Bentonite seepage-control layers mitigate the transport of antibiotics and antibiotic resistance genes during reclaimed water infiltration.

Reclaimed water reuse is crucial for alleviating water scarcity, yet poses risks of disseminating antibiotics and antibiotic resistance genes (ARGs) into river-groundwater systems. Although clay minerals and bentonite are known to adsorb extracellular DNA, the performance of a bentonite seepage-control layer (SCL) in simultaneously controlling intracellular ARGs (iARGs) and extracellular ARGs (eARGs) during reclaimed-water infiltration has not been systematically evaluated. We employed large-scale (3 m) soil column simulators to investigate bentonite SCL interception performance during a 40-day reclaimed-water infiltration experiment. By integrating hydraulic characterization with chemical analysis using high-performance liquid chromatography (HPLC) and molecular analyses using quantitative real-time polymerase chain reaction (qPCR) and 16S rRNA sequencing, we quantified the transport, retention, and microbial ecological responses. Results showed the bentonite SCL acted as a robust physicochemical barrier, enhancing the apparent attenuation of antibiotics by 48.51% and reducing ARG migration. The bentonite SCL reduced antibiotic and ARG transport through the coupled effects of hydraulic retardation and sorptive retention. Independent adsorption experiments further demonstrated that bentonite had a substantially higher capacity for retaining plasmid DNA than native soil, while eARGs remained the dominant ARG fraction in both column systems (p < 0.001). Furthermore, the SCL achieved ARG interception without causing significant ecological disturbance: microbial α-diversity was preserved and the indigenous community structure remained stable. These findings demonstrate that bentonite SCLs provide a robust, cost-effective, and passive strategy for mitigating emerging biological contaminant transport, offering a practical pathway for reducing the ecological risks associated with reclaimed water reuse.