Centro de Protección Vegetal y Biotecnologia, IVIA, Moncada, Valencia 46113, Spain.
Mol Plant Microbe Interact. 2011 Oct;24(10):1119-31. doi: 10.1094/MPMI-05-11-0110.
Citrus tristeza virus (CTV) naturally infects only some citrus species and relatives and within these it only invades phloem tissues. Failure to agroinfect citrus plants and the lack of an experimental herbaceous host hindered development of a workable genetic system. A full-genome cDNA of CTV isolate T36 was cloned in binary plasmids and was used to agroinfiltrate Nicotiana benthamiana leaves, with or without coinfiltration with plasmids expressing different silencing-suppressor proteins. A time course analysis in agroinfiltrated leaves indicated that CTV accumulates and moves cell-to-cell for at least three weeks postinoculation (wpi), and then, it moves systemically and infects the upper leaves with symptom expression. Silencing suppressors expedited systemic infection and often increased infectivity. In systemically infected Nicotiana benthamiana plants, CTV invaded first the phloem, but after 7 wpi, it was also found in other tissues and reached a high viral titer in upper leaves, thus allowing efficient transmission to citrus by stem-slash inoculation. Infected citrus plants showed the symptoms, virion morphology, and phloem restriction characteristic of the wild T36 isolate. Therefore, agroinfiltration of Nicotiana benthamiana provided the first experimental herbaceous host for CTV and an easy and efficient genetic system for this closterovirus.
甜橙衰退病毒（CTV）自然感染仅局限于一些柑橘属物种及其近缘种，而且仅侵染韧皮部组织。由于无法对柑橘属植物进行农杆菌感染，以及缺乏实验性的草本宿主，阻碍了一个可行的遗传系统的开发。CTV 分离物 T36 的全长 cDNA 被克隆到双元质粒中，并被用于农杆菌浸润烟草原生质体，浸润时可以或不与表达不同沉默抑制蛋白的质粒共浸润。在农杆菌浸润叶片中的时间进程分析表明，CTV 至少在接种后 3 周内积累并在细胞间移动，然后系统移动并在有症状表达的上部叶片中感染。沉默抑制蛋白加速了系统感染，并且常常增加了感染力。在系统感染的烟草原生质体植物中，CTV 首先侵染韧皮部，但在 7 周后，也在其他组织中发现了 CTV，并在上部叶片中达到高病毒滴度，从而允许通过茎劈接种将其有效传播至柑橘属植物。受感染的柑橘植物表现出与野生 T36 分离物相同的症状、病毒形态和韧皮部限制特征。因此，烟草原生质体的农杆菌浸润为 CTV 提供了第一个实验性草本宿主，以及用于该科病毒的简单而有效的遗传系统。
