Molecular characterization of carbapenem-resistant Klebsiella pneumoniae isolates from a university hospital in Brazil

Abstract

Introduction: The emergence of Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae (KPC-Kpn) isolates is attracting significant attention in nosocomial infection settings. K. pneumoniae is the main pathogen that harbours blaKPC genes. Methodology: This study evaluated 54 K. pneumoniae carbapenem-resistant isolates from patients hospitalized at the University Hospital of Londrina, between July 2009 and July 2010. The isolates were phenotypically screened for carbapenemase production and submitted for genotypic confirmation by polymerase chain reaction (PCR) for KPC, metallo-β-lactamases, OXA-48, and extended-spectrum beta-lactamase genes. The absence of outer membrane proteins (OMP) was investigated by SDS-PAGE. The susceptibility profile was determined by broth microdilution, according to Clinical and Laboratory Standards Institute protocol. Results: All isolates were phenotypically positive for class A carbapenemase production, but negative for metallo-β-lactamase activity. PCR analysis demonstrated that all isolates carried blaKPC genes and sequencing showed that all strains belonged to KPC-2 subtype. Four strains did not show porin expression, and all isolates were resistant to ertapenem, meropenem, and imipenem. Susceptibility rates reached 35.2% for gentamicin, 85.2% for polymixyn B, 87% for colistin, and 98.1% for both tigecycline and fosfomycin. Pulsed-field gel electrophoresis showed six clones, and three of them predominated among the isolates. Conclusions: KPC-2-producing K. pneumoniae is becoming predominant among carbapenem-resistant K. pneumoniae isolates at the hospital. The association of the enzyme KPC with other resistance determinants, such as loss of porins, may increase the severity of the situation of nosocomial infections. There is an urgent need to develop strategies for infection control and prevention.