comparison of phenotypic methods and pcr for the detection
TRANSCRIPT
Comparison of phenotypic methods and PCR for the detection of
carbapenemase production in clinical Klebsiella pneumoniae isolates
Tekintas Y*1, Cilli FF2, Erac B3, Yasar M2, Aydemir SS2, Hosgor-Limoncu M 3 1Izmir Katip Celebi University, Faculty of Pharmacy, Department of Pharmaceutical Microbiology, Izmir,Turkey
2Ege University, Faculty of Medicine, Department of Medical Microbiology, Izmir, Turkey 3Ege University, Faculty of Pharmacy, Department of Pharmaceutical Microbiology, Izmir, Turkey
*E-mail: [email protected]
BACKGROUND
Multidrug resistant Klebsiella pneumoniae isolates are a major problem both in our country and the World.
Carbapenems are frequently used in the treatment of nosocomial K. pneumoniae infections. Nowadays, OXA and
Metallo beta lactamase (MBL) group enzymes are detected in the carbapenem resistant Klebsiella isolates. The
aim of this study is to compare phenotypic and genotypic methods for the detection of OXA-48 and MBLs, which
are thought to be responsible for carbapenem resistance of K. pneumoniae strains.
MATERIAL/METHODS
Carbapenemase presence and types were investigated in carbapenem resistant 54 K. pneumoniae strains isolated
from the Ege University Hospital Bacteriology Laboratory of Medical Microbiology Department. VITEK MS
and VITEK 2 Compact® automated systems were used for identification and antibiotic susceptibilities of the
strains, respectively. In carbapenem resistant strains, minimum inhibitor concentration (MIC) values of
meropenem were determined by gradient test method in the direction of EUCAST recommendations.
Phenotypic enzyme typing was done with the '' MASTDISCS ™ ID carbapenemase (Enterobacteriaceae)
detection disc set ''. The presence of OXA-48 and MBL (IMP, VIM, SIM, NDM) genes in the isolates was
investigated by polymerase chain reaction (PCR). Carbapenem Inactivation Method (CIM) was also used to
detect carbapenemase activity.
RESULTS
According to the VITEK 2 Compact® and gradient test results, 54 K. pneumoniae isolates were found to be
resistant to at least one carbapenem. Imipenem, meropenem and ertapenem MIC50 and MIC90 values were
determined as 32 μg / ml. Only OXA-48 was found in 33 of the strains and only NDM gene was detected in
two strains, but 19 strains were found to contain both genes by PCR. SIM, VIM, IMP genes were not
encountered in any of the isolates.
Table. Compatibility of phenotypic methods with PCR
CONCLUSIONS
MBL and OXA-48 genes were detected at high levels in carbapenem resistant K. pneumoniae strains isolated in
our hospital. Our results suggested that MASTDISC is highly compatible with PCR for single carbapenemase
gene harboring isolates, For isolates carrying two carbapenemase genes (OXA-48+NDM), MASTDISC should be
verified with PCR for understanding type of the enzyme. CIM has showed a low detection level in isolates only
containing OXA-48 (21%). The success rate is much higher in strains which carries NDM and OXA-48 genes
together (89%).
MASTDISC CIM
Carbapenemase Phenotype Number of strains Phenotype Number of strains
gene
blaOXA-48 n=33 MBL 0 (+) 7 (21%)
OXA-48 33 (100%) (-) 26 (79%)
blaOXA-48+blaNDM n=19 MBL 18 (95%) (+) 17 (89%)
OXA-48 1 (5%) (-) 2 (11%)
blaNDM n=2 MBL 2 (100%) (+) 1 (50%)
OXA-48 0 (-) 1 (50%)
MBL: Metallo Beta Lactamase, CIM: Carpabenemase Inactivation Method