A novel, multiplex, real-time PCR-based approach for the detection of the commonly occurring pathogenic fungi and bacteria

Ádám Horváth, Zoltán Pet, E. Urbán, C. Vágvölgyi, F. Somogyvári

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Background: Polymerase chain reaction (PCR)-based techniques are widely used to identify fungal and bacterial infections. There have been numerous reports of different, new, real-time PCR-based pathogen identification methods although the clinical practicability of such techniques is not yet fully clarified.The present study focuses on a novel, multiplex, real-time PCR-based pathogen identification system developed for rapid differentiation of the commonly occurring bacterial and fungal causative pathogens of bloodstream infections. Results: A multiplex, real-time PCR approach is introduced for the detection and differentiation of fungi, Gram-positive (G+) and Gram-negative (G-) bacteria. The Gram classification is performed with the specific fluorescence resonance energy transfer (FRET) probes recommended for LightCycler capillary real-time PCR. The novelty of our system is the use of a non-specific SYBR Green dye instead of labelled anchor probes or primers, to excite the acceptor dyes on the FRET probes. In conjunction with this, the use of an intercalating dye allows the detection of fungal amplicons.With the novel pathogen detection system, fungi, G + and G- bacteria in the same reaction tube can be differentiated within an hour after the DNA preparation via the melting temperatures of the amplicons and probes in the same tube. Conclusions: This modified FRET technique is specific and more rapid than the gold-standard culture-based methods. The fact that fungi, G + and G- bacteria were successfully identified in the same tube within an hour after the DNA preparation permits rapid and early evidence-based management of bloodstream infections in clinical practice.

Original languageEnglish
Article number300
JournalBMC Microbiology
Volume13
Issue number1
DOIs
Publication statusPublished - Dec 23 2013

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Fluorescence Resonance Energy Transfer
Real-Time Polymerase Chain Reaction
Fungi
Bacteria
Coloring Agents
Mycoses
DNA
Infection
Gram-Negative Bacteria
Bacterial Infections
Gold
Freezing
Polymerase Chain Reaction
Temperature

Keywords

  • Bloodstream infections
  • Candida
  • Clinically relevant bacteria
  • Fluorescence resonance energy transfer
  • Melting point analysis

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

Cite this

A novel, multiplex, real-time PCR-based approach for the detection of the commonly occurring pathogenic fungi and bacteria. / Horváth, Ádám; Pet, Zoltán; Urbán, E.; Vágvölgyi, C.; Somogyvári, F.

In: BMC Microbiology, Vol. 13, No. 1, 300, 23.12.2013.

Research output: Contribution to journalArticle

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