The emergence and spread of antibiotic resistance genes (ARGs) have become a pressing issue in global agricultural production. However, understanding how these ARGs spread across different spatial scales, especially when exposed to both antibiotics and pesticides, has remained a challenge. Here, metagenomic assembly and binning methodologies were used to determine the spread pathway of ARGs in the soil-lettuce system exposed to both antibiotics and fungicides. The results showed that the selective pressure from these agrochemicals not only facilitates the spread of ARGs from soil to lettuce but also significantly elevates the risk of developing multi-antibiotic resistance among bacteria. Our findings indicate that ARGs migrate through distinct mechanisms, including both vertical and horizontal gene transfer (HGT), with plasmids playing a crucial role in facilitating this migration. This process has enabled key pathogenic bacteria such as Acinetobacter, Pseudomonas, and Pantoea to acquire resistance and remain recalcitrant, posing the potential risk to crop health and food safety. In summary, our findings highlighted that antibiotic and fungicide could drive upward migration of ARGs in the soil-plant system and reduced the quality and safety of agricultural products.
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