First Report of Carnation Italian ringspot virus in Chrysanthemum zawadskii var. latilobum in Korea.

Chrysanthemum zawadskii var. latilobum (CZ), known as “Gujeolcho” in Korea, is a traditional herbal remedy for inflammatory diseases, and is widely used in Asia as a tea. CZ is also a popular ornamental flower in Korea. In June 2014, during a survey conducted in commercial CZ greenhouses in Jincheon City, Korea, virus-like symptoms of mosaic and yellowing were observed in many plants. Leaf samples were collected from 20 symptomatic plants in a commercial greenhouse. To identify the causal agent(s), total RNA was extracted from one of the samples, and a transcriptome library was generated using the TruSeq Stranded Total RNA with Ribo-Zero Plant Kit (Illumina, San Diego, CA) according to the standard protocol. Next-generation sequencing (NGS) was performed using an Illumina HiSeq2000 sequencer. De novo assembly of the quality filtered NGS reads (101-bp paired-end reads) was performed using the Trinity pipeline and the assembled contigs were analyzed against the viral reference genome database in Genbank by BLASTn and BLASTx searches (Schelhorn et al. 2013). The entire NGS procedure was performed by Macrogen Inc. (Seoul, Korea). Among the analyzed contigs, only one large contig (4617 bp) was clearly of viral origin. Nucleotide blast search showed that the contig has maximum identities of 88% (with 96% coverage) to the isolate Gyp1 (Genbank Accession No. GQ259476) of Carnation Italian ringspot virus (CIRV), which was isolated fromGypsophila paniculata. Therefore, all of the collected samples were tested by reverse transcription-PCR (RT-PCR) using the CIRV-specific primer pair designed using the CIRV contig sequence obtained from NGS (CIRV-Dec-Fw 5′-CGGTGGGGACGTTTATGTTGTTT-3′ and CIRV-Dec-Rv 5′-TGAAGTTTCCGTCCCGTCGTATCT-3′). All samples yielded PCR fragments with an estimated size of 564 bp. To confirm the RT-PCR result, all samples were tested using the CIRV commercial double-antibody sandwich (DAS)-ELISA kit (DSMZ, Braunschweig, Germany). CIRV was detected by DAS-ELISA in all samples. To determine the complete genome sequence of an isolate of CIRV from CZ, the 4617-bp contig sequence obtained by NGS was amplified by RT-PCR using the specific primer pair (CIRV-Amp-Fw 5′-TGCGCTAACTGTTGCTTTGCGTA-3′ and CIRV-Amp-Rv 5′-TGTCGTCAGTAGCTTCCACAAGT-3′) and the sequence was confirmed by de novo sequencing. Terminal sequences of the virus genome were analyzed by the 5′ and 3′ rapid amplification of cDNA ends (RACE) method as described previously (Kwak et al. 2013). The assembled complete genome sequence of CIRV isolated from CZ was 4745 nt long and deposited as GenBank Accession No. KP888563. CIRV belongs to the genus Tombusvirus in the family Tombusviridae. CIRV was first identified by Hollings et al. (Hollings et al. 1970) from carnation plants (Dianthus caryophyllus), and a few more isolates of CIRV were later obtained from cherry trees, Gypsophila, surface water, and Limonium sinuatum (Koenig et al. 2009). To our knowledge, this is the first report of the CIRV isolate from CZ. The plant could serve as a reservoir for CIRV in Korea. Because CZ is widely distributed in Korea and CIRV can cause serious diseases in other ornamental crops such as carnation and chrysanthemum, further surveys of the CIRV incidence in ornamental crops in Korea may be required. CIRV is readily eliminated by thermotherapy (Hollings et al. 1970), and could help recover healthy stocks of the affected chrysanthemum varieties. © 2015 American Phytopathological Society. All rights reserved.