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Rapid bioassay of cervid prions in PrP transgenic Drosophila: addressing the threat to animal and human health from Chronic Wasting Disease

PrP 转基因果蝇中鹿病毒的快速生物测定：解决慢性消耗性疾病对动物和人类健康的威胁

基本信息

批准号：
BB/T00343X/1
负责人：
Raymond Bujdoso
金额：
$ 60.61万
依托单位：
University of Cambridge
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2020
资助国家：
英国
起止时间：
2020 至无数据
项目状态：
未结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FT00343X%2F1
关键词：
Rapid bioassay cervid prions PrP

项目摘要

Prion diseases are fatal neurodegenerative disorders of humans and other animal species. These conditions are transmissible both within and between species. The pathogen that causes these transmissible brain diseases is an aggregated form of a normal protein found in neurons and is referred to as a prion. The only reliable method to detect prion infectivity is by bioassay in experimental animals. Animal prion diseases are a threat to human health because of their zoonotic potential, highlighted by the BSE epizootic in cattle and subsequent emergence of vCJD in humans. Chronic wasting disease (CWD) is a highly contagious and increasingly prevalent fatal prion disease of deer, and related species. CWD is a potential zoonosis since infectious prions are present in cervid skeletal muscle, which is consumed by humans. This view is supported by studies that show non-human primates can be experimentally infected with CWD-infected cervid brain tissue. CWD was considered to be restricted to N. West America but has now been found in Europe with cases detected in Norway and Finland. In attempts to control the disease Norway initiated the large-scale cull of wild reindeer, while Finland banned the export of live deer, two acts that had a significant impact upon their respective cervid industries. Emergence of these new foci of CWD infection poses significant new unmet challenges to cervid and human health across Europe, including the UK. The BSE epizootic in the UK highlighted the critical need for early development of a tractable prion bioassay for preparedness in assessment of the risk to animal and human health from new animal prion diseases, or as in the case of CWD, new reservoirs of an existing prion disease. It is important to develop a tractable prion bioassay that measures CWD prion infectivity in tissues and blood from affected cervids in order to formulate risk-based control measures that curb the spread of the disease amongst animals. In addition, it is necessary to carry out further CWD prion bioassays in experimental animals that are more ethically acceptable than primates in order to help predict the ability of this disease to infect humans. Both of these issues have been attempted using mice transgenic for cervid or human PrP, which provided murine models of deer and humans for CWD bioassays. These mouse prion bioassays involve injecting CWD-infected material into experimental mice and waiting to see if these animals develop prion disease. The mouse bioassay may take years to complete and is very expensive because of the cost of keeping large numbers of mice for a long period of time. Consequently, there is a need to develop a faster, cheaper, animal bioassay to assess CWD prion infectivity, that is as sensitive as the mouse prion bioassay. In our lab, we model prion disease in the fruit fly Drosophila melanogaster because they are relatively easy to work with, enable more rapid data collection and are less expensive to keep compared to large experimental animals and mice. Here, we will use Drosophila to develop a sensitive bioassay to detect CWD prions. To do so, we have introduced the cervid or human PrP gene into the flies that allows them to produce the protein that aggregates in the brain of deer with CWD or humans with vCJD, respectively. We already know, from our previously published studies, that this mammalian protein will aggregate and cause prion disease in transgenic Drosophila when flies are fed infectious prions. We will determine the sensitivity of our fly-based CWD prion bioassay, including its ability to bioassay cervid blood, and investigate its ability to model human susceptibility to CWD prions. We estimate that PrP transgenic Drosophila can be used to bioassay prion inocula at 10% (or less) of the cost of the mouse prion bioassay. Furthermore, our fly-based prion bioassay is completed within 6 weeks, whereas the mouse bioassay may require one or more years to assess the same prion inocula.

朊病毒病是人类和其他动物的致命性神经退行性疾病。这些疾病在物种内部和物种之间都可以传播。导致这些传染性脑部疾病的病原体是在神经元中发现的正常蛋白质的聚集形式，被称为朊病毒。检测朊病毒感染性的唯一可靠方法是通过实验动物的生物测定。动物朊病毒疾病是对人类健康的威胁，因为它们具有人畜共患的潜力，突出表现为牛的疯牛病流行和随后在人类中出现的vCJD。慢性消耗性疾病（CWD）是鹿及其相关物种的一种高度传染性和日益流行的致命性朊病毒疾病。CWD是一种潜在的人畜共患病，因为感染性朊病毒存在于人类消耗的鹿骨骼肌中。这一观点得到了一些研究的支持，这些研究表明，非人类灵长类动物可以通过实验感染感染CWD感染的鹿脑组织。CWD仅限于N.但现在已在欧洲发现，挪威和芬兰也发现了病例。为了控制这种疾病，挪威开始大规模扑杀野生驯鹿，而芬兰则禁止出口活鹿，这两项行动对各自的鹿产业产生了重大影响。这些新的慢性消耗病感染疫源地的出现对包括英国在内的整个欧洲的鹿科动物和人类健康构成了重大的新的未解决的挑战。英国的疯牛病流行突出了早期开发易于处理的朊病毒生物测定的迫切需要，以备评估新的动物朊病毒疾病对动物和人类健康的风险，或在慢性消耗病的情况下，现有朊病毒疾病的新宿主。重要的是要开发一种易于处理的朊病毒生物测定法，测量受影响的鹿科动物的组织和血液中的慢性消耗病朊病毒感染性，以制定基于风险的控制措施，遏制疾病在动物中的传播。此外，有必要在伦理上比灵长类动物更可接受的实验动物中进行进一步的CWD朊病毒生物测定，以帮助预测这种疾病感染人类的能力。这两个问题已经尝试使用转基因的鹿或人类PrP，这提供了慢性消耗病生物测定鹿和人类的小鼠模型的小鼠。这些小鼠朊病毒生物测定包括将感染了CWD的材料注射到实验小鼠体内，并等待观察这些动物是否会患上朊病毒疾病。小鼠生物测定可能需要数年才能完成，并且由于长时间保持大量小鼠的成本而非常昂贵。因此，需要开发一种更快、更便宜的动物生物测定法来评估慢性消耗病朊病毒感染性，其与小鼠朊病毒生物测定法一样敏感。在我们的实验室中，我们在果蝇Drosophila melanogaster中模拟朊病毒疾病，因为它们相对容易使用，能够更快速地收集数据，并且与大型实验动物和小鼠相比，保持成本更低。在这里，我们将使用果蝇来开发一种灵敏的生物测定法来检测慢性消耗病朊病毒。为了做到这一点，我们已经将鹿或人类PrP基因引入果蝇，使它们能够产生蛋白质，分别聚集在患有CWD的鹿或患有vCJD的人类的大脑中。我们已经知道，从我们以前发表的研究，这种哺乳动物蛋白质将聚集，并导致朊病毒疾病的转基因果蝇时，苍蝇喂养传染性朊病毒。我们将确定基于苍蝇的CWD朊病毒生物测定的灵敏度，包括其生物测定鹿血的能力，并研究其模拟人类对CWD朊病毒易感性的能力。我们估计，PrP转基因果蝇可用于生物测定朊病毒接种物在10%（或更少）的小鼠朊病毒生物测定的成本。此外，我们的基于苍蝇的朊病毒生物测定在6周内完成，而小鼠生物测定可能需要一年或多年来评估相同的朊病毒接种物。