Anaerobic ammonium oxidation coupled to iron(III) reduction catalyzed by a lithoautotrophic nitrate-reducing iron(II) oxidizing enrichment culture

The last two decades have seen nitrogen/iron-transforming bacteria at the forefront of new biogeochemical discoveries, such as anaerobic ammonium oxidation coupled to ferric iron reduction (feammox) and lithoautotrophic nitrate-reducing ferrous iron-oxidation (NRFeOx). These emerging findings continue to expand our knowledge of the nitrogen/iron cycle in nature and also highlight the need to re-understand the functional traits of the microorganisms involved. Here, as a proof-of-principle, we report compelling evidence for the capability of an NRFeOx enrichment culture to catalyze the feammox process. Our results demonstrate that the NRFeOx culture predominantly oxidizes NH4+ to nitrogen gas, by reducing both chelated nitrilotriacetic acid (NTA)-Fe(III) and poorly soluble Fe(III)-bearing minerals (γ-FeOOH) at pH 4.0 and 8.0, respectively. In the NRFeOx culture, Fe(II)-oxidizing bacteria of Rhodanobacter and Fe(III)-reducing bacteria of unclassified_Acidobacteriota coexisted. Their relative abundances were dynamically regulated by the supplemented iron sources. Metagenomic analysis revealed that the NRFeOx culture contained a complete set of denitrifying genes along with hao genes for ammonium oxidation. Additionally, numerous genes encoding extracellular electron transport-associated proteins or their homologs were identified, which facilitated the reduction of extracellular iron by this culture. More broadly, this work lightens the unexplored potential of specific microbial groups in driving nitrogen transformation through multiple pathways and highlights the essential role of microbial iron metabolism in the integral biogeochemical nitrogen cycle.