Jeong-Soo Kim

Complete genome sequence of motherwort yellow mottle virus, a novel putative member of the genus Torradovirus.

The complete genome sequence of a new virus isolated from a motherwort plant exhibiting yellow mottle, mild mosaic, and stunting symptoms in Andong, Korea, was determined. The genome of this virus is composed of two single-stranded RNAs (7068 and 4963 nucleotides in length, respectively) carrying poly(A) tails. RNA1 contains one large open reading frame (RNA1-ORF1), while two potential ORFs (RNA2-ORF1 and RNA2-ORF2) were found in RNA2. BLAST searches of protein databases showed that RNA1-ORF1 and RNA2-ORF2 have maximum amino acid sequence identities of 53xa0% and 57xa0% to the RNA1-ORF1 and RNA2-ORF2, respectively, of lettuce necrotic leaf curl virus (LNLCV, a recently identified torradovirus). Phylogenetic analysis provided further evidence that the virus identified in this study is probably a member of a new species in the genus Torradovirus. The name “motherwort yellow mottle virus” (MYMoV) is proposed for this new virus.

The complete genome sequence of a novel virus, bellflower veinal mottle virus, suggests the existence of a new genus within the family Potyviridae

A new virus was isolated from a bellflower (Campanula takesimana) plant showing veinal mottle symptoms, and its complete genome sequence was determined. The viral genome consists of a positive-sense single-stranded RNA of 8,259 ribonucleotides. Electron microscopic observation revealed that the viral genome is packaged as a filamentous particle with an average length of approximately 760 nm. BLAST searches of protein databases showed that the encoded polyprotein has a maximum amino acid sequence identity of 34.1% (with 95% coverage) to that of the isolate AD of Chinese yam necrotic mosaic virus (CYNMV; genus Macluravirus). Phylogenetic analysis and comparison of the encoded amino acid sequences with those of other viruses demonstrated that the identified virus shows minimal sequence similarity to known viruses and should therefore be considered a member of a new genus in the family Potyviridae. The name bellflower veinal mottle virus (BVMoV) is proposed for this new virus.

First Report of Sweet potato golden vein associated virus Infecting Sweet Potato in Korea

Sweet potato (Ipomoea batatas) is one of the most important crops in eastern Asia, including Korea. Consumption of sweet potato is increasing gradually because of its growing reputation as a health food. Recently, outbreaks of viruses infecting sweet potatoes have increased all over the world, probably because sweet potatoes are produced via vegetative propagation (1,2). In Korea, most sweet potatoes in fields have been infected by a begomovirus, Sweet potato leaf curl virus (SPLCV), and other viruses such as Sweet potato feathery mottle virus, Sweet potato virus G, and Sweet potato latent virus (3). Many countries have monitored sweet potato virus infections in fields as well as in germplasm collections to select virus-free stocks. In 2013, 20 sweet potato plants showing leaf roll symptoms in Muan, South Korea, were collected and analyzed. Total DNA was isolated from sweet potato leaves (Viral Gene-spin Viral DNA/RNA Extraction Kit, iNtRON Biotechnology, Seongnam, Korea) and viral DNA was amplified by rolling circle amplification (RCA, TempliPhi Amplification Kit, GE Healthcare Life Sciences, Uppsala, Sweden) following the manufacturers instructions. Amplicons were digested by restriction enzyme SacI (TaKaRa Bio, Shiga, Japan) and products were run on a 1.5% agarose gel. A 2.8-kb DNA fragment was purified from a gel, ligated into a pGEM-T easy vector (Promega, Madison, WI), and sequenced (Macrogen, Seoul, Korea). Based on a BLAST search, most of the sequences (36/38) were identified as SPLCV, but two independent clones 2,824 nt in length from sweet potato cv. Sincheonmi were similar to Sweet potato golden vein associated virus (SPGVaV) isolate US:MS:1B-3 (94.38%, GenBank Accession No. HQ333143). The complete genome sequence of the SPGVaV-Korea isolate contained six ORFs, as expected for a typical monopartite begomovirus. The sequence was deposited in GenBank under accession number KF803170. SPGVaV is a whitefly (Bemisia tabaci)-transmitted virus (genus Begomovirus, family Geminiviridae). A phylogenetic analysis that included other begomoviruses that infect sweet potato showed SPGVaV-Korea to segregate with other SPGVaV isolates. SPGVaV has previously only been reported in Brazil and the United States (1). This is the first report of SPGVaV in sweet potato outside of the Americas. References: (1) L. C. Albuquerque et al. Virol. J. 9:241, 2012. (2) E. Choi et al. Acta Virol. 56:187, 2012. (3) H. R. Kwak et al. Plant Pathol. J. 22:239, 2006.

First report of Peanut stripe virus (Family Potyviridae) in South Korea.

