Evaluation of peptide nucleic acid-mediated multiplex real-time PCR kits for rapid detection of carbapenemase genes in gram-negative clinical isolates

Seri Jeong, Jung Ok Kim, Seokhoon Jeong, Il Kwon Bae, Wonkeun Song

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Abstract

The emergence of clinical isolates of carbapenemase-producing microbes confers multidrug-resistance to these bacteria and renders them difficult to treat. This study was performed to evaluate peptide nucleic acid (PNA) probe-based multiplex real-time PCR kits used to detect carbapenemase genes. In total, 324 carbapenemase genes, collected from 318 gram-negative clinical isolates in 36 different hospital laboratories, were assayed to evaluate multiplex real-time PCR kits (PANAGENE; Daejeon, Korea). The nine most prevalent carbapenemase genes (. KPC, OXA-48, GES, IMP, VIM, NDM, IS. Aba1-OXA-51, OXA-23, and OXA-58) were included in this study. The sensitivity and specificity of the multiplex real-time PCR assay to all of the carbapenemase genes were above 99.0%, except for IS. Aba1-OXA-51. The detection limit of the assay was 100 target copies per 25. μL of reaction volume for all of the nine genetic types of carbapenemases, and the genes were all detected in a single three-hour PCR. The assay also showed considerable efficiency (above 80.0%), stable reproducibility (coefficient of variation, below 5.0%) and a long shelf-life (more than eight months) with no cross reactivity. The developed PNA-mediated multiplex real-time PCR assay was useful for the rapid, accurate and simultaneous identification of nine carbapenemase genes in gram-negative clinical isolates, suggesting its potential to help choose the appropriate antibiotics and aid the control of carbapenemase genes.

Original languageEnglish
Pages (from-to)4-9
Number of pages6
JournalJournal of Microbiological Methods
Volume113
DOIs
Publication statusPublished - 2015 Jun 1

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All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology
  • Microbiology (medical)

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