Development of rationally designed live-attenuated lumpy skin disease vaccines

合理设计的块状皮肤病减毒活疫苗的开发

• 批准号: BB/R008833/1
• 负责人: Philippa Beard
• 金额: $ 77.12万
• 依托单位: The Pirbright Institute
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2018
• 资助国家: 英国
• 起止时间: 2018 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FR008833%2F1
• 关键词: Development; rationally; designed; live; attenuated

The poxvirus genus Capripoxvirus (CPPV) contains three viral species which each cause high consequence, transboundary disease in ruminants. Research into CPPVs is very timely as the three species are spreading from their traditional geographic ranges into new regions. In particular LSDV has spread since 2011 throughout Eastern Mediterranean and Western Asian countries into Turkey (2013) and in August 2015 entered Europe (Greece) for the first time. It is now present throughout the Balkans, Caucasus and south-eastern Russia and threatens other European and Asian countries. Despite their importance there has been little progress in the last 30 years in improving CPPV vaccines.The newest types of poxvirus vaccines currently being developed in human medicine, such as NYVAC, are genetically engineered. In brief, comparative analyses of viral genomes are used to identify specific target regions of DNA for manipulation in order to produce a safe and effective "live" vaccine. In contrast all currently available CPPV vaccines are randomly mutated and some contain a mix of viruses of variable genotypes and virulence. There have been reports from the field of poor efficacy and safety of these randomly mutated vaccines, indicating the need for better CPPV vaccines. This project will follow the approach used in human medicine to develop safer and more reliable genetically engineered CPPV vaccines. Full genome sequencing of wildtype and attenuated strains of CPPV by Pirbright researchers and others has enabled bioinformatic analysis of these strains to identify CPPV genes which associate with virulence. We have additionally identified orthologues of known virulence factors of other poxvirus genera and combined the datasets to single out four putative virulence factors of CPPV, genes most likely to encode proteins which contribute to CPPV virulence. Deletion of these genes is likely to result in attenuated CPPV strains.The initial work (objective 1) will develop CRISPR/Cas9 technology to overcome difficulties in manipulating the CPPV genome. Following on from this, the four putative CPPV virulence factors will be mutated in a wildtype strain of LSDV. The function of the virulence factors will be assessed in isolation (objective 3) and in the context of a viral infection (objective 4). The data will then be assessed. Each mutant strain that (i) replicates to normal or near normal levels in cell culture, and (ii) is supported by evidence of a function consistent with a role as a virulence factor from the in vitro studies in objectives 3 and 4, will be taken forward into animal trials to experimentally measure the influence of the protein on viral virulence (objective 5). Cattle will be infected with a mutant, wildtype or repaired LSDV strain and physiological, behavioural, virological and immunological measurements taken over the following 28 days to assess the severity of the resultant disease. In the final study of the project (objective 6) the data from objectives 3, 4 and 5 will be used to design two mutated LSDV strains which will be assessed as live-attenuated vaccines in a challenge model of LSD. The hypothesis is that the targeted mutation/s will substantially reduce the ability of the virus to cause disease, while retaining strong poxviral immunogenicity.The main outputs of this project will be (a) identification of virulence factors encoded by LSDV, (b) an improved method for genetically manipulating CPPVs, and (c) rationally designed, genetically engineered vaccines to prevent LSD.

痘病毒属（CPPV）包含三种病毒，每种病毒在反刍动物中引起高度后果的跨界疾病。研究cppv是非常及时的，因为这三种物种正在从传统的地理范围向新的地区传播。特别是自2011年以来，LSDV在东地中海和西亚国家蔓延到土耳其（2013年），并于2015年8月首次进入欧洲（希腊）。它现在遍布巴尔干、高加索和俄罗斯东南部，并威胁着其他欧洲和亚洲国家。尽管它们很重要，但在过去30年中，在改进CPPV疫苗方面几乎没有取得进展。目前在人类医学中开发的最新类型的痘病毒疫苗，如NYVAC，是经过基因工程改造的。简而言之，利用病毒基因组的比较分析来确定DNA的特定目标区域，以便进行操作，以生产安全有效的“活”疫苗。相比之下，目前所有可用的CPPV疫苗都是随机突变的，有些疫苗含有不同基因型和毒力的病毒混合物。有报道称这些随机突变疫苗的有效性和安全性较差，这表明需要更好的CPPV疫苗。该项目将遵循人类医学中使用的方法，开发更安全、更可靠的基因工程CPPV疫苗。Pirbright等研究人员对CPPV野生型和减毒株进行了全基因组测序，使这些菌株的生物信息学分析能够确定与毒力相关的CPPV基因。此外，我们还鉴定了其他痘病毒属已知毒力因子的同源物，并结合数据集，挑选出CPPV的四种推定毒力因子，这些基因最有可能编码有助于CPPV毒力的蛋白质。这些基因的缺失可能导致CPPV毒株的减毒。最初的工作（目标1）将开发CRISPR/Cas9技术来克服操纵CPPV基因组的困难。在此基础上，四种假定的CPPV毒力因子将在LSDV野生型株中发生突变。将在单独（目标3）和病毒感染背景下（目标4）评估毒力因子的功能。然后将对数据进行评估。每个突变株(i)在细胞培养中复制到正常或接近正常水平，以及（ii）有证据表明其功能与目标3和4中体外研究中作为毒力因子的作用一致，将被用于动物试验，以实验测量蛋白质对病毒毒力的影响（目标5）。牛将感染突变型、野生型或修复的LSDV菌株，并在接下来的28天内进行生理、行为、病毒学和免疫学测量，以评估所产生疾病的严重程度。在该项目的最后研究（目标6）中，目标3、4和5的数据将用于设计两种LSDV突变株，它们将在LSD攻毒模型中作为减毒活疫苗进行评估。假设是，靶向突变/s将大大降低病毒引起疾病的能力，同时保持强痘病毒免疫原性。该项目的主要产出将是(a)鉴定LSDV编码的毒力因子，(b)改良的cppv基因操纵方法，以及(c)合理设计基因工程疫苗以预防LSD。

项目成果

Financial impact of sheeppox and goatpox and estimated profitability of vaccination for subsistence farmers in selected northern states of Nigeria.

• DOI: 10.1016/j.prevetmed.2021.105503
• 发表时间: 2022-01
• 期刊: Preventive veterinary medicine
• 影响因子: 2.6
• 作者: Rawlins ME;Limon G;Adedeji AJ;Ijoma SI;Atai RB;Adole JA;Dogonyaro BB;Joel AY;Beard PM;Alarcon P
• 通讯作者: Alarcon P

A novel strain of lumpy skin disease virus causes clinical disease in cattle in Hong Kong

• DOI: 10.1111/tbed.14304
• 发表时间: 2021-09-16
• 期刊: TRANSBOUNDARY AND EMERGING DISEASES
• 影响因子: 4.3
• 作者: Flannery, John;Shih, Barbara;Beard, Philippa M.
• 通讯作者: Beard, Philippa M.

Household and animal factors associated with sheeppox and goatpox sero-prevalence and identification of high-risk areas in selected States of northern Nigeria.

• DOI: 10.1016/j.prevetmed.2021.105473
• 发表时间: 2021-11
• 期刊: Preventive veterinary medicine
• 影响因子: 2.6
• 作者: Adedeji AJ;Ijoma SI;Atai RB;Dogonyaro BB;Adole JA;Maurice NA;Osemeke OH;Waziri IA;Atuman YJ;Lyons NA;Stevens KB;Beard PM;Limon G
• 通讯作者: Limon G