Tick Vaccine Prevention of Lyme Borreliosis

描述（由申请人提供）：我们的策略是通过干扰蜱虫的进食能力来防止莱姆病的传播。拟议的项目试图确定和表征一种抗凝血剂家族，这种抗凝血剂被硬蜱用来捕食它们的哺乳动物宿主。这种抗凝剂对蜱虫取食和蜱虫传播病原体的成功至关重要。我们将通过从蜱的唾液腺和唾液中进行生化分离，并利用合适的引物片段探测cDNA和基因组文库来探索蜱的抗凝血剂家族。在我们的实验室中，通过检测免疫动物的抗体cDNA文库来分离蜱虫唾液蛋白的尝试获得了40个克隆；其中22种是我们称之为SALP-14的抗凝剂。在目前的提议中，我们将用先前鉴定的克隆同源部分衍生的引物来探测文库，以表征我们现在认识到的对蜱虫喂养和莱姆病传播至关重要的抗凝血剂家族。我们将表达和纯化重组抗凝血剂，并对其活性进行表征。为了确定活性位点，我们将生成这些抗凝血剂的缺失突变体，并确定有效催化活性和抗原性所必需的最小区域。目的是通过针对抗凝血分子的功能表位的免疫反应来干扰喂养。吃饱是蜱完成其生命周期和传播它们所携带的病原体的必要条件。

DESCRIPTION (provided by applicant): Our strategy is to prevent transmission of Lyme borreliosis by interfering with the tick's ability to feed to repletion. The proposed project attempts to identify and characterize a family of anticoagulants used by Ixodes ticks needed to permit feeding on their mammalian hosts. Such anticoagulants are vital to the success of tick feeding and in the transmission on tick borne pathogens. We shall explore the family of Ixodes tick anticoagulants by biochemical separation from tick salivary gland and saliva, and by probing cDNA and genomic libraries with appropriate primer fragments. Previous attempts in our laboratories to isolate tick salivary proteins by probing cDNA libraries with antibodies from animals made tick immune resulted in 40 clones; 22 of which were represented by an anticoagulant that we have called SALP-14. In the present proposal we will probe libraries with primers derived from homologous portions of the clones previously identified in order to characterize what we now recognize as a family of anticoagulants vital to tick feeding and transmission of Lyme borreliosis. We will express and purify the recombinant anticoagulants and characterize their activities. To determine active sites, we will generate deletion mutants of these anticoagulants and identify the smallest region essential for effective catalytic activity, as well as antigenicity. The goal is to interfere with feeding by the immune response directed against the functional epitope(s) of the anticoagulant molecules. Feeding to repletion is essential for ticks to complete their life cycle and for the transmission of the pathogens they harbor.