Fan Xudong, Zhang Zunping, Ren Fang, Hu Guojun, Li Chen, Zhang Baodong, Dong Yafeng
National Center for Eliminating Viruses from Deciduous Fruit Trees, Research Institute of Pomology, Chinese Academy of Agriculture Sciences, Xingcheng 125100, China.
Plants (Basel). 2020 Oct 11;9(10):1340. doi: 10.3390/plants9101340.
Grapevine berry inner necrosis virus (GINV) belongs to the genus Trichovirus in the family Betaflexiviridae. The GINV isolate LN_BETA_RS was obtained from a "Beta" grapevine () exhibiting chlorotic mottling and ring spot in Xingcheng, Liaoning Province, China. To verify the correlation between GINV and grapevine chlorotic mottling and ring spot disease, we constructed an infectious cDNA clone of GINV isolate LN_BETA_RS using the seamless assembly approach. Applied treatments of agroinfiltration infectious cDNA confirmed systemic GINV infection of the 37B by reverse transcription polymerase chain reaction (RT-PCR) and transmission electron microscopy, exhibiting chlorotic mottling symptoms on leaves. Infectious cDNA was also transmitted to new healthy plants through rub-inoculation. Moreover, the cDNA clone was agroinfiltrated into "Beta" and "Thompson Seedless" grapevine plantlets, and the inoculated grapevines exhibited leaf chlorotic mottling and ringspot during the two years of observation. GINV-inoculated "Beta" grapevines had serious leaf chlorotic mottling and ringspot symptoms on the whole plant, while relatively few symptoms were observed on the leaves of agroinoculated "Thompson Seedless" grapevines in early spring and only weak ring spot gradually appeared later in the top young leaves. Our experiments fulfilled Koch's postulates and revealed the causative role of GINV in grapevine chlorotic mottling and ring spot disease.
葡萄浆果内部坏死病毒（GINV）属于β-柔线病毒科的毛形病毒属。GINV分离株LN_BETA_RS取自中国辽宁省兴城一株表现出褪绿斑驳和环斑症状的“贝塔”葡萄（）。为了验证GINV与葡萄褪绿斑驳和环斑病之间的相关性，我们采用无缝组装方法构建了GINV分离株LN_BETA_RS的感染性cDNA克隆。通过农杆菌浸润感染性cDNA的应用处理，利用逆转录聚合酶链反应（RT-PCR）和透射电子显微镜证实了37B葡萄植株被GINV系统感染，叶片上出现了褪绿斑驳症状。感染性cDNA也通过摩擦接种传播到新的健康葡萄植株上。此外，将该cDNA克隆农杆菌浸润接种到“贝塔”和“汤普森无核”葡萄幼苗中，在两年的观察期内，接种的葡萄植株均表现出叶片褪绿斑驳和环斑症状。接种GINV的“贝塔”葡萄植株整株叶片出现严重的褪绿斑驳和环斑症状，而农杆菌接种的“汤普森无核”葡萄植株在早春时叶片上观察到的症状相对较少，仅在顶部幼叶上后期逐渐出现轻微的环斑。我们的实验满足了科赫法则，揭示了GINV在葡萄褪绿斑驳和环斑病中的致病作用。
