First Report of Coleus blumei viroid 5 from Coleus blumei in India and Indonesia.

Coleus blumei, which was found originally in Indonesia, is an ornamental plant grown worldwide. It can be infected by several viroids of the genus Coleviroid, family Pospiviroidae. Six main viroids that infect coleus have been reported: Coleus blumei viroid 1 through 6 (CbVd-1 ~ CbVd-6). Although CbVd-1 was first reported in a commercial coleus in Brazil in 1989 (1), and then in Germany, Japan, Canada, Korea, China, and India, CbVd-5 was reported only in China in 2009 (2). Symptoms caused by CbVd-5 varied depending on different cultivars, and in case of an unknown cultivar of "Red with dark green edge," are very clear albino symptoms. From 2010 to 2011, 60 and 3 leaf samples of coleus were collected from Hyderabad, India, and Java, Indonesia, respectively, and subjected to low molecular weight RNA extraction according to Li et al. (3). The results of dot-blot hybridization using CbVd-5 cRNA probes and RT-PCR using CbVd-5 specific primers (CbVd-5-P: 5'-TGACTAGAACAGTAGTAAAG-3' / CbVd-5-P: 5'-AATTGAGGTCAAACCTCTTT-3') demonstrated that 28 out of the 60 samples from India and all three samples from Indonesia were positive for CbVd-5. The resulting RT-PCR fragments from one sample selected randomly from each country were cloned into the pMD18-T vector (Takara) and transformed into E. coli DH5α competent cells. Five positive clones of each sample were sequenced. The result of sequence analysis revealed that the similarities of CbVd-5 between the sequences we obtained and the reference sequence (GenBank Accession No. NC003683) were 97.8 to 100%. Bioassay using nine viroid-free coleus plants from three cultivars (three from each cultivar), inoculated with CbVd-5 infectious clones by stem slashing, demonstrated that CbVd-5 could induce albino symptom on the leaves of the unknown cultivar "Red with dark green edge" 2 months after inoculation. To our knowledge, this is the first report of CbVd-5 from India and Indonesia, and the second report of CbVd-5 in the world. Considering the effect of CbVd-5 on the appearance of coleus and its recombination ability, a certification program may be needed to control the spread of this viroid. References: (1) M. E. N. Fonseca et al. Fitopatol. Bras. 14:94, 1989. (2) W. Y. Hou et al. Arch. Virol. 154:315, 2009. (3) S. F. Li et al. Ann. Phytopathol. Soc. Jpn. 61:381, 1995.