感染地黃(Rehmannia glutinosa)菸草嵌紋病毒(Tobacco mosaic virus)之鑑定

Rehmannia glutinosa being a kind of medicinal plant cultivated in Taiwan was found to be infected by viruses. A plant with leaf mosaic was collected from Puli, and from which a pure line of virus isolate (RG1) was obtained through series of single lesion isolation. By host range test, electron microscope observation, immunosorbent electron microscopy, Ouchterlony immunodiffusion, coat protein electrophoresis, western blot analysis, and nucleotide sequencing, and analyses, the virus was identified as a isolate of Tobacco mosaic virus (TMV). The virus systemically infects Rehmannia glutinosa and causes mosaic symptom, but locally infects Chenopodium amaranticolor, C. quinoa and Nicotiana tabacum cv. Vam-Hicks and caused necrotic lesions. Direct negative staining by electron microscopy observation of diseased tissue revealed the presence of rod-shaped virus particles, around 300-310×16-18 nm. The virus was purified from inoculated leaves of C. quinoa by isopycnic centrifugation in cesium sulfate and the purified preparation was used to inject rabbit for antiserum production. The virus was found consisting of one species of coat protein subunit with an estimated molecular weight of 19 kDa. In Ouchterlony immunodiffusion and western blot analyses, the prepared antiserum reacted with homologous TMV antigens. The 3'-terminal 1044 nucleotides of this virus were cloned, sequenced and compared with corresponding sequences of representative Tobacco mosaic virus strains listed in public databases. It was found that this virus is most closely related to the Tobacco mosaic virus reported in China (Accession No. AY555269), with nucleotide and amino acid sequence identities of 97.1% and 99.4%, respectively. This is the first report of Tobacco mosaic virus infecting Rehmannia glutinosa in Taiwan.