Abstract：
BACKGROUND: Rhipicephalus microplus poses a significant problem for livestock worldwide and is primarily controlled with synthetic acaricides. The continuous use of acaricides results in the selection of resistance and causes environmental harm. Vaccination presents an alternative solution to this problem, although searching for the suitable antigen is still a work in progress. Salivary proteins hold promise for inclusion in vaccine formulation due to their roles in modulating host responses, assisting blood feeding and pathogen transmission. Serpins are a class of proteinase inhibitors and are among the molecules found in tick saliva that modulate host blood coagulation, inflammation, and adaptive immune responses. Previous studies have demonstrated the potential of R. microplus serpin 17 (RmS-17) to interfere with the host\'s defenses, and antibodies have been shown to neutralize its effects. This makes RmS-17 an putative target for vaccine development.
METHODS: Epitope mapping of RmS-17 was achieved using in silico approach combining linear B-cell epitope and antigenicity predictor. In addition, epitope mapping using overlapping peptides in an ELISA screening was used. The serpin tridimensional structure and the epitopes spatial location within the molecule were determined. Peptides were synthetized based on the predictions and used for the production of rabbit anti-sera. Purified IgG\'s were used to assess the antibodies capacity to neutralize RmS-17.
RESULTS: Through in silico mapping, nine potential B cell epitope regions were screened, with p1RmS-17 and p2RmS-17 selected for the experiment based on antigen prediction. In the ELISA screening using overlapping peptides, eight antibody-binding regions were identified, and p3RmS-17 and p4RmS-17 were chosen. Antibodies raised against p3RmS-17 and p4RmS-17 partially neutralized RmS-17 activity.
CONCLUSIONS: It was found that antibodies against a single epitope are sufficient to partially neutralize RmS-17 activity. These findings support the possibility of using an epitope-based vaccine for immunization against R. microplus.
METHODS: Epitope mapping of RmS-17 was achieved using in silico approach combining linear B-cell epitope and antigenicity predictor. In addition, epitope mapping using overlapping peptides in an ELISA screening was used. The serpin tridimensional structure and the epitopes spatial location within the molecule were determined. Peptides were synthetized based on the predictions and used for the production of rabbit anti-sera. Purified IgG\'s were used to assess the antibodies capacity to neutralize RmS-17.
RESULTS: Through in silico mapping, nine potential B cell epitope regions were screened, with p1RmS-17 and p2RmS-17 selected for the experiment based on antigen prediction. In the ELISA screening using overlapping peptides, eight antibody-binding regions were identified, and p3RmS-17 and p4RmS-17 were chosen. Antibodies raised against p3RmS-17 and p4RmS-17 partially neutralized RmS-17 activity.
CONCLUSIONS: It was found that antibodies against a single epitope are sufficient to partially neutralize RmS-17 activity. These findings support the possibility of using an epitope-based vaccine for immunization against R. microplus.
摘要：
背景：微小根孢对世界范围内的家畜造成重大问题，主要用合成杀螨剂控制。连续使用杀螨剂会导致抗性的选择并造成环境危害。疫苗接种是解决这个问题的另一种方法，尽管寻找合适的抗原仍在进行中。唾液蛋白由于其在调节宿主反应中的作用而有望包含在疫苗制剂中。协助血液喂养和病原体传播。Serpin是一类蛋白酶抑制剂，并且是在tick唾液中发现的调节宿主血液凝固的分子之一。炎症,和适应性免疫反应。以前的研究已经证明了R.microplusserpin17（RmS-17）干扰宿主防御的潜力，和抗体已被证明可以中和其作用。这使得RmS-17成为疫苗开发的推定靶标。
方法:使用结合线性B细胞表位和抗原性预测因子的计算机模拟方法实现RmS-17的表位定位。此外,使用在ELISA筛选中使用重叠肽的表位作图。确定了serpin三维结构和分子内的表位空间位置。基于预测合成肽并用于生产兔抗血清。纯化的IgG用于评估抗体中和RmS-17的能力。
结果：通过计算机绘图，筛选了9个潜在的B细胞表位区域，根据抗原预测选择p1RmS-17和p2RmS-17进行实验。在使用重叠肽的ELISA筛选中，鉴定了8个抗体结合区,选择p3RmS-17和p4RmS-17。针对p3RmS-17和p4RmS-17产生的抗体部分中和RmS-17活性。
结论:发现针对单个表位的抗体足以部分中和RmS-17活性。这些发现支持使用基于表位的疫苗免疫针对R.microplus的可能性。
方法:使用结合线性B细胞表位和抗原性预测因子的计算机模拟方法实现RmS-17的表位定位。此外,使用在ELISA筛选中使用重叠肽的表位作图。确定了serpin三维结构和分子内的表位空间位置。基于预测合成肽并用于生产兔抗血清。纯化的IgG用于评估抗体中和RmS-17的能力。
结果：通过计算机绘图，筛选了9个潜在的B细胞表位区域，根据抗原预测选择p1RmS-17和p2RmS-17进行实验。在使用重叠肽的ELISA筛选中，鉴定了8个抗体结合区,选择p3RmS-17和p4RmS-17。针对p3RmS-17和p4RmS-17产生的抗体部分中和RmS-17活性。
结论:发现针对单个表位的抗体足以部分中和RmS-17活性。这些发现支持使用基于表位的疫苗免疫针对R.microplus的可能性。
