Enrofloxacin metabolism and antibiotic resistance in Monopterus albus differs between pond cage and greenhouse micro-flow aquaculture systems.

Enrofloxacin (ENR) is a fluoroquinolone antibiotic widely used in aquaculture, yet its metabolic fate and resistance selection dynamics remain poorly characterized. Here, we investigated ENR metabolism, tissue distribution, withdrawal periods, gut microbial communities, and antibiotic resistance genes (ARGs) in Monopterus albus reared under pond net cage (PNC) and greenhouse micro-flow (GMF) systems following a 5-day medicated feed treatment (20 mg ENR/kg body weight). Eight ENR metabolites were tentatively identified, with ciprofloxacin (CIP) as the predominant metabolite. The GMF system significantly shortened the estimated withdrawal period to 4050 °C·d compared to 5550 °C·d for PNC (about 30% reduction). Critically, the PNC withdrawal period exceeded the current Chinese regulatory standard of 500 °C·d by more than 10 folds, revealing a substantial gap between existing guidelines and on-farm food safety requirements. Sediment analysis showed that 7.8% of the administered ENR dose persisted as a long term environmental reservoir. Metagenomic sequencing revealed that even a single ENR treatment induced an approximately 10-fold increase in fluoroquinolone resistance genes by day 60 post-treatment, with cross-resistance extending to multiple other antibiotic classes. Aeromonadaceae and Enterobacteriaceae were identified as the primary ARG hosts, and ARG abundance was significantly correlated with mobile genetic element prevalence, suggesting enhanced horizontal gene transfer potential. These findings demonstrate that aquaculture system design profoundly influences antibiotic fate and resistance selection, and that current withdrawal standards are inadequate for scaleless species. System-specific withdrawal guidelines and strengthened antimicrobial stewardship are urgently needed to mitigate environmental and food safety risks from aquaculture antibiotic use.