Anti-GLRaV-1 coat protein polyclonal antibody

Grapevine leafroll-associated virus 1 (GLRaV-1) and Grapevine rupestris stem pitting-associated virus (GRSPaV) were eradicated from Vitis vinifera L cv. Agiorgitiko by combining in vitro thermotherapy and tissue culture. GRSPaV is known to be quite recalcitrant to elimination, whereas GLRaV-1 is more easily knocked out. In this study, the effectiveness of two different virus elimination techniques, including meristem- and shoot-tip culture, was evaluated. Results showed that meristem-tip culture combined with thermotherapy was the most effective for eliminating both viruses as confirmed by nested RT-PCR assays. Success rate for GLRaV-1 (91.2%) was higher than for GRSPaV-1 (67.6%). The ratio of virus elimination to survival was higher for meristem-tip culture than for shoot tips (1.108 and 0.469 respectively). The effect of six basal media on in vitro shoot proliferation of virus-free explants of cv. Agiorgitiko was also studied and woody plant medium (WPM) proved to be the most effective. The presence of cytokinin 6-benzyladenine (BA) alone resulted in chlorotic plantlets, while Supplementation with the auxin naphthaleneacetic acid (NAA) enhanced proliferation rate. Root induction at two temperature regimes (22 +/- 2 degrees C and 26 +/- 21 degrees C) revealed that higher temperature was more effective in the presence of IBA (indole-3-butyric acid) rather than NAA.

The genetic diversity of 34 isolates of Grapevine leafroll-associated virus 1 (GLRaV-1) from different wine, table, and ornamental grape cultivars in California, New York, and Washington States in the United States was investigated. Segments of the heat-shock protein 70 homolog (HSP70h) gene, coat protein (CP) gene, coat protein duplicate 2 (CPd2) gene, and open reading frame 9 (p24) were amplified by reverse-transcription polymerase chain reaction, cloned, and sequenced. A pairwise comparison of nucleotide sequences revealed intra- and interisolate sequence diversity, with CPd2 and HSP70h being the most and the least divergent, respectively, among the four genomic regions studied. The normalized values for the ratio of nonsynonymous substitutions per nonsynonymous site to synonymous substitutions per synonymous site indicated different purifying selection pressures acting on each of the four genomic regions, with the CP and CPd2 being subjected to the strongest and weakest functional constraints, respectively. A global phylogenetic analysis of sequences from the four genomic regions revealed segregation of GLRaV-1 isolates into three major clades and a lack of clearly defined clustering by geographical origin. In contrast, only two lineages were apparent when the CP and CPd2 gene sequences were used in phylogenetic analyses. Putative recombination events were revealed among the HSP70h, CP, and p24 sequences. The genetic landscape of GLRaV-1 populations presented in this study provides a foundation for better understanding of the epidemiology of grapevine leafroll disease across grape-growing regions in the United States. In addition, this study will benefit grape clean plant programs across the country in improving the sanitary status of planting materials provided to nurseries and grape growers.