A unique approach to identify markers for congenital syphilis and neurosyphilis

描述（由申请人提供）：广泛挑战领域-转化科学（15）特定挑战主题- 15-OD（ORR）-101 *：预防、早期发现和治疗罕见疾病的试点项目，梅毒是一种慢性、多系统疾病，由螺旋体，梅毒螺旋体梅毒亚种（T。苍白球）。螺旋体通过性接触获得，在世界范围内流行。先天性梅毒和神经梅毒的表现在西方国家已经变得罕见，但在非洲、中亚和东欧的不发达国家仍然是一个问题。先天性梅毒是毁灭性的，可导致死胎，流产，出生体重低或早产。神经梅毒也在世界各地重新出现。T. pallidum序列显示其自身是细菌中最小的基因组之一（1.1Mb），具有一些潜在的表面蛋白，这些蛋白可以决定螺旋体在感染期间定殖各种组织的能力。螺旋体是细胞外病原体，它们对哺乳动物细胞的粘附似乎在组织定殖中起关键作用。然而，一些挑战导致了对T.苍白球发病机制(i)T.苍白球不能在体外生长。这限制了彻底调查这种生物的能力。(ii)遗传技术对这种螺旋体是不可行的。(iii)T.梅毒螺旋体表现出极端的宿主特异性，人类是其唯一的天然宿主。因此，迫切需要开发新的策略来研究T。梅毒发病机制，特别是相关的先天性梅毒和神经梅毒使用小动物模型系统。T.苍白球与引起莱姆病的伯氏疏螺旋体密切相关。这两种螺旋体在结构和生理学上是相似的生物体，并且共享几种同源蛋白质。T.苍白球和B.伯氏螺旋体在其宿主中表现出不同的疾病阶段，包括各种组织的播散性感染，随后是潜伏性和慢性疾病。为B开发遗传工具。在过去的十年里，burgdorferi的研究导致了对莱姆病发病机制的理解的重大进展。在这个提案中，我们将采用一种高度创新和非传统的方法，使用生物发光B。burgdorferi作为替代系统，以研究选定的T.苍白球分子在细胞特异性相互作用和发病机制。我们的假设是一个或多个T.生物发光B中的苍白球蛋白。burgdorferi将促进特定地点的入侵和定居。可导致先天性传播的胎盘侵入和中枢神经系统侵入可通过体内成像系统（IVIS 200）在小鼠中检测到。以下研究将检验这一假设。具体目的1：检测T. B中的苍白球蛋白。burgdorferi，并评估它们在体外粘附特定哺乳动物细胞系中的作用。具体目标2：确定B.在表达T. burgdorferi后，获得了在小鼠脑和胎盘中有效定殖的能力。梅毒螺旋体蛋白质，也促进莱姆病螺旋体从胎盘到幼崽的先天性传播。一种新型生物发光B的研制burgdorferi代理模型系统将有助于可视化小鼠胎盘和神经元组织的播散性定植，T.苍白球蛋白这种获得功能的方法将有助于填补对这种历史性的、不可培养的螺旋体的理解的差距。此外，迄今为止还没有确定有希望的抗梅毒候选疫苗。我们希望我们在B的经验。burgdorferi和我们的合作者，希拉·卢克哈特博士和阿图罗·百夫长的专业知识，在T。梅毒螺旋体的致病机制将有助于我们实现我们的目标，确定关键毒力因子的T。苍白球，并确定其作为诊断标志物和新的疫苗候选人的潜力。 公共卫生相关性：梅毒是一种慢性多系统疾病，先天性梅毒和神经梅毒的表现在西方国家是罕见的。先天性梅毒是毁灭性的，可导致死胎、流产、低出生体重或早产，神经梅毒也在世界各地重新出现。利用发光细菌作为替代系统来表达不能培养的梅毒螺旋体蛋白，将有助于研究小鼠胎盘和神经组织的定殖，也有助于鉴定新的诊断标志物和候选疫苗。

DESCRIPTION (provided by applicant): Broad Challenge Area-Translational Science (15) Specific Challenge Topic- 15-OD(ORDR)-101*: Pilot projects for prevention, early detection and treatment of rare diseases, Syphilis is a chronic, multisystemic disease caused by the spirochete, Treponema pallidum subspecies pallidum (T. pallidum). The spirochete is acquired by sexual contact and is prevalent worldwide. Congenital syphilis and neurosyphillis manifestation of the disease have become rare in the Western countries but they remain a problem in underdeveloped countries of Africa, Central Asia and Eastern Europe. Congenital syphilis is devastating and can result in stillbirths, abortions, low birth weights or premature births. Neurosyphillis is also re-emerging around the world. T. pallidum sequence revealed itself to be one of the smallest genomes (1.1Mb) among bacteria with a few potential surface proteins that can determine the spirochete's ability to colonize various tissues during infection. Spirochetes are extracellular pathogens and their adherence to mammalian cells appears to play a critical role in tissue colonization. However, several challenges have resulted in the poor understanding of T. pallidum pathogenesis. (i) T. pallidum cannot be grown in vitro. This limits the ability to investigate this organism thoroughly. (ii) Genetic techniques are not feasible for this spirochete. (iii) T. pallidum exhibits extreme host specificity with humans as its only natural host. Therefore, there is a desperate need to develop novel strategies to investigate the molecular basis of T. pallidum pathogenesis especially