TY - JOUR
T1 - Development and Analysis of qPCR for the Identification of Arthroconidial Yeasts of the Genus Magnusiomyces
AU - Kaplan, Engin
AU - Aktaş, Deniz
AU - Döğen, Aylin
AU - Hilmioğlu-Polat, Süleyha
AU - Gümral, Ramazan
AU - Hagen, Ferry
AU - Ilkit, Macit
AU - de Hoog, G. Sybren
N1 - Publisher Copyright:
© 2021, The Author(s), under exclusive licence to Springer Nature B.V. part of Springer Nature.
PY - 2021/3
Y1 - 2021/3
N2 - The arthroconidial yeasts Magnusiomyces capitatus and M. clavatus are emerging opportunistic pulmonary pathogens. They are closely related and difficult to distinguish based on morphological and physiological traits. We applied an SYBR® green-based quantitative PCR (qPCR) assay to identify the species. We analyzed 30 reference strains originating from clinical and environmental sources by targeting the Rpb2 gene encoding the second largest subunit of RNA polymerase II. The qPCR assays were tested by direct identification of M. capitatus and M. clavatus in spiked sputum and household dishwasher swabs, respectively, as models for clinical and environmental samples. The assays were proved to be reliable for species-level identification of both species, with 100% sensitivity and 100% specificity, lowest inter-assay deviations (RSDr ≤ 1.65%, R2 values >0.99), detection limit of 10 theoretical copy number of target DNA, and detection cell limit of ≥5000 yeast cells from spiked sputum samples. The developed qPCR assay is a practical molecular approach for the detection of M. capitatus and M. clavatus that can be used as a stand-alone assay or in conjunction with culture-dependent approaches.
AB - The arthroconidial yeasts Magnusiomyces capitatus and M. clavatus are emerging opportunistic pulmonary pathogens. They are closely related and difficult to distinguish based on morphological and physiological traits. We applied an SYBR® green-based quantitative PCR (qPCR) assay to identify the species. We analyzed 30 reference strains originating from clinical and environmental sources by targeting the Rpb2 gene encoding the second largest subunit of RNA polymerase II. The qPCR assays were tested by direct identification of M. capitatus and M. clavatus in spiked sputum and household dishwasher swabs, respectively, as models for clinical and environmental samples. The assays were proved to be reliable for species-level identification of both species, with 100% sensitivity and 100% specificity, lowest inter-assay deviations (RSDr ≤ 1.65%, R2 values >0.99), detection limit of 10 theoretical copy number of target DNA, and detection cell limit of ≥5000 yeast cells from spiked sputum samples. The developed qPCR assay is a practical molecular approach for the detection of M. capitatus and M. clavatus that can be used as a stand-alone assay or in conjunction with culture-dependent approaches.
KW - Arthroconidial yeasts
KW - Molecular diagnostics
KW - Quantitative real-time PCR
UR - https://www.scopus.com/pages/publications/85098554606
U2 - 10.1007/s11046-020-00510-4
DO - 10.1007/s11046-020-00510-4
M3 - Article
C2 - 33392857
AN - SCOPUS:85098554606
SN - 0301-486X
VL - 186
SP - 41
EP - 51
JO - Mycopathologia
JF - Mycopathologia
IS - 1
ER -