Pathogenesis of Haemophilus Ducreyi Infections

描述（由申请方提供）：杜克雷嗜血杆菌是一种严格的人类病原体，可引起软下疳，这是一种生殖器溃疡疾病（GUD），可促进人类免疫缺陷病毒（HIV-1）的传播。为了研究H. ducreyi，我们开发了一个人类挑战模型，接近模拟自然感染。在感染过程中，H. ducreyi在脓肿的恶劣环境中被发现并抵抗吞噬作用。我们发现H. ducreyi对宿主有广泛的转录反应，表明H. ducreyi感知并响应宿主环境。H. ducreyi只包含两个已知对细胞质外应激有反应的系统：2组分调节因子CpxRA和替代性σ因子RpoE。几个H. CpxRA特异性效应子的ducreyi同源物在脓疱中上调，而RpoE特异性效应子的许多同源物在脓疱中下调，表明CpxRA和RpoE系统在H. ducreyi和以协调的方式起作用以响应由宿主引起的压力。我们发现CpxRA控制了H. ducreyi和CpxRA的不受控制的激活削弱了生物体感染人类志愿者的能力。我们假设H. ducreyi通过CpxRA和RpoE感知宿主环境，CpxRA和RpoE在感染过程中发挥作用以对抗体内遇到的应激并控制毒力决定簇的产生，CpxRA或RpoE系统中的组成型活性表达子或缺失突变体的毒力将被减弱，并且已知或新的毒力决定簇在感染过程中将受到CpxRA和RpoE的差异调节。为了验证这些假设，我们的具体目标包括：1）评估cpxR、rpoE、rseA和rseC缺失突变体和表达组成型激活的CpxR和RpoE的菌株在人类志愿者中的毒力; 2）确定CpxRA和RpoE系统在人类感染相关模型中致病的潜在机制; 3）鉴定由CpxRA和RpoE途径控制的新型毒力决定因子，并评估它们在体内（Aim 1）和体外（Aim 2）模型中的感染作用。我们的建议提供了独特的机会来研究两个相互关联的应激反应系统对人类病原体生存的贡献，将导致新的毒力决定因素的鉴定，并已导致一种新的抗菌策略的发展。经NIAID批准，本次修订的目的是获得资金，以支持与资助申请的目标1和3相关的人类接种实验。 公共卫生相关性：杜克雷嗜血杆菌是一种导致生殖器溃疡疾病的细菌，有助于艾滋病毒传播。H.当它感染人类宿主时，ducreyi开启了它的许多基因的表达。我们的研究旨在探讨如何H。ducreyi利用压力反应系统来适应人类宿主。H. ducreyi仅被三类抗生素杀死，并且是开发新疗法的高度优先生物体。该项目将为H. ducreyi和其他细菌感染。

DESCRIPTION (provided by applicant): Haemophilus ducreyi is a strict human pathogen that causes chancroid, a genital ulcer disease (GUD) that facilitates the transmission of the human immunodeficiency virus (HIV-1). To study the biology of H. ducreyi, we developed a human challenge model that closely simulates natural infection. During infection, H. ducreyi is found in the hostile environment of an abscess and resists phagocytosis. We found that H. ducreyi had a broad transcriptional response to the host, suggesting that H. ducreyi senses and responds to the host environment. H. ducreyi contains only two systems known to respond to extracytoplasmic stress: the 2- component regulator CpxRA and the alternative sigma factor, RpoE. Several H. ducreyi homologues of CpxRA-specific effectors were upregulated, while many homologues of RpoE-specific effectors were downregulated in pustules, suggesting that that CpxRA and RpoE systems are linked in H. ducreyi and function in a coordinated fashion to respond to stresses mounted by the host. We found that CpxRA controlled the expression of several major virulence determinants of H. ducreyi and that uncontrolled activation of CpxRA impaired the ability of the organism to infect human volunteers. We hypothesize that H. ducreyi senses the host environment via CpxRA and RpoE, that both CpxRA and RpoE function during infection to combat stresses encountered in vivo and control the production of virulence determinants, that constitutively active expressers or deletion mutants in the CpxRA or RpoE systems will be attenuated for virulence and that known or novel virulence determinants will be differentially regulated by CpxRA and RpoE during infection. To test these hypotheses, our specific aims include: 1) evaluation of cpxR, rpoE, rseA and rseC deletion mutants and strains that express constitutively activated CpxR and RpoE for virulence in human volunteers; 2) determination of the mechanism(s) underlying the contributions of the CpxRA and RpoE systems to pathogenesis in models relevant to human infection; 3) identification of novel virulence determinants controlled by the CpxRA and RpoE pathways and evaluation of their role in infection in the in vivo (Aim1) and in vitro (Aim 2) models. Our proposal offers the unique opportunity to study the contributions of two interrelated stress response systems to the survival of a pathogen in humans, will lead to the identification of novel virulence determinants, and has already led to the development of a novel antimicrobial strategy. As approved by NIAID, the purpose of this revision is to obtain funds to support the human inoculation experiments related to Aims 1 and 3 of the funded application. PUBLIC HEALTH RELEVANCE: Haemophilus ducreyi is a bacterium that causes a genital ulcer disease that facilitates HIV transmission. H. ducreyi turns on the expression of many of its genes when it infects the human host. Our study is designed to examine how H. ducreyi uses stress response systems to adapt to the human host. H. ducreyi is only killed by three classes of antibiotics and is a high priority organism for the development of new therapies. This project should yield targets of vaccines or alternative treatments for H. ducreyi and other bacterial infections.