Performance recovery and antibiotic resistance gene risk mitigation in wastewater anaerobic digestion under long-term Ciprofloxacin stress via zero-valent iron-biochar.

Ciprofloxacin (CIP) imposes persistent stress on upflow anaerobic sludge blanket (UASB) reactor, leading to suppressed methanogenic activity and heightened antibiotic resistance risks. Although zero-valent iron-biochar has been reported to effectively alleviate CIP stress, its effectiveness in restoring performance and maintaining stable operation under long-term CIP exposure remains poorly understood. In this study, the recovery effects of straw biochar (SBC), nanoscale zero-valent iron-SBC (nZVI-SBC), and microscale ZVI-SBC were systematically evaluated in UASB reactors subjected to long-term CIP stress. The results demonstrate that only nZVI-SBC effectively restored system performance, increasing CH4 production by 38.7%. Mechanistically, nZVI-SBC promoted the enrichment of electroactive bacteria and key methanogenesis-related functional genes. Concurrently, nZVI-SBC achieved efficient CIP removal (89.7%) and reduced the predicted toxicity of transformation intermediates. Furthermore, nZVI-SBC suppressed fluoroquinolone antibiotic resistance genes (ARGs) and high-risk subtypes by inhibiting horizontal gene transfer-related pathways. Overall, these findings establish nZVI-SBC as an effective strategy for recovering UASB performance while simultaneously mitigating ARGs dissemination under long-term CIP stress.