使用Cas13和多个CRISPRRNAs无放大检测动物感染病毒
Caitlin H Lamb1, Aartjan J W Te Velthuis1, Cameron Myhrvold1,2,3,4
1Department of Molecular Biology, Princeton University, Princeton, NJ 08544.
bioRxiv : the preprint server for biology
|June 4, 2025
概括
一种新的CRISPR-Cas13测定方法提供了一种快速,灵敏和具有成本效益的方法,用于检测动物传播病毒,如汉塔病毒和流感A病毒,而无需放大. 这种可在现场部署的工具显示出在各种样本中早期检测感染的前景.
科学领域:
- 分子生物学分子生物学
- 病毒学 病毒学
- 生物技术是生物技术.
背景情况:
- 动物传播病毒,包括汉塔病毒和A型流感病毒,对人类和动物健康构成重大风险.
- 目前对这些病毒的诊断方法可能是缓慢的,昂贵的,或需要专门的实验室设备.
- 迫切需要快速,灵敏和易于使用的诊断工具来早期检测动物性病毒感染.
研究的目的:
- 开发和评估一种基于CRISPR-Cas13的无放大测定方法,用于检测汉塔病毒和A型流感病毒RNA.
- 评估使用多个CRISPRRNAs (crRNAs) 对无放大测定灵敏度的影响.
- 确定使用这种试验在实验室和临床样本中检测病毒RNA的可行性.
主要方法:
- 一种无放大CRISPR-Cas13测定被设计用于检测汉塔病毒 (图拉病毒 - TULV,安第斯病毒 - ANDV) 和流感A病毒的基因组RNA.
- 测试的灵敏度是使用不同病毒的不同数量的crRNA测试的.
- 用组织培养样本和临床鼻抽样中的病毒RNA验证了试验的性能.
主要成果:
- 对于UTLV和ANDV检测,使用两个或三个crRNA显著提高了无放大试验的灵敏度.
- 额外的crRNAs没有提高流感A病毒RNA检测的灵敏度.
- 在临床样本中,CRISPR-Cas13测定成功检测了细胞培养中的TULV和ANDVRNA以及临床样本中的流感A病毒RNA,显示了与RT-qPCR对阳性样本鉴定的85%一致性.
结论:
- 无放大CRISPR-Cas13技术为动物性病毒RNA的快速和敏感检测提供了一个可行的策略.
- 优化crRNA组合可以提高针对特定病毒点的测试灵敏度.
- 这种测定具有作为现场部署的诊断工具的潜力,用于早期识别动物传播病毒感染,帮助公共卫生监测和应对.
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