Severe mosaic symptoms resembling those reported for a blotch isolate of Peanut stripe virus (PStV) (1) were observed in the year 1998 in Suwon, South Korea, on several peanut cultivars. The incidence of the virus was as high as 100% in cv. Daekwang. The virus was seed transmitted to varying degrees depending on the cultivar and a maximum seed transmission of 15.7% was observed in cv. Aul. Electron microscopic examination of leaf dip preparations showed filamentous rods having modal length of 720 nm. Viral inclusion bodies in infected cells were of pinwheel, scroll, and laminated aggregates. The 3 terminal region of the viral genome was amplified using degenerate primers (2) and the resulting approximately 700 bp fragment was cloned and sequenced. GenBank searches using the 709 nucleotides consisting of the complete 3-untranslated region and a part of the coat protein gene showed that the virus shared 98% sequence identity with the currently known PStV isolates. To our knowledge, this is the first report of PStV in the Republic of Korea. References: (1) J. W. Demski et al. Ann. Appl. Biol. 105:495, 1984. (2) S. S. Pappu et al. J. Virol. Methods 41:9, 1993.

A determinant of disease symptom severity is located in RNA2 of broad bean wilt virus 2.

Broad bean wilt virus 2 (BBWV2), which belongs to the genus Fabavirus, is a destructive pathogen of many economically important horticultural and ornamental crops. In this study, we constructed infectious full-length cDNA clones of two distinct isolates of BBWV2 under control of the cauliflower mosaic virus 35S promoter. BBWV2-PAP1 isolated from paprika (Capsicum annuum var. gulosum) induces severe disease symptoms in various pepper varieties, whereas BBWV2-RP1 isolated from red pepper (Capsicum annuum L.) causes mild symptoms. Agrobacterium-mediated inoculation of the infectious cDNA clones of BBWV2-PAP1 and RP1 resulted in the same symptoms as the original virus isolates. The infectious cDNA clones of BBWV2-PAP1 and RP1 were used to examine the symptoms induced by pseudorecombinants between the two isolates to localize in which of the two genomic RNAs are the symptom severity determinants in BBWV2. The pseudorecombinant of RP1-RNA1 and PAP1-RNA2 induced severe symptoms, similar to those caused by the parental isolate PAP1, whereas the pseudorecombinant of PAP1-RNA1 and RP1-RNA2 induced mild symptoms, similar to those caused by the parental isolate RP1. Our results suggest that BBWV2 RNA2 contains a symptom determinant(s) capable of enhancing symptom severity.

First Report of Broad bean wilt virus 2 in Leonurus sibiricus in Korea

Leonurus sibiricus L. (family Lamiaceae) has been used as a traditional herbal remedy to treat various gynecologic diseases. Although it is a widely distributed subtropical weed in Southeast Asia, L. sibiricus have been commercially cultivated on a small scale in many geographic areas of Korea. In August 2012, field-grown L. sibiricus plants showing mosaic, yellowing, and stunting symptoms were collected near a pepper field in Andong, Korea. Since L. sibiricus is only consumed as a raw material of traditional medicine in Korea, symptomatic plants lose commercial value entirely. To identify the causal agent(s) of the virus-like symptoms, total RNA was extracted from the symptomatic leaves, 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) were performed using the Trinity pipeline and the assembled contigs (92,329 contigs) were analyzed against the viral reference genome database in GenBank by BLASTn and BLASTx searches (3). The entire NGS procedure was performed by Macrogen Inc. (Seoul, South Korea). Among the analyzed contigs, only two large contigs were clearly of viral origin. Nucleotide blast searches showed that the first and second contigs (5,914 and 3,534 bp, respectively) have maximum identities of 91 and 95% to RNA1 of the isolate RP3 (GenBank Accession No. JX183225) and RNA2 of the isolate RP7 (JX183234) of Broad bean wilt virus 2 (BBWV-2), which were isolated from pepper in Korea. The NGS results were confirmed by analyzing the sequences of the fragments covering the entire BBWV-2 genome amplified by RT-PCR using specific primers for BBWV-2 as described previously (1). To obtain the complete genome sequence, terminal sequences of both RNA segments were analyzed by the 5 and 3 rapid amplification of cDNA ends (RACE) method as described previously (1). The assembled full-length sequences of BBWV-2 RNA1 and RNA2 isolated from L. sibiricus were 5,951 and 3,575 nucleotides in length, respectively, and deposited in GenBank under the accessions KM076648 and KM076649, respectively. BBWV-2 belongs to the genus Fabavirus in the family Secoviridae and it is known to have a wide host range. To investigate the host range of the BBWV-2 isolated from L. sibiricus, sap from the symptomatic leaves of L. sibiricus was inoculated to the test plants including Nicotiana benthamiana, Capsicum annuum (red pepper), and C. annuum var. gulosum (Paprika). RT-PCR detection and sequencing of the amplicons showed that all the inoculated test plants were infected with the BBWV-2 isolated from L. sibiricus. Currently, BBWV-2 is epidemic in pepper fields in Korea (1,2). Because BBWV-2 is easily transmitted by various aphids, and L. sibiricus is widely distributed in both wild and cultivated fields in Korea, this host might serve as a potential source of BBWV-2 to other crops such as pepper. To the best of our knowledge, this is the first report of BBWV-2 in L. sibiricus. References: (1) H.-R. Kwak et al. Plant Pathol. J. 29:274, 2013. (2) H.-R. Kwak et al. Plant Pathol. J. 29:397, 2013. (3) S.-E. Schelhorn et al. PLoS Comput. Biol. 9:e1003228, 2013.

