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Novel multiplex real-time quantitative PCR detecting system approach for direct detection of Candida auris and its

Amir Arastehfar1, Wenjie Fang1,2,3, Farnaz Daneshnia1

  • 1Westerdijk Fungal Biodiversity Institute, Utrecht 3584, The Netherlands.

Future Microbiology
|December 13, 2018
PubMed

Insights

A new multiplex quantitative PCR (qPCR) assay was developed for rapid detection of multidrug-resistant Candida species, including Candida auris, in clinical samples. This assay offers high specificity and sensitivity for early identification and improved patient outcomes in healthcare settings.

Area of Science:

  • Medical Mycology
  • Molecular Diagnostics
  • Infectious Diseases

Background:

  • Candida auris and related species are multidrug-resistant opportunistic pathogens causing significant healthcare-associated infections globally.
  • Current diagnostic methods lack specificity and speed for timely detection of these yeasts in clinical settings.
  • There is a critical need for rapid and accurate diagnostic tools to combat the spread of these resistant fungi.

Purpose of the Study:

  • To develop and validate a novel multiplex quantitative PCR (qPCR) assay for the simultaneous detection of Candida auris, Candida haemulonii, Candida duobushaemulonii, and Candida pseudohaemulonii.
  • To assess the analytical specificity, sensitivity, and reproducibility of the developed multiplex qPCR assay.
  • To evaluate the assay's performance in detecting target species from spiked serum samples.

Main Methods:

  • Development of a multiplex qPCR assay targeting specific genes of Candida auris, C. haemulonii, C. duobushaemulonii, and C. pseudohaemulonii.
  • Analytical validation including specificity testing against non-target organisms and human DNA, and sensitivity testing using serial dilutions of target genomes.
  • Performance evaluation using serum samples spiked with known quantities of target Candida species DNA.

Main Results:

  • The multiplex qPCR assay demonstrated 100% analytical specificity against a panel of relevant microorganisms and human DNA.
  • The assay achieved high sensitivity, detecting as few as ten genomes of target species with excellent reproducibility (R² >0.99).
  • In spiked serum samples, the assay successfully detected up to ten genomes of C. auris and one genome of the closely related species (R² >0.98).

Conclusions:

  • The developed multiplex qPCR assay is highly specific and sensitive for the direct detection of Candida auris and closely related species in serum samples.
  • This assay holds significant potential for the rapid and accurate diagnosis of infections caused by these multidrug-resistant yeasts in clinical settings.
  • Further clinical validation studies are recommended to confirm the assay's utility in routine patient management.
Keywords:
bloodstream infectionmelting curve analysisqPCR

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