Abstract
Carbapenem-resistant Enterobacteriaceae is one of the emergent medical concerns. In particular, we have to pay attention to carbapenemase-producing bacteria because carbapenemase production is plasmid-encoded and can spread to various species. Since it is difficult to detect carbapenemase producers by phenotypical methods such as the antibiotic sensitivity test, we have developed a new multiplex PCR method. We selected six primer pairs from previous reports and developed a multiplex PCR method to detect six plasmid-mediated carbapenemase genes (blaIMP, blaVIM, blaOXA-48-like, blaNDM, blaKPC and blaGES) in only one tube. We evaluated three types of PCR master mix reagent in terms of sensitivity and specificity, using 5, 1, 2, 2, 1 and 2 strains of plasmid-mediated carbapenemase-producing gram-negative rods with IMP, VIM, OXA-48-like, NDM, KPC and GES, respectively. Moreover, we investigated 25 species (30 strains) of gram-negative rods that do not produce carbapenemase. Platinum® Multiplex PCR Master Mix (Applied Biosystems, Life Technologies Corporation, USA) showed the best performance in screening for six plasmid-mediated carbapenemase genes. Our newly developed multiplex PCR method can discriminate carbapenemase genes in 3.5 hours using only one tube. We conclude that our newly developed assay may contribute to the detection of carbapenemase-producing bacteria with good reactivity and rapidity.
