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Cefotaxime (CTX) is a third-generation cephalosporin (3GC) commonly used to treat infections caused by  Escherichia coli . Two genetic mechanisms have been associated with 3GC resistance in  E. coli . The first is the conjugative transfer of a plasmid harbouring antibiotic-resistance genes. The second is the introduction of mutations in the promoter region of the  ampC  β-lactamase gene that cause chromosome-encoded β-lactamase hyperproduction. A wide variety of promoter mutations related to AmpC hyperproduction have been described. However, their link to CTX resistance has not been reported. We recultured 172 cefoxitin-resistant  E. coli isolates with known CTX minimum inhibitory concentrations and performed genome-wide analysis of homoplastic mutations associated with CTX resistance by comparing Illumina whole-genome sequencing data of all isolates to a PacBio sequenced reference chromosome. We mapped the mutations on the reference chromosome and determined their occurrence in the phylogeny, revealing extreme homoplasy at the −42 position of the  ampC  promoter. The 24 occurrences of a T at the −42 position rather than the wild-type C, resulted from 18 independent C&gt;T mutations in five phylogroups. The −42 C&gt;T mutation was only observed in  E. coli lacking a plasmid-encoded  ampC  gene. The association of the −42 C&gt;T mutation with CTX resistance was confirmed to be significant (false discovery rate  T mutation of the  ampC  promotor as significantly associated with CTX resistance and underlines the role of recurrent mutations in the spread of antibiotic resistance.

Genome-wide analysis in Escherichia coli unravels a high level of genetic homoplasy associated with cefotaxime resistance