Application of IVIAT to Bacillus anthracis

IVIAT在炭疽杆菌中的应用

• 批准号: 6681644
• 负责人: Edward T. Ryan
• 金额: $ 27.3万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2005-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6681644
• 关键词: Bacillus anthracis; Escherichia coli; animal tissue; anthrax; antiserum; bacterial genetics; bioterrorism /chemical warfare; functional /structural genomics; gene expression; genetic library; genetic screening; human tissue; laboratory rabbit; virulence

DESCRIPTION (provided by applicant): Bacillus anthracis is the cause of cutaneous, inhalation, and gastrointestinal anthrax. B. anthracis has also recently been used as an agent of bioterrorism. The identification of bacterial genes expressed uniquely in vivo during infection with B. anthracis would improve our understanding of the molecular bacterial events that occur during anthrax, and could lead to improved therapeutics and (less likely) an improved anthrax vaccine. One recently developed technique that permits identification of bacterial genes expressed uniquely in vivo is IVIAT (in vivo induced antigen technology). In this procedure, convalescent serum collected from humans or animals infected with a pathogen of interest is absorbed against bacteria grown in vitro. Absorbed serum is then used to probe a genomic DNA expression library of the pathogen of interest in an E. coli host system. Reactive clones express an antigen expressed uniquely in vivo, and reactive genes and their products can be further identified and analyzed. Our hypothesis is that B. anthracis contains genes that are expressed uniquely in vivo during anthrax, and that identification of such genes and their products will lead to improved understanding of the pathogenic events that occur during anthrax. Our specific aim, therefore, is to apply IVIAT to B. anthracis using two sets of already collected convalescent sera: one from rhesus macaques that survived inhalational challenge with virulent Ames strain B. anthracis and cleared documented B. anthracis bacteremia as part of a fully approved anthrax study at the Centers for Disease Control and Prevention; the other from humans surviving naturally acquired cutaneous anthrax (collected as part of a fully approved collaborative study between Kazakhstani and CDC researchers). We designed our study to take into account the high lethality of anthrax and the presence of a B. anthracis capsule, and in our project, we will specifically not evaluate well characterized virulence factors of B. anthracis (such as exotoxin and capsule), but will focus our efforts on identifying previously unrecognized genes uniquely expressed in vivo. IVIAT is an established protocol in our laboratory, and this preliminary collaborative study should lay a foundation for subsequent analysis of identified B. anthracis genes and their products.

描述（由申请人提供）：炭疽芽孢杆菌是皮肤，吸入和胃肠道炭疽病的原因。 B.炭疽病最近也被用作生物恐怖主义的剂。 在炭疽芽孢杆菌感染期间，在体内唯一表达的细菌基因的鉴定将改善我们对炭疽期间发生的分子细菌事件的理解，并可能导致改善的治疗疗法，并且（较少）改善了炭疽疫苗。 最近开发的技术允许鉴定在体内独特表达的细菌基因是iviat（体内诱导的抗原技术）。 在此过程中，从人类或感染感兴趣病原体的动物收集的康复血清被吸收在体外生长的细菌。 然后，吸收的血清用于探测大肠杆菌宿主系统中感兴趣的病原体的基因组DNA表达文库。 反应性克隆表达了一种抗原在体内独特的抗原，可以进一步鉴定和分析反应性基因及其产物。 我们的假设是，炭疽芽孢杆菌包含在炭疽期间在体内独特表达的基因，并且对这种基因及其产物的鉴定将导致人们对炭疽期间发生的致病事件的理解得到改善。 因此，我们的具体目的是使用两组已经收集的康复血清将iviat应用于炭疽芽孢杆菌：一组来自恒河猕猴的猕猴，一种与毒力型AMES菌株B.炭疽病生存，并清除了已记录的嗜血芽孢杆菌细菌，作为在中心疾病控制和疾病控制和预言中得到充分认可的植物研究的一部分。另一名来自人类自然获得的皮肤炭疽（作为全面批准的哈萨克斯坦和CDC研究人员之间的合作研究的一部分）。 我们设计了我们的研究来考虑炭疽芽孢杆菌的高致死性和存在的炭疽芽孢杆菌胶囊的存在，在我们的项目中，我们将特别不会评估炭疽芽孢杆菌的毒力性良好的毒力因子（例如Exotoxin和capsule），但会将我们的精力集中在识别先前未识别的基因中，以识别在Vivo中的独特表现出来。 Iviat是我们实验室中建立的协议，这项初步的协作研究应为随后分析已鉴定的炭疽基因芽孢杆菌及其产品奠定基础。