First Report of Little cherry virus 1 and 2 in Sweet Cherry in Korea

Abstract

HomePlant DiseaseVol. 102, No. 5First Report of Little cherry virus 1 and 2 in Sweet Cherry in Korea PreviousNext DISEASE NOTES OPENOpen Access licenseFirst Report of Little cherry virus 1 and 2 in Sweet Cherry in KoreaS.-Y. Cho, H. Kim, and S.-I. YiS.-Y. ChoSearch for more papers by this author, H. KimSearch for more papers by this author, and S.-I. Yi†Corresponding author: S.-I. Yi; E-mail: E-mail Address: seedin@korea.krSearch for more papers by this authorAffiliationsAuthors and Affiliations S.-Y. Cho H. Kim S.-I. Yi † , Seed Testing and Research Center, Korea Seed and Variety Service, Gimcheon, Republic of Korea. Published Online:9 Mar 2018https://doi.org/10.1094/PDIS-06-17-0783-PDNAboutSections ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat Sweet cherry, Prunus avium L. of the family Rosaceae, is a minor fruit crop in Korea whose cultivation area is steadily increasing through regional specialization. Recently, Apple chlorotic leaf spot virus, Apple mosaic virus, Cherry necrotic rusty mottle virus (CNRMV), Cherry virus A (CVA), and Cherry green ring mottle virus have been reported in sweet cherries in Korea (Cho et al. 2013, 2014). In July 2016, eight samples from eight Napoleon cherry trees showing virus-like symptoms, such as vein chlorosis, protruding veins, and reddened leaves, were harvested in Gyeongju-si, Gyeongsangbuk-do, Republic of Korea. Total RNA was extracted from these symptomatic leaves using TRI Reagent (Molecular Research Center, Cincinnati, OH) and the extracted total RNA were analyzed by next-generation sequencing using an Illumina HiSeq2500 (Theragen Bio Institute, Suwon, Korea) and raw sequence data were analyzed using SeqGenesis (Daejeon, Korea). BlastX and BlastN analyses of de novo assembled contigs confirmed the presence of Little cherry virus 1 (LChV-1, contigs of lengths of 3,521 and 725 bases), LChV-2 (362, 359, and 276 bases), CVA (555, 443, and 317 bases), and CNRMV (531, 377, and 339 bases). These contigs showed 74 to 99% nucleotide identity with reference genome sequences (accession nos. KR736335, AF416335, EU188438, and LN879389). Each sample was analyzed by reverse transcription (RT)-PCR using specific primers designed from the contig sequences obtained in this study, LChV-1 865F/1164R (5′-TGGCAAAGAGGTCCAGTGAC-3′)/(5′-TCCTTTCGAGCTAGTCGTATCA-3′), LChV-2 27F/276R (5′-GAGGGTCATACCGTCAGTAAAGT-3′)/(5′-ACCCGATCAATACATTGAAGCAC-3′), CVA TR10 F/R (5′-GGTCCACCACGCTCAACAAG-3′)/(5′-TTCCAAGGCCCCTTCTTATCTC-3′), and CNRMV 32F/331R (5′-GAACCTTCAAACTCGACGCTG-3′)/(5′-ATGTTTCAGACTCGGGCACC-3′). All samples were infected by one (CVA, 2/8 samples), two (LChV-1 and CVA, 2/8), three (LChV-1, CVA, and CNRMV, 2/8), or four viruses (LChV-1, 2, CVA, and CNRMV, 2/8). To amplify the full length of the LChV-1 coat protein (CP, 1,215 base pairs), specific primers were used for RT-PCR based on sequences deposited in GenBank (KR736335): LChV-1 CP NTR F/R (5′-AAACAGCTTGATATAAATTAATTTGTTTCT-3′)/(5′-TTTACTTGAATAGCATCGAAAGGTAC-3′). Sequences of the complete or partial viral genes were deposited in GenBank under the names LChV-1 CP isolate Gyeongju, and LChV-2 isolate Gyeongju (MF083705 and MF083706) and theses isolates showed 98 and 97% nucleotide identity with those of an LChV-1 isolate from China (KR736335) and LChV-2 isolate from Germany (AF531505). Crude leaf extracts of symptomatic virus infected leaves were mechanically inoculated into Nicotiana benthamiana, N. tabacum, N. glutinosa, and Chenopodium amaranticolor. All inoculated plants were tested by RT-PCR at 14 days postinfection with specific primer pairs (LChV-1 865F/1164R, LChV-2 27F/276R, CVA TR10 F/R, and CNRMV 32F/331R). Virus infections were confirmed in tested N. benthamiana (CNRMV and CVA), N. tabacum (LChV-1, CNRMV, and CVA), N. glutinosa (LChV-1, CNRMV, and CVA), and C. amaranticolor (LChV-1). LChV-2 was not detected in any tested plants, perhaps because LChV-2 is transmitted to plants by an insect (apple mealybug) (Raine et al. 1986). To our knowledge, this is the first report on the occurrence and sequences of LChV-1 and LChV-2 in sweet cherry trees in Korea.