中国幽门螺杆菌基因组特征及种群结构研究

发布时间：2018-06-09 21:02

本文选题：幽门螺杆菌 + 比较基因组　；参考：《中国疾病预防控制中心》2017年博士论文

【摘要】：幽门螺杆菌(Helicobacter pylori,缩写为H.pylori或HP)是一种定植于人体胃部的病原菌,可引起慢性胃炎、消化性溃疡,并与胃癌发生密切相关。人体一旦感染HP如不经规范治疗可终生带菌,因此HP常被作为一种可靠的生物标识来推测人类种群结构与历史迁徙。据推算,HP已伴随人类走出非洲向全球各地迁移、共进化了六万年以上,逐步形成了与人类种群相对应的七个群。以往的HP种群结构研究主要是基于7个管家基因序列多态性,不能准确反映我国HP的种群结构。本研究通过整合国际公共数据及对我国八个地区分离自不同消化道疾病、不同民族菌株的基因组测序,建立了包括491株全球HP的分层基因组数据集,其中包括107株东亚HP、65株中国HP。分析了中国HP的基因组特征及其在东亚、全球范围内的基因交流情况,以及中国HP种群中新发现的前噬菌体基因组特点。利用三代测序技术针对胃内多重感染相关的7个克隆构建了基因组完成图,并评价了三代与二代测序数据对于噬菌体预测的准确性。从群体和个体角度分析了 HP前噬菌体与宿主的共进化规律。第一部分:中国幽门螺杆菌比较基因组分析65株中国HP共鉴定出3072个泛基因,其中核心基因1147个,附属基因1925个,菌株特异基因0～47个。拼接序列经reads校正获得237250个核心基因组SNPs。不同菌株间显示出高度的遗传异质性,发现两个DNA转移相关Ⅳ型分泌系统是塑造中国菌株遗传多样性的主要因素。从85个毒力基因中发现76个序列较保守,主要编码尿素酶、鞭毛蛋白、Cag致病岛及VacA毒素。6个外膜蛋白编码基因存在较大变异,其中sabA、sabB将中国菌株分成了 2个距离较远的进化支,表明在中国不同人群中HP通过不同的表面抗原识别模式逃避宿主免疫。分层分析初步发现在整个种群水平上HP具有高度开放的基因库,鉴定出697个核心基因,为进一步筛选种属鉴定相关特异基因奠定基础。中国HP的序列多样性不能直接预测宿主疾病结局,反映了疾病发生过程中宿主免疫、胃内微环境、宿主外环境等多种其他因素与HP交互作用的复杂性。第二部分:中国幽门螺杆菌种群结构分析利用群体结构分析软件Chromopainter、fineSTRUCTURE和系统发育树,在中国HP中初步发现地域聚集性特征。在云南少数民族菌株中发现民族聚集性特点。从东亚亚群(hspEAsia)中鉴定出16个亚组,其中10个亚组为中国HP。中、日、韩、马来西亚华裔HP各亚组间基因交流频繁,遗传关系紧密,与其它亚群基因交流较少;在全球HP层面上的基因流分析发现东亚亚群、美洲印第安亚群、南印度群及欧洲群各自具有相对独立的基因库。这些地域特异性的亚组可能经历遗传漂变和瓶颈效应。根据HP与宿主平行共进化的特点,推测中国HP种群结构的形成可能与近代历史上的人群迁徙有关。对中国HP群体结构的初步鉴定使得未来开展中国HP与胃癌的GWAS研究中能够从地域、民族角度准确界定同质人群,扣除人群遗传背景的干扰,具有重要公共卫生意义。第三部分:中国幽门螺杆菌前噬菌体遗传多样性分析HP噬菌体研究在国际上报道很少,对其遗传特点及在HP中的作用认识十分有限。本研究首次在中国HP基因组中发现完整的前噬菌体YN4-84P,与最近日本分离的溶源性噬菌体KHP30基因组高度相似,但编码门户蛋白的基因被iS605插入,可能影响其生物合成。中国HP、全球整个种群HP的噬菌体携带率分别为13.07%和8.2%。噬菌体可能在塑造HP种群多样性方面发挥一定作用,且参与HP与人类的共进化。发现前噬菌体中携带黏附定植相关外膜蛋白基因,未发现毒素基因和耐药基因。通过对一名慢性胃炎病人分离7个克隆的基因组完成图分析,发现噬菌体与HP基因组具有一致的分化模式。评价了基于二代和三代测序数据进行前噬菌体预测的准确性。
[Abstract]:Helicobacter pylori (Helicobacter pylori, abbreviated as H.pylori or HP) is a pathogenic bacteria colonized in the stomach of the human body, which can cause chronic gastritis, peptic ulcers, and is closely related to gastric cancer. Once the human body is infected with HP without standard treatment, it can be used for life, so HP is often used as a reliable biomarker to speculate on the human population. It is reckon that HP has been associated with the migration of human beings out of Africa to all over the world, and has evolved over sixty thousand years and has gradually formed seven groups corresponding to the human population. The previous study on the structure of HP population is based on the polymorphism of 7 housekeeping gene sequences, which can not accurately reflect the population structure of our country's HP. The international public data and the genome sequencing of eight regions of our country, which are isolated from different digestive tract diseases and different ethnic strains, have set up a stratified genome data set including 491 global HP, including 107 East Asian HP and 65 Chinese HP. to analyze the genomic characteristics of Chinese HP and its genetic exchange in East Asia and around the world. The characteristics of the newly discovered phage genome in the HP population of China. The genome completion map was constructed by three generation sequencing technology for 7 clones associated with intragastric multiple infection, and the accuracy of the three and two generation sequencing data for phage prediction was evaluated. The co evolution of the pre HP phage with the host from the group and the individual angles was analyzed. The first part: 65 Chinese HP strains of Helicobacter pylori were analyzed. 3072 pan genes were identified, including 1147 core genes, 1925 accessory genes and 0~47 strain specific genes. The splice sequence was corrected by reads to obtain the high genetic heterogeneity among the different strains of SNPs., and found two The DNA transfer related type IV secretory system was the main factor in the genetic diversity of the Chinese strain. 