中华皮肤科杂志 ›› 2004, Vol. 37 ›› Issue (4): 203-205.

• 论著 • 上一篇    下一篇

种特异性引物鉴定申克孢子丝菌

王晓慧1, 冀朝辉2, 李若瑜1   

  1. 1. 北京大学第一医院皮肤科真菌和真菌病研究中心;
    2. 北京大学基础医学院生物化学与分子生物学系
  • 收稿日期:2003-04-26 出版日期:2004-04-15 发布日期:2004-04-15

Identification of Sporothrix schenckii Based on the Nucleotide Sequences in Internal Transcribed Spacer Region 2 of Ribosomal DNA Genes

WANG Xiao-hui1, JI Zhao-hui2, LI Ruo-yu1   

  1. Department of Dermatology, First Hospital of Peking University, Beijing 100034, China
  • Received:2003-04-26 Online:2004-04-15 Published:2004-04-15

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摘要： 目的 研究申克孢子丝菌的分子鉴定方法,为孢子丝菌感染的分子诊断奠定基础.方法 对22株申克孢子丝菌和12种12株暗色真菌临床分离株的核糖体DNA(rDNA)内转录间隔区(ITS)进行聚合酶链反应扩增.对10株来源于中国不同地区及1株来源于美国的申克孢子丝菌的扩增产物测序并进行分析,以获得的ITS2区序列为靶序列设计出申克孢子丝菌的种特异性引物(SSP),并用该引物扩增全部受试菌株.结果 序列分析显示,申克孢子丝菌rDNA的ITS2区相当保守,特异性引物对22株申克孢子丝菌可扩增出一条300bp的片段,其他受试菌株均为阴性.结论 应用设计出的种特异性引物,结合PCR方法,鉴定申克孢子丝菌特异、敏感、可靠,可用于临床诊断.

关键词: 孢子丝菌属, DNA引物, DNA,核糖体间隔区

Abstract: Objective To study the molecular identification method of Sporothrix schenckii based on the nucleotide sequences in internal transcribed spacer region 2 of ribosomal DNA (rDNA) genes. Methods Species-specific primers were used to amplify the internal transcribed spacer region 2 of rDNA of 22 clinical isolates of Sporothrix schenckii and 12 strains of dematiaceous fungi. Totally 11 strains of Sporothrix schenckii were sequenced and analyzed, in which 1 strain came from the US and the others were isolated from different areas in China. A pair of species-specific oligonucleotide primers (SSP) were designed based on the ITS2 sequence. With the species-specific primers, rDNA of Sporothrix schenckii and dematiaceous fungi were amplified by PCR. Results Sequencing and analysis showed that internal transcribed spacer region 2 of rDNA gene was conservative in Sporothrix schenckii. A 300 bp fragment was obtained from 22 strains of Sporothrix schenckii, but not from the other species. Conclusions This method is specific, sensitive and reliable for the identification of Sporothrix schenckii and could be used for clinical molecular diagnosis.

Key words: Sporothrix, DNA primers, DNA, ribosomal spacer