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目的探讨ITS基因测序对须癣毛癣菌进行分子生物学鉴定的可行性,以确定致病菌株的种属。方法从病人头皮及头发分离出一株真菌,采用直接涂片镜检和培养的方法,观察其镜下形态和菌落特征,同时取纯培养物进行普通PCR,PCR产物经1%琼脂糖凝胶电泳分析,Tanon-3500凝胶成像系统进行鉴定,收集目的片段进行ITS基因测序。结果本菌的镜下形态和培养特征与其他毛癣菌属极其相似,难以直接鉴定分离菌为须癣毛癣菌,将其纯培养后用真菌通用引物ITS4、ITS5进行PCR扩增,所得的扩增片段大小约700 bp,将目的片段送至公司进行双向测序,测序结果经Blast比对,须癣毛癣菌与之相符率达99.5%。结论 ITS基因测序可以准确鉴定从临床分离的须癣毛癣菌。
Objective To investigate the feasibility of ITS gene sequencing for the identification of Trichophyton mentagrophytes molecular biology to identify the species of pathogenic strains. Methods A strain of fungus was isolated from the scalp and hair of the patient. Microscopic examination and culture were used to observe the microscopic morphology and colony characteristics. The normal culture was used for PCR. The PCR products were separated on 1% agarose gel Electrophoresis analysis, Tanon-3500 gel imaging system for identification, collection of the target fragment ITS gene sequencing. Results The microscopic morphology and culture characteristics of this bacterium were very similar to those of other Trichophyton species. It was difficult to directly identify the isolates as Trichophyton mentagrophytes. After pure culture, the fungi common primers ITS4 and ITS5 were amplified by PCR. The size of amplified fragment was about 700 bp. The target fragment was sent to the company for bi-directional sequencing. The results of Blast comparison showed that T. punctatus matched 99.5%. Conclusion ITS gene sequencing can accurately identify Trichophyton mentagrophytes isolated from clinic.