3D fluorescence microscope for virus-host interaction studies

用于病毒-宿主相互作用研究的 3D 荧光显微镜

• 批准号: RTI-2022-00621
• 负责人: Mossman, Karen
• 金额: $ 10.93万
• 依托单位: McMaster University
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments
• 财政年份: 2021
• 资助国家: 加拿大
• 起止时间: 2021-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=741741
• 关键词: 3D; fluorescence; microscope; virus; host

Bats are ecologically important as pollinators and insect controllers. They are natural reservoirs for several emerging and re-emerging viruses, including coronaviruses such as SARS-CoV-2, that are causative agents of important human and agricultural infections. These viruses are speculated to have spilled over or "jumped" from bats to humans and other animals, causing severe and often fatal disease. Despite evidence of bats harboring these viruses, they do not appear to show overt symptoms or clinical signs of infection. Little is understood why bats are not susceptible to these viral infections and the physiological characteristics that allow them to be carriers or reservoirs of these viruses. The Mossman lab has over 20 years of experience studying virus-host interactions and along with clinical colleagues in Toronto, were the first to isolate and characterize SARS-CoV-2 from two patients, which has led to over 20 publications. Our Discovery Grant has generated a unique and innovative program that leverages our expertise (virus-host interactions and innate immunity) and that of Dr. Faure who runs the world's only bat colony containing wild caught and bred animals (McMaster Bat Lab). We have developed cell lines and tools with which to study mechanisms of virus-bat interactions, including primary cultures and 3D organoids, the latter composed of multiple cell types from intestinal stem cells. We routinely use fluorescence microscopy to monitor how viruses invade cells and how cells respond to viral invasion. As our current inverted fluorescence microscope is obsolete due to lack of availability of components for repair (it was purchased 19 years ago), we are requesting a new fluorescence microscope. The requested infrastructure, a THUNDER 3D Imaging System, uses new computational clearing technologies to generate high resolution and high contrast images from thick samples, such as organoid cultures. It allows for acquisition of images taken deep within tissues such as organoids. The THUNDER system is a Leica technology, as is our current microscope, allowing for recycling of some components. This technology will allow us to monitor the kinetics and complexity of virus-host interactions in real time, while maintaining the integrity of sensitive samples. Understanding how bats harbor and spread viruses with an apparent lack of symptoms has implications for disease control in animals and humans. As big brown bats naturally occur in Canada, our findings will directly impact Canadian agricultural and health sectors. Moreover, findings will inform global strategies for disease control and development of novel antiviral therapies, particularly for new emerging viruses that bats appear to carry. Finally, this program will lay the foundation for training of highly qualified personnel in a multidisciplinary environment, working at the forefront of science and impacting on critical areas such as pandemic preparedness and understanding.

蝙蝠作为传粉者和昆虫控制者在生态学上具有重要意义。它们是几种新出现和重新出现的病毒的天然宿主，包括SARS-CoV-2等冠状病毒，这些病毒是重要的人类和农业感染的病原体。据推测，这些病毒从蝙蝠身上溢出或“跳跃”到人类和其他动物身上，导致严重且往往致命的疾病。尽管有证据表明蝙蝠携带这些病毒，但它们似乎没有表现出明显的感染症状或临床体征。很少有人知道为什么蝙蝠对这些病毒感染不敏感，以及使它们成为这些病毒的携带者或储存者的生理特征。Mossman实验室拥有超过20年的研究病毒-宿主相互作用的经验，并与多伦多的临床同事一起沿着，首次从两名患者中分离并鉴定了SARS-CoV-2，这导致了20多篇出版物。我们的发现补助金产生了一个独特的创新计划，利用我们的专业知识（病毒-宿主相互作用和先天免疫）和Faure博士的专业知识，Faure博士经营着世界上唯一一个包含野生捕获和繁殖动物的蝙蝠殖民地（McMaster蝙蝠实验室）。我们已经开发了细胞系和工具，用于研究病毒-蝙蝠相互作用的机制，包括原代培养物和3D类器官，后者由来自肠干细胞的多种细胞类型组成。我们通常使用荧光显微镜来监测病毒如何入侵细胞以及细胞如何对病毒入侵作出反应。由于我们目前的倒置荧光显微镜因缺乏维修组件而过时（19年前购买），我们正在申请一台新的荧光显微镜。所要求的基础设施，THUNDER 3D成像系统，使用新的计算清除技术，从厚样品（如类器官培养物）生成高分辨率和高对比度图像。它允许采集在组织（如类器官）深处拍摄的图像。THUNDER系统采用徕卡技术，与我们目前的显微镜一样，允许回收某些组件。这项技术将使我们能够真实的监测病毒-宿主相互作用的动力学和复杂性，同时保持敏感样品的完整性。了解蝙蝠如何携带和传播明显缺乏症状的病毒，对动物和人类的疾病控制具有重要意义。由于大棕蝙蝠自然发生在加拿大，我们的研究结果将直接影响加拿大的农业和卫生部门。此外，研究结果将为疾病控制和新型抗病毒疗法的开发提供全球战略信息，特别是针对蝙蝠似乎携带的新病毒。最后，该计划将为在多学科环境中培养高素质人才奠定基础，在科学的前沿工作，并对大流行的准备和理解等关键领域产生影响。