Grapevine leafroll-associated virus 2 (GLRaV-2)- mechanical transmission, purification, production and properties of antisera, detection by ELISA

Grapevine leafroll-associated closterovirus 2 (GLRaV-2) was transmitted from grape cv. Muscat d'Alexandrie to Nicotiana benthamiana following inoculation with a grapevine leaf petiole extract conc. by ultracentrifugation. Virus-infected N. benthamiana showed symptoms of chlorotic local lesions, systemic vein clearing followed by yellowing, stem necrosis and death of the plant. The plants were only infected with GLRaV-2. Antisera produced to GLRaV-2 strongly decorated particles of the virus in immuno-EM and clearly and specifically detected GLRaV-2 in conc. extracts of infected grapevines in Western blots. The antisera were used with success for the specific and sensitive detection of GLRaV-2 by ELISA. Treatment of purified GLRaV-2 preparations with glutaraldehyde before immunization markedly improved the quality of the antisera