Energy-mediated vs. ammonium-regulated gene expression in the obligate ammonia-oxidizing bacterium, Nitrosococcus oceani.

Ammonia serves as the source of energy and reductant and as a signaling molecule that regulates gene expression in obligate ammonia-oxidizing chemolithotrophic microorganisms. The gammaproteobacterium, Nitrosococcus oceani, was the first obligate ammonia-oxidizer isolated from seawater and is one of the model systems for ammonia chemolithotrophy. We compared global transcriptional responses to ammonium and the catabolic intermediate, hydroxylamine, in ammonium-starved and non-starved cultures of N. oceani to discriminate transcriptional effects of ammonium from a change in overall energy and redox status upon catabolite availability. The most highly expressed genes from ammonium- or hydroxylamine-treated relative to starved cells are implicated in catabolic electron flow, carbon fixation, nitrogen assimilation, ribosome structure and stress tolerance. Catabolic inventory-encoding genes, including electron flow-terminating Complexes IV, FoF1 ATPase, transporters, and transcriptional regulators were among the most highly expressed genes in cells exposed only to ammonium relative to starved cells, although the differences compared to steady-state transcript levels were less pronounced. Reduction in steady-state mRNA levels from hydroxylamine-treated relative to starved-cells were less than five-fold. In contrast, several transcripts from ammonium-treated relative to starved cells were significantly less abundant including those for forward Complex I and a gene cluster of cytochrome c encoding proteins. Identified uneven steady-state transcript levels of co-expressed clustered genes support previously reported differential regulation at the levels of transcription and transcript stability. Our results differentiated between rapid regulation of core genes upon a change in cellular redox status vs. those responsive to ammonium as a signaling molecule in N. oceani, both confirming and extending our knowledge of metabolic modules involved in ammonia chemolithotrophy.

Nitrate ammonification by Nautilia profundicola AmH: experimental evidence consistent with a free hydroxylamine intermediate.

The process of nitrate reduction via nitrite controls the fate and bioavailability of mineral nitrogen within ecosystems; i.e., whether it is retained as ammonium (ammonification) or lost as nitrous oxide or dinitrogen (denitrification). Here, we present experimental evidence for a novel pathway of microbial nitrate reduction, the reverse hydroxylamine:ubiquinone reductase module (reverse-HURM) pathway. Instead of a classical ammonia-forming nitrite reductase that performs a 6 electron-transfer process, the pathway is thought to employ two catalytic redox modules operating in sequence: the reverse-HURM reducing nitrite to hydroxylamine followed by a hydroxylamine reductase that converts hydroxylamine to ammonium. Experiments were performed on Nautilia profundicola strain AmH, whose genome sequence led to the reverse-HURM pathway proposal. N. profundicola produced ammonium from nitrate, which was assimilated into biomass. Furthermore, genes encoding the catalysts of the reverse-HURM pathway were preferentially expressed during growth of N. profundicola on nitrate as an electron acceptor relative to cultures grown on polysulfide as an electron acceptor. Finally, nitrate-grown cells of N. profundicola were able to rapidly and stoichiometrically convert high concentrations of hydroxylamine to ammonium in resting cell assays. These experiments are consistent with the reverse-HURM pathway and a free hydroxylamine intermediate, but could not definitively exclude direct nitrite reduction to ammonium by the reverse-HURM with hydroxylamine as an off-pathway product. N. profundicola and related organisms are models for a new pathway of nitrate ammonification that may have global impact due to the wide distribution of these organisms in hypoxic environments and symbiotic or pathogenic associations with animal hosts.

Model of the molecular basis for hydroxylamine oxidation and nitrous oxide production in methanotrophic bacteria.

Many methane-oxidizing bacteria (MOB) have been shown to aerobically oxidize ammonia and hydroxylamine (NH(2)OH) to produce nitrite and nitrous oxide (N(2)O). Genome sequences of alphaproteobacterial, gammaproteobacterial, and verrucomicrobial methanotrophs revealed the presence of haoAB, cytL, cytS, nirS or nirK, and norCB genes that may be responsible for N(2)O production, and additional haoAB genes were sequenced from two strains of Methylomicrobium album. The haoAB genes of M. album ATCC 33003 were inducible by ammonia and NH(2)OH, similar to haoAB induction by ammonia in Methylococcus capsulatus Bath. Increased expression of genes encoding nitric oxide reductase (cNOR; norCB) was measured upon exposure of M. capsulatus Bath to NaNO(2) and NO-releasing sodium nitroprusside. Only incubations of M. capsulatus Bath with methane, ammonia, and nitrite produced N(2)O. The data suggest a possible pathway of nitrite reduction to NO by reversely operating NH(2)OH oxidoreductase and NO reduction to N(2)O by cNOR independently or in conjunction with ammonia-induced enzymes (i.e. HAO or cytochrome c'-ß). Results of this study show that MOB likely have diverse mechanisms for nitrogen oxide metabolism and detoxification of NH(2)OH that involve conventional and unconventional enzymes.

Nitrosococcus watsonii sp. nov., a new species of marine obligate ammonia-oxidizing bacteria that is not omnipresent in the world's oceans: calls to validate the names 'Nitrosococcus halophilus' and 'Nitrosomonas mobilis'.

Local associations between anammox bacteria and obligate aerobic bacteria in the genus Nitrosococcus appear to be significant for ammonia oxidation in oxygen minimum zones. The literature on the genus Nitrosococcus in the Chromatiaceae family of purple sulfur bacteria (Gammaproteobacteria, Chromatiales) contains reports on four described species, Nitrosococcus nitrosus, Nitrosococcus oceani, 'Nitrosococcus halophilus' and 'Nitrosomonas mobilis', of which only N. nitrosus and N. oceani are validly published names and only N. oceani is omnipresent in the world's oceans. The species 'N. halophilus' with Nc4(T) as the type strain was proposed in 1990, but the species is not validly published. Phylogenetic analyses of signature genes, growth-physiological studies and an average nucleotide identity analysis between N. oceani ATCC19707(T) (C-107, Nc9), 'N. halophilus' strain Nc4(T) and Nitrosococcus sp. strain C-113 revealed that a proposal for a new species is warranted. Therefore, the provisional taxonomic assignment Nitrosococcus watsonii is proposed for Nitrosococcus sp. strain C-113(T) . Sequence analysis of Nitrosococcus haoAB signature genes detected in cultures enriched from Jiaozhou Bay sediments (China) identified only N. oceani-type sequences, suggesting that different patterns of distribution in the environment correlate with speciation in the genus Nitrosococcus.