Phylogenetic analysis of viruses in Tuscan Vitis vinifera sylvestris (Gmeli) Hegi

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Abstract
The health status of the native grapevine Vitis vinifera subsp. sylvestris (Gmeli) Hegi in natural areas in Europe has received little attention. A survey was carried out on wild grapevines in Tuscany (Italy), where isolates of the Grapevine rupestris stem pitting virus (GRSPaV), Grapevine leafroll-associated virus 1 and 3 (GLRaV-1 and GLRaV-3) and Grapevine virus A (GVA) were detected. The complete coat protein (CP) region of these isolates was sequenced to investigate the relationship of the viral variants from Tuscan wild grapevines with isolates from different geographical origins. According to the phylogenetic analyses, GLRaV-1 and GLRaV-3 isolates from Tuscan wild grapevines clustered with isolates from cultivated grapevines with nucleotide sequence identities ranging from 66% to 87% and from 72.5% to 99% respectively, without any correlation between the distribution and geographical origin. Conversely, GRSPaV and GVA isolates clustered together with other Italian isolates from V. vinifera with nucleotide sequence identities ranging from 71.14% to 96.12% and from 73.5% to 92%, respectively. Our analysis of the whole amino acid sequences revealed a high conservation level for the studied proteins explained by a selective pressure on this genomic region, probably due to functional constraints imposed on CP, such as specific interactions with cellular receptors in the insect vectors necessary for successful transmission. In addition, analyses of genetic recombination suggest no significant point mutations that might play a significant role in genetic diversification. The dN/dS ratio also estimated a low number of non-silent mutations, highlighting the purifying selective pressure. The widespread distribution of the Rugose wood complex (GRSPaV and GVA associated disease) in comparison with the Grapevine Leafroll associated viruses (GLRaV-1 and -3) could explain the major geographical correlation found for the viral variants detected in Tuscany.

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