肺炎克雷伯菌Ⅵ型分泌系统的比较基因组学研究

发布时间：2018-05-29 00:54

  本文选题：肺炎克雷伯菌 + 细菌Ⅵ型分泌系统　； 参考：《上海交通大学》2015年博士论文

【摘要】：肺炎克雷伯菌(Klebsiella pneumoniae)是院内感染最重要的多重耐药条件致病菌之一。11个不同来源的肺炎克雷伯菌基因组已被全测序,包括耐碳青霉烯类抗生素的、属于ST11型中国流行克隆株的临床菌株HS11286。近期发现的Ⅵ分泌系统(type Ⅵ secretion system,T6SS)是一种细胞接触依赖性的多组件跨膜通道,在细菌致病机制及细菌间拮抗作用中扮演着重要角色。T6SS已被发现存在于铜绿假单胞菌和霍乱弧菌等多个革兰阴性细菌中;但是在肺炎克雷伯菌中鲜有研究。本论文推测T6SS与肺炎克雷伯菌的致病性、耐药性、自然生存和传播性等关键生物学特性相关,利用生物信息学方法对已测序肺炎克雷伯菌基因组的T6SS基因簇和分泌效应物进行了系统地识别和比较,以辅助实验研究来解析T6SS的生物学功能。本文首先预测了肺炎克雷伯菌的T6SS基因簇。通过文献挖掘和生物信息学预测,本文识别了498个细菌菌株中的906个T6SS基因簇数据,主要包括:(i)经过对708篇PubMed收录文献的检索和人工校验,收集的57个菌株中64个T6SS、92个T6SE和25个免疫蛋白及127个调控因子;(ii)基于T6SS保守组件氨基酸序列的高度相似性预测的T6SS。SecReT6根据T6SS核心组件的序列相似性和基因排布,将这些T6SS系统地分为3种类型:i,ii和iii。利用PostgreSQL关系型数据库管理系统,本文建立了开放数据库SecReT6。SecReT6整合了多种异源数据,提供了基于Web的T6SS核心组件、T6SE和免疫蛋白、调控因子等功能和分类信息的浏览和查询服务。SecReT6还提供了相似性搜索、基因簇比对和系统发生学分析等在线工具。此外,基于不同类型T6SS核心组件的隐马尔科夫模型特征谱,开发了T6SS基因簇识别工具T6SS-HMMER。利用T6SS-HMMER工具,本文在11个已全测序的肺炎克雷伯菌基因组中识别了26个T6SS。其中,在肺炎克雷伯菌HS11286中识别2个T6SS基因簇:编码了12个T6SS核心组件的T6SS-1(KPHS_22970..KPHS_23190),及编码9个T6SS组件的T6SS-2(KPHS_32450..KPHS_32770)。其次,本文采用了三种方法来预测肺炎克雷伯菌T6SS分泌效应物(T6SE):(i)SecReT6收录了和已知T6SE高序列相似性的蛋白;(ii)通过T6SS基因簇比较分析发现,插入T6SS保守组件基因中的可变区可能携带了新的T6SE和相邻的免疫蛋白;(iii)基于已知T6SE的多重序列特征,提出一个基于支持向量机分类器的算法来预测零散分布在基因组的新T6SE。本文在肺炎克雷伯菌HS11286中识别了T6SS-1基因簇编码的效应蛋白KPHS_23105和4个免疫蛋白KPHS_23060-KPHS_23090。最后,本文将T6SS保守基因簇的识别工作扩展到细菌III型和IV型等多组件分泌系统,以及原噬菌体、整合性接合元件、整合子、插入序列和基因组岛等可移动遗传元件的识别。这些分泌系统和可移动遗传元件常携带致病因子和获得性耐药基因,可视为病原细菌感染的风险基因。针对细菌基因组序列中这些致病和耐药相关风险基因的准确和快速识别需求,本文开发了基于高性能计算机群的在线预测服务STeP。STeP提供的VRprofile工具可在10分钟内完成肺炎克雷伯菌HS11286的5.3Mb染色体序列的分析任务;对预测出来的T3SS/T4SS/T6SS/T7SS和可移动遗传元件,VRprofile还提供了对后台数据库MobilomeDB收录已知元件的可视化比较分析服务。STeP服务器还提供了对完全或部分测序细菌基因组快速注释工具CDSeasy,细菌保守基因簇识别工具CGCfinder,COG基因集的可视化共定位工具COGviewer。总之,本论文以肺炎克雷伯菌为切入点,对已测序细菌基因组中T6SS及分泌效应物的分布进行了系统地考查。所构建的T6SS开放系统数据库和T6SE识别工具,有助于实验研究肺炎克雷伯菌及其他重要的革兰阴性病原细菌中T6SS与致病机制及细菌间拮抗作用之间的关系。细菌致病和耐药相关基因簇的生物信息学分析工具集将有助于从基因组序列水平快速预测病原细菌的潜在致病和耐药能力。
[Abstract]:Klebsiella pneumoniae (Klebsiella pneumoniae) is one of the most important multidrug resistant condition pathogenic bacteria in hospital infection. The genome of Klebsiella pneumoniae, one of the different sources of.11, has been fully sequenced, including the carbapenems, which belongs to the ST11 type Chinese popular clone strain HS11286. recently found in the VI secretory system (type VI). Secretion system, T6SS) is a cell contact dependent multicomponent transmembrane channel, which plays an important role in the pathogenic mechanism of bacteria and the antagonism of bacteria..T6SS has been found in many Gram-negative bacteria such as Pseudomonas aeruginosa and Vibrio cholerae, but there are few studies in Klebsiella pneumoniae. This paper speculated that T6SS It is related to the key biological characteristics of Klebsiella pneumoniae, such as the pathogenicity, resistance, natural survival and transmission, and systematically identifies and compares the T6SS gene cluster and secretory effector of the genome of Klebsiella pneumoniae by bioinformatics method, in order to assist the analysis of the biological function of T6SS. The T6SS gene cluster of Klebsiella pneumoniae was predicted. Through literature mining and bioinformatics prediction, this paper identified 906 T6SS gene cluster data of 498 bacterial strains, including: (I) after retrieving and manually checking 708 PubMed collections, 64 T6SS, 92 T6SE, 25 immunoglobulin and 127 of the collected strains were collected. Regulatory factors; (II) T6SS.SecReT6 based on the high similarity prediction based on the amino acid sequence of the T6SS conservative component, T6SS.SecReT6 is divided into 3 types based on the sequence similarity and gene arrangement of the T6SS core components: