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Identification of Candida auris and related species by multiplex PCR based on unique GPI protein‐encoding genes
Author(s) -
Alvarado María,
Bartolomé Álvarez Joaquín,
Lockhart Shawn R.,
Valentín Eulogio,
RuizGaitán Alba Cecilia,
Eraso Elena,
Groot Piet W. J.
Publication year - 2021
Publication title -
mycoses
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.13
H-Index - 69
eISSN - 1439-0507
pISSN - 0933-7407
DOI - 10.1111/myc.13204
Subject(s) - candida auris , amplicon , biology , multiplex polymerase chain reaction , gene , polymerase chain reaction , multiplex , genetics , identification (biology) , fungal pathogen , microbiology and biotechnology , computational biology , antifungal , botany
Background The pathogen Candida auris is rapidly gaining clinical importance because of its resistance to antifungal treatments and its persistence in hospital environments. Early and accurate diagnosis of C. auris infections is crucial, and however, the fungus has often been misidentified by commercial systems. Objectives To develop conventional and real‐time PCR methods for accurate and rapid identification of C. auris and its discrimination from closely related species by exploiting the uniqueness of certain glycosylphosphatidylinositol (GPI)‐modified protein‐encoding genes. Methods Species‐specific primers for two unique putative GPI protein‐encoding genes per species were designed for C. auris , C. haemulonii, C. pseudohaemulonii, C. duobushaemulonii , C. lusitaniae and C. albicans . Primers were blind tested for their specificity and efficiency in conventional and real‐time multiplex PCR set‐up. Results All primers combinations showed excellent species specificity. In multiplex mode, correct identification was aided by different‐sized amplicons for each species. Efficiency of the C. auris primers was validated using a panel of 155 C. auris isolates, including all known genetically diverse clades. In real‐time multiplex PCR, different melting points of the amplicons allowed the distinction of C. auris from four related species. C. auris limit of detection was 5 CFU/reaction with a threshold value of 32. The method was also able to detect C. auris in spiked blood and serum. Conclusions PCR identification based on unique GPI protein‐encoding genes allows for accurate and rapid species identification of C. auris and related species without need for expensive equipment when applied in conventional PCR set‐up.