# Tracking ESBL-Producing Escherichia coli Across Municipal Wastewater and Farm Ecosystems: A One Health Investigation

## TL;DR

This study tracks antibiotic-resistant E. coli in wastewater and farms, finding poultry as a major source and highlighting the need for integrated health monitoring.

## Contribution

Identifies poultry as a key reservoir of ESBL-producing E. coli and reveals genetic links between farm and wastewater isolates.

## Key findings

- ESBL-EC was detected in 27.4% of samples, with highest prevalence in poultry (31.5%).
- Resistance genes like blaCMY-2, blaCTX-M-1, and blaOXA frequently co-occurred in poultry isolates.
- Phylogenetic analysis showed clonal relatedness between poultry and cattle isolates.

## Abstract

Antimicrobial-resistant Escherichia coli producing extended-spectrum β-lactamases (ESBL-EC) pose a growing public health concern because they can spread across humans, animals, and shared environments. Here, we investigated how common ESBL-EC are in municipal wastewater treatment plants and farm settings in southeastern North Carolina, and examined their resistance genes and genetic relatedness. ESBL-EC were found in more than one-quarter of all samples tested, with the highest levels detected in poultry, followed by wastewater and cattle. Key resistance genes were identified, often co-occurring in poultry isolates. Genetic analysis showed that some bacteria from poultry and cattle were closely related, suggesting possible transmission between animal reservoirs. Overall, these findings identify poultry as an important reservoir of ESBL-EC and underscore the need for integrated One Health surveillance to reduce the spread of antimicrobial resistance across environmental and agricultural systems.

Extended-spectrum β-lactamase-producing E. coli (ESBL-EC) threatens public health by driving widespread antimicrobial resistance transmission in environmental and agricultural settings. This study examined the prevalence, genetic determinants, and phylogenetic relationships of ESBL-EC isolated from municipal wastewater treatment plants (WWTPs) and farm environments in southeastern North Carolina. A cross-sectional survey was conducted between May and September 2025 across two WWTPs and two farms (cattle and poultry). We sampled influent and effluent wastewater, plus fecal and water specimens collected from chickens, ducks, and cattle. Antimicrobial susceptibility testing was performed using the Kirby–Bauer disk diffusion method against nine drugs, while PCR and sequencing were used for genotypic characterization. Phylogenetic analysis assessed genetic relatedness among isolates. ESBL-EC was detected in 27.4% (n = 124) of 452 samples, with the highest prevalence in chickens (31.5%), followed by WWTP influents (28.2%), ducks (18.5%), and cattle (12.1%). Dominant resistance genes included blaCMY-2 (71.8%), blaCTX-M-1 and blaOXA (54% each), and blaSHV (29.8%). Co-occurrence of blaCMY-2 with blaCTX-M-1 and blaOXA was observed in poultry isolates. Phylogenetic analysis revealed clonal relatedness between poultry and cattle isolates. These findings highlight poultry as a key reservoir and emphasize the need for One Health surveillance to mitigate cross-reservoir transmission of resistant E. coli.