I, II and III. use PostgreSQL relational database management system, and the integration of the open database SecReT6.SecReT6 is established in this paper. A variety of heterogeneous data, providing Web based T6SS core components, T6SE and immunoglobulin, regulatory factors and other functions and classification information browsing and query services.SecReT6 also provide online tools such as similarity search, gene cluster comparison and phylogenetic analysis. In addition, hidden Markov model based on different types of T6SS core components The T6SS gene cluster identification tool, T6SS-HMMER., was developed using the T6SS-HMMER tool to identify 26 T6SS. in 11 fully sequencing Klebsiella pneumoniae genome, and 2 T6SS gene clusters were identified in Klebsiella pneumoniae HS11286: T6SS-1 (KPHS_22970..KPHS_23190) of 12 T6SS core components and 9 T6SS groups T6SS-2 (KPHS_32450..KPHS_32770). Secondly, three methods are used to predict the T6SS secretory effector of Klebsiella pneumoniae (T6SE): (I) SecReT6 includes a protein that is similar to the known high sequence of T6SE sequences; (II) it is found that the variable region inserted into the T6SS conservative component gene may carry a new T6SE and a new T6SE sequence. Adjacent immune proteins; (III) based on the multiple sequence characteristics of known T6SE, an algorithm based on the support vector machine classifier is proposed to predict the scattered distribution in the genome. The effect protein KPHS_23105 and the 4 immune protein KPHS_23060-KPHS_23090., which are encoded by the T6SS-1 gene cluster, are identified in the Klebsiella pneumoniae HS11286. This paper extends the identification of T6SS conserved gene clusters to multi component secretory systems such as bacterial III and IV, as well as the identification of removable genetic elements such as original phage, integrated conjugation elements, integrons, insertion sequences and genome islands. These secretory systems and removable relic elements often carry pathogenic factors and acquired resistance genes, In view of the accurate and rapid identification requirements of these pathogenic and drug-resistant risk genes in the bacterial genome sequence, the VRprofile tool provided by the online prediction service STeP.STeP based on high performance computer group can complete the 5.3Mb dyeing of Klebsiella pneumoniae HS11286 within 10 minutes. The analysis task of the sequence of color bodies; for the predicted T3SS/T4SS/T6SS/T7SS and removable genetic components, VRprofile also provides a visual comparative analysis service for a background database MobilomeDB that contains known components..STeP server also provides a rapid annotation tool for complete or partial sequencing of bacterial gene groups CDSeasy, a bacterial conservative gene cluster. The identification tool CGCfinder, the Visual Co location tool of the COG gene set COGviewer., in this paper, the distribution of the T6SS and the secretory effectors in the genome of the sequenced bacteria was systematically investigated with Klebsiella pneumoniae as the breakthrough point. The T6SS open system database and the T6SE identification tool were used to study the experimental study of pneumonia Klein. The relationship between T6SS and the pathogenic mechanism and the antagonism among the other important gram-negative pathogenic bacteria. The bioinformatics analysis tool set for bacterial and drug-resistant gene clusters will help to quickly predict the potential pathogenicity and drug resistance of pathogenic bacteria from the genome sequence level.
【学位授予单位】：上海交通大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R378
，

本文编号：1948867