76 of the 85 virulence genes were found to be more conservative. The main coding genes encoding urease, flagellin, Cag pathogenicity island and VacA toxin.6 outer membrane protein were large variation, and sabA and sabB divided the Chinese strain into 2 distances. The distant evolutionary branch indicates that HP is escaping from host immunity through different surface antigen recognition patterns in different populations of China. Stratification analysis has preliminarily found that HP has a highly open gene pool at the whole population level, and 697 core genes are identified, which lays the foundation for further screening of specific genes for identification of the species. The sequence of HP in China is diverse. Sex does not directly predict the outcome of the host disease, reflecting the complexity of host immunity, microenvironment in the stomach, and the external environment of the host in the course of the disease. The second part: the structure analysis of Helicobacter pylori in China, Chromopainter, fineSTRUCTURE and phylogenetic tree, are used in the analysis of the structure of Helicobacter pylori strains in China. The characteristics of regional aggregation were found in the national HP. 16 subgroups were identified in the ethnic minority strains in Yunnan. From the East Asian subgroup (hspEAsia), 16 subgroups were identified, of which 10 subgroups were in the Chinese HP., Japan, Korea, and Malaysia Chinese Asian HP subgroups with frequent genetic exchanges, and less genetic communication with other subgroups; Genetic flow analysis at the global HP level shows that East Asia subsets, Indian subgroups, southern India groups and European groups have relatively independent gene banks. These regional specific subgroups may experience genetic drift and bottleneck effects. According to the characteristics of parallel coevolution between HP and host, it is presumed that the formation of Chinese HP population structure may be in modern times. The preliminary identification of the population migration in history. The preliminary identification of the HP population structure in China makes it possible to define the homogenous population from the region and the national angle in the GWAS study of HP and gastric cancer in the future, and to deduct the interference of the genetic background of the population. Third parts: the genetic diversity of the phage before Helicobacter pylori in China. HP phage analysis is rarely reported in the world and has limited understanding of its genetic characteristics and its role in HP. This study was the first to discover the complete pre phage YN4-84P in the Chinese HP genome, which is highly similar to that of the soluble phage KHP30 genome isolated from recent Japan, but the gene encoding the portal protein is inserted by iS605 and may be inserted. The bacteriophages of HP in the whole population of China HP, 13.07% and 8.2%. phages, may play a role in shaping the diversity of HP population, and participate in the co evolution of HP and human. The former phage found that the adhesion and colonization related epicin gene was carried in the pre phage, and the toxin gene and the drug resistance gene were not found. The genome completion map analysis of 7 clones isolated from a chronic gastritis patient showed that the phage has a consistent pattern of differentiation with the HP genome. The accuracy of pre phage prediction based on the two and three generation sequencing data was evaluated.
【学位授予单位】：中国疾病预防控制中心
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R378

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