Transmission and migration of antibiotic resistance genes following agricultural fertilization in sloping croplands

Livestock manure, a major anthropogenic source of antibiotic resistance genes (ARGs) in agricultural soils due to residual veterinary antibiotics, is commonly used as a nutrient-rich fertilizer on sloping cropland. However, the role of landscape features, particularly topographic heterogeneity in shaping ARG transmission and migration remains poorly understood. In this study, we analyzed 76 metagenomes from five environmental habitats collected along three sloping cropland routes in the Dongting Lake region of China. Soil shared 276 ARG subtypes with other habitats, indicating manure fertilization on slopes facilitates ARGs diffusion across ecosystem. ARG abundance exhibited strong spatial patterns in soil samples, associated with distance from fertilized zones and buffer strips. In fertilized highland soils, mobile genetic elements (MGEs), such as transposases and Insertion Sequence Common Region (ISCRs), were significantly correlated with ARG abundance, indicating active horizontal gene transfer. In unfertilized-lowland soils, ARG composition was primarily influenced by heavy metals, particularly arsenic and cadmium. Source-tracking analysis showed that up to 70.3% of microbes migrated downslope via gravitational runoff, facilitating long-distance ARG dispersal. Risk assessment revealed higher ecological than human health risks, with high-risk ARGs linked to crop pathogens. Our findings highlight the need for landscape-based ARG management strategies within the One Health framework.