Zhejiang Academy of Science and Technology for Inspection and Quarantine, Hangzhou, 310016, China.
College of Animal Science and Technology, Zhejiang Agriculture & Forestry University, Hangzhou, 311300, China.
BMC Vet Res. 2024 Sep 28;20(1):440. doi: 10.1186/s12917-024-04245-y.
Nipah virus (NiV) is a zoonotic pathogen that poses a significant threat because of its wide host range, multiple transmission modes, high transmissibility, and high mortality rates, affecting both human health and animal husbandry. In this study, we developed a one-step reverse transcription droplet digital PCR (RT-ddPCR) assay that targets the N gene of NiV.
Our RT-ddPCR assay exhibited remarkable sensitivity, with a lower limit of detection of 6.91 copies/reaction. Importantly, it displayed no cross-reactivity with the other 13 common viruses and consistently delivered reliable results with a coefficient of variation below 10% across different concentrations. To validate the effectiveness of our RT-ddPCR assay, we detected 75 NiV armored RNA virus samples, mimicking real-world conditions, and negative control samples, and the RT-ddPCR results perfectly matched the simulated results. Furthermore, compared with a standard quantitative real-time PCR (qPCR) assay, our RT-ddPCR assay demonstrated greater stability when handling complex matrices with low viral loads.
These findings show that our NiV RT-ddPCR assay is exceptionally sensitive and provides a robust tool for quantitatively detecting NiV, particularly in stimulated field samples with low viral loads or complex matrices. This advancement has significant implications for early NiV monitoring, safeguarding human health and safety, and advancing animal husbandry practices.
尼帕病毒（NiV）是一种人畜共患病病原体，由于其广泛的宿主范围、多种传播模式、高传染性和高死亡率，对人类健康和畜牧业构成重大威胁。在本研究中，我们开发了一种针对 NiV 的 N 基因的一步法逆转录液滴数字 PCR（RT-ddPCR）检测方法。
我们的 RT-ddPCR 检测方法表现出出色的灵敏度，检测下限为 6.91 拷贝/反应。重要的是，它与其他 13 种常见病毒没有交叉反应，并且在不同浓度下始终提供变异系数低于 10%的可靠结果。为了验证我们的 RT-ddPCR 检测方法的有效性，我们检测了 75 个模拟实际情况的 NiV 装甲 RNA 病毒样本和阴性对照样本，RT-ddPCR 结果与模拟结果完全匹配。此外，与标准定量实时 PCR（qPCR）检测方法相比，我们的 RT-ddPCR 检测方法在处理低病毒载量或复杂基质的复杂样本时表现出更高的稳定性。
这些发现表明，我们的 NiV RT-ddPCR 检测方法具有极高的灵敏度，为定量检测 NiV 提供了一种强大的工具，特别是在低病毒载量或复杂基质的现场样本中。这一进展对早期 NiV 监测具有重要意义，有助于保护人类健康和安全，并推进畜牧业实践。
