Dirammox Is Widely Distributed and Dependently Evolved in Alcaligenes and Is Important to Nitrogen Cycle

Nitrogen cycle is an essential process for environmental health. Dirammox (direct ammonia oxidation), encoded by the dnfT1RT2ABCD cluster, was a novel pathway for microbial N₂ production defined in Alcaligenes ammonioxydans HO-1. Here, a copy of the cluster dnfT1RT2ABCD as a whole was proved to have existed and very conserved in all Alcaligenes genomes. Phylogenetic analyses based on 16S rRNA gene sequences and amino acid sequences of DnfAs, together with G + C content data, revealed that dnf cluster was evolved associated with the members of the genus Alcaligenes. Under 20% O₂ conditions, 14 of 16 Alcaligenes strains showed Dirammox activity, which seemed likely taxon-related. However, the in vitro activities of DnfAs catalyzing the direct oxidation of hydroxylamine to N₂ were not taxon-related but depended on the contents of Fe and Mn ions. The results indicated that DnfA is necessary but not sufficient for Dirammox activity. The fact that members of the genus Alcaligenes are widely distributed in various environments, including soil, water bodies (both freshwater and seawater), sediments, activated sludge, and animal-plant-associated environments, strongly suggests that Dirammox is important to the nitrogen cycle. In addition, Alcaligenes species are also commonly found in wastewater treatment plants, suggesting that they might be valuable resources for wastewater treatment.