Microplastics (MPs) and antibiotic resistance genes (ARGs) are two classes of emerging and prevalent contaminants in terrestrial environments. To date, effects of MPs on the occurrence of ARGs in terrestrial invertebrates remain uncertain. Here we exposed earthworms to a soil amended with polystyrene MPs at two environmentally relevant concentrations to elucidate the occurrence and mechanisms of ARGs in earthworms impacted by MPs with different sizes. Nano-size and 10 mg/kg of 100 mu m MPs slightly affected the occurrence of ARGs in earthworms. Highest abundance of ARGs was found in the presence of 10 mg/kg of 10 mu m MPs, whereas 100 mg/ kg of 10 mu m MPs significantly changed the profile of ARGs. Metagenomics sequencing and toxicity tests indicated that MPs caused toxicity and influenced the abundance of microbial community in earthworms, resulting in the changes of ARGs. Results of proteomics and metabolomics demonstrated that 100 mg/kg of 10 mu m MPs changed the microenvironment of earthworm gut, built a new homeostatic process, and thus increased the abundance of key bacterial that carried a variety of ARGs. This study highlights the size-dependent toxic effects of MPs and their impacts on the transfer of ARGs in terrestrial environments.

Microplastics (MPs) and antibiotic resistance genes (ARGs) are two classes of emerging and prevalent contaminants in terrestrial environments. To date, effects of MPs on the occurrence of ARGs in terrestrial invertebrates remain uncertain. Here we exposed earthworms to a soil amended with polystyrene MPs at two environmentally relevant concentrations to elucidate the occurrence and mechanisms of ARGs in earthworms impacted by MPs with different sizes. Nano-size and 10 mg/kg of 100 mu m MPs slightly affected the occurrence of ARGs in earthworms. Highest abundance of ARGs was found in the presence of 10 mg/kg of 10 mu m MPs, whereas 100 mg/ kg of 10 mu m MPs significantly changed the profile of ARGs. Metagenomics sequencing and toxicity tests indicated that MPs caused toxicity and influenced the abundance of microbial community in earthworms, resulting in the changes of ARGs. Results of proteomics and metabolomics demonstrated that 100 mg/kg of 10 mu m MPs changed the microenvironment of earthworm gut, built a new homeostatic process, and thus increased the abundance of key bacterial that carried a variety of ARGs. This study highlights the size-dependent toxic effects of MPs and their impacts on the transfer of ARGs in terrestrial environments.