Ammonia Oxidation and Nitrite Reduction in the Verrucomicrobial Methanotroph Methylacidiphilum fumariolicum SolV

The Solfatara volcano near Naples (Italy), the origin of the recently discovered verrucomicrobial methanotroph Methylacidiphilum fumariolicum SolV was shown to contain ammonium ([Formula: see text]) at concentrations ranging from 1 to 28 mM. Ammonia (NH₃) can be converted to toxic hydroxylamine (NH₂OH) by the particulate methane monooxygenase (pMMO), the first enzyme of the methane (CH₄) oxidation pathway. Methanotrophs rapidly detoxify the intermediate NH₂OH. Here, we show that strain SolV performs ammonium oxidation to nitrite at a rate of 48.2 nmol [Formula: see text].h^-1.mg DW^-1 under O₂ limitation in a continuous culture grown on hydrogen (H₂) as an electron donor. In addition, strain SolV carries out nitrite reduction at a rate of 74.4 nmol [Formula: see text].h^-1.mg DW^-1 under anoxic condition at pH 5-6. This range of pH was selected to minimize the chemical conversion of nitrite ([Formula: see text]) potentially occurring at more acidic pH values. Furthermore, at pH 6, we showed that the affinity constants (K _s ) of the cells for NH₃ vary from 5 to 270 microM in the batch incubations with 0.5-8% (v/v) CH₄, respectively. Detailed kinetic analysis showed competitive substrate inhibition between CH₄ and NH₃. Using transcriptome analysis, we showed up-regulation of the gene encoding hydroxylamine dehydrogenase (haoA) cells grown on H₂/[Formula: see text] compared to the cells grown on CH₄/[Formula: see text] which do not have to cope with reactive N-compounds. The denitrifying genes nirk and norC showed high expression in H₂/[Formula: see text] and CH₄/[Formula: see text] grown cells compared to cells growing at micro_max (with no limitation) while the norB gene showed downregulation in CH₄/[Formula: see text] grown cells. These cells showed a strong upregulation of the genes in nitrate/nitrite assimilation. Our results demonstrate that strain SolV can perform ammonium oxidation producing nitrite. At high concentrations of ammonium this may results in toxic effects. However, at low oxygen concentrations strain SolV is able to reduce nitrite to N₂O to cope with this toxicity.