Emergence of blaKPC-3-Tn4401a associated with a pKPN3/4-like plasmid within ST384 and ST388 Klebsiella pneumoniae clones in Spain

KPC carbapenemase-producing Klebsiella pneumoniae isolates have been increasingly detected in Europe. We report the first KPC-producing isolates characterized in Madrid, Spain. Twelve K. pneumoniae isolates recovered from clinical and surveillance cultures from eight patients (September 2009 to February 2010) that were resistant to carbapenems and resulted in a positive modified Hodge test, were screened for carbapenemase genes (PCR, sequencing and hybridization). Clonal relatedness was established by PFGE and multilocus sequence typing. Plasmid characterization included incompatibility group assay and XhoI/HindIII restriction pattern comparison. The genetic environment was characterized by PCR based on the Tn4401 sequence. All 12 isolates were resistant to all beta-lactams, including imipenem and meropenem (MIC >= 8 mg/L). Eleven of them, fully susceptible to aminoglycosides and fluoroquinolones, showed related PFGE patterns and belonged to sequence type (ST) 384 and harboured the bla(KPC-3), bla(OKP-5), bla(OXA-9) and bla(TEM-1) genes, whereas one isolate resistant to quinolones belonged to ST388, and also harboured the bla(CTX-M-10) gene. The bla(KPC-3), bla(OXA-9) and bla(TEM-1) genes were located on an similar to 85 kb non-transferable plasmid that was a derivative of pKPN3/pKPN4 previously described in K. pneumoniae. The bla(KPC-3) gene was located on the Tn4401 'isoform a' variant, which is usually linked to bla(KPC-2), but rarely to bla(KPC-3). The emergence of Tn4401-bla(KPC-3) within a pKPN3/4-like plasmid and its novel association with ST384 and ST388 K. pneumoniae clones in Spain is reported. Although bla(KPC-3) has been scarcely reported in Europe, the location of this Tn4401a in a widespread K. pneumoniae plasmid supports the possibility of broader future dissemination.