Duplication of blaCTX-M-1 and a class 1 integron on the chromosome enhances antimicrobial resistance in Escherichia coli isolated from racehorses in Japan

Abstract

Extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae have become a cause for great concern. Although some studies have reported the prevalence of ESBL-producing bacteria and ESBL-encoding genes in horses worldwide, the genetic structure surrounding the ESBL gene has not been analysed in detail. In the present study, we isolated two ESBL-producing Escherichia coli strains from diseased racehorses in Japan and demonstrated the mechanisms underlying the acquisition of their antimicrobial resistance (AMR) genes. Two ESBL-producing E. coli strains (E148 and E189) were isolated from the heart and liver of horses with endocarditis and sepsis in 2014 and 2016, respectively, in Japan. Complete genomic sequences of the two strains were analysed using a PacBio RSII sequencer. Antimicrobial susceptibility testing was performed by the agar dilution method. The two isolates possessed a chromosomal AMR gene cluster containing blaCTX-M-1 that was similar to the pEQ1 plasmid found in E. coli isolated from a racehorse in the Czech Republic. In one of the two strains, tandem duplication of the 16-kb region containing blaCTX-M-1 and a class 1 integron, which occurred via IS26-mediated recombination, increased minimum inhibitory concentrations (MICs) associated with the duplicated AMR genes. Chromosomal blaCTX-M-1 possibly derived from the pEQ1 or pEQ1-like plasmid was found in Japanese equine E. coli isolates. In Japanese strains, many AMR genes containing blaCTX-M-1 and the class 1 integron are highly accumulated in one region on the chromosome, and the AMR of E. coli was enhanced via the IS26-mediated duplication of the AMR gene cluster.