Three-dimensional synergistic mechanism ofphysical injury, microbiota dysbiosis, and gene transfer in the gut of Cipangopaludina cathayensisunder microplastics and roxithromycin exposure.

Microplastics (MPs) and antibiotics pose a combined threat to aquatic organisms by impairing gut health and promoting the spread of antibiotic resistance genes (ARGs). In this study, Cipangopaludina cathayensis was exposed for 28 days to polystyrene MPs, roxithromycin (ROX), and their combination to assess impacts on intestinal barrier integrity, microbiota composition, and ARG proliferation. MPs alone caused significant mucosal damage, villus atrophy, epithelial shedding, and reduced digestive enzyme activities. ROX exposure altered microbiota structure by increasing Bacteroidetes and reducing Firmicutes. Co-exposure (CM group) exacerbated epithelial injury and enzyme inhibition but partially restored balance through enrichment of SCFA-producing, anti-inflammatory bacteria. ARG levels in the CM group rose by over 1000 %, with notable increases in multidrug resistance genes (e.g., blaOXA10) and integrons (e.g., cIntI-1), mainly linked to Bacteroides and Proteobacteria. Transcriptomic data indicated oxidative stress and epithelial disruption under MPs, and upregulation of efflux and integron genes with ROX. Combined exposure triggered DNA repair and SOS pathways, facilitating horizontal gene transfer. These findings highlight a three-dimensional synergistic mechanism-physical damage, microbial dysbiosis, and gene transfer-that amplifies ARG dissemination and intestinal toxicity, underscoring the need to assess ecological risks of composite pollutants in freshwater systems.These processes form a self-reinforcing loop in which physical epithelial damage promotes microbial dysbiosis, which in turn facilitates ARG proliferation through increased permeability and immune disruption.