Differential variations of intracellular and extracellular antibiotic resistance genes between treatment units in centralized sewage sludge treatment plants

Centralized sludge treatment plants (CSTPs) are implicated as strong hotspots of antibiotic resistance genes (ARGs). However, the knowledge gap on the fate of intracellular and extracellular ARGs (iARGs and eARGs), and the functionality of resistant hosts limit risk assessment and management of CSTP resistome. Here, the flow of iARGs and eARGs across treatment units and analyses of ARG hosts were systematically explored in three full-scale CSTPs using quantitative metagenomic approaches. We found that 29% of sludge ARGs could be removed, with iARGs being dominant in the produced biosolids. The treatment process significantly affected the variations of iARG and eARG abundance while no significant difference in composition between iARGs and eARGs was observed in CSTPs. 15% of 295 recovered genomes were identified as antibiotic-resistant hosts, among which Actinobacteriota tended to encode multiple resistance. The key functions of ARG hosts were relative to the biological organic removal (e.g., carbohydrates). There also existed relationships between certain resistance mechanisms and functional traits, indicating that ARGs might take part in the physiological process of microorganisms in the sludge treatment. These findings provide important insight into the differential resistome variations and host functionality, which would be crucial in the management of antibiotic resistance in CSTPs.