Rapid identification of Trichophyton tonsurans by PCR-RFLP analysis of ribosomal DNA regions

Background: Culture morphology of Trichophyton (T) tonsurans, an emerging pathogen of dermatophytosis in Japan, varies widely and species level. identification is sometimes very difficult. Reliable molecular markers are expected to be introduced for their identification. Objective: The present study was conducted to evaluate the efficacy of restriction fragment length polymorphism (RFLP) analysis of PCR amplified ribosomal (r) DNA including internal transcribed spacers (ITS), as an identification toot. Methods: Total cellular DNA was extracted from 26 Japanese isolates of T tonsurans, along with several taxa, of the members in the T mentagrophytes complex, T rubrum, T violaceum and Epidermophyton floccosum, using a mini-preparation method. PCR amplicons were digested with restriction enzymes Mva I or Hinf I, then electrophoresed on 5% polyacrylamide get. Results: The banding profiles were observed about 8 h from initiating DNA extraction. Intraspecies potymorphism was not detected among T tonsurans isolates, and their profiles obtained using Mva I digestion were clearly different from those of the other dermatophyte species. The restriction profiles evaluated from nucleotide sequence of the regions by a computer analysis were compatible with the etectrophoresed profiles on get. Conclusion: PCR-RFLP analysis is a rapid and reliable toot for the identification of T tonsurans. (C) 2003 Japanese Society for Investigative Dermatology. Published by Elsevier Science Ireland Ltd. All rights reserved.