须癣毛癣菌基因分型的研究

目的探讨ITS区域的探针与DNA印迹杂交法对须癣毛癣菌的基因分型,并分析基因型与来源地区的相关性。方法用溴化十六烷基三甲铵(CTAB)法提取DNA,以皮肤癣菌的特异性引物NS5[5-′AACTTAAAGGAATT-GACGGAAG-3′]与ITS4[5-′TCCTCCGCTTATTGATATGC-3′]为引物,以须癣毛癣菌的标准菌株为模板,用PCR法扩增rD-NA部分18S区、ITSⅠ区、5.8 s区和ITSⅡ区为探针,随机引物法将探针标记,EcoR1酶切基因组DNA,按ECL试剂盒标准流程将酶切的基因组DNA与探针杂交;根据杂交的不同带型而分型。结果所试35株须癣毛癣菌分为14型(A~N型),A,B,C,D四型占62.86%。南方株以A和C型为主,北方株以B和D型为主。结论用ITS区域探针与DNA印迹杂交法对须癣毛癣菌基因组分型敏感性强、分辨力较高;南北方须癣毛癣菌DNA分型存在一定差异;DNA分型对须癣毛癣菌病的流行病学调查和菌学的生物学研究具有重要价值。

DNA Typing of Trichophyton Mentagrophytes

Objective To explore the DNA typing of Trichophyton mentagrophytes by Southern blotting,analyze the relationship between genotype and different geographical areas.Methods Thirty five strains of T.Mentagrophytes(14fromDalian,3fromShenyang,5fromshanghai,5frombeijing,3fromnanjing,2fromhebei,2fromkunming,and1fromhangzhou)were studied.DNA was extracted using cetyltrimethyl ammonium bromide(CTAB).The polymorphisms were detected by hybridization of EcoR1-digested T.Mentagrophyes genomic DNA with a probe amplified from the small-subunit rDNA and adjacent internal transcribed spacer(ITS) regions.The probe was made using a pair of primers NS5[5′-AACTTAAAGGAATTGACGGAAG-3′]and ITS4[5′-TCCTCCGCTTATTGATATGC-3′] and was labeled by ECL.The band patterns obtained by Southern blotting were used as the basis of the genotyping of T.Mentagrophytes.Results Fifteen individual patterns(DNA type A to N) were recognized among 35 strains of T.mentagrophytes.Type A to D accounted for 62.86% of all strains.The DNA patterns of Southern strains were represented by type A and C,those of northern strains were represented by type B and D.Conclusion The DNA typing of T.Mentagrophytes using Southern blotting was sensitive,highly distinguishable.The DNA patterns of southern strains are obviously different from those of northern strains.