Cloning and sequencing of the β-lactamase gene and surrounding DNA sequences ofCitrobacter braakii,Citrobacter murliniae,Citrobacter werkmanii,Escherichia fergusoniiandEnterobacter cancerogenus

Abstract

To further identify the origins of plasmid-mediated cephalosporinases that are currently spreading worldwide, the chromosomal β-lactamase genes of Citrobacter braakii, Citrobacter murliniae, Citrobacter werkmanii reference strains and of Escherichia fergusonii and Enterobacter cancerogenus clinical isolates were cloned and expressed into Escherichia coli and sequenced. These β-lactamases had all a single pI value >8 and conferred a typical AmpC-type resistance pattern in E. coli recombinant strains. The cloned inserts obtained from genomic DNAs of each strain encoded Ambler class C β-lactamases. The AmpC-type enzymes of C. murliniae, C. braakii and C. werkmanii shared 99%, 96% and 95% amino acid sequence identity, respectively, with chromosomal AmpC β-lactamases from Citrobacter freundii. The AmpC-type enzyme of E. cancerogenus shared 85% amino acid sequence identity with the chromosomal AmpC β-lactamase of Enterobacter cloacae OUDhyp and the AmpC-type enzyme of E. fergusonii shared 96% amino acid sequence identity with that of E. coli K12. The ampC genes, except for E. fergusonii, were associated with genes homologous to regulatory ampR genes of other chromosomal class C β-lactamases that explain inducibility of β-lactamase expression in these strains. This work provides further evidence of the molecular heterogeneity of class C β-lactamases.