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

基于核和线粒体核糖体 DNA 区域多态性的皮肤癣菌鉴定

基本信息

批准号：
10670811
负责人：
MOCHIZUKI Takashi
金额：
$ 1.79万
依托单位：
Kanazawa Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10670811/
关键词：
Dermatophytes Identification Molecular technique ribosomal DNA ITS リボゾームDNA

项目摘要

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

本研究旨在评估分子技术鉴定从人类病变中分离出的皮肤癣菌的功效。研究中使用的技术是线粒体 (mt) DNA 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 间隔区的 RFLP 区域，以及基因组 DNA 中 rDNA 内转录间隔区 (ITS) 的序列分析。使用 rDNA 区域 PCR 扩增子的 RFLP mtDNA，用限制性酶 HinA III 消化产生的图谱显示 T. rubrum 和 T. mentagrophytes 之间的差异，但当扩增子用 Hae III 和 Msp I 消化时，这两个物种之间的 RFLP 图谱没有差异。可以用来区分红色毛癣菌和须毛癣菌，但该技术获得的信息相当少。对基因组DNA中rDNA的ITS1进行序列分析有很多优点。此外，一旦确定了序列，数据就可以在其他地方使用，并参考存放在基因库中的数据，但该方法的费用是实际应用的缺点。对须癣菌复合体、红色须癣菌和絮状桉中的几个物种进行了 ITS 区域的 PCR-RFLP 分析。通过 Mva I 或 I 消化了 ITS 1+2 区域（约 700 bp）。提示：我指出这些谱是物种特异性的，而且该技术便宜且节省时间，因此该方法也适合同时处理多个分离株。总之，我建议在实践中使用Mva I或Hinf I对ITS 1+2区域进行PCR-RFLP分析来鉴定皮肤癣菌，如果结果不清楚或不可接受，则确定ITS 1的核苷酸序列，并进行分析。参考保存在基因库中的数据较少。