Alson H. Smith Jr. Agricultural Research and Extension Center, Virginia Polytechnic Institute and State University (Virginia Tech), Winchester, VA 22602, USA.
Department of Law and Economics, Shinshu University, Nagano 390-8621, Japan.
Viruses. 2023 Oct 17;15(10):2102. doi: 10.3390/v15102102.
We developed a NitroPure Nitrocellulose (NPN) membrane-based method for sampling and storing grapevine sap for grapevine virus detection. We devised an efficient nucleic acid extraction method for the NPN membrane, resulting in 100% amplification success for grapevine leafroll-associated virus 2 (GLRaV2) and 3 (GLRaV3), grapevine rupestris stem pitting-associated virus (GRSPaV), grapevine virus A, grapevine virus B, and grapevine red blotch virus (GRBV). This method also allowed the storage of recoverable nucleic acid for 18 months at room temperature. We created a sampling kit to survey GLRaV2, GLRaV3, and GRBV in Japanese vineyards. We tested the kits in the field in 2018 and then conducted mail-in surveys in 2020-2021. The results showed a substantial prevalence of GLRaV3, with 48.5% of 132 sampled vines being positive. On the other hand, only 3% of samples tested positive for GLRaV2 and none for GRBV.
我们开发了一种基于 NitroPure 硝酸纤维素（NPN）膜的方法，用于采样和储存葡萄藤汁液，以检测葡萄病毒。我们设计了一种针对 NPN 膜的高效核酸提取方法，该方法使 100%成功扩增了葡萄卷叶伴随病毒 2（GLRaV2）和 3（GLRaV3）、葡萄根瘤蚜伴随病毒（GRSPaV）、葡萄病毒 A、葡萄病毒 B 和葡萄红斑病毒（GRBV）。该方法还允许在室温下储存可回收核酸长达 18 个月。我们创建了一个采样试剂盒，用于调查日本葡萄园中的 GLRaV2、GLRaV3 和 GRBV。我们于 2018 年在实地测试了这些试剂盒，然后在 2020-2021 年进行了邮件调查。结果显示 GLRaV3 的流行率很高，132 株采样葡萄中有 48.5%呈阳性。另一方面，只有 3%的 GLRaV2 样本呈阳性，没有 GRBV 阳性样本。
