Virulence genes Haemophilus influenzae

描述(申请人提供)：流感嗜血杆菌是一种 人类限制的细菌，可以作为上呼吸道共生菌存在， 但也会引起局部性和/或系统性疾病。超过95%的人 鼻咽分离株缺乏包膜，不能分型，并引起局灶性 感染，虽然包膜(和罕见的非分型)菌株会导致败血症和 脑膜炎。共生菌和病原菌的基因组均为250kb(On)。 平均)比无毒的实验室毒株(RD KW2O)大，其基因组 《序列》于1995年发表。我们已经发现RD KW2O菌株的基因组 是一个“脚手架”，上面有基因、基因簇和操纵子编码毒力 因子被插入，通常是在嗜血杆菌特异的DNA摄取部位。 使用差异杂交，我们将识别那些允许 通过共生体的鼻咽定植，以及其他推定的 病原菌的毒力基因座。每一个DNA片段和 侧翼DNA，将进行测序，直到RDKW2O支架被确定。全 序列信息将在密歇根大学基因组中心的网站上公布。vbl.使用 同源性(非常规标准)推定毒力的严格标准 基因将被删除或突变，突变株的毒力 在相关的体外和/或体内感染模型中与父母进行比较。 例如，假设粘附素存在于下呼吸道分离物中 可以用人类呼吸道上皮细胞株和原代细胞进行测试 组织中生长在气液界面上的人呼吸道上皮 文化。一种独特的分离株，一种导致脑膜炎的无法分型的菌株 接种Hib结合疫苗的具有免疫能力的儿童似乎是一种 一个支系的成员；我们希望确认初步发现并启动 全球数据库(与MLST嗜血杆菌中心合作) 疫苗失败的分离株。在了解其分子机制的基础上 殖民与感染，预防和干预战略可以 是被设计出来的。

DESCRIPTION (provided by applicant): Haemophilus influenzae is a human-restricted bacterium which can exist as an upper respiratory commensal, but also cause focal and/or systemic disease. More than 95 percent of the nasopharyngeal isolates lack capsules and are not serotypeable, and cause focal infections, while encapsulated (and rare nontypeable) strains cause sepsis and meningitis. The genome of both commensal and pathogenic isolates is 250 kb (on average) larger than the avirulent laboratory strain (Rd KW2O) whose genome sequence was published in 1995. We have found that the genome of strain Rd KW2O is a "scaffold" on which genes, gene clusters and operons encoding virulence factors are inserted, usually at the Haemophilus-specific DNA uptake sites. Using differential hybridization we will identify those DNA loci that permit nasopharyngeal colonization by commensals, and the additional putative virulence loci in pathogenic isolates. Each of these DNA fragments and the flanking DNA, will be sequenced until the Rd KW2O scaffold is identified. All sequence information will be posted on the UW Genome Center web site. Using strict criteria for homology (not conventional criteria) putative virulence genes will either be deleted or mutated and the virulence of the mutant compared to the parent in a relevant in vitro and/or in vivo infection model. For example a putative adhesin present in a lower respiratory tract isolate could be tested with human respiratory epithelial cell lines, and with primary human respiratory epithelium growing at an air-liquid interface in tissue culture. One unique isolate, an untypeable strain causing meningitis in an immunocompetent child immunized with a Hib-conjugate vaccine appears to be a member of a clade; we wish to confirm the preliminary finding and initiate a global database (in collaboration with the MLST Haemophilus Center) for these vaccine-failure isolates. With an understanding of the molecular mechanisms of colonization versus infection, strategies for prevention and intervention can be devised.