A diagnostic tool to determine whether grapevines are infected with the viral Grapevine red blotch virus (GRBV) has been developed by University of California researchers. Dr. Monica Cooper, UCCE viticulture advisor, noted that early detection of this incurable grapevine disease allows for removal of infected vines and reduces the level of inoculum in the vineyard.
Grapevine red blotch disease was first detected at the UC Davis Oakville research station in 2007 but was not formally identified until 2012. Cooper, speaking in a UC Experts Talk webinar, said the disease may have evolved in California from a latent virus found in native grapes. The virus infects plants in the Vitaceae family, including cultivated grape varieties. The disease has been found in many wine grape varieties and is believed to infect table grapes, raisin grapes and rootstock.
Vines infected with GRBV show symptoms similar to vine leafroll disease, with leaves turning red in early fall, primarily at the base of shoots. Unlike leafroll, red blotchinfected vines have red or pink veins in the leaves, red blotches on the leaves and no leafroll. Visual mapping of infected vines can be tricky, Cooper said, because symptoms can vary by cultivar, location or season.
Disease impacts fruit quality
Infected vines have an economic impact on vineyards, producing fruit that is often unsuitable for market. The disease reduces sugar accumulation, increases malic acid and less consistently increases pH and titratable acidity. Cluster weight can be reduced.
Cooper noted there are two ways the disease spreads in vineyards: through grafts using infected plant material or via the threecornered alfalfa hopper (Spissistilus festinus). Currently the only treatment is vine removal.
In the webinar, Cooper discussed the loopmediated isothermal amplification (LAMP) tool. Growers, vineyard managers or crop consultants can use this inhome assay to detect the presence of GRBV in vines without sending samples to a diagnostic lab. The LAMP tool detects and amplifies viral DNA from GRBVinfected vines. The amplification causes a color change used to interpret results.
Sampling steps
Plant material can be collected from petioles, canes or vine trunks. Petioles can be sampled for the LAMP assay from veraison to just before presenescence. Once leaves begin to yellow, they are not suitable sample material. Canes can be sampled from presenescence to pruning. Cooper said canederived samples can be stored in a freezer for a short time, and petiole samples can be refrigerated for up to a week before use. Trunk material can be sampled at any time during the year.
Location on the plant where the sample is taken matters. Cooper noted that GRBV is unevenly distributed within grapevines. Studies have shown that basal tissue or older tissue is more reliable and less likely to result in false negatives. For example, samples should be taken from both sides of a bilateral cordon. She recommended collecting samples from multiple locations on a vine for best representation of disease status.
With trunk or cane material, Cooper said to peel back the outer bark and use a pipette to collect a sample from the vascular tissue. The sample placed in a test tube serves as the sample template. The next step is to mix the reagents and dispense them into PCR tubes. The reagents include primers, distilled water and Master Mix, which is used to elicit the color change. The final step is to add the samples to each tube and close them tightly. The tubes are then placed in a heat block for about 35 minutes. During that time the amplification takes place, resulting in the color change.
Once removed from the heat block, any tube with a yellow color indicates the sample is positive for GRBV. All negative controls remain pink.
This method of testing grapevine tissue samples for the presence of GRBV requires attention to detail, Cooper said, because there is a high risk of contamination. Cleaning workspaces and all equipment thoroughly with a bleach solution can help prevent contamination. Cooper noted there is a learning curve with this testing method, but with careful technique it can be mastered.
