Serological Detection of Grapevine Associated Closteroviruses in Infected Grapevine Cultivars

Western blot immunoassay and enzyme-linked immunosorbent assay using different monoclonal antibodies (MAb) and polyclonal antisera (PA) revealed mixed infections of serologically related and unrelated grapevine leafroll associated viruses (GLRaVs) and grapevine corky bark associated virus (GCBaV) in symptomatic grapevines. A PA designated rootstock-scion incompatibility (RSI)-24 kDa, grapevine corky bark PA, and GLRaV-2b MAb reacted to polypeptides of approximately 24 kDa isolated from grapevines exhibiting rootstock-scion incompatibility, leafroll, and corky bark disease symptoms, suggesting that these isolates are infected with closely related viruses. A PA designated GLRaV-2 US detected virus specific polypeptides of 38, 37, 36, and 24 kDa, while a polyclonal antiserum designated GLRaV-2 FR detected a single virus-specific polypeptide of approximately 24 kDa. The reactivity of GLRaV-2 US to various polypeptides suggests that the immunogen used to produce this antiserum was a mixture of viruses. Apical meristems were excised and cultured to eliminate the infection of viruses in the grapevines showing RSI symptoms and in the cultivar French Colombard infected with GLRaV-1. The elimination of these viruses was confirmed by Western blot assay. These studies show that the Western blot assay can be used to detect and differentiate grapevine disease-associated closteroviruses.