Metagenomic Profiling\nof Internationally Sourced Sewage\nInfluents and Effluents Yields Insight into Selecting Targets for\nAntibiotic Resistance Monitoring

It has been debated whether wastewater treatment plants\n(WWTPs)\nprimarily act to attenuate or amplify antibiotic resistance genes\n(ARGs). However, ARGs are highly diverse with respect to their resistance\nmechanisms, mobilities, and taxonomic hosts and therefore their behavior\nin WWTPs should not be expected to be universally conserved. We applied\nmetagenomic sequencing to wastewater influent and effluent samples\nfrom 12 international WWTPs to classify the behavior of specific ARGs\nentering and exiting WWTPs. In total, 1079 different ARGs originating\nfrom a variety of bacteria were detected. This included ARGs that\ncould be mapped to assembled scaffolds corresponding to nine human\npathogens. While the relative abundance (per 16S rRNA gene) of ARGs\ndecreased during treatment at 11 of the 12 WWTPs sampled and absolute\nabundance (per mL) decreased at all 12 WWTPs, increases in relative\nabundance were observed for 40% of the ARGs detected at the 12th WWTP.\nAlso, the relative abundance of mobile genetic elements (MGE) increased\nduring treatment, but the fraction of ARGs known to be transmissible\nbetween species decreased, thus demonstrating that increased MGE prevalence\nmay not be generally indicative of an increase in ARGs. A distinct\nconserved resistome was documented in both influent and effluent across\nsamples, suggesting that well-functioning WWTPs generally attenuate\ninfluent antibiotic resistance loads. This work helps inform strategies\nfor wastewater surveillance of antibiotic resistance, highlighting\nthe utility of tracking ARGs as indicators of treatment performance\nand relative risk reduction.