Comprehensive insights into the role of extracellular polymeric substances of Geobacter in the horizontal transfer of antibiotic resistance genes

Extracellular polymeric substances (EPS) in bioelectrochemical systems play a pivotal role in the removal of antibiotics and regulation of the dissemination of antibiotic resistance genes (ARGs). However, a comprehensive analysis of the EPS secreted by electroactive bacteria is still lacking to fully understand its role in regulating ARGs transmission mechanisms. In this study, the transformation system using plasmid pBBR1MCS-3 and Geobacter to investigate the impact of EPS on transformation frequency under roxithromycin (ROX) stress. Compared with the absence of EPS, the transformation frequency of pBBR1MCS-3 in the presence of EPS was reduced by 66.8 %. Results showed that EPS could decrease ROX content, thereby reducing Geobacter intracellular reactive oxygen species levels, SOS response, and cell membrane permeability. The interaction mechanism between EPS and ARGs was further elucidated through fluorescence quenching, X-ray photoelectron spectroscopy, fouriertransform infrared spectroscopy, and theoretical calculations. EPS forms a novel complex with ARGs through static quenching. Carboxyl and amine functional groups of protein in EPS could interact with ARGs via hydrogen bonding. Furthermore, PLS-PM analysis demonstrated that under ROX pressure, the secretion of EPS by Geobacter primarily reduced the horizontal genes transfer (HGT) of ARGs by alleviating oxidative stress induced by ROX. This study unveils the mechanism underlying the impact of EPS secreted by Geobacter under ROX pressure on the HGT of ARGs while highlighting its potential role in slowing down the spread of ARGs.