Single-step PCR using (GACA)4 primer: utility for rapid identification of dermatophyte species and strains.
Dermatophytes are fungi that belong to three genera: Epidermophyton, Microsporum, and Trichophyton. Identification of dermatophyte species is essential for appropriate diagnosis and treatment of dermatophytosis. Routine identification depends on macroscopic and microscopic morphology, which is time-consuming and does not identify dermatophyte strains. In this study, two PCR-based methods were compared for their abilities to identify 21 dermatophyte isolates obtained from Egyptian patients to the species and strain levels. The first method employed a two-step method: PCR amplification, using ITS1 and ITS4 as primers, followed by restriction enzyme digestion using the endonuclease MvaI. The second method employed a one-step approach employing the repetitive oligonucleotide (GACA)(4) as a primer. Dermatophyte strains were also identified using a conventional culture method. Our results showed that the conventional culture method identified four species: Microsporum canis, Trichophyton mentagrophytes, Trichophyton rubrum, and Trichophyton violaceum. Moreover, both PCR methods agreed with the diagnosis made using the conventional approach. Furthermore, ITS1/ITS4-based PCR provided no strain differentiation, while (GACA)(4)-based PCR identified different varieties among the T. mentagrophytes isolates. Taken together, our results suggest that (GACA)(4)-based PCR has utility as a simple and rapid method for identification of dermatophyte species as well as utility for differentiation of T. mentagrophytes variants.
Shehata
Atef S
AS
Microbiology Department, Faculty of Medicine, Suez Canal University, Ismailia, Egypt.
Mukherjee
Pranab K
PK
Aboulatta
Hassan N
HN
el-Akhras
Atef I
AI
Abbadi
Said H
SH
Ghannoum
Mahmoud A
MA
D017931:DNA Primers / Q000235:genetics*; D004271:DNA, Fungal / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D003881:Dermatomycoses / Q000382:microbiology*; D004534:Egypt; D004821:Epidermophyton / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008864:Microsporum / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D014249:Trichophyton / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification
D017931:DNA Primers; D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer