Differential fates of antibiotics in sulfur-driven autotrophic denitrification: Ciprofloxacin transformation linked to sulfide oxidation.

The coexistence of residual antibiotics in wastewater raises concerns about the functional stability of sulfur-driven autotrophic denitrification (SDAD) mediated by sulfur-oxidizing bacteria (SOB) and the fate of antibiotics during treatment. This study investigated the functional tolerance of SOB to three representative antibiotics-ciprofloxacin (CIP), oxytetracycline (OTC), and sulfamonomethoxine (SMM)-under 360-min batch exposure and elucidated their removal behaviors during SOB-mediated sulfide (S2-) oxidation and NO3-/NO2- reduction. Batch experiments revealed distinct fates among the three antibiotics. OTC was removed to a limited extent via adsorption, and SMM remained largely stable. In contrast, CIP exhibited selective transformation closely associated with active S2- oxidation, rather than NO3-/NO2- reduction, with removal efficiency increasing from 11.6 ± 1.7% to 38.1 ± 0.3% as the initial S2- concentration rose from 0 to 12 mg-S/L. Notably, none of the three antibiotics significantly inhibited SOB-mediated S2- oxidation or NO3-/NO2- reduction under 360-min batch exposure, indicating appreciable short-term functional tolerance of the enriched SOB community. Five CIP transformation products were tentatively identified, with predicted lower ecotoxicity than the parent compound. A proof-of-concept sequential-substrate experiment further demonstrated that reintroducing S2- after denitrification substantially enhanced CIP removal, supporting the validity of a phased sulfur-based treatment strategy. To gain mechanistic insight, AlphaFold 3-based molecular docking analyses were performed, which suggested that S2- enhanced the association between CIP and sulfide-quinone oxidoreductase, with calculated binding energy decreasing from -5.2 to -7.0 kcal/mol in the presence of S2-, whereas no such change was observed for OTC or SMM. These findings advance the understanding of antibiotic removal in SDAD systems and provide a basis for mechanism-guided process optimization in the treatment of antibiotic-containing low C/N wastewaters.