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A Closterovirus (Family: Closteroviridae) Isolated from Tobacco Crops in Northern Greece (Macedonia)

In 1997, viruslike symptoms similar to those caused by potato potyvirus Y (PVY) were observed in tobacco (Nicotiana tabacum ) plants of the cv. Basmas in the area of Drama (Macedonia) in northern Greece. Diseased plants showed vein clearing and necrosis of the petioles and main veins, and were severely stunted. Serological tests with polyclonal antibodies prepared against PVY, cucumber mosaic cucumovirus (CMV), alfalfa mosaic alfamovirus (AMV), and tomato spotted wilt tospovirus (TSWV) gave negative results. The virus was mechanically transmitted from infected field tobacco plants to plants of N. tabacum cvs. White Burley, Samsun NN, Xanthi, Christie, and Basmas, and to plants of the species N. benthamiana, N. clevelandii, N. glutinosa, N. roselata, and N. rustica. All tobacco species and cultivars generally reacted with chlorotic local lesions on the inoculated leaves followed by systemic vein clearing, often veinal necrosis, leaf curling, and severe stunting. Similar symptoms were produced on Petunia hybrida. Other plant species such as beet, faba beans, lettuce, tomato, Datura stramonium, and watermelon were not infected by the virus. Aphid transmission studies with Myzus persicae showed that the virus was transmitted in a semi-persistent manner. Electron microscopic observations of leaf squash preparations from infected plants revealed the presence of long, filamentous, virus particles of 1,600 to 1,800 nm in length, typical for the family Closteroviridae. The virus was partially purified in a process in which the extract was treated with Triton for 1 h at 4°C, followed by two cycles of high-speed centifugation on a 20-40-60 % sucrose cushion. The virus was denaturated on CsCl or CsSO ⁴ gradients. The results show that the virus isolated is the first member in the Closteroviridae reported to naturally infect N. tabacum.