Francisella tularensis intracellularly-induced outer membrane proteins

描述(由申请人提供)： 摘要 = 注意：此摘要摘自应用程序，未经SRA校对。如果应用程序扫描过程有问题，提取的文本可能不正确或不完整。 = 图拉氏方济氏菌是图拉热症的病原体，是一种革兰氏阴性的胞内球菌，擅长在包括人类在内的许多哺乳动物物种中引发急性致命疾病。图拉氏杆菌是公认的最危险的细菌病原体之一，因为它的感染量低(10个生物体)，易于气雾化，感染途径多样，并有能力导致严重的发病率和死亡率。关于图拉氏丝虫用来入侵宿主细胞并随后导致疾病的分子，人们知之甚少，但外膜蛋白(OMP)可能在这一过程中起着重要作用。事实上，OMP已被证明是许多其他细菌病原体的毒力因子，因此是理想的疫苗靶标。在我的博士后研究期间，我开发了一种提取和纯化图拉夫酵母OMPS的方法，该方法已经鉴定了近30个新的OMPS。在随后的研究中，我证明了一种基于OMP的疫苗可以保护小鼠免受A型图拉氏菌强毒力的肺部攻击。虽然这些结果令人信服，并极大地推动了图拉热症研究领域的发展，但用于这些研究的细菌是在富媒体条件下培养的。到目前为止，关于OMP在宿主感染过程中的表达的报道很少，但有理由假设图拉氏F菌在体内显著改变其OMP谱，以促进宿主细胞的侵袭和疾病。因此，这项研究计划将通过以下方面扩展我之前的发现：(1)使用RRT-PCR检测已知的OMP在哺乳动物宿主中是否上调或下调；(2)发现仅在巨噬细胞中表达的新的OMP；(3)通过产生OMP缺乏的突变体，确定新的毒力因子，并在小鼠肺部感染模型中评估这些突变体的传染性；以及(4)鉴定针对OMPS的抗体介导和细胞介导的免疫反应，从而对图拉氏丝虫肺部攻击具有保护作用。这些项目子目标旨在回答有关图拉氏肺吸虫毒力以及保护性免疫相关的基本问题。此外，这项建议在开发安全、有效的疫苗以预防图拉氏丝虫感染和疾病方面具有实际应用价值。土拉方杆菌之所以重要，不仅是因为它被归类为A类选择剂，还因为它在细胞内的快速致病作用。人们对图拉氏丝虫的表面分子知之甚少，但这些信息可能为开发安全、有效的图拉热症疫苗提供重要信息。这项提案中概述的项目寻求识别与感染有关的细菌分子，并表征保护性免疫反应。

DESCRIPTION (provided by applicant): Abstract ================== NOTICE: THIS ABSTRACT WAS EXTRACTED FROM APPLICATION AND HAS NOT BEEN PROOFED BY AN SRA.WHEN THERE ARE PROBLEMS WITH THE APPLICATION SCANNING PROCESS, THE EXTRACTED TEXT MAY BE INCORRECT OR INCOMPLETE. ================== Francisella tularensis, the etiological agent of tularemia, is a Gram-negative intracellular coccobacillus that is adept at inducing acute, fatal disease in a number of mammalian species, including humans. F. tularensis is well-recognized as one of the most dangerous bacterial pathogens known because of its low infectious dose (< 10 organisms), ease of aerosolization, multiple routes of infection, and capacity for inducing severe morbidity and mortality. Little is known about the molecules that F. tularensis uses to invade host cells and subsequently cause disease, but outer membrane proteins (OMPs) are likely instrumental in this process. Indeed, OMPs have been shown to be virulence factors in many other bacterial pathogens and, for this reason, are ideal vaccine targets. During my postdoctoral research, I developed a method to extract and purify F. tularensis OMPs, which has led to the identification of nearly 30 new OMPs. In subsequent studies, I demonstrated that an OMP-based vaccine protected mice against virulent Type A F. tularensis pulmonary challenge. While these results are compelling and have greatly advanced the tularemia research field, the bacteria used for these studies were grown under rich media conditions. To date, little has been reported about OMP expression during host infection, yet it is reasonable to hypothesize that F. tularensis dramatically alters its OMP profile in vivo to facilitate host cell invasion and disease. As such, this research proposal will extend my previous findings by: (1) Using rRT-PCR to examine whether known OMPs are up- or down-regulated in the mammalian host; (2) Discovering new OMPs that are exclusively expressed in macrophages; (3) Identifying new virulence factors through the generation of OMP-deficient mutants and assessing the infectivity of these mutants in a mouse pulmonary infection model; and (4) Characterizing both the antibody-mediated and cell- mediated immune responses against OMPs that confer protection against F. tularensis pulmonary challenge. These project subaims are designed to answer fundamental questions about F. tularensis virulence as well as the correlates of protective immunity. Additionally, this proposal has practical applications for the development of safe, efficacious vaccines to prevent F. tularensis infection and disease. Francisella tularensis is important not only because of its classification as a Category A Select Agent, but because of its rapid intracellular pathogenesis. Little is known about the surface molecules of F. tularensis, yet such information could provide vital information for the development of a safe, efficacious vaccine against tularemia. The projects outlined in this proposal seek to identify bacterial molecules involved in infection and characterize protective immune responses.