Molecular interference and ecological restructuring: The acute strikes of nitroaromatics on anammox granular sludge

Nitroaromatic compounds, widely used in the pharmaceutical, dye, and pesticide industries, often coexist with high-strength ammonium in industrial effluents, posing a dual stress on biological nitrogen removal systems. Anaerobic ammonium oxidation (anammox) represents a promising technology due to its high efficiency, low energy consumption, and minimal sludge yield. However, the acute response mechanism of anammox granular sludge (AnGS) to nitroaromatic compounds remains poorly understood. This study systematically investigated the acute effects of nitrobenzene (NB) and its reduction product aniline (AN) on AnGS by integrating nitrogen removal performance, extracellular polymeric substances (EPS) characterization, microbial community evolution, and functional gene prediction. Results demonstrated a concentration-dependent inhibition of anammox activity, with specific anammox activity (SAA) decreasing by 57.91 ± 2.44 % and 25.06 ± 1.38 % at 10 mg L −1 of NB and AN, respectively, and nearly complete suppression at 50 mg L −1 . NB exhibited stronger toxicity than AN, consistent with greater shifts in EPS composition, surface hydrophilicity, and microbial community structure. Molecular docking revealed that NB exhibited a stronger disruptive potential due to its superior affinity and specific interactions with active-site residues. High-throughput sequencing revealed severe inhibition of anammox bacteria like Candidatus_Brocadia , while fermentative genera like Acetoanaerobium were significantly enriched, indicating a functional trade-off under stress. Functional gene analysis further confirmed the downregulation of key anammox genes ( hzs , hdh ). These findings highlight the compound-specific toxicity mechanisms and the resilience of microbial communities through metabolic plasticity, providing new insights into the ecological risks of nitroaromatics in anammox systems.