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进食至饱腹的能力来预防莱姆病的传播。拟议的项目试图确定和表征一个家庭的抗凝血剂所使用的硬蜱蜱需要允许喂养他们的哺乳动物宿主。这种抗凝剂对于蜱虫喂养的成功和蜱虫携带的病原体的传播至关重要。我们将探讨家庭的硬蜱蜱抗凝剂从蜱唾液腺和唾液的生化分离，并通过探测cDNA和基因组文库与适当的引物片段。在我们的实验室以前的尝试，分离蜱唾液蛋白的探针cDNA文库与抗体从动物蜱免疫导致40个克隆，其中22个代表的抗凝剂，我们称之为SALP-14。在本提案中，我们将探针库与引物来自同源部分的克隆以前确定，以表征我们现在认识到的一个家庭的抗凝剂至关重要的蜱喂养和传播莱姆病。我们将表达和纯化重组抗凝剂并鉴定其活性。为了确定活性位点，我们将产生这些抗凝剂的缺失突变体，并确定有效催化活性以及抗原性所必需的最小区域。目标是通过针对抗凝剂分子的功能表位的免疫应答来干扰进食。喂食至饱对于蜱完成其生命周期和传播其携带的病原体至关重要。

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.