基于mtLSU rRNA和ITS1-5.8SrRNA-ITS2基因序列的肺孢子虫分子系统发育学研究

发布时间：2018-04-17 10:12

本文选题：肺孢子虫 + 分子系统发育　；参考：《中国人民解放军军事医学科学院》2005年博士论文

【摘要】：目的:以mt LSU rRNA和ITS1-5.8SrRNA-ITS2两种基因为分子标记,构建系统发育树,探讨肺孢子虫与宿主的进化关系以及属的分类地位。方法:采用按动物实际体重定量皮下注射地塞米松法诱导实验动物(SD大鼠、ICR小鼠、长爪沙鼠、新西兰白兔、C57BL/6N小鼠和豚鼠)感染肺孢子虫。收集实验动物肺泡灌洗液、肺组织、临床疑似病例痰液样本。提取肺孢子虫总DNA。设计合成扩增2个目的基因PCR引物,检验其特异性和敏感性。扩增目的基因,产物直接或克隆测序,分析比较序列的同源性和进化距离。结合GenBank中相关基因序列构建分子系统发育树。结果:从建立的PCP动物模型及临床病例样本获得了肺孢子虫DNA。获得了4种宿主该虫mt LSU rRNA基因5个序列,其中人1和长爪沙鼠肺孢子虫的该基因序列未见报道。获得4种宿主该虫的ITS1-5.8SrRNA-ITS2基因4个序列,其中长爪沙鼠和新西兰白兔肺孢子虫的该基因序列未见报道。构建了肺孢子虫系统发育树。结论:按动物实际体重定量皮下注射地塞米松方法建立了低死亡率PCP动物模型。PCR检测mt LSU rRNA基因法对PCP检测可行。肺孢子虫具有遗传多样性和宿主特异性,与其宿主之间呈协同进化关系。长爪沙鼠和新西兰白兔肺孢子虫可能是该属中独立的“种”。肺孢子虫属归属于真菌界的子囊菌门。
[Abstract]:Objective: to construct phylogenetic tree by using mt LSU rRNA and ITS1-5.8SrRNA-ITS2 as molecular markers, and to investigate the evolutionary relationship between Pneumocystis and hosts and the taxonomic status of genera.Methods: Pneumocystis were induced by subcutaneous injection of dexamethasone (Dexamethasone) on the basis of actual animal weight in SD rats ICR mice C57BL / 6N mice and guinea pigs.The samples of alveolar lavage fluid, lung tissue and sputum of clinical suspected cases were collected.Extraction of total DNA from Pneumocystis pneumocystis.Two target gene PCR primers were designed and synthesized to test their specificity and sensitivity.The target gene was amplified, the product was sequenced directly or cloned, and the homology and evolutionary distance of the sequence were analyzed and compared.Molecular phylogenetic tree was constructed by combining the sequence of related genes in GenBank.Results: Pneumocystis pneumocystis DNA was obtained from PCP animal model and clinical samples.Five sequences of Mt LSU rRNA gene were obtained from four species of host, of which human 1 and Pneumocystis longhaurus were not reported.Four sequences of ITS1-5.8SrRNA-ITS2 gene were obtained from four host species of Pneumocystis chinensis, including gerbils of gerbils and Pneumocystis rabbits of New Zealand, which were not reported.The phylogenetic tree of Pneumocystis was constructed.Conclusion: the animal model of low mortality PCP was established by quantitative subcutaneous injection of dexamethasone according to the actual body weight of animals. The detection of mt LSU rRNA gene by PCR was feasible for PCP detection.Pneumocystis have genetic diversity and host specificity, and there is a coevolutionary relationship between Pneumocystis and their hosts.The gerbils and Pneumocystis of New Zealand rabbits may be independent "species" of the genus.Pneumocystis of the genus Pneumocystis belonging to the Ascomystis of the Fungi.
【学位授予单位】：中国人民解放军军事医学科学院
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R384

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