Single-Step PCR Using (GACA)₄ Primer: Utility for Rapid Identification of Dermatophyte Species and Strains

• Published in: Journal of Clinical Microbiology 2008-08, Vol.46 (8), p.2641-2645
• Main Authors: Shehata, Atef S, Mukherjee, Pranab K, Aboulatta, Hassan N, El Akhras, Atef I, Abbadi, Said H, Ghannoum, Mahmoud A
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Dermatomycoses - microbiology; DNA Primers - genetics; DNA, Fungal - genetics; DNA, Ribosomal Spacer - genetics; Egypt; Epidermophyton - classification; Epidermophyton - genetics; Epidermophyton - isolation & purification; Fundamental and applied biological sciences. Psychology; Human mycoses; Humans; Infectious diseases; Medical sciences; Microbiology; Microsporum - classification; Microsporum - genetics; Microsporum - isolation & purification; Mycology; Mycoses; Mycoses of the skin; Polymerase Chain Reaction - methods; Polymorphism, Restriction Fragment Length; Trichophyton - classification; Trichophyton - genetics; Trichophyton - isolation & purification
• ISSN: 0095-1137; 1098-660X
• DOI: 10.1128/JCM.00697-08

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.