Antigen Discovery and Vaccine Development for Human Babesia Parasites

人类巴贝斯虫病是一种由扁虱传播的类似疟疾的疾病，在世界各地分布，并在美国流行。 各州。该疾病在2011年加入了CDC-国家报告疾病名单，原因是 已报告的病例。在美国，大多数临床病例都与微小巴贝斯虫有关，而巴贝斯虫 邓卡尼病例主要来自华盛顿、俄勒冈和加利福尼亚州。然而，由于缺乏 由于具有免疫能力的个体可能是无症状的，所以真正的 美国和世界各地巴贝斯虫病病例的发病率很可能被低估了。目前还没有疫苗可以预防 人类巴贝斯虫病和目前的治疗方法是无效的，需要高剂量，其中一些是相关的 有很大的副作用。即使有了这些药物和其他支持措施，病死率也超过了 在敏感宿主中为20%。在过去的8年里，我们对生物学有了很大的了解， 巴贝斯虫感染人类的致病机制及药物敏感性：(1)我们完成了第一个基因组 对B.microti和B.duncani进行了转录分析和注释；(2)探讨了B.microti和B.duncani的基因组多样性。 (3)证实微小杆菌和邓肯芽胞杆菌具有天然的耐受性 临床推荐的治疗方法；(4)发现了一种能根治微小杆菌感染的联合疗法 (5)完成了对微小芽孢杆菌的首次免疫蛋白质组学分析，鉴定了主要的 该寄生虫的免疫优势抗原，并建立了第一个诊断方法来检测活性B。 微体感染；以及(6)发现了寄生虫使用的一种基于囊泡的分泌系统，以输出一些 它的蛋白质传给宿主。与当前的RFA特别相关的是，我们使用了免疫蛋白质组学和蛋白质组学 鉴定微小分枝杆菌和邓肯分枝杆菌分泌和表面展示蛋白的方法 30微杆菌抗原决定其在受感染的小鼠和人的血清中的血清学特征。主 这一建议的假设是，出口的一些微小杆菌和邓肯杆菌的抗原可以作为 非法抗体疫苗，以阻止巴贝斯虫入侵宿主红细胞或促进消除巴贝斯虫- 被宿主巨噬细胞感染的红细胞。这个应用程序的主要目标是进一步描述 这些分泌抗原的免疫原性分析，并在小鼠身上进行疫苗接种研究，以鉴定这些 可被开发为疫苗的抗原，以对抗微小巴贝斯虫和邓肯巴贝斯虫感染。在目标1中，我们 将产生微小杆菌和邓肯杆菌分泌和表面展示蛋白的功能性重组形式 并用它们来确定小鼠体液免疫反应的血清学特征和动力学 和人类血清。在目标2中，我们将进行细胞生物学研究，以确定分泌物和细胞 候选出口抗原在这些寄生虫的红细胞内生命周期中的分布。在《目标3》中， 我们将在感染微小分枝杆菌和邓肯分枝杆菌的小鼠模型上进行免疫研究，以确定这些 在与人类相容的佐剂中注射时有效的疫苗的抗原。

Human babesiosis is a tick-borne malaria-like illness with a worldwide distribution and endemic in the United States. The disease joined the list of CDC-nationally notifiable diseases in 2011 due to the increasing number of reported cases. The majority of clinical cases in the US have been linked to Babesia microti whereas Babesia duncani cases have been reported primarily from Washington, Oregon, and California. However, due to the lack of specific diagnostic assays and because immune competent individuals can be asymptomatic, the true incidence of babesiosis cases in the US and worldwide is most likely underestimated. There is no vaccine against human babesiosis and current therapies are not effective, require high doses and some of them are associated with major side effects. Even with these drugs and other supportive measures, the case fatality rate exceeds 20% in susceptible hosts. During the past 8 years, we gained significant understanding of the biology, pathogenesis and drug susceptibility of Babesia parasites that infect humans: (1) we completed the first genomic and transcriptomic analyses and annotations of B. microti and B. duncani; (2) probed the genomic diversity of B. microti in human blood and ticks; (3) demonstrated that B. microti and B. duncani are inherently tolerant to clinically recommended therapies; (4) discovered a combination therapy that results in radical cure of B. microti infection in mice; (5) completed the first immunoproteomic analysis of B. microti, identified the main immunodominant antigens of this parasite, and developed the first diagnostic assay for detection of active B. microti infection; and (6) discovered a vesicular-based secretion system used by the parasite to export some of its proteins to the host. Of particular relevance to the current RFA, we used immunoproteomic and proteomic approaches to identify secreted and surface-displayed proteins of B. microti and B. duncani, and for more than 30 B. microti antigens determine their serological profile in sera from infected mice and humans. The main hypothesis of this proposal is that some of the exported antigens of B. microti and B. duncani could serve as vaccines to illicit antibodies to block Babesia invasion of host erythrocytes or to facilitate elimination of Babesia- infected erythrocytes by host macrophages. The primary goals of this application are to further characterize the immunogenicity profile of these secreted antigens, and to conduct vaccination studies in mice to identify those antigens that could be developed as vaccines against Babesia microti and B. duncani infections. In Aim 1, we will produce functional recombinant forms of secreted and surface displayed proteins of B. microti and B. duncani and use them to determine the serological profile and kinetics of the humoral immune responses using mouse and human sera. In Aim 2. We will conduct cell biological studies to determine the secretion and cellular distribution of the candidate exported antigens during the intraerythrocytic life cycle of these parasites. In Aim 3, we will conduct immunization studies in mouse models of B. microti and B. duncani infections to identify those antigens that are effective vaccines when administered in human-compatible adjuvants.