Impacts of Environmental Pollutants on Antimicrobial Resistance Gene Transfer: A Comparative Analysis

Horizontal gene transfer is a major driver of antimicrobial resistance gene (ARG) dissemination in the environment. Although the influence of individual environmental pollutants on ARG transfer has been widely studied, comprehensive comparisons across different pollutants remain limited due to the absence of high-throughput detection methods. Herein, we developed a high-throughput screening (HTS) platform to systematically evaluate the effects of environmental pollutants on ARG transfer. We established a transfer-responsive fluorescence reporter system by genetically engineering an <i>Escherichia coli</i> strain as a donor, incorporating a conjugative RP4 plasmid carrying three ARGs. Following the horizontal transfer of the RP4 plasmid into the recipient bacterium, the nonfluorescent recipient will emit green fluorescence. The HTS platform provided faster, more efficient, and reproducible analysis than traditional colony-forming unit assays. Of the 136 environmental pollutants tested, only four antibiotics significantly enhanced ARG transfer, while others showed negligible effects at environmentally relevant concentrations. A mechanistic analysis revealed that these antibiotics induced asymmetric pressure, activating the type IV secretion system in donor cells, and thereby facilitating conjugation transfer. Overall, the HTS platform provides a robust and efficient method for evaluating the impact of pollutants on ARG transfer, thereby enhancing our comprehension of environmental risks and facilitating targeted regulatory interventions.