Development of Agrobacterium tumefaciens Infiltration of Infectious Clones of Grapevine Geminivirus A Directly into Greenhouse-Grown Grapevine and Nicotiana benthamiana Plants

Grapevine virus infectious clones are important tools for fundamental studies, but also because of their potential for translational applications for grapevine improvement. Although several grapevine virus infectious clones have been developed, there has been difficulty in directly infecting mature grapevine plants, and many of the viruses used still cause disease symptoms in grapevine plants, making them less likely candidates for biotechnological applications in grapes. Here, we developed an improved Agrobacterium tumefaciens infiltration method that can be used to deliver DNA plasmids and viral infectious clones directly into approximately 20- to 40-cm-high (above soil) greenhouse-grown grapevine plants. We also developed infectious clones for two isolates of grapevine geminivirus A (GGVA): Longyan (China; GenBank accession KX570611; GGVA-76) and Super Hamburg (Japan; GenBank accession KX570610; GGVA-93). Neither virus caused any obvious symptoms when inoculated to plants of grapevine varieties Colombard, Salt Creek, Cabernet Sauvignon, and Vaccarese. However, the two GGVA isolates induced different symptom severity and viral titer in Nicotiana benthamiana plants. The two GGVA isolates used here were found to accumulate to different titers in different parts/branches of the infected grapevine plants. The GGVA infectious clones and the improved grapevine infiltration technique developed here provide new, valuable tools that can be applied to grapevine plants, possibly even for translational applications such as disease management and desired trait improvements.

Automated summary

Developed an improved injection method for delivering viral infectious clones into greenhouse-grown grapevine plants. Developed infectious clones for two isolates of grapevine geminivirus A and found that they accumulated to different viral titers in different parts of the infected grapevine plants. These tools can be applied to grapevine plants, potentially for disease management and trait improvements.