Strain Differentiation of Trichophyton rubrum Using Polymorphism of rDNA Intergenic Spacer

Choi, Jong Soo; Shin, Dong Hoon; Seung, Jun Je; Kim, Ki Hong

Abstract

BACKGROUND
In Korea, Trichophyton (T.) rubrum is occupied more than 80% of causative fungi of dermatophytosis. Strain differentiation of T. rubrum is essential for epidermiologic study. OBJECTIVE: The aim of this study is to develope the primers for amplification of rDNA intergenic spacer (IGS) to detect polymorphism of T. rubrum. METHODS: The primers were designed from 25S and 18S of rDNA, and were applied to 20 strains of T. rubrum, which included 1 standard strain and 19 clinical isolates. RESULTS: Primers for amplification of polymorphic rDNA IGS were designed from the 3'end of the 25S (primer ANID25-3, 5'-GACAGGTTAGTTTTACCCTACTGA-3') and the 5'end of the 18S (TR18-2R, 5'- ATCTAATAAATACACCCCTTCCGA-3'). PCR condition was adjusted for detecting polymorphism. Best results were obtained at 55degrees C for annealing temperature and 3 minutes for extension time. Eight bands sized as 1.1, 2.4, 2.7, 2.9, 3.2, 3.8, 5 kb were amplified with PCR using the primers. With 4 bands sized 2.7, 2.9, 3.2 and 3.8 kb, 20 strains of T. rubrum could be grouped into 6 subtypes. CONCLUSION: The PCR fingerprinting with the primers for rDNA IGS was able to differentiated strains of T. rubrum, and it can be applied in clinical and epidemiologic studies. The primer could be applied to other fungi with unknown sequences.