relevant to congenital syphilis and neurosyphillis using a small animal model system. T. pallidum is closely related to Borrelia burgdorferi, which causes Lyme disease. Both of these spirochetes are structurally and physiologically similar organisms and share several homologous proteins. T. pallidum and B. burgdorferi exhibit different phases of disease in their hosts, including disseminated infection of various tissues followed by latent and chronic disease. Development of genetic tools for B. burgdorferi in the past decade has led to a significant progress in the understanding of Lyme disease pathogenesis. In this proposal, we will employ a highly innovative and unconventional approach of using bioluminescent B. burgdorferi as a surrogate system to study the role of selected T. pallidum molecules in cell-specific interactions and pathogenesis. Our hypothesis is that expression of one or more T. pallidum proteins in bioluminescent B. burgdorferi will promote invasion and colonization of specific sites. Invasion of the placenta, which may lead to congenital transmission and of the central nervous system will be detectable in mice by an in vivo imaging system (IVIS 200). The following studies will test this hypothesis. Specific Aim 1: To examine the expression and localization of T. pallidum proteins in B. burgdorferi and assess their roles in adherence to specific mammalian cell lines in vitro. Specific Aim 2: To determine if B. burgdorferi acquires the ability to colonize brain and placenta of mice efficiently after expression of T. pallidum protein(s) and also promotes congenital transmission of Lyme spirochetes from placenta to the litter. Significance Development of a new bioluminescent B. burgdorferi surrogate model system in this proposal will help visualize disseminated colonization of the mouse placenta and neuronal tissues facilitated by the expression of T. pallidum proteins. This gain of function approach will help fill the gap in the understanding of this historic, uncultivable spirochete. Furthermore, there are no promising vaccine candidates identified against syphilis to date. We expect that our experience in B. burgdorferi and expertise of our collaborators, Drs. Sheila Lukehart and Arturo Centurion, in T. pallidum pathogenesis will help us achieve our objective to identify critical virulence factors of T. pallidum and determine their potential as diagnostic markers and novel vaccine candidates. PUBLIC HEALTH RELEVANCE: Syphilis is a chronic multisystemic disease and congenital syphilis and neurosyphillis manifestation of the disease are rare in the Western countries. Congenital syphilis is devastating and can result in stillbirths, abortions, low birth weights or premature births and neurosyphillis is also re-emerging around the world. Using bioluminescent bacteria as surrogate system to express uncultivable syphilis-causing spirochete proteins in this proposal will facilitate examination of colonization of mouse placenta and neuronal tissues and will also help in identification of new diagnostic markers and vaccine candidates.