EAGER: An innovative approach for quantification and prospective isolation of Nosema ceranae life stages from host cells with potential for application to diverse pathogen species

EAGER：一种从宿主细胞中定量和前瞻性分离蜜蜂微孢子虫生命阶段的创新方法，具有应用于多种病原体物种的潜力

• 批准号: 2125981
• 负责人: Jonathan Snow
• 金额: $ 9.71万
• 依托单位: Barnard College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2125981&HistoricalAwards=false
• 关键词: EAGER; innovative; approach; quantification; prospective

Some microbial pathogens are obligate intracellular pathogens. They are hard to study because of their complex life cycles which occur inside the cells of their hosts. In other areas of biology, a technique called fluorescence activated cell sorting (FACS) is used to identify and separate different cell types from a complex mixture to near homogeneity based on differential fluorescent-based staining. This proposal will use this technique to measure and isolate different life stages of a specific parasite (Nosema ceranae) that infects honey bees and negatively impacts colony health. In the work funded by this proposal, use of this technique is expected to increase the understanding of the course of infection in honey bees. This new system can also be used to increase knowledge of the biology of other intracellular pathogens in bees or other animals. AS such, this project serves the NSF mission to promote the progress of science. In turn, findings from using this technique can be the first step in identifying means to cure or prevent infections. Thus, this technology can be used in the bioeconomy by researchers who want to stabilize honey bee populations and pollination.The study of obligate intracellular pathogens is difficult because they cannot easily be reproduced outside the host in axenic conditions. Many avenues of inquiry necessitate that cells of the pathogen be separated from those of the host. In addition, to study intracellular pathogen species with complex life cycles, it is critical individual life stages can also be separated. The experimental challenges of such manipulations and separations are compounded in situations where both host and pathogen are non-model organisms for which there are limited or no species-specific molecular tools. The obstacles presented can be addressed using flow cytometry-based techniques for relative quantification and localization of different cell types from honey bees that contain different life stages of the Nosema ceranae microsporidian, a key pathogen of honey bees. Specialized cell dyes measuring general cell features, in conjunction with flow cytometry to isolate distinct cell populations, are expected to develop a new system to advance the understanding of host-pathogen interactions and infection dynamics. Establishment of this technique promises to facilitate a number of new directions in microsporidia research. As some of the cellular features assessed by this technique are widespread in eukaryotes, there is a high probability that the technique can be adapted for use with other pathogen species from diverse phylogenetic groups alone or in combination with other available dyes. This proposal will also train an undergraduate student in microbiology research. As such, this proposal is training the next generation of the science workforce.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

一些微生物病原体是强制性细胞内病原体。它们很难研究，因为它们的宿主细胞内部发生了复杂的生命周期。在其他生物学领域，一种称为荧光活化细胞分选（FACS）的技术用于鉴定和将不同的细胞类型与基于差异荧光染色的近似混合物分离到近乎均匀性。该建议将使用该技术来测量和分离特定寄生虫（Nosema ceranae）的不同生命阶段，该寄生虫感染蜜蜂并对菌落健康产生负面影响。在该提案资助的工作中，预计该技术的使用将增加对蜜蜂感染过程的理解。该新系统也可用于增加蜜蜂或其他动物中其他细胞内病原体的生物学知识。因此，该项目为NSF促进科学进步的任务服务。 反过来，使用此技术的发现可能是识别治愈或防止感染的手段的第一步。因此，这项技术可以由想要稳定蜜蜂种群和授粉的研究人员在生物经济学中使用。强制性细胞内病原体的研究很困难，因为在轴承条件下，它们不能轻易地在宿主之外复制。许多查询途径都必须将病原体的细胞与宿主分离。此外，为了研究具有复杂生命周期的细胞内病原体物种，也可以分离至关重要的个体生命阶段。在宿主和病原体都是非模型生物的情况下，这种操纵和分离的实验挑战是复杂的，而这些生物的生物有限或没有物种特异性的分子工具。可以使用基于流式细胞术的技术来解决呈现的障碍，以相对定量和蜜蜂的不同细胞类型的定位，这些蜂蜜蜜蜂包含不同的生命阶段，蜜蜂是蜜蜂的关键病原体。预计，测量一般细胞特征的专门细胞染料与流式细胞仪结合使用，以分离不同的细胞群体，有望开发出一种新系统，以提高对宿主 - 病原体相互作用和感染动态的理解。建立该技术有望促进微孢子虫研究中的许多新方向。由于该技术评估的某些细胞特征在真核生物中广泛存在，因此，该技术很有可能与单独的系统发育基团或与其他可用染料结合的其他病原体进行调整。该建议还将培训一名微生物研究的本科生。因此，该提案正在培训下一代科学劳动力。该奖项反映了NSF的法定任务，并被认为是值得通过基金会的知识分子优点和更广泛影响的评论标准来评估值得支持的。