2005 Annual Report 4d.Progress report. This report serves to document research conducted under a specific cooperative agreement between ARS and Washington State University. Additional details of research can be found in the report for the parent project 5358-21000-034-00D Production Systems to Promote Yield and Quality of Grapes in the Pacific Northwest. Project objective: Determine the impact of leaf roll disease on Vitis vinifera L. horticultural traits as indicated by vineyard productivity and fruit quality; vegetative growth; fruit set; fruit maturation; fruit quality at harvest; and vine cold hardiness. This will be related to the development of cytopathology associated with disease. Progress to date: A vineyard was established on-site at Washington State University - Prosser. The research block contains the wine grape cultivars ‘Lemberger’, ‘Merlot’ and ‘Pinot noir’. The ‘Merlot’ section contains both virus-free vines and vines infected with GLRaV-3. The ‘Lemberger’ block contains virus-free vines and those infected with GLRaV-1 and/or -3. Since these are mature, fruiting vines and the virus status of each vine has been determined, measurement of physiological parameters can be initiated immediately. The non-infected ‘Merlot’ vines in this block have been used in the development of baseline measurements of the effect of heating and cooling on berry development (Tarara et al., Am. J. Enol. Vitic. 51(2000)182). This method will be applied to measure the effect of viruses under different conditions imposed by the chamber-free environmental control system. We have confirmed the presence of GLRaV-8 in the ‘Pinot noir’ block. This is the first identification of GLRaV-8 in Washington. We have developed a molecular assay that is being used for the detection of this virus. These vines provide source material for the further characterization of the virus and for the development of more robust assay systems. The development of an ELISA for GLRaV-8 is advanced. This study will examine and characterize the cytopathology induced by virus infection. Phloem and companion tissues of plants affected by leafroll disease contain vesicles characteristic of virus infection; the affected cells become dysfunctional. The link between the development of vesicles and physiological events of the vine has not been established. We have identified three candidate genes within the genome of our local strain of GLRaV-3 that could produce proteins associated with vesicles. The gene for each of these non-structural proteins were cloned and sequenced, and the gene inserted into a bacterial expression vector to enable the production and recovery of significant amounts of the protein. Antibodies have been produced to these proteins and the antibodies will be used for the immunolocalization of the viral proteins within diseased cells. The accumulation and distribution of viral proteins will be related to physiological events within diseased grapevines, including the loss of phloem function and arrested fruit development and maturation. The vines in this research plot have been used to demonstrate that infection of ‘Merlot’ by GLRaV-3 increases its susceptibility to cold injury. Data from the block provided growers with a clear demonstration of another dimension of leaf roll disease and its deleterious effects on grape production. Extension of this project is the basis of a Ph.D. student graduate research program. A recent graduate from a plant virology Master’s program has been recruited and has commenced his Ph.D. program in the Washington State University Department of Plant Pathology. His research will examine the physiological and horticultural effects of leaf roll disease on grapevines using the research block described above and the tools that have been developed in the early stages of this program.