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The anammox-based technologies are generally inhibited by refractory dissolved organic matter (rDOM), which is ubiquitous in real wastewater. In this study, a novel cost-effective approach, namely, zero-valent iron (ZVI) treatment, was presented to alleviate such inhibitory effects. The results showed that ZVI mitigated the inhibition of fulvic acids (FA) to anammox. Due to the 2 g/L ZVI addition, the nitrogen removal efficiency (NRE) increased from 83.53 to 90.06% when the FA concentration increased from 0 to 160 mg/L in wastewater. Additionally, the co-occurrence network linking Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and 16S rRNA sequencing results revealed the mechanism of ZVI mitigating inhibition by promoting the reduction of nitro and nitroso groups of FA and reshaping the metabolic division of functional bacteria. Hydrolytic acidifying bacteria (e.g., Anaerolineaceae and Ignavibacterium) were enriched for FA degradation, which was ultimately beneficial for denitrifying bacteria and anaerobic ammonium oxidation bacteria (AnAOB). Furthermore, metagenomics revealed that ZVI stimulated multiple nitrogen removal formed by anammox, denitrification, and DNRA by accelerating extracellular electron transfer resorting to FA serving as an electron shuttle and upregulating functional genes encoding electron generation, transport, and consumption processes.

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