Cloning and characterization of bla(VIM), a new integron-borne metallo- β-lactamase gene from a Pseudomonas aeruginosa clinical isolate

Production of a metallo-β-lactamase activity was detected in a carbapenem-resistant Pseudomonas aeruginosa clinical isolate (isolate VR- 143/97) from an Italian inpatient at the Verona University Hospital (northern Italy). The metallo-β-1actamase determinant was isolated from a genomic library of VR-143/97, constructed in an Escherichia coli plasmid vector, by screening for clones with reduced susceptibility to imipenem. Sequencing of the cloned gene revealed that it encoded a new class B β-1actamase that was named VIM-1. At the sequence level VIM-1 was rather divergent from the other class B enzymes (16.4 to 38.7% identity), overall being more similar to members of subclass B1 including the β-1actamase II of Bacillus cereus (Bc- II), the Bacteroides fragilis CcrA, the Chryseobacterium meningosepticum BlaB, and the cassette-encoded IMP-1 enzymes. Among these, VIM-1 showed the highest degree of similarity to Bc-II. Similarly to bla(IMP), bla(VIM) was also found to be carried on a gene cassette inserted into a class 1 integron. The bla(VIM)-containing integron was located on the chromosome of P. aeruginosa VR-143/97, and the metallo-β-1actamase-encoding determinant was not transferable to E. coli by conjugation. Expression of the integron-borne bla(VIM) gene in E. coli resulted in a significant decrease in susceptibility to a broad array of β-lactams (ampicillin, carbenicillin, piperacillin, mezlocillin, cefotaxime, cefoxitin, ceftazidime, cefoperazone, cefepime, and carbapenems), revealing a very broad substrate specificity of the VIM-1 enzyme.