Erratum to: Complete genome sequence of longan witches’ broom-associated virus, a novel member of the family Potyviridae

The prediction of cleavage sites in the polyprotein derived from the genomic sequence of Longan witches’ broomassociated virus (LWBaV), as depicted in Fig. 1C of the article by Seo and colleagues [1], contains a site which does not comply with the standard consensus for the NIaPro mediated processing of potyvirus mature gene products [2]. As already mentioned by authors in the text, the site proposed for the cleavage at the NIb/CP junction is located close to another possible site, which could lead to a slightly larger CP with a N-terminal sequence starting with the residues AGEMTD..., and a predicted size of 311 amino acids. Until further experimental evidence can be provided, the annotation of the CP of LWBaV should preferably be derived from the predicted standard processing event (LVIETQ/AG) at position 2775 in the viral polyprotein. In addition, the numbering corresponding to the position for the P1/HC-Pro cleavage site (EPITYY/SG) in the same Fig. 1C should be corrected to 396. The predicted sizes of the two first gene products of LWBaV should be 396 and 456 amino acids for P1 and HC-Pro, respectively.

First report of Clover yellow vein virus on Glycine max in Korea.

Glycine max (Soybean) is the most important edible crop in Korea. In Korea, eight viruses have been reported to infect soybean, including Alfalfa mosaic virus (AMV), Cowpea mosaic virus (CPMV), Cucumber mosaic virus (CMV), Soybean dwarf virus (SbDV), Soybean mosaic virus (SMV), Soybean yellow common mosaic virus (SYCMV), Soybean yellow mottle virus (SYMMV), and Peanut stunt virus (PSV) (1). In 2012, Glycine max were observed in Daegu, South Korea, with mosaic and mottling symptoms on leaves. Samples with virus-like symptoms (n = 151) were collected from Daegu including legume genetic resource field. Virus particles were filamentous rod shaped, average length 760 nm, and were analyzed by RT-PCR using specific primers for several Potyviruses and previously reported viruses infecting soybean. Only two samples showing mosaic and mottling symptoms were identified as Clover yellow vein virus (ClYVV) based on RT-PCR using primers specific for ClYVV (5-GTTGGCTTGGTTGACACTGA-3 and 5-CTTCGATCATGGATGCACA-3). The sequences of amplified fragments were 97 to 98% similar with ClYVV. ClYVV is a distinct species in the genus Potyvirus and family Potyviridae. ClYVV is transmitted by several species of aphids and by mechanical inoculation (2). ClYVV was first reported on Gentiana scabra, and the disease has never been reported in soybean fields in Korea. The biological properties and full genome sequence of the selected ClYVV isolate of apparent virus symptoms between two samples were analyzed. The ClYVV isolate was inoculated to local lesion plants, re-isolated from local lesions three times, and propagated in Nicotiana benthamiana, and then named ClYVV-Gm. The ClYVV-Gm induced local lesions on inoculated leaves of N. tabacum cv. Xanthi-nc, Tetragonia expansa, and systemic symptoms on upper leaves of Chenopodium amaranticolor, C. quinoa, and N. clevelandii. The ClYVV-Gm caused mosaic and mottling symptoms on Glycine max cv. Kwangan and Phaseolus vulgaris. The genome of ClYVV-Gm was determined to be 9,584 nucleotides in length (GenBank Accession No. KF975894), and it shared 83% to 97% nucleotide identity with the sequences of 27 previously reported ClYVV isolates including Vicia fava and Pisum sativum. Despite low occurrence of ClYVV in Glycine max, ClYVV has a broad host range including tobacco, weed species, and soybean, which can lead to spreading of the virus. Our results indicate that emergence of ClYVV could become a problem to Leguminosae in Korea. To our knowledge, this is the first biological and molecular report of ClYVV infecting Glycine max in Korea. References: (1) Y. H. Lee et al. Korea Soybean Digest 29:7, 2012. (2) T. Sasaya et al. Phytopathology 87:1014, 1997.