Acetochlor and sulfamethoxazole co-selection alter soil microbial nitrogen metabolism and resistome in agroecosystem

Agricultural soils increasingly face co-contamination by herbicides and antibiotics, yet the ecological impacts of such multipollutant exposure remain poorly understood. This study employed a soil-plant microcosm combined with metagenomic sequencing to investigate the co-selective effects of acetochlor (ACE) and sulfamethoxazole (SMX) on soil microbiomes and antibiotic resistance genes (ARGs). The results showed that SMX functioned as the dominant ecological filter, significantly reducing microbial diversity and restructuring community composition via suppressing Pseudomonadota while enriching Acidobacteriota. Co-exposure further decreased diversity and shifted nitrogen metabolic pathways: SMX inhibited denitrification and nitrogen fixation, whereas co-combination synergistically enhanced the potential of nitrous oxide emission. Critically, herbicide-antibiotic co-exposure drove the emergence of clinically relevant ARGs (e.g., CMY-80, MCR-2.5) and enhanced their dissemination by increasing network complexity among host microorganisms. Moreover, ACE acted as an 'antibiotic adjuvant', accelerating resistance evolution through stress-induced physiological responses and mobility activation. ACE dose-dependent responses revealed the dual ecological role of agrochemicals: signaling molecules at low concentrations (2.5 mg/kg) and stressors at elevated levels (5.0 mg/kg). Genomic analysis further showed a higher chromosomal than plasmid-borne ARG abundance, reflecting a dynamic equilibrium between persistent and mobile resistance under fluctuating environmental pressures. These findings underscore the necessity of incorporating multipollutant scenarios into risk assessment, as single-contaminant evaluations underestimate the ecological and public health risks in agricultural ecosystems.