First Report of Grapevine yellow speckle viroid 1 and Hop stunt viroid Infecting Grapevines (Vitis vinifera) in Korea

Viroids are the smallest pathogens, and they are composed of a circular single-stranded RNA genome (Hammann et al. 2012). The grapevine (Vitis vinifera L.) is host to several viroids. In June 2016, grapevine leaf samples from two popular cultivars in Korea, Campbell Early and Jaok, and rootstock 188-08 obtained from the Grape Research Institute, Okcheon, Korea, and the Sangju National Agricultural Cooperative Federation, Sangju, Korea, were collected and tested to determine their viroid status. None of the grapevines showed apparent disease symptoms. Six different libraries from three different genotypes were prepared and subjected to high-throughput sequencing using Illumina’s HiSeq 2000 followed by bioinformatics analyses (Jo et al. 2016). The six libraries, except those obtained with cv. Jaok from the Grapevine Research Institute, contained 74 contigs associated with Grapevine yellow speckle viroid 1 (GYSVd1) from the genus Apscaviroid and Hop stunt viroid (HSVd) from the genus Hostuviroid. The complete genome sequence for GYSVd1 (GenBank KY244035) and HSVd (KY244036) was obtained from rootstock 188-08. BLASTN revealed that GYSVd1 from rootstock 188-08 with 366 nt shared 99% identity with the clone Boonmang-1 from Thailand (KP010005.1), while HSVd from rootstock 188-08 with 297 nt shared 100% identity with the clone chi100Y-02-4-5 from China (AB219944.1). To confirm coinfection of GYSVd1 and HSVd, RT-PCR was conducted using GYSVd1-specific primers (5′-CGGATCACTTTCCTGTGGTT-3′ [position 1-20] and 5′-AGGACCTCTTTGCAGGGGT-3′ [position 348-366]) based on the GYSVd1 reference sequence (NC_001920.1) and HSVd-specific primers (5′-GCCCCGGGGCTCCTTTCTCAGGTAAG-3′ [position 85-60] and 5′-CCCGGGGCAACTCTTCTCAGAATCC-3′ [position 78-102]) based on the HSVd reference sequence (NC_001351.1) (Astruc et al. 1996). The amplified PCR products were cloned in pGEM-T-Easy Vector (Promega, Wisconsin, U.S.A.) and sequenced. The results showed that five samples, except Jaok from the Grapevine Research Institute, were coinfected by the two viroids, confirming the high-throughput sequencing data. The genome sequences of the two viroids in this study were compared with those identified from Ixeridium dentatum in Korea (Lee et al. 2015). Only one nucleotide at the 344 position between the two GYSVd1 genomes was changed, while the two HSVd genomes from grapevine and I. dentatum were 100% identical. This result indicates that I. dentatum is a natural host for the two viroids and that it might be involved in the transmission of the two viroids. The two viroids were also found in the newly developed grapevine cultivars Jarang and Jungrang. To obtain the exact infection rate for the two viroids in various grapevine cultivars in Korea, a nationwide assay should be conducted. This is the first report of GYSVd1 and HSVd infecting grapevines in Korea. © 2017, American Phytopathological Society. All rights reserved.