Antibiotic resistance genes (ARGs) are a growing problem that is widespread in river-lake ecosystems, where they pose a threat to the aquatic environment's health and public safety. These systems serve as critical nodes in water management, as they facilitate the equitable allocation of water resources through long-term and frequent water diversions. However, hydrological disturbances associated with water-regulation practices can influence the dynamics of their potential host microorganisms and associated resistance genes. Consequently, identifying the key ARGs and their resistance mechanisms in heavily regulated waters is vital for safeguarding human health and that of river-lake ecosystems. In this study, we examined the impact of water-regulation factors on ARGs and their hosts within a river-lake continuum using 16S rRNA and metagenomic sequencing. We found that a significant increase in ARG abundance during regulation periods (p < 0.05), especially in the aquatic environment. Key resistance genes were macB, tetA, evgS, novA, and msbA, with increased efflux pinpointed as their principal resistance mechanism. Network analysis identified Flavobacteriales, Acinetobacter, Pseudomonas, Burkholderiaceae, and Erythrobacter as key potential host microorganisms, which showed increased abundance within the water column during regulation periods (p < 0.05). Flow velocity and water depth both drove the host microorganisms and critical ARGs. Our findings underscore the importance of monitoring and mitigating the antibiotic resistance risk during water transfers in river-lake systems, thereby supporting informed management and conservation strategies.

Antibiotic pollution from manured farmland soils is a major public concern, and their potential interaction with manure particles and/or manure–dissolved organic matter (DOM) often complicates their leaching behaviour. This study investigated the leaching of sulfadiazine (SDZ) and florfenicol (FFC) with manure-DOM in undisturbed field lysimeters and repacked soil columns under natural and simulated rainfall conditions. The results showed that manure-DOM reduced SDZ mass flux, but soil hydrological processes and heavy rainfall events led to accelerated leaching. FFC was more prone to leaching in a manured plot (0.48 µg m⁻² h⁻¹) compared to the control (0.12 µg m⁻² h⁻¹), suggesting DOM facilitated transport of FFC in the field lysimeter study via cotransport mechanisms favored by abundant macropores in the study site. In contrast, SDZ and FFC mobility were reduced in repacked soil columns under manure-DOM conditions, suggesting complexation and adsorption in matrix pores. Two kinetic site models and two-site nonequilibrium adsorption models revealed the existence of nonequilibrium conditions and kinetic sorption processes in the repacked column. FFC exhibited lower leaching potential compared to SDZ in both the repacked column and natural field conditions. Redundancy analyses revealed that FFC had a close relationship with humic-like components (C1 and C3), but SDZ was more related to the protein-like components (C2) of DOM. The presence of manure-DOM may decrease the ecological risks of highly mobile antibiotics under matrix flow through complexations and adsorption. However, a similar scenario in macroporous flow under heavy rainfalls may lead to accelerated leaching.