Deciphering antibiotic resistance gene transfer in activated sludge systems for piggery wastewater: behaviors, hosts and drivers

Understanding the transfer and driving mechanisms of antibiotic resistance genes (ARGs) in activated sludge is essential for mitigating environmental risks, particularly during real wastewater treatment where these processes remain poorly characterized. This study investigated the prevalence of ARGs in a sequencing batch reactor (SBR) – up-flow microaerobic sludge reactor (UMSR) system treating high-risk piggery wastewater and revealed critical pathways for resistance propagation. Ten prevalent ARG subtypes, categorized into three types, were selected as target genes, exhibiting a total relative abundance of 0.52 copies per 16S rRNA in raw wastewater. The SBR-UMSR system reduced total ARGs by 0.04 log in wastewater, with half of subtypes decreasing 0.14–1.30 log, despite 0.23–0.58 log enrichment in sludge. By integrating correlation analysis with partial least-squares path modeling, this study identified Burkholderiaceae as the primary potential host of ARGs and pinpointed other high-risk hosts. It further revealed two crucial mechanisms: i) conjugation-mediated horizontal gene transfer dominated ARG propagation, and ii) bacterial community succession served as the main driving force for ARG transfer. This study advances mechanistic understanding of ARG transmission in real wastewater systems, providing critical insights for optimizing sludge management to mitigate antibiotic resistance risks.