira briensis C-128 was found to be 77.2 %. Multiple copies of modules encoding chemolithotrophic metabolism were identified in their genomic context. The gene inventory supports chemolithotrophic metabolism with implications for function in soil environments.

D1FHS, the Type Strain of the Ammonia-Oxidizing Bacterium Nitrosococcus wardiae spec. nov.: Enrichment, Isolation, Phylogenetic, and Growth Physiological Characterization.

An ammonia-oxidizing bacterium, strain D1FHS, was enriched into pure culture from a sediment sample retrieved in Jiaozhou Bay, a hyper-eutrophic semi-closed water body hosting the metropolitan area of Qingdao, China. Based on initial 16S rRNA gene sequence analysis, strain D1FHS was classified in the genus Nitrosococcus, family Chromatiaceae, order Chromatiales, class Gammaproteobacteria; the 16S rRNA gene sequence with highest level of identity to that of D1FHS was obtained from Nitrosococcus halophilus Nc4(T). The average nucleotide identity between the genomes of strain D1FHS and N. halophilus strain Nc4 is 89.5%. Known species in the genus Nitrosococcus are obligate aerobic chemolithotrophic ammonia-oxidizing bacteria adapted to and restricted to marine environments. The optimum growth (maximum nitrite production) conditions for D1FHS in a minimal salts medium are: 50 mM ammonium and 700 mM NaCl at pH of 7.5 to 8.0 and at 37°C in dark. Because pertinent conditions for other studied Nitrosococcus spp. are 100-200 mM ammonium and <700 mM NaCl at pH of 7.5 to 8.0 and at 28-32°C, D1FHS is physiologically distinct from other Nitrosococcus spp. in terms of substrate, salt, and thermal tolerance.

The nitrogen cycle.

Nitrogen is the fourth most abundant element in cellular biomass, and it comprises the majority of Earth's atmosphere. The interchange between inert dinitrogen gas (N2) in the extant atmosphere and 'reactive nitrogen' (those nitrogen compounds that support, or are products of, cellular metabolism and growth) is entirely controlled by microbial activities. This was not the case, however, in the primordial atmosphere, when abiotic reactions likely played a significant role in the inter-transformation of nitrogen oxides. Although such abiotic reactions are still important, the extant nitrogen cycle is driven by reductive fixation of dinitrogen and an enzyme inventory that facilitates dinitrogen-producing reactions. Prior to the advent of the Haber-Bosch process (the industrial fixation of N2 into ammonia, NH3) in 1909, nearly all of the reactive nitrogen in the biosphere was generated and recycled by microorganisms. Although the Haber-Bosch process more than quadrupled the productivity of agricultural crops, chemical fertilizers and other anthropogenic sources of fixed nitrogen now far exceed natural contributions, leading to unprecedented environmental degradation.

Pathways and key intermediates required for obligate aerobic ammonia-dependent chemolithotrophy in bacteria and Thaumarchaeota.

Chemolithotrophic ammonia-oxidizing bacteria and Thaumarchaeota are central players in the global nitrogen cycle. Obligate ammonia chemolithotrophy has been characterized for bacteria; however, large gaps remain in the Thaumarchaeotal pathway. Using batch growth experiments and instantaneous microrespirometry measurements of resting biomass, we show that the terrestrial Thaumarchaeon Nitrososphaera viennensis EN76(T) exhibits tight control over production and consumption of nitric oxide (NO) during ammonia catabolism, unlike the ammonia-oxidizing bacterium Nitrosospira multiformis ATCC 25196(T). In particular, pulses of hydroxylamine into a microelectrode chamber as the sole substrate for N. viennensis resulted in iterative production and consumption of NO followed by conversion of hydroxylamine to nitrite. In support of these observations, oxidation of ammonia in growing cultures of N. viennensis, but not of N. multiformis, was inhibited by the NO-scavenger PTIO. When based on the marginal nitrous oxide (N2O) levels detected in cell-free media controls, the higher levels produced by N. multiformis were explained by enzyme activity, whereas N2O in N. viennensis cultures was attributed to abiotic reactions of released N-oxide intermediates with media components. Our results are conceptualized in a pathway for ammonia-dependent chemolithotrophy in Thaumarchaea, which identifies NO as an essential intermediate in the pathway and implements known biochemistry to be executed by a proposed but still elusive copper enzyme. Taken together, this work identifies differences in ammonia-dependent chemolithotrophy between bacteria and the Thaumarchaeota, advances a central catabolic role of NO only in the Thaumarchaeotal pathway and reveals stark differences in how the two microbial cohorts contribute to N2O emissions.

Draft Genome Sequences of Gammaproteobacterial Methanotrophs Isolated from Marine Ecosystems.

The genome sequences of Methylobacter marinus A45, Methylobacter sp. strain BBA5.1, and Methylomarinum vadi IT-4 were obtained. These aerobic methanotrophs are typical members of coastal and hydrothermal vent marine ecosystems.

Draft genomes of gammaproteobacterial methanotrophs isolated from terrestrial ecosystems.

Genome sequences of Methylobacter luteus, Methylobacter whittenburyi, Methylosarcina fibrata, Methylomicrobium agile, and Methylovulum miyakonense were generated. The strains represent aerobic methanotrophs typically isolated from various terrestrial ecosystems.