The gut as a reservoir of drug-resistant pathogens: Mechanisms of ENR-driven horizontal gene transfer in aquaculture.

Enrofloxacin (ENR), commonly used in aquaculture, plays a role in the development and dissemination of antibiotic resistance genes (ARGs). While most research on ARGs has focused on the environment, the gut, the host's largest microbial habitat, remains underexplored. Accordingly, this research investigates the gut microbiome, aiming to assess the potential mobility of ARGs after ENR exposure. Additionally, ENR exposure alters short-chain fatty acid (SCFAs) levels. Subsequent conjugation transfer experiments demonstrated that ENR exposure modifies SCFA levels, and this alteration facilitates the spread of ARGs. Both plasmid- and phage-mediated ARGs transmission were observed. ENR exerted selective pressure on the gut microbiota, significantly promoting plasmid-mediated conjugation as a key driver of ARGs dissemination. Simultaneously, environmental stress triggered the release of progeny phages carrying ARGs, further facilitating their spread. Conjugation experiments confirmed that ENR and SCFAs interact with bacterial outer membrane proteins, inducing the production of ROS. As a result of ROS production, membrane integrity is disrupted and membrane permeability is increased, ultimately causing an increase in the frequency of conjugative transfer and facilitating the horizontal delivery of ARGs. Therefore, ENR not only directly influences the transmission of ARGs but also indirectly promotes their transmission by altering SCFA levels. The study findings underscore the risks posed by excessive use of ENR in aquaculture to public health, providing scientific evidence to prevent food safety hazards from market entry of aquatic products carrying drug-resistant pathogens.