Relapsing Fever Spirochete Protein Production within the Vector

描述（由申请人提供）：了解微生物在不同自然环境中的适应性仍然是新兴传染病研究的知识空白。蜱传疾病继续危害人类健康，鉴于媒介具有很高的繁殖潜力、在极端条件下生存的能力以及传播寄生虫、病毒和细菌病原体的能力，了解病原体在蜱传媒介中的定植与公共卫生特别相关。其中一种病原体是蜱传回归热（RF）螺旋体。RF螺旋体对人类健康的全球分布和经济负担未得到充分认识，特别是考虑到该疾病对贫困国家人口的流行和影响，但我们不了解病原体如何在蜱虫媒介中适应。这种病媒的一个独特特征是，蜱虫的寿命非常长，每次进食之间可以间隔数年，但细菌仍然存活并具有传染性。此外，螺旋体的摄食速度和随后的传播速度与其他蜱传疾病有很大不同，这进一步强调了了解RF螺旋体如何在蜱内生存的必要性。阻碍疾病预防疗法发展的一个限制是对导致RF螺旋体定植蜱病媒介的分子事件了解不足。为了解决这个问题，我们建立了一个体内系统来研究蜱-哺乳动物的完整感染周期，这是一种螺旋体。我们鉴定了螺旋体在蜱感染过程中上调的一系列20个连续的质粒定位基因，并通过计算机分析表明这些基因编码了假定的外膜蛋白（Omps）。我们已经在大肠杆菌中扩增、克隆和表达了15个基因，并提出了针对重组蛋白的兔血清，使我们能够确定蛋白质是否表面暴露。以确认的Omps为重点，我们将确定在蜱的两个区域（中肠和唾液腺）中Omps的差异产生，这两个区域是turicatae必须定殖以确保螺旋体继续生命周期的区域。在完成本研究后，我们将更好地了解蜱感染过程中turicatae表面蛋白质组，并确定在蜱媒介的中肠和唾液腺中是否有差异产生Omps。我们的研究结果将为竞争性R01应用提供必要的数据，以研究中肠和唾液腺中独特产生的Omps是否对turicatae在这些组织中的定植是必要的。

DESCRIPTION (provided by applicant): Understanding microbial adaption within different natural environments remains a gap in knowledge in emerging infectious disease research. Tick-borne diseases continue to afflict human health, and given the vectors high reproductive potential, the ability to survive in extreme conditions, and the ability to transmit parasitic, virl, and bacterial pathogens, understanding pathogen colonization within the tick vector is particularly relevant to public health. One such pathogen is tick-borne relapsing fever (RF) spirochetes. The global distribution and economic burden of RF spirochetes on human health is underappreciated, especially considering the prevalence and impact of the disease on populations in impoverished countries, yet we do not understand how the pathogen adapts within the tick vector. A unique characteristic of the vector is that the ticks are extremely long-lived and can go years between feedings, yet the bacteria remain viable and transmissible. Additionally, the rapidity of feeding and subsequent transmission of the spirochetes differs significantly from other tick-borne diseases, further emphasizing the need to understand how RF spirochetes survive within the tick. One limitation that has prevented the development of disease preventing therapeutics has been a poor understanding of the molecular events contributing to RF spirochete colonization of the tick vector. To address this, we have established an in vivo system to study the complete tick-mammalian infectious cycle for Borrelia turicatae, a species of RF spirochete. We have identified a series of 20 consecutive plasmid localized genes that the spirochetes up- regulate during tick infection, and an in silico analysis indicated that the genes encode for putative outer membrane proteins (Omps). We have amplified, cloned, and expressed 15 of the genes in E. coli and propose to generate rabbit serum against the recombinant proteins, allowing us to determine if the proteins are surface exposed. Focusing on the confirmed Omps, we will determine the differential production of the Omps within two regions of the tick that B. turicatae must colonize to ensure the spirochetes continued life cycle, the midgut and salivary glands. At the completion of this proposal, we will better understand the surface proteome of B. turicatae during tick infection and we will determine if Omps are differentially produced within the midgut and salivary glands of the tick vector. Our findings will generate the data necessary for a competitive R01 application investigating if the Omps uniquely produced within the midgut and salivary glands are necessary for B. turicatae colonization of these tissues.