Unveiling hidden risks of chiral fungicide benzovindiflupyr: Stereoselectivity in soil antibiotic resistance gene transmission.

Antibiotic resistance gene (ARG) dissemination is closely associated with modern agricultural practices. However, the stereoselective effects of widely applied chiral pesticides on resistance evolution remain insufficiently investigated. This study systematically explored the differential effects of benzovindiflupyr enantiomers on transmission of ARGs through long-term soil incubation experiments combined with metagenomic and in vitro studies. Results demonstrated that 1S,4R-enantiomer exhibited significantly longer half-life than 1 R,4S-enantiomer. 1 R,4S-enantiomer induced extreme enrichment of a few ARGs. 1S,4R-enantiomer persistently increased abundance of multiple ARGs. Compared with 1 R,4S-enantiomer, 1S,4R-enantiomer more consistently enhanced abundance of mobile genetic elements (MGEs) related to conjugative transfer. Moreover, 1 R,4S-enantiomer primarily enriched specific genera within Pseudomonadota. 1S,4R-enantiomer simultaneously promoted abundance of multiple genera across both Pseudomonadota and Bacteroidota, driving cross-phylum genera to correlate with shared ARGs. Genomic analysis confirmed that Pseudomonadota under 1S,4R-enantiomer treatment carried more ARGs and MGEs. In vitro transformation experiments ultimately validated that 1S,4R-enantiomer significantly enhanced transformation efficiency across multiple ARGs consistently, substantially exceeding 1 R,4S-enantiomer effects. Overall, 1S,4R-enantiomer poses more significant risks for horizontal transfer of ARGs. This study elucidates enantioselective effects of chiral pesticides on transmission of ARGs, providing a foundation for improving chiral agrochemical risk assessment.