2006 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. Dr. Ken Eastwell and collaborators conducted the following research towards the agreement: 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; vine cold hardiness. This will be related to the development of cytopathology associated with disease. Background: Plants possess a natural defense mechanism by which they attempt to reduce the replication of viruses. Many viruses, in turn, developed various strategies for suppressing this response to insure their “survival”. Thus, the development of disease symptoms is a delicate balance between these compensating factors. Infection of grapevines with Rupestris stem pitting associated virus (RSPaV) is reported to increase the severity of leafroll disease in vineyards; this project explores this interaction. Early studies of RSPaV were significantly impaired by the extreme genetic variability of RSPaV isolates. A better understanding of this variability will be assessed during the initial stages of this project. Progress to date: Several field isolates of RSPaV were identified and the 3’-terminus of their genomes sequenced. A comparison of these sequences led to the identification of consensus sequences and the development of a reliable molecular assay (RT-PCR). The coat protein from a reference strain was expressed in bacteria and the resulting protein was purified and is currently being used to solicit the production of antibodies. Antibodies will be used to develop serological assays for detection of RSPaV since such assays are less sensitive to minor differences in gene sequences relative to RT-PCR. Antibodies and the techniques built upon them will be used throughout the project to confirm presence of virus and to examine the distribution of RSPaV within infected grape tissue. RSPaV is so named because of the development of wood cylinder abnormalities induced in Vitis rupestris. This virus-associated pitting is not evident in most infected wine grape (Vitis vinifera) selections. We have selected eight grape rootstocks that have V. rupestris in their parentage. These are being propagated and will be inoculated with RSPaV to determine if the development of wood cylinder abnormalities was inherited during the breeding process. Based on analogy to other members of the family Closteroviridae, we identified three genes in the genome of Grapevine leafroll associated virus 3 (GLRaV-3) that may modify the replication of RSPaV. Each of these genes has been cloned and the product of each will be assayed for its ability to interact with RSPaV. Antibodies produced to the proteins whose syntheses are directed by these genes will be used to localize the accumulation of gene products within diseased cells. A recent graduate from a plant virology Master’s program was recruited and began the Ph.D. program of study at Washington State University, Department of Plant Pathology in September 2005. His research program is based on the above project description.