Genomic Epidemiology of Antibiotic-Resistant Bacteria Sampled from Metropolitan Wastewater

Wastewater surveillance is an effective approach for monitoring populations of antibiotic-resistant bacteria and tracking the spread of antimicrobial resistance (AMR) across different settings. In this study, hospital and municipal wastewater were collected monthly for 12 months from multiple locations in the greater Pittsburgh area to quantify the presence of antibiotic-resistant bacteria and investigate their genomic diversity. After quantitative culturing on six different selective media types, a total of 150 isolates were speciated by 16S rRNA sequencing, which revealed diverse pathogenic and non-pathogenic taxa, including Klebsiella spp. (n = 28), Pseudomonas spp. (n = 20) and Aeromonas spp. (n = 37). A subset of isolates (n = 46) underwent whole genome sequencing, which identified several antibiotic resistance genes of clinical concern, such as bla_KPC (n = 17), bla_NDM (n = 6) and bla_IMP (n = 6), and revealed genetic similarities between wastewater isolates and clinical isolates collected from infected patients at a Pittsburgh-area medical center. In addition, analysis of plasmids carried by wastewater isolates revealed closely related plasmids present in isolates from different species and sampling locations. Overall, these findings suggest that both hospital and municipal wastewater act as interconnected reservoirs of antimicrobial resistance. Integrating wastewater surveillance with clinical and genomic data could enable the early detection of emerging resistance threats and support proactive infection-control strategies.