Analysis of grapevine leafroll-associated virus 3 (GLRav-3) isolates by single-strand conformation polymorphism (SSCP)

Project Number
WW 07-12

Project title
Analysis of grapevine leafroll-associated virus 3 (GLRav-3) isolates by single-strand conformation polymorphism (SSCP)

Project leader
Goszczynski, D E

Institution
ARC Plant Protection Research Institute

Team members
Jooste, A E C
Goszczynski, T
Kasdorf, G G F

Project description
GLRaV-3 is one of the most widespread closteroviruses in vineyards worldwide and is considered to be a genuine agent of grapevine leafroll disease. The disease affects the quality of vines by delaying ripening of grapes and decreasing the accumulation of sugar. As GLRaV-3 is transmitted by insect vectors, the South African grapevine industry experiences a serious problem of re-infection of virus-free propagation material. Up to date, five insect vectors capable of transmitting GLRaV-3 have been identified. Of these, mealybug Planococcus ficus is the most common in South African vineyards.

Although there is no scientific data, it seems that re-infection of leafroll disease is especially fast in South Africa, and this probably relates to spreading of GLRaV-3. There can be many reasons for this situation, but the most probable is the existence of an, as yet, unidentified insect vector in South African vineyards, especially effective in GLRaV-3 transmission, or the existence of strains of GLRaV-3, which are especially efficiently transmitted by mealybugs. We also cannot rule out the possibility that certain strains of GLRaV-3 survive standard virus elimination procedure used by the industry.

This illustrates the importance of closer analysis of GLRaV-3 isolates to get firm data on whether we are dealing with different strains of this virus in South African vineyards. There are reports suggesting the existence of ?mild? and ?severe? strains of GLRaV-3. Single strand conformation polymorphism (SSCP) is an excellent technique, which allows quick detection of differences in nucleotide sequences of viruses. The technique includes standard RT-PCR, special treatment of amplified product and electrophoresis; thus it can be applied for analysis of many virus isolates with relatively low cost. Single-strand conformation polymorphism (SSCP) analysis of GLRaV-3 isolates will be done to determine whether nucleotide sequence variants of GLRaV-3 exists in South African vineyards. If sequence variants of GLRaV-3 are found, the project will be extended to SSCP study of GLRaV-3 isolates from different vineyards, focusing on farms where spread of leafroll is especially fast. Representatives of sequence variants will be cloned and primers for their specific detection by RT-PCR designed.

Presentation(s)
Goszczynski, D E and Jooste, A E C. 2002. Molecular heterogeneity of virus associated with leafroll (LR), corky bark (CB) and Shiraz (SD) diseases of grapevines. Presentation at the Winetech Grapevine Virus Workshop, 6 May, Simondium, South Africa

Goszczynski, D E and Jooste, A E C. 2003. Are South African GLRaV-3 isolates molecularly unique? Presentation at the Winetech Grapevine Virus Workshop. 5 May, ARC Infruitec-Nietvoorbij, Stellenbosch, South Africa.

Goszczynski, D E and Jooste, A E C. 2004. Sequence results confirm the existence of two clearly divergent molecular groups of grapevine leafroll-associated virus 3 (GLRaV-3) in South African vineyards. Presentation at the Winetech Grapevine Virus Workshop. 11 May, ARC Infruitec-Nietvoorbij, Stellenbosch, South Africa

Goszczynski, D E and Jooste, A E C. 2004. Do some grapevine virus A (GVA) variants multiply under control of Shiraz and Merlot, and do those able to evade anti-virus defences induce Shiraz-disease? Presentation at the Winetech Grapevine Virus Workshop. 11 May, ARC Infruitec-Nietvoorbij, Stellenbosch, South Africa

Goszczynski, D E and Jooste, A E C. 2005. Further characterisation of the two divergent variants of GLRaV-3.Presentation at the Winetech Grapevine Virus Workshop. 11 May, ARC Infruitec-Nietvoorbij, Stellenbosch, South Africa

Article
Jooste, A E C, Goszczynski, D E. 2005. Single-strand conformation polymorphism (SSCP), cloning and sequencing reveals two major group of divergent molecular variants of grapevine leafroll-associated virus 3 (GLRaV-3) Vitis, v. 44 (1) (p. 39-43)

Meng, B, Li, C, Wang, W, Goszczynski, D E, Gonsalves, D. 2005. Complete sequences and structures of two new variants of grapevine rupestris stem pitting associated virus and comparative analyses, Journal of General Virology, v. 86 (p. 1555-1560)

Meng, B, Li, C, Goszczynski, D E, Gonsalves, D. 2005. Genome sequences and structures of two biologically distinct strains of grapevine leafroll-associated virus 2 and sequence analysis, Virus Genes, v. 31 (p. 31-41)

Haviv, S, Galiakparov, N, Goszczynski, D E, Batuman, O, Czosnek, H, Mawassi, M. 2006. Engineering the genome of grapevine virus A into a vector for expression of proteins in herbaceous plants, Journal of Virological methods, v. 132 (p. 227-231)

FinalReport.pdf

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