Genotyping of Candida albicans using length fragment and high-resolution melting analyses together with minisequencing of a polymorphic microsatellite locus.
Microsatellite length polymorphism (MLP) typing is a PCR-based method used for genotyping of the diploid yeast Candida albicans. However, MLP is subject to homoplasia which can hamper the accuracy of the results. We combined fragment length analysis, high-resolution DNA melting (HRM) analysis, and SNaPshot minisequencing after a single amplification of the CDC3 locus to study 95 epidemiologically independent C. albicans isolates. HRM analysis for a given electrophoretic group led to a maximum of three different curves due to the presence of a SNP upstream of the tandem repeat which could be characterized using the SNaPshot assay. The combination of the three methods had a discriminatory index of 0.88 in complete congruence with previous MLP typing (Mantel test R=0.99, P<10(-)(4)). HRM is a useful tool of adding resolving power to MLP genotyping in identifying SNPs.
Costa
Jean-Marc
JM
Groupe hospitalier Chenevier-Mondor, APHP, Laboratoire de Parasitologie-Mycologie, Créteil, France.
Garcia-Hermoso
Dea
D
Olivi
Martine
M
Cabaret
Odile
O
Farrugia
Cécile
C
Lecellier
Gaël
G
Dromer
Françoise
F
Bretagne
Stéphane
S
D002176:Candida albicans / Q000235:genetics* / Q000302:isolation & purification; D002177:Candidiasis / Q000382:microbiology*; D004271:DNA, Fungal / Q000032:analysis / Q000235:genetics; D014644:Genetic Variation; D016681:Genome, Fungal; D006801:Humans; D018895:Microsatellite Repeats; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D017422:Sequence Analysis, DNA
D004271:DNA, Fungal