人腺病毒分型标准和重组进化机制研究

There are many kinds of adenovirus (Adv) including 5 genera, 43 species and 149 types. The host spectrum of Adv is broad. The host of Adv infection are as many as 22 kinds including human beings，domestic and wild animals and birds. There have been emergent 46 types categorized in 22 species since 2005. Human adenoviruses (HAdVs) are common pathogens that cause several diseases, such as pneumonia, acute gastroenteritis, and epidemic keratoconjunctivitis. HAdV infection is also associated with a serious adenovirus syndrome in immunocompromised patients. And some types of HAdV possess the potential of oncogenicity. Recombination is a well-known feature of AdV genetics. However, the recombination mechanism is not clear. There are some criterion for HAdV typing. But all of them were not perfect and convinent. The genomic or main functional gene sequences of all 68 types of HAdV will be download from GenBank and subjected bioinformatic analysis in this project. A rational and practical typing criteria will be proposed via the phylogenic analysis and the measurement of phylogenic distance et al. Nevertheless, all the recombinant strains will be screened and the common recombinant characteristics, recombinant condition, recombinant hotspot and the sequences will be analyzed to elucidate the function and the mechanism of HAdv recombination. Meanwhile, the clinical HAdv strains will be isolated, identified and sequenced from clinical sample. The identified strains will be typed using the new typing criteria to confirm the feasibility of the new criteria. And the strains preserved in the laboratory will subject induced mutation and to provide artificial recombination condition to confirm the adenovirus recombination theory.

腺病毒种类多，共有5个属，43个种，149个型；感染人、畜、禽和野生动物等22种宿主, 存在着跨物种传播的危险性；新的型别不断涌现，2005年以来，腺病毒增加了22个种，46个型。人腺病毒引起人类肺炎、肠炎和结膜炎等疾病，有些型别还具有潜在的致癌性。腺病毒重组现象普遍，但重组机制不明，分型标准不完善，易造成混乱。本课题拟对68个型别的人腺病毒基因组及各主要功能基因进行生物信息学分析，建立种系发生进化树，测出种系发生距离。在现有的分型方法的基础上提出人腺病毒合理、实用、可行的分型标准并找出所有发生重组变异的人腺病毒毒株，研究其重组变异的共同特点、条件、作用、重组热点及分子结构基础，阐明重组机制等。同时，对临床标本分离株进行鉴定和主要功能基因测序，利用本课题提出的分型标准对分离株进行分型并对分型标准进行验证。对已分离鉴定的人腺病毒进行人工诱变或提供重组热点序列，对重组进化机制进行验证。

本项目从GenBank等公开数据库搜集整理到233株人腺病毒全基因组序列数据，结合本实验室从临床样品中分离培养到的腺病毒毒株的测序结果，通过序列比对和人工检核，分别采用距离法和极大似然法构建腺病毒基因组进化树，证实了采用腺病毒基因组数据对腺病毒进行分类的合理性。通过系统地梳理和比较腺病毒基因组数据的保守区和变异区分布特征，将腺病毒基因组划分为34个保守区和35个高变区，并根据序列比对结果从中筛选到七个区域作为腺病毒分型的优先测序区域。通过理论分析和临床标本实际测序验证，证实了该优先测序方案是可行的。本项目提出的腺病毒优先测序方案和分型标准分别在依托单位和当地市级疾病预防控制中心的疫情防控中获得了成功应用。本项目采集临床样品三千六百余份，从中分离到腺病毒毒株31株。本项目在执行期间发表学术论文3篇，主办国际学术会议1次，外出参加国际学术会议6人次，培养（已毕业）博士后1名，博士1人，硕士7人。