| Summary: | Despite its importance in veterinary medicine, there is little information about antimicrobial resistance (AMR) and its transmission in dairy cattle. The aim of this work is to compare AMR phenotypes and genotypes in resistant <i>Escherichia coli</i> and to determine how the resistance genes spread among the <i>E. coli</i> population on dairy farms in Québec, Canada. From an existing culture collection of <i>E. coli</i> isolated from dairy manure, a convenient selection of the most resistant isolates (a high level of multidrug resistance or resistance to broad-spectrum β-lactams or fluoroquinolones) was analyzed (<i>n</i> = 118). An AMR phenotype profile was obtained for each isolate. Whole genome sequencing was used to determine the presence of resistance genes, point mutations, and mobile genetic elements. In addition, a subset of isolates from 86 farms was taken to investigate the phylogenetic relationship and geographic distribution of the isolates. The average agreement between AMR phenotypes and genotypes was 95%. A third-generation cephalosporin resistance gene (<i>bla<sub>CTX-M-15</sub></i>), a resistance gene conferring reduced susceptibility to fluoroquinolones (<i>qnrS</i>1), and an insertion sequence (ISKpn19) were detected in the vicinity of each other on the genome. These genes were harbored in one triplet of clonal isolates from three farms located >100 km apart. Our study reveals the dissemination of resistant <i>E. coli</i> clones between dairy farms. Furthermore, these clones are resistant to broad-spectrum β-lactam and fluoroquinolone antimicrobials.
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