Enantioselective effects of chiral antibiotics on antibiotic resistance gene dissemination and risk in activated sludge

Misuse of antibiotics drives the spread of antibiotic resistance genes (ARGs). Although reducing residual antibiotic concentrations can help curb ARG proliferation, the biodegradation and transformation of antibiotic stereoisomers may exacerbate resistance development. However, the impact of antibiotic enantiomers on ARG proliferation remains poorly understood. This study employed metagenomic analysis to investigate the enantiomer-specific selection and resistance risks of chiral antibiotic ofloxacin (OFL) and its ( S )-enantiomer, levofloxacin (LEV), in activated sludge. Results showed that LEV primarily promoted the enrichment of ARGs related to aminoglycoside and mupirocin resistance by selecting for pathogenic bacteria carrying virulence factors under high toxicity stress. OFL-driven ARG proliferation involved more diverse mechanisms, including increased gene mobility, co-selection with heavy metals, broader host range, and elevated pathogenicity. The antibiotic resistome risk index (ARRI) further demonstrated a higher environmental risk under OFL treatment than LEV. These findings offer critical insights into the enantioselective resistance risks posed by chiral antibiotics.