Phenotypic and Molecular Detection of the Metallo-Beta-Lactamases in Carbapenem-Resistant <i>Pseudomonas aeruginosa</i> Isolates from Clinical Samples

Abstract

Background: Pseudomonas aeruginosa is an opportunistic pathogen that causing high morbidity and mortality, and responsible for a high proportion of hospital infections. Recently, P. aeruginosa infections having carbapenem group antibiotic-resistant have been increasing and serious problems are being experienced in treatment. Carbapenem resistance mechanisms are the loss of OprD porin protein, excretion of drug with the efflux system and production of metallo-beta-lactamase (MBL). Objectives: The aim of this study was to investigate the production of MBL in carbapenem resistant P. aeruginosa isolates phenotypically and genotypically. Study design: In this study, 58 P. aeruginosa isolates were included from various clinical samples between May 2014 and March 2015 at the Microbiology Laboratory of Tertiary Care Hospital of Mersin University, Faculty of Medicine. Methods: The strains were identified by automated system (Vitek 2, BioMerieux, France); and antibiotic susceptibilities were performed by using the disc diffusion method and the automated system (Vitek 2). Gradient test was performed for imipenem (IPM) and meropenem (MEM) to confirm the carbapenem resistance. Combined disc test (CDT) and MBL E-test were used for the detection of MLB phenotypically. The most common MBL genes, blaIMP-1, blaIMP-2, blaVIM1, blaVIM-2, blaSPM, blaGIM-1 were investigated with polymerase chain reaction (PCR). Direct DNA sequence analysis was performed to PCR positive isolates. Results: Of the 58 isolates, 57 (99%) were found resistant to IPM, 43 (74%) to MEM, one was intermediate susceptible to IPM and 15 (25%) to MEM by automated system. Of the 58 isolates, 57 were found to be resistant and one was intermediate susceptible to IPM, 34 isolate were found to be resistant, 17 were intermediate susceptible and 7 were susceptible to MEM by gradient test. MBL positivity was detected on 37 (63.7%) isolates by CDT and 16 (27%) by E-test. MBL gene region was detected on 8 isolates (13.7%) by PCR and 6 isolates were found to be VIM-1 gene region positive and 2 isolates were GIM-1 gene region positive. This study is the first study showing the GIM-1 gene in P. aeruginosa in Turkey. DNA sequence analysis data was compared with the reference NCBI sequence data in the PubMed-BLAST program to confirm gene regions. Conclusions: Phenotypic tests can be used as screening test for the detection of MBL activity in carbapenem resistant P. aeruginosa isolates. However, confirmation of the results by molecular tests and determination of the gene regions responsible for resistance are very important epidemiologically.