Dissecting the neutralising antibody response to the porcine reproductive and respiratory syndrome virus to identify novel vaccine targets

猪繁殖与呼吸综合征（PRRS）是影响全球养猪业的最重要的传染病。猪繁殖与呼吸综合征病毒（PRRSV）是对动物福利和食品安全的主要威胁，正如猪高热病爆发所证明的那样，该疾病迅速蔓延到东南亚并造成毁灭性后果。据估计，美国和欧洲每年因PRRSV造成的损失分别超过6亿美元和15亿欧元。在英国，几乎一半的猪群感染了PRRSV，成本超过100英镑/头母猪。PRRSV作为两种遗传和抗原性不同的物种存在，PRRSV-1和-2，它们都在迅速进化。来自这两个物种的高致病性菌株的出现以及目前的减毒活疫苗未能提供针对不断扩大的病毒菌株多样性的广泛保护，对全球范围内的有效疾病控制提出了重大挑战。因此，迫切需要探索疫苗开发的替代方法来对抗PRRSV。中和抗体赋予针对PRRSV的保护作用，并且最近的研究表明抗体应答可以中和多种多样的PRRSV毒株。保守的抗原靶点的进一步了解将使新的vaccine.Identification的广泛中和抗体识别的表位的设计是一个激烈的背景下，一些高度可变的人类病毒的最近的研究领域。其核心是产生和分析天然存在的单克隆抗体（mAb）的特异性的方法。分析抗原特异性B细胞及其免疫球蛋白基因的方法学的最新进展为设计新型免疫原提供了大量的人单克隆抗体，新一代测序技术的出现为分析免疫个体的功能性抗体库开辟了新的途径。已经建立了将来自单个B细胞的免疫球蛋白重链和轻链的可变序列配对的方法，从而提供关于天然mAb的信息。然后，生物信息学可用于创建B细胞应答的全面视图，并鉴定用于表达重组mAb的克隆或克隆家族及其功能表征。然而，这些方法尚未在猪的背景下建立和开发。本项目旨在利用这些尖端技术来鉴定和鉴定能够中和PRRSV的天然存在的猪单克隆抗体，并绘制它们与病毒的次要（GP 2a，GP 3和GP 4）和主要（GP 5和M）包膜蛋白的反应性。它与BBSRC战略高度相关;针对动物健康的优先事项，生物科学的系统方法和管理动物的福利。拟议的工作结合了尖端技术和互补学科;在PRRS免疫学（格雷厄姆），免疫遗传学（哈蒙德），结构病毒学（格里姆斯）和蛋白质表达（欧文斯）所需的专业知识。该项目得到了全球最大的动物保健公司硕腾公司（Raue）的支持，该公司将提供行业视角，以确保项目完成后能够顺利进入下一个开发阶段。

Porcine reproductive and respiratory syndrome (PRRS) is the most important infectious disease affecting the global pig industry. PRRS viruses (PRRSV) are a major threat to both animal welfare and food security, as demonstrated by the pig high fever disease outbreak that rapidly spread across Southeast Asia with devastating consequences. Annual losses to PRRSV in the USA and Europe are estimated to exceed US$600 million and 1.5 billion euros, respectively. In the UK almost half of pig herds are infected with PRRSV with costs exceeding £100/sow. PRRSV exists as two genetically and antigenically distinct species, PRRSV-1 and -2, which are both rapidly evolving. The emergence of highly pathogenic strains from both species and the failure of current live attenuated vaccines to provide broad protection against an ever-expanding diversity of viral strains pose significant challenges to effective disease control world-wide. There is therefore an urgent requirement to explore alternative approaches to vaccine development to combat PRRSV. Neutralizing antibodies confer protection against PRRSV and recent studies have shown antibody responses can neutralize a wide diversity of PRRSV strains. An improved understanding of conserved antigenic targets would enable the design of novel vaccines.Identification of the epitopes recognised by broadly neutralising antibodies is an area of intense recent research in the context of a number of highly variable human viruses. Central to this are methods to generate and analyse the specificity of naturally occurring monoclonal Abs (mAbs). Recent advances in methodologies to analyse antigen-specific B cells and their immunoglobulin genes are now providing large numbers of human mAbs for potential application in the design of novel immunogens.The advent of next-generation sequencing technologies has opened new approaches to analyse the functional antibody repertoire from immune individuals. Methods have been established that pair the variable sequences of immunoglobulin heavy and light chains from individual B cells thus providing information on native mAbs. Bioinformatics can then be used to create a comprehensive view of the B cell response and to identify clones or clonal families for expression of recombinant mAbs and their functional characterisation. However, these approaches have yet to be established and exploited in the context of the pig. This project aims to adapt these cutting edge technologies to identify and characterise naturally occurring porcine mAbs capable of neutralising PRRSV and to map their reactivity to the minor (GP2a, GP3 and GP4) and major (GP5 and M) envelope proteins of the virus.The proposed project addresses the urgent need to explore novel approaches to PRRS vaccine development. It is highly relevant to BBSRC strategy; targeting the priorities of animal health, systems approaches to the biosciences and welfare of managed animals. The proposed work combines cutting edge techniques and complimentary disciplines; with the required expertise in PRRS immunology (Graham), immunogenetics (Hammond), structural virology (Grimes) and protein expression (Owens). The project is support by Zoetis (Raue), the largest global animal health company, who will provide an industrial perspective to ensure that upon completion the project is well placed to proceed to the next stages of development.