Metallo-β-lactamases in clinical Pseudomonas isolates in taiwan and identification of VIM-3, a novel variant of the VIM-2 enzyme

A total of 209 clinical isolates Pseudomonas (193 Pseudomonas aeruginosa, 10 P. putida, 4 P. stutzeri, and 2 P. fluorescens isolates) with reduced susceptibilities to imipenem and/or ceftazidime were subjected to PCR assays with primers specific for bla(IMP-1). bla(IMP-2), bla(VIM-1), and bla(VIM-2) and sequence analysis to identify the metallo-beta -lactamases (MBLs) prevalent among these organisms in Taiwan; and 21 isolates gave positive results. Five isolates including two P. putida and three P. stutzeri isolates were found to carry bla(IMP-1) and six isolates including five P. putida and one P. stutzeri isolates harbored bla(VIM-2). The remaining 10 isolates were P. aeruginosa and all were found to carry a novel variant of bla,,,, designated bla,,,. There are only two nucleotide differences between bla(VIM-2) and bla(VIM-3), leading to two amino acid alterations. Our findings indicate that VIM-2 and its variant have become the most prevalent metalloenzymes in Pseudomonas in Taiwan. Southern hybridization with the bla(VIM-2)-, bla(VIM-3)-, and bla(IMP-1) -specific probes revealed that only two VIM-2-producing P. putida isolates appeared to carry the MBL gene on plasmids. Pulsed-held gel electrophoresis showed that six VIM-3-producing P. aeruginosa isolates and two IMP-1-producing P. stutzeri isolates were genetically related, suggesting that the spread of these MBL genes in Taiwan could be due to clonal dissemination as well as genetic exchange between different clones.