ITS-RFLP optimization for dermatophyte identification from clinical sources in Alagoas (Brazil) versus phenotypic methods

Abstract

Introduction: Dermatophytoses are superficial mycoses, and the identification of their etiological agents is routinely performed by culture and microscopic features, which is time-consuming and relies on personnel expertise. Molecular approaches have been developed to provide faster and reliable results; therefore, this study aimed to identify dermatophytes isolated from Alagoas state patients, employing phenotypical and molecular methods.
 Methodology: Clinical samples for morphological identification were collected from private and public laboratories and cultivated on Sabouraud dextrose agar. DNA extraction was followed by ITS amplicon analysis after restriction enzyme digestion DdeI (ITS-RFLP).
 Results: Out of fourteen representative strains, ITS-RFLP with DdeI efficiently identified Microsporum canis, Nannizzia gypsea, and Trichophyton rubrum, while species of the complex T. tonsurans/T. mentagrophytes presented the same restriction pattern. After genotyping, 2 T. tonsurans and 1 Microsporum sp. strain were reclassified as T. rubrum.
 Conclusions: RFLP of ITS-region followed by DdeI digestion produced faster and relatively reliable results than classic methods; however, this method has not been as efficient for closely related dermatophytes cryptic species.

Identification of Chromoblastomycosis and Phaeohyphomycosis Agents through ITS-RFLP

“…Recent studies have shown that this technique serves as a valuable tool for identifying a variety of pathogenic fungal species, such as Cryptococcus sp., Dermatophytes, Paracoccidioides sp., and Candida sp. [20,[29][30][31].…”

“…Furthermore, a recent study published by Fonseca in 2022 [30] highlighted the efficiency of the ITS region in a simple digestion RFLP assay for identifying species of dermatophytes of the genera Trichophyton sp. and Microsporum sp.…”

Diagnostic ability of Peptidase S8 gene in the Arthrodermataceae causing dermatophytoses: A metadata analysis

“…Molecular diagnosis for the dermatophytes is often by nucleic acid amplification techniques targeting the ITS and CHS1 regions [ 32 , 38 – 40 ]. Several modifications of PCR like multiplex techniques [ 41 – 43 ], nested PCR [ 44 ], PCR-ELISA [ 45 ], restriction digestion of the PCR products [ 46 , 47 ] or real time detection [ 48 , 49 ] for these targets are reported. The use of ITS and chitinase based molecular detection methods have proven valuable [ 50 ].…”