Pathogenic Escherichia coli Isolated from Recreational Waters in Northwestern Wyoming Display Multidrug Resistance

To determine the prevalence, antimicrobial resistance, and genomic characteristics of antimicrobial-resistant Escherichia coli isolated from recreational freshwater in the Greys-Hoback watershed, Wyoming, USA.

A total of 263 presumptive E. coli isolates were recovered using membrane filtration (EPA Method 1603), of which 218 (83%) were confirmed as E. coli by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Antimicrobial susceptibility testing by broth microdilution identified 20 (∼9%) isolates with clinically relevant resistance phenotypes, including 6 multidrug-resistant (MDR; ≥3 antimicrobial classes). Whole-genome sequencing of the resistant subset identified blaEC alleles in all genomes, along with additional resistance determinants, including sul2, tet, aph variants, and floR genes. Plasmid replicons were detected in most isolates, with IncF-type plasmids predominating and associated with MDR gene clusters. Contig-level analysis identified plasmid-associated MDR regions in selected isolates, although plasmid reconstruction was limited by short-read assemblies. Phylogenetic analyses revealed multiple sequence types and O: H serotypes, demonstrating substantial genetic diversity and suggesting that the resistant E. coli population originated from multiple sources rather than from clonal expansion. All sequenced isolates carried multiple virulence-associated genes, indicating co-occurrence of resistance and virulence determinants.

By integrating antimicrobial susceptibility testing, whole-genome sequencing, and plasmid reconstruction, we show that recreational freshwater in northwestern Wyoming harbors genetically diverse antimicrobial-resistant pathogenic E. coli, including MDR isolates associated with putative mobile resistance elements. These findings support the inclusion of recreational water systems in environmental AMR surveillance frameworks and highlight the value of integrating phenotypic and genomic approaches for isolate characterization.