Molecular characterization of AmpC-producing Escherichia coli clinical isolates recovered at two Belgian hospitals

Objective. - To characterize the genetic determinants associated with an AmpC phenotype in clinical Escherichia coli isolates. Methods. - E. coli strains recovered at two Belgian hospitals between 2004 and 2006 were selected on the basis of an AmpC-producing phenotype. Plasmid-mediated cephalosporinases coding genes and the sequence of chromosomal ampC genes were identified by PCR/sequencing. The isolates were submitted to phylotyping and genotyping analysis using rep-PCR (Diversilab) and PFGE. A novel chromosomal ampC gene was cloned. Results. - Eighty-three out of 6850 E. coli isolates were selected. Seventy-two isolates were found to overexpress their chromosomal cephalosporinases while 11 contained plasmid-mediated cephalosporinases. Among chromosomal AmpC overproducers, 12 were extended-spectrum AmpC (ESAC) expressing isolates which all displayed reduced susceptibility to cefepime. Cloning of a new ESAC allele suggested that L293P mutation was responsible of the extension of the hydrolysis spectrum to cefepime. AmpC overproducers, including ESAC producers, predominantly belonged to phylogenetic group A and B1, while plasmid-mediated AmpC-producing isolates preferentially belong to phylogroup B2 and D. According to rep-PCR, the majority of the E. coli isolates belonging to phylogroup A were clonally related which was further confirmed by PFGE for the 11 ESAC expressing isolates. Conclusions. - Chromosomal AmpC overproduction was the most common resistance mechanism, and the occurrence of ESAC was found to be as frequent as plasmid-mediated cephalosporinases. The detection of a new ESAC allele, of an ESAC producing strain belonging to phylogroup D and the existence of a clonal relationship between ESAC producing strains underline the need for study of the clinical relevance of this mechanism of resistance. (C) 2009 Elsevier Masson SAS. All rights reserved.