两种被忽视的正布尼亚病毒（Oya 病毒和伊宁市 Lake 病毒）的 RT-qPCR 检测方法的建立与评估。
Development and evaluation of RT-qPCR assays for two neglected orthobunyaviruses: Oya virus and Ebinur Lake virus.
机构信息
Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei, China; University of Chinese Academy of Sciences, Beijing, China.
Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei, China; University of Chinese Academy of Sciences, Beijing, China; Sino-Africa Joint Research Center, Chinese Academy of Sciences, Wuhan, Hubei, China.
出版信息
Virus Res. 2024 Jan 2;339:199265. doi: 10.1016/j.virusres.2023.199265. Epub 2023 Nov 16.
OBJECTIVES
Oya virus (OYAV) and Ebinur lake virus (EBIV) belong to the genus Orthobunyavirus within the Peribunyaviridae family, and both are recognized as the novel virus with potential threat to the animal or public health. Given their potential to cause outbreaks and their detection in diverse samples across different regions, the need for a reliable and efficient molecular detection method for OYAV and EBIV becomes imperative.
METHODS
The S-segment of OYAV and EBIV was used for designing specific primer and probe sets, which were employed in a real-time reverse transcription quantitative PCR (RT-qPCR) assay. The analytical performance of these assays, encompassing specificity, sensitivity, reproducibility, and fitness for purpose, was thoroughly evaluated across various sample matrices.
RESULTS
The developed RT-qPCR assays were very specific to their respective targets. Both assays were highly reproducible (%CV<3) and sensitive with the 95% limit of detection (LOD) of 0.80 PFU/mL for OYAV primer probe set and 0.37 PFU/mL for EBIV primer probe set. Furthermore, the assays fitness for purpose was good as it could detect the specific viruses in virus-spiked serum samples, virus-inoculated mosquito samples, field caught mosquitoes and biting midge samples.
CONCLUSIONS
Our study has successfully developed specific, sensitive, and reliable RT-qPCR assays for the detection of OYAV and EBIV. These assays hold great promise for their potential application in clinical and field samples in the future.
目的
Oya 病毒（OYAV）和伊宁市湖病毒（EBIV）属于布尼亚病毒科 Peribunyaviridae 属中的正布尼亚病毒属，两者均被认为是对动物或公共卫生具有潜在威胁的新型病毒。鉴于它们有可能引发疫情，并且在不同地区的各种样本中都有检测到，因此需要一种可靠且高效的 OYAV 和 EBIV 分子检测方法。
方法
使用 OYAV 和 EBIV 的 S 片段设计了特异性引物和探针集，用于实时逆转录定量 PCR（RT-qPCR）检测。通过对各种样本基质进行全面评估，对这些检测方法的分析性能（包括特异性、灵敏度、重现性和适用性）进行了评估。
结果
开发的 RT-qPCR 检测方法对其各自的靶标具有很高的特异性。两种检测方法均具有高度重现性（%CV<3）和灵敏度，OYAV 引物探针组的 95%检测限（LOD）为 0.80 噬菌斑形成单位/毫升，EBIV 引物探针组的 LOD 为 0.37 噬菌斑形成单位/毫升。此外，这些检测方法的适用性良好，因为它们可以在病毒感染的血清样本、病毒接种的蚊子样本、野外捕获的蚊子和吸血蠓样本中检测到特定的病毒。
结论
我们的研究成功开发了用于检测 OYAV 和 EBIV 的特异性、灵敏性和可靠的 RT-qPCR 检测方法。这些检测方法在未来的临床和现场样本中具有很大的应用潜力。
Zotero 插件