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Within the remaining 44 genotypes, a self-seedling of Cabernet Franc and first generation hybrids of V. vinifera were identified We were also able to identify the parent cultivar of six hybrids. Species designations were assigned to the 38 remaining unique genotypes by calculating the probability of a sample being V. vinifera rather than V. californica because there is no database of V. californica profiles. In this manner, a decreasing V. vinifera contribution was illustrated (Figure 1).
Results III: RT-PCR assays
The Vitis and non-Vitis samples collected in Fall 2008 were assayed using both conventional RT-PCR and realtime RT-PCR.
175 Vitis samples had positive CT values between 17.1 and 35.6, which for all samples indicated a nucleic acid amount within a 105-fold difference, relative to positive controls.
Results IV: Virus incidence
In six out of nine sites adjacent to vineyards (Sites 1, 5, 6, 7, 8, 9), all wild Vitis tested negative for the viruses included in this study. At three sites, (Sites 2, 3, 4), which were within 2 km of each other along the Napa River, 100% of the wild Vitis collected in 2008 tested positive for GVA and/or GLRaV-3 (samples circled in yellow in Figure 3). We returned to these three sites in 2010 and collected 164 additional samples; the incidence of GLRaV-3 was 50% to 80% in wild Vitis and 28% to 80% in V. vinifera within vineyards (Figure 3).
In the two sites not adjacent to vineyards (Sites 10 and 11), 4 out of 21 (19%) of the wild Vitis tested positive for virus. Three samples tested positive for all four viruses GLRaV- 2, -3, GVA, and GVB and one tested positive for GVA only (data not shown).
All Vitis samples tested negative for GLRaV-1, -4, -5, -9, and GVD. All non-Vitis tested negative for all viruses investigated.
Leafroll symptoms in wild vines were observed sometimes, but often not.
Results V: Viral sequences
Partial CP gene sequence data was collected for five GLRaV-2, 17 GLRaV- 3, five GVA, and five GVB isolates from non-cultivated Vitis and adjacent V. vinifera. The GLRaV-2 RT-PCR products were 88% to 96% identical at the nucleotide level to GLRaV-2 gene sequences in GenBank; GLRaV-3 products were 88% to 100% identical to GLRaV-3 gene sequences; GVA products were 91% to 93% identical to GVA gene sequences; and GVB products were 91% to 97% identical to GVB gene sequences.
A high incidence of GLRaV-3 and/ or GVA in V. californica and V. californica x V. vinifera hybrids was identified in one localized area of the Napa Valley, establishing that these wild Vitis are GLRaV-3 and GVA hosts.
Wild grapes at six other sites were not positive for the viruses included in this study, despite the fact that most sites had many wild vines within several meters of virus-infected vineyards. This suggests that GLRaV-3 and GVA incidence is not widespread in V. californica and hybrids in Napa Valley.
Analysis of GLRaV-3 gene sequences indicated that the GLRaV-3 isolates from wild and cultivated grapes were closely related. This suggests that these wild isolates probably do not represent a major source of diversity that could lead to major epidemics in V. vinifera.
GLRaV-2 and GVB were detected in wild grape samples collected in riparian areas not adjacent to vineyards. Analysis of the GLRaV-2 gene sequences suggested that GLRaV-2 was probably introduced into riparian areas relatively recently, via V vinifera infected with GLRaV-2.
GLRaV-2 is the only leafroll-associated virus included in the genus Closterovirus, whose other members are transmitted semi-persistently by aphids. No GLRaV-2 vectors have been identified, however, and spread within vineyards has not been observed.
Aphids are not common on grapes in California, but have been periodically reported. It is possible that they could be much more abundant in riparian areas and serve as a GLRaV-2 vector.
Heartfelt thanks to the American Vineyard Foundation and the North Coast Viticulture Research Group for funding this project; numerous growers in Napa County; K. Lowe and P. Saldivar for assistance in locating study sites; J. Roncoroni, J. DiTomaso, and E. Dean for plant identification assistance; J. Yang, FPS, for assistance in DNA identification; and numerous FPS staff for assistance in sample collection and testing.
For more information:
Klaassen, V.A., S.T. Sim, G.S. Dangl, F. Osman, M. Al Rwahnih, A. Rowhani, and D. Golino. 2011 Vitis californica and Vitis californica x Vitis vinifera hybrids are hosts for Grapevine leafroll-associated virus-2 and -3 and Grapevine virus A and B. Plant Dis. 95:657-665. online here.