High-frequency secondary mutations after suicide-driven allelic exchange mutagenesis in extraintestinal pathogenic Escherichia coli

Frequent unintended secondary mutations occurred in extraintestinal pathogenic Escherichia coli strains CP9, CFT073, and RS218 during suicide plasmid-mediated, putatively specific deletions of hlyA, papG allele III, and iha. Pulsed-field gel electrophoresis and PCR analyses demonstrated genomic alterations and/or unintended loss of defined virulence genes (papG, the F7-2 papA allele, iutA, sat, hlyD, and cnf). Caution is warranted when attributing the observed phenotypic changes to the intended mutation.