Application of PCR-RFLP to Rapid Identification of the Main Pathogenic Dermatophytes from Clinical Specimens
Message:
Abstract:
Background
In the present study, a PCR-RFLP based molecular technique was designed to rapid identification of der matophytes in clinical specimens. Skin scrapings obtained from human cases suspected to dermatophytosis were studied in or der to identify involved etiological fungi.
Methods
In this experimental study, the specimens (skin scrapings) of patients referred to Mycology Department of Pas teur Institute of Iran were inoculated on Petri dishes contained selective agar for pathogenic fungi (SAPF) and incubated at 25º C until visible growth of fungal colonies. The colonies were examined for standard morphological characteristics after visi ble growth on the agar medium. A small portion of each fungal colony was further studied by restriction fragment length poly morphism (RFLP) analysis of the PCR-amplified internal transcribed spacer (ITS) region of ribosomal DNA (rDNA). PCR amplicons were electrophoresed on 2% agarose gel after digesting by different restriction enzymes including MvaI, HinfI and HaeIII.
Results
Among 160 clinical samples examined, 6 dermatophyte species including Trichophyton mentagrophytes, T. ru brum, T. verrucosum, T. tonsurans, Microsporum canis and Epidermophyton floccosum were finally identified based on the col ony morphology and microscopic criteria. Specific PCR products and RFLP patterns for MvaI, HinfI and HaeIII en zymes allowed the rapid identification and reliable differentiation of isolated dermatophytes at the genus or species level for 5-10 day-old colonies.
Conclusions
The results showed that PCR-RFLP analysis of the ITS region of rDNA is a rapid and reliable tool which al lows identification of major pathogenic dermatophytes isolated in this study at species level in young 5-10 day-old colonies.
Language:
English
Published:
Iranian Journal of Public Health, Volume:38 Issue: 1, Spring 2009
Page:
18
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