Arthropod saliva /midgut transcripts for vector vaccines

用于载体疫苗的节肢动物唾液/中肠转录本

• 批准号: 6987110
• 负责人: Jesus Valenzuela
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6987110
• 关键词: Leishmania; Psychodidae; antigen antibody reaction; arthropod borne communicable disease; biotechnology; complementary DNA; disease vectors; enzyme linked immunosorbent assay; flow cytometry; functional /structural genomics; hamsters; high throughput technology; host organism interaction; human tissue; laboratory mouse; microorganism antigen; nucleic acid sequence; polymerase chain reaction; proteomics; salivary glands; ticks; vaccine development; vector vaccine; western blottings

The Vector Molecular Biology Unit focuses on the molecular aspects of salivary and midgut transcripts and proteins in vector/host and vector/parasite interactions. We are interested in the use of salivary genes as vaccines to block parasite or pathogen transmission and in the use of midgut proteins as transmission blocking vaccines. We have previously identified and isolated a salivary gene from the sand fly Phlebotomus papatasi that confers protection against Leishmania major infection in mice. The approach to use vector salivary proteins or genes to block parasite infection may be applicable to other species of Leishmania and also to other vector borne diseases. By using high-throughput approaches we have isolated and sequenced a large number of cDNAs from various species of sand flies and from the tick Ixodes scapularis, the vector of Lyme disease. We have selected a number of these salivary genes and prepared DNA vaccines using a high-throughput cloning approach. We tested salivary DNA constructs from the sand flies P. argentipes, P. ariasi and L. longipalpis in animals and identified salivary proteins that produced a strong delayed skin response, salivary proteins that produce a strong antibody response and salivary proteins that produced both types of immune responses. Future work is directed to test these molecules for protection against pathogen transmission and to understand the mechanism of this protection. Another interest of the unit is the identification of the Leishmania major attachment site on the Phlebotomus papatasi sand fly midgut. Classical biochemical attempts have failed to identify and isolate this attachment site in the vector midgut. We have isolated and sequenced a large set of full-length genes expressed on the midgut of P. papatasi and identified a transcript coding for a tandem repeat galectin (galactose binding protein) and named PpGalec. Confocal microscopy studies demonstrated that PpGalec is expressed in the midgut surface of P. papatasi. Invitro and invivo binding experiments show that PpGalec is used by Leishmania major as a receptor for mediating specific binding to the insect midgut, an event that is crucial for parasite survival and that accounts for species-specific vector competence. These studies demonstrate the feasibility of using midgut receptors for parasite ligands as target antigens for transmission-blocking vaccines.

载体分子生物学单元侧重于唾液和中肠和中肠和蛋白质的分子方面，以及载体/宿主以及载体/寄生虫相互作用。我们对使用唾液基因作为疫苗的使用感兴趣，以阻止寄生虫或病原体传播以及将中肠蛋白用作传播疫苗的使用。我们以前已经从沙蝇植物帕帕塔西（Papatasi）中鉴定出并分离了唾液基因，该基因赋予了对小鼠利什曼原虫重大感染的保护。使用载体唾液蛋白或基因阻断寄生虫感染的方法可能适用于其他利什曼尼亚物种以及其他载体传播疾病。通过使用高通量方法，我们已经从各种沙蝇和tick ixodes capapularis（莱姆病的向量）分离并测序了大量cDNA。我们选择了许多这样的唾液基因，并使用高通量克隆方法制备了DNA疫苗。我们在动物中测试了来自沙蝇P. argentipes，Ariasi和L. longipalpis的唾液DNA构建体，并鉴定出产生强延迟皮肤反应的唾液蛋白，唾液蛋白，唾液蛋白产生强大的抗体反应和唾液蛋白产生了两种类型的免疫反应。未来的工作将用于测试这些分子以保护病原体传播，并了解这种保护的机制。该单元的另一个兴趣是鉴定利什曼原虫的主要附件位于phlebotomus papatasi sand蝇中心。经典的生化尝试未能识别和隔离矢量中心中的该附件位点。我们已经分离并测序了一组在P. p. p. p. p. p. p.的中肠上表达的大型全长基因，并鉴定了编码串联重复乳糖素（半乳糖结合蛋白）的转录本，并命名为ppgalec。共聚焦显微镜研究表明，PPGALEC在P. papatasi的中肠表面表达。 Invitro和Invivo结合实验表明，Leishmania Major将PPGALEC用作介导特异性结合与昆虫Midgut的受体，这一事件对于寄生虫生存至关重要，并且考虑了物种特异性载体的能力。这些研究表明，使用中肠受体将寄生虫配体作为透射阻塞疫苗的靶抗原具有可行性。