A simple multiplex PCR assay for rapid identification of common pathogenic dermatophytes

Background: The most common etiological agents of human dermatophytosis in various parts of world are Trichophyton rubrum, Trichophyton interdigitale, and Epidermophyton floccosum. The main aim of this study was designing and evaluating a simple and straight forward multiplex PCR assay for reliable identification/differentiation of these species in clinical isolates. Methods: The reliable sequences of several molecular targets of dermatophytes species were used to design a multiplex PCR for identification of common pathogenic dermatophytes. The isolates and clinical specimens examined in this study are included 7 standard strains of dermatophytes, 101 isolates of dermatophytes and non-dermatophyte molds/yeasts which had already been identified by sequencing or PCR-RFLP, and 155 clinical samples from patients suspected of cutaneous mycoses. Results: Species-specific primer pairs for T. rubrum and T. interdigitale/T. mentagrophytes were designed based on the sequence data of the translation elongation factor 1-alpha gene, and the primers for E. floccosum targeted the specific sequence of the ITS region. The multiplex PCR successfully detected T. rubrum, T. interdigitale/T. mentagrophytes and E. floccosum strains that were identified by sequencing or PCR-RFLP. However, the primer pairs selected for T. interdigitale/T. mentagrophytes, cross-reacted with T. tonsurans. In testing the PCR system directly for clinical samples, the proportion of positive multiplex PCR was higher than positive culture (68.1% vs. 55.4%, respectively). Conclusion: The multiplex assay could detect three common out of several causal agents of dermatophytosis i.e. T. rubrum, T. interdigitale and E. floccosum, therefore, by adding a pan-dermatophyte primers it can be used as a comprehensive detection/identification test