Antibiotic resistance evolution driven synergistically by antibiotics and typical organic pollutants in antibiotic production wastewater

A major concern regarding the risk of antibiotic production wastewater (APW) for the transmission of antibiotic resistance (AR) stems from the residual antibiotics. However, APW also contains high concentrations of organic pollutants, many of which have severe biological toxicity and joint toxicity with antibiotics. The contribution of these organic pollutants to the development of AR in the APW treatment system is unknown. In this study, a wild-type Escherichia coli strain was exposed to six typical organic pollutants in APW individually and synergistically with the antibiotic ampicillin (AMP). Independent exposure to organic compounds had negligible effects on the evolution of AR, whereas they synergistically induced AR mutations and increased antibiotic persistence with AMP, especially the raw material d-p-hydroxyphenylglycine (DHPG), at relevant concentrations in APW. Combined exposure to 1–500   mg/L DHPG and 1   mg/L AMP synergistically increased the mutation frequencies against multiple antibiotics by up to 2928.9-fold in a dose–time pattern, and the combination index reached 445.7. Phenotypic and genotypic analyses revealed that the synergism between DHPG and AMP was associated with increased antibacterial activity, enhanced oxidative stress, and stimulation of efflux pump expression. Overall, our results highlight the elevated risk of AR induction caused by antibiotics and organic pollutants in APW.