Padronização de uma PCR-RFLP e comparação com métodos tradicionais para identificação das principais espécies de dermatófitos em amostras clínicas de cães e gatos

Abstract

The Objective of this study was to standardize a PCR-RFLP and compare it with traditional methods of identification of dermatophytes in clinical specimens from dogs and cats. We analyzed 150 clinical samples from animals with skin diseases. In samples of hair and / or crusts direct microscopic exams were performed with KOH (30%) as well as culture on Sabouraud dextrose agar supplemented with yeast extract, chloramphenicol and cycloheximide. DNA was extracted from the colonies, after the evaluation of macroscopic and microscopic features as well as of clinical specimens with the DNeasy Blood & Tissue extraction kit (QIAGEN ®). The primers used were ITS 1 and ITS 4. The amplicons were digested by restriction enzyme BseBI. To study the correlation between the tests we used the Kappa coefficient (K). After analyzing the results we observed patterns different that identify Microsporum canis, Microsporum gypseum and Trichophyton mentagrophytes. There was also excellent agreement between the results of culture and PCR-RFLP of 87,5% (7/8). Another observation made was that negative samples in traditional methods (direct examination and culture), but were contaminated with the agent, were identified by molecular technique, indicating a higher sensitivity of the same. It concluded, then, that the PCR-RFLP method is more sensitive and as specific as the routine methods, allowing it to be used for a fastest identification of these species of dermatophytes in clinical samples from dogs and cats.