A Closterovirus from a Stem-Pitting-Diseased Grapevine

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Abstract
A closterovirus was isolated in Nicotiana clevelandii following mechanical inoculation from a grapevine with stem-pitting symptoms. The virus was easily mechanically transmissible thereafter between N. clevelandii plants, in which it caused vein clearing and stunting. Leaf crinkle symptoms appeared in N. megalosiphon, but the virus was not transmitted to 22 other herbaceous plant spp. In N. clevelandii sap, the virus lost infectivity when heated to 50.degree. C for 10 min, diluted with water to 10-5, or stored for 6 days at 20.degree. C or for 15 days at 5.degree. C. The virus was not seed transmitted in N. clevelandii, and was not transmitted by Myzus persicae or Macrosiphum euphorbiae. It was purified best by extraction in 0.05 M tris-HCl buffer, pH 7.8, containing 0.01 M MgSO4, clarification with bentonite, precipitation with polyethylene glycol and rate-zonal density gradient centrifugation in Cs2SO4. In uranyl acetate negative stain, the very flexous particles had a diameter of 11-12 nm, a clearly visible pitch of 3.6-4.0 nm, and a modal length near 800 nm. The modal length was similar in neutral sodium phosphotungstate. The particle structure was typical of closteroviruses. Three antisera (bleedings from 1 rabbit) gave titers up to 1/64 in slide precipitin tests, and 1/512 in immunoelectron microscopic decoration tests. The virus was not serologically related to apple chlorotic leafspot, beet yellows, carnation yellow fleck, heracleum latent, lilac chlorotic leafspot, apple stem grooving, or potato T viruses.