Metformin drives the antibiotic resistome in activated sludge by reshaping microbial communities and promoting...

Aerobic granular sludge (AGS) serves as a major reservoir and dissemination hotspot for human bacterial pathogens (HBPs) and antibiotic resistance genes (ARGs). Metformin (MET) as an emerging contaminant, which exacerbates antibiotic resistance and poses a problem for the stable operation of the activated sludge process in wastewater treatment plants. However, the specific mechanisms underlying the effects of MET stress on microbial communities and ARGs propagation in activated sludge remain poorly understood. In this study, we employed metagenomic analysis to investigate the effects of MET exposure, under a composite antibiotic background, on microbial community dynamics and resistome profiles in AGS systems and interpreted these effects from the perspectives of energy metabolism and community competition. Our findings demonstrate that MET exposure significantly enriched HBPs and multidrug resistance-related ARGs. Co-occurrence network analysis further identified that, among all sludge samples, 27 high-risk HBPs were strongly correlated with ARGs, virulence factor genes, and mobile genetic elements. Additionally, MET was also found to enhance ATP production in specific HBPs, conferring a competitive edge that facilitates ARG accumulation. Furthermore, the natural transformation and conjugation experiments further demonstrated the key role of MET in promoting horizontal gene transfer. In summary, this study underscores the role of MET in exacerbating the ecological risk of antibiotic resistance in AGS systems by concurrently enriching pathogenic bacteria and facilitating the horizontal transfer of ARGs, thereby highlighting the potential environmental impacts of MET as a pervasive contaminant on the propagation of resistance within wastewater treatment ecosystems.