Perturbation and mechanism of non-antibiotic drug in regulating resistome and metabolome of anammox consortia: An overlooked and underrated cause of multiresistance

Abstract

Given that antibiotic resistance genes (ARGs) become emerging pollutants, their accumulation and transfer caused by extensive use of antibiotic and non-antibiotic drugs raise concerns. In contrast, the impacts of non-antibiotic drugs have been largely overlooked, and their regulation mechanism in anammox system remains unclear. In this study, the single and combined effects of typical antibiotic (doxycycline) and non-antibiotic drug (propranolol) on anammox consortia were evaluated and compared from process performance, community succession, metabolic and gene regulation perspectives. Significant reduction in the nitrogen removal efficiency (49.0 ± 0.5%) was only observed in the anammox process stressed by propranolol and doxycycline, but it was reversible. Metabolome results showed that anammox consortia upregulated amino acid metabolism under propranolol stress, while the abundance of pyrimidine and purine metabolic pathways significantly increased under doxycycline and multidrug stress. Microorganisms adapted to these stresses by changing the utilization preference for metabolites. Metagenomic analyses indicated that doxycycline and propranolol induced the high frequency of gene transfer through increasing oxidative stress level, membrane permeability, pili and efflux pumps. Significant correlations between ARGs and mobile genetic elements (MGEs), especially intI1, demonstrated the conjugative transfer of ARGs. Furthermore, structural equation models showed that microbial community, bacterial metabolism, MGEs, reactive oxygen species (ROS) and corresponding SOS response were key driving factors for ARGs transfer in the anammox systems. These findings fill the gap in the effect of non-antibiotic drugs on anammox system and their regulation mechanism, which further provides a theoretical guidance for anammox-based process to treat multidrug-containing wastewater.