A novel zebrafish embryo model to define virulence factors of Bartonella henselae

描述(由申请人提供)：人类感染亨氏巴尔通体(Bh)可导致一系列临床症状和疾病，包括与猫抓病中观察到的局限性Bh感染相关的淋巴结病，以及由全身疾病引起的心内膜炎和菌血症。在一些患者中，全身感染Bh包括细菌性血管瘤病，其特征是由于小血管的增殖，皮肤或内脏出现红色/紫色结节。这种不寻常的血管生成宿主对感染的反应在某些巴尔通体物种的细菌中是独一无二的。体外研究已经确定了Bh中重要的毒力因子，这些因子在确定感染和/或引发宿主反应方面发挥了作用。第一个是VIRB/VirD4 IV型分泌系统，负责运送巴尔通体效应蛋白(BEPS A-G)，作用于内皮细胞的能力各不相同。第二个主要毒力因子是不依赖于VIRB的第二型IV型分泌系统TRW，它编码多种蛋白质，包括小黏附素TrwL1-TrwL-8，已被证明在宿主细胞黏附中发挥重要作用。最后，高分子量三聚体自转运蛋白粘附素Bada在体外已被广泛研究，它在自凝集、与宿主细胞的黏附、与细胞外基质蛋白结合、抑制吞噬和诱导血管生成等方面起着至关重要的作用。虽然体外研究已被证明对Bh致病机制的研究具有重要价值，但Bh毒力因子在体内尚未得到证实，而且对编码这些毒力因子的基因是如何调控的知之甚少。为了解决这个问题，我们利用转基因斑马鱼(ZF)胚胎开发了一种新的Bh感染体内模型，使我们能够同时监控感染和宿主反应，包括血管生成。我们假设我们的ZF胚胎模型将使我们能够：更严格地测试已在体外定义的毒力因子的作用；在体内识别新的毒力因子和毒力调节因子；并评估这些基因产物在宿主对Bh感染的反应中的作用。为了验证这些假设，我们提出了以下具体目标：1)定义毒力因子和毒力调节因子在Bh感染中的作用 2)在实验感染的转基因ZF胚胎中，确定Bh毒力因子和调节蛋白在诱导宿主对感染的反应中的作用。迫切需要体内模型来更好地了解这种细菌的独特发病机制和宿主反应，以便开发新的治疗策略。

DESCRIPTION (provided by applicant): Infection of humans with the zoonotic bacterium Bartonella henselae (Bh) can result in a range of clinical symptoms and disease including lymphadenopathy associated with localized Bh infection observed in cat-scratch disease, to endocarditis and bacteremia resulting from systemic disease. In some patients, systemic infection with Bh includes bacillary angiomatosis which is characterized by red/purple nodules on the skin or internal organs due to proliferation of the small blood vessels. This unusual angiogenic host response to infection is unique among bacteria to certain Bartonella species. In vitro studies have defined important virulence factors in Bh that play a role in establishing infection and/or eliciting a host response. The first is the VirB/VirD4 type IV secretion system that is responsible for delivering the bartonella effector proteins (Beps A-G) that act on endothelial cells in varying capacities. The second major virulence factor is a VirB-independent second type IV secretion system Trw, which encodes multiple proteins including small adhesins TrwL1-TrwL-8 which have been shown to play an important role in host cell adherence. Lastly, the high molecular weight trimeric auto transporter adhesin BadA has been studied extensively in vitro and is critical for auto agglutination, adhesion to host cells, binding to extracellular mtrix proteins, inhibition of phagocytosis, and induction of angiogenesis. While in vitro studies have proven invaluable for the study of Bh pathogenesis, no Bh virulence factors have been confirmed in vivo and remarkably little is known about how the genes encoding these virulence factors are regulated. To address this problem, we have developed a novel in vivo model of Bh infection using transgenic zebra fish (ZF) embryos that permits us to both monitor infection and host response including angiogenesis. We hypothesize that our ZF embryo model will enable us to: more rigorously test the role of virulence factors that have been defined in vitro; identify ne virulence factors and regulators of virulence in vivo ; and assess the role of these gene products in the host response to Bh infection. To test these hypotheses we propose the following specific aims: 1) Define the role of virulence factors and regulators of virulence in Bh infection using the transgenic ZF embryo model, and 2) Determine the role of Bh virulence factors and regulatory proteins in eliciting a host response to infection in experimentally infected transgenic ZF embryos. An in vivo model is urgently needed to better understand the unique pathogenesis and host response to this bacterium in order to develop novel treatment strategies.