Identification of Dermatophytes based on polymorphysms of nuclear and mitochondrial ribosomal DNA regions

本研究旨在评价分子技术对人体皮损分离的皮肤真菌的鉴定效果。采用线粒体DNA （mt）中核糖体(r) DNA区域的限制性片段长度多态性（RFLP）、基因组DNA中rDNA间隔区RFLP分析和基因组DNA中rDNA内转录间隔区（ITS）序列分析技术。利用mtDNA rDNA区PCR扩增片段的RFLP分析，用HinA III酶切得到的图谱显示红绒t和红绒t存在差异。用Hae III和Msp i酶切扩增子后，两种植物间的RFLP谱无明显差异，该技术可用于区分红藓和红藓，但获得的信息较少。基因组DNA中rDNA的ITS1序列分析具有许多优点。该方法重现性好，灵敏度高。此外，一旦序列确定，数据可以在其他地方使用，并参考存放在基因库中的数据。然而，这种方法的费用是在实际应用中的缺点。采用PCR-RFLP分析方法，对棕藓藓复合体、红藓藓和絮凝藓中的几种植物进行了ITS区分析。Mva I或Hint酶切ITS 1+2区（约700 bp）。我指出这些资料是针对特定物种的。灵敏度和重现性足以满足种级鉴定。该方法成本低，省时，适用于同时处理多个分离物。综上所述，我建议在实践中使用Mva I或Hinf I对ITS 1+2区域进行PCR-RFLP分析，用于皮肤真菌的鉴定。如果结果不明确或不可接受，则确定ITS 1的核苷酸序列，并参考存放在基因库中的数据。少

The present study was conducted to evaluate the efficacy of molecular techniques for the identification of dermatophytes isolated from human lesions. The techniques used in the study were, restriction fragment length polymorphisms (RFLP) of ribosomal (r) DNA regions in mitochondrial (mt) DNA, RFLP analysis of spacer regions of rDNA in genomic DNA, and sequence analysis of internal transcribed spacer (ITS) of rDNA in genomic DNA.Using RFLP of PCR amplicons of rDNA region of mtDNA, the profiles generated by digestion with restriction enzyme HinA III showed differences between T. rubrum and T. mentagrophytes. But there was no difference between the RFLP profiles between these two species when the amplicons were digested with Hae III and Msp I. The technique could be used for the differenciation between T. rubrum and T. mentagrophytes, but the information obtained by the technique was rather minimum.There were many advantages of sequence analysis of ITS1 of rDNA in genomic DNA. The method … More was highly reproducible and highly sensitive. Moreover, once the sequence is determined, the data could be used elsewhere and referred to data deposited in gene banks. The expense of the method was, however, the drawback in the application in practice.PCR-RFLP analysis of ITS regions were performed on several species in the T. mentagrophytes complex, T. rubrum and E. floccosum. Digestion of ITS 1+2 regions (ca. 700 bp) by Mva I or Hint. I indicated these profiles to be species specific. The sensitivity and reproducibility was enough for the species level identification. Also the technique was inexpensive and time saving, therefore, the method is also suitable for treatment of a number of isolate at the same time.In conclusion, I propose to use PCR-RFLP analysis of ITS 1+2 regions by Mva I or Hinf I for the identification of dermatophytes in practice. If the result is unclear or unacceptable, then determine the nucleotide sequence of ITS 1, and refer the data to those deposited in a genebanks. Less