微流芯片鉴定并分析马拉色菌属菌种基因型的初步研究

发布时间：2018-04-08 14:41

本文选题：马拉色菌属　切入点：微流芯片　出处：《复旦大学》2012年硕士论文

【摘要】：第一部分：马拉色菌属的分类及其在常见皮肤病的分布情况目的：研究马拉色菌属的分类及其在二种常见马拉色菌相关疾病中的分布情况。方法：以标准株作对照,采用PCR扩增rDNAITS2区并测序的分子生物学方法,将来源于花斑糠疹和马拉色菌毛囊炎的马拉色菌进行分类鉴定,分析各菌种在这二种疾病的分布情况。结果：从72例花斑糠疹和11例马拉色菌毛囊炎患者皮损处分离到7个菌种,共83株马拉色菌。鉴定出合轴马拉色菌41株(49.40%)、糠秕马拉色菌26株(31.33%)、球形马拉色菌10株(12.05%)、日本马拉色菌2株(2.41%)、限制马拉色菌1株(1.20%)、大和马拉色菌1株(1.20%)、皮炎马拉色菌1株(1.20%)及合轴与糠秕的混合感染1株(1.20%)。二种疾病菌种构成与疾病类型无明显关系,近半数花斑糠疹患者皮损中分离出合轴马拉色菌,而马拉色菌毛囊炎患者皮损中分离出的菌种无显著差异。结论：不同疾病马拉色菌菌种构成可能不同,合轴马拉色菌与花斑糠疹密切相关；马拉色菌rDNA的ITS2区PCR扩增与测序可用于马拉色菌的分类鉴定。第二部分：利用微流芯片鉴定并分析马拉色菌属菌种基因型目的：探讨微流芯片在马拉色菌鉴定与分型中的应用优势。方法：采用DNA测序的方法,将临床收集的菌株鉴定至种；选取临床最常见的糠秕、合轴和球形马拉色菌等7种马拉色菌分别进行RAPD-PCR扩增,将PCR产物进行微流芯片的菌种基因型鉴定。结果：大多数菌株均可被2种随机引物(S22、S24)扩增而获得清晰条带,但以S22引物扩增的条带更为稳定、清晰,作为本研究主要引物。不同种马拉色菌通过微流芯片基因型定量分析得到不同大小的固定阳性条带,所有菌株均可见种间和种内多态性。使用RAPD结合微流芯片方法,基本可将8种马拉色菌(糠秕、合轴、球形、厚皮、斯洛菲、日本、大和及皮肤马拉色菌)区分开来。结论：RAPD结合微流芯片方法作为一种快速、高通量、高灵敏性的分析技术,在马拉色菌种间菌株遗传多样性、亲缘关系的分析及新种鉴定中显示出一定优越性,适合用于流行病学分析。
[Abstract]:Part I: classification of Malassezia and its distribution in common skin diseasesObjective: to study the classification of Malassezia and its distribution in two common malassezia related diseases.Methods: Malassezia derived from pityriasis florescens and Malassezia folliculitis were classified and identified by using PCR amplification of rDNAITS2 region and sequencing, and the distribution of each strain in the two diseases was analyzed.Results: totally 83 strains of Malassezia were isolated from the lesions of 72 cases of furfuria macularis and 11 cases of Malassezia folliculitis.The mixed infection of chaff was 1. 20%.There was no significant relationship between the composition of the two diseases and the type of the disease. Malassezia was isolated from the skin lesions of nearly half of the patients with furfuriasis, but there was no significant difference between the two kinds of bacteria isolated from the skin lesions of the patients with Malassezia folliculitis.Conclusion: the species composition of Malassezia may be different, and Malassezia is closely related to pityriasis flowered, and PCR amplification and sequencing of the ITS2 region of Malassezia rDNA can be used for classification and identification of Malassezia.Part two: identification and analysis of Malassezia genotypes by microfluidic microarrayObjective: to investigate the advantages of microfluidic microarray in the identification and typing of Malassezia.Methods: by DNA sequencing, the collected strains were identified into species, and 7 strains of Malassezia malassezia, including chaff, axon and malassezia, were selected for RAPD-PCR amplification.The PCR products were identified by microfluidic microarray genotyping.Results: most strains could be amplified by two random primers (S22 S24), but the bands amplified with S22 primers were more stable and clear, so they were the main primers in this study.Fixed positive bands of different sizes were obtained by microfluidic microarray genotyping in different stallions, and interspecific and intraspecific polymorphisms were observed in all strains.Using RAPD and microfluidic chip method, eight species of Malassezia (chaff, axon, spherical, thick skin, Slofi, Japan, and Malassezia grandis) can be basically distinguished.Suitable for epidemiological analysis.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R756

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