RT-PCR Based Detection of Rupestris stem pitting associated virus Within Field-Grown Grapevines Throughout the Year

Area of Science:

• Plant Virology
• Molecular Diagnostics
• Viticulture

Background:

• Rupestris stem pitting associated virus (RSPaV) often shows no symptoms unless other viruses are present.
• Reverse transcription-polymerase chain reaction (RT-PCR) is a detection method, but can be unreliable due to interfering compounds or low virus titers.
• Identifying reliable grapevine tissues for RSPaV detection is crucial for accurate diagnostics.

Purpose of the Study:

• To evaluate grapevine tissues for reliable virus detection using a new extraction method and internal control.
• To survey RSPaV presence in various tissues of different Vitis vinifera varieties and rootstocks in Ontario, Canada.
• To determine optimal tissue sources and sampling times for RSPaV detection.

Main Methods:

• Utilized a novel RNA extraction method with an internal control for RT-PCR assays.
• Collected samples from multiple grapevine tissues (buds, shoot tips, seeds, canes) of various cultivars and rootstocks.
• Conducted surveys across different seasons in Ontario, Canada.

Main Results:

• Amplifiable RNA extracts were consistently obtained from buds, shoot tips, seeds, and canes.
• RSPaV was detected in most tissues of infected plants, with exceptions in young summer buds.
• Dormant cane buds (three single buds) proved to be a reliable and less labor-intensive source for detection compared to cane shavings.

Conclusions:

• A new extraction method enhances the reliability of RT-PCR for RSPaV detection in grapevines.
• Dormant cane buds are recommended as a practical and effective tissue source for RSPaV diagnosis.
• The study provides valuable insights for grapevine virus disease management and certification programs.