P-612 - Enhancing RNAi-Based Control of Grapevine Red Blotch Virus Using Spray-Induced Gene Silencing

Approaches to control grapevine red blotch virus (GRBV) are limited, posing a significant challenge to viticulture, with costly vine removal being the only available option to date. RNA interference (RNAi) is a conserved plant defense mechanism triggered by viral infection, but its role in GRBV suppression remains poorly documented. RNAi can be enhanced through spray-induced gene silencing (SIGS), where double-stranded RNA (dsRNA) targeting the pathogen is ectopically applied to the plant, offering a non-transgenic and sustainable solution for viral control.

Our previous work identified nine GRBV genomic hotspots targeted by RNAi during early infection. Based on these findings, we evaluated the silencing efficacy of hotspot-derived dsRNA (HS-dsRNA) in GRBV(+) tissue-cultured plants, and showed a significant reduction in viral gene expression at 12- and 24-days post-application. We further investigated systemic uptake and silencing potential in greenhouse plants. Fluoro-labeled HS-dsRNA, applied via low-pressure spray, was detected in vascular bundles within 48 hours, with systemic movement observed through 7 and 14 days.

To assess silencing efficiency, we treated GRBV(+) plants with HS-dsRNA via spray, and viral gene expression was quantified by qPCR at 1, 2, and 3 months, with long-term evaluations (6–12 months) ongoing to determine application frequency for sustained suppression.

These results provide the first experimental validation of dsRNA-induced RNAi for viral suppression in grapevine and demonstrate the potential of SIGS as a sustainable GRBV control strategy.