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 tool.


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 gel.


RESULTS
The banding profiles were observed about 8 h from initiating DNA extraction. Intraspecies polymorphism 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 electrophoresed profiles on gel.


CONCLUSIONS
PCR-RFLP analysis is a rapid and reliable tool for the identification of T. tonsurans.
Mochizuki
Takashi
T
Department of Dermatology, Kanazawa Medical University, Uchinada, Ishikawa 920-0293, Japan. mocizuki@kanazawa-med.ac.jp
Tanabe
Hiroshi
H
Kawasaki
Masako
M
Ishizaki
Hiroshi
H
Jackson
Colin J
CJ
D000293:Adolescent; D000368:Aged; D001483:Base Sequence / Q000235:genetics; D002648:Child; D004275:DNA, Ribosomal / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D015183:Restriction Mapping; D013997:Time Factors; D014249:Trichophyton / Q000235:genetics* / Q000302:isolation & purification*
D004275:DNA, Ribosomal