• Research in Plant Disease
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• v.16 no.3
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• pp.326-330
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• 2010

Three pome fruit viruses, Apple chlorotic leaf spot virus (ACLSV), Apple stem grooving virus (ASPV) and Apple stem pitting virus (ASGV) were detected in pear trees (Pyrus pyrifolia) using double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) in Ansung, Naju and Ulsan provinces of Korea. Infection rate of three viruses was 35.2% from 452 leaf samples of the three cultivars of pear trees. Also, each of three viruses was detected by reverse transcription polymerase chain reaction (RT-PCR) for a limited number of samples. Infection rate of three viruses was 86.3% from 233 leaf samples of the three pear cultivars. The virus infection rates by RT-PCR were much higher than ELISA. ASGV was prevailing on pear with 74.2%, whereas ASPV and ACLSV were found in 34.8% and 0.4% of tested samples, respectively. Symptoms caused by ASGV showed black spots of infected Niitaka cultivar leaves. The ACLSV, ASPV and ASGV isolates showed 83~94% sequence identity at a nucleotide level to other pome fruit virus isolates when analyzed by NCBI BLAST. Pome fruit viruses occurring in pear were ACLSV, ASPV and ASGV. This is the first report of pear trees infected ASPV in Korea.

• Research in Plant Disease
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• v.23 no.4
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• pp.355-362
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• 2017

The 4 viruses, the Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), and Apple mosaic virus (ApMV) and 1 viroid, Apple scar skin viroid (ASSVd) are known major viral pathogens of apple trees in Korea. Infection degree of the 5 viral pathogens in the commercial nursery trees of major apple cultivars, 'Hongro', 'Fuji' and bud mutation of 'Fuji' was investigated. Infection ratio of the ACLSV, ASPV and ASGV for scion of an apple cultivar 'Hongro' were 100%, 81.3% and 100%, respectively. On the other hand, no infection for either ApMV and ASSVd detected. For the root stock of the cultivar, infection ratio of ACLSV, ASPV and ASGV showed 87.5%, 81.3% and 100% as well as ApMV and ASSVd were 12.5% and 6.3%, respectively. From the scion of apple cultivars 'Fuji' and bud mutation of 'Fuji', infection ratio of ACLSV, ASPV and ASGV showed 86.7%, 86.7% and 100%, respectively. Whereas, no infection for either ApMV or ASSVd detected. From the root stock of the cultivars, infection ratio of ACLSV, ASPV and ASGV showed 86.7%, 93.3% and 93.3% as well as ApMV and ASSVd were 12.5% and 6.3%, respectively. Result of our study indicates that most of commercial nursery apple trees were supplied with multiple infections by apple viruses causing potential losses for apple growers and, henceforth, agricultural policy for supply of the virus-free trees should be employed as soon as possible.

• The Plant Pathology Journal
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• v.30 no.2
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• pp.195-199
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• 2014

Apple stem pitting virus (ASPV), of the Foveavirus genus in the family Betaflexiviridae, is one of the most common viruses of apple and pear trees. To examine variability of the coat protein (CP) gene from ASPV, eight isolates originating from 251 apple trees, which were collected from 22 apple orchards located in intensive apple growing areas of the North Gyeongsang and North Jeolla Provinces in Korea, were sequenced and compared. The nucleotide sequence identity of the CP gene of eight ASPV isolates ranged from 77.0 to 97.0%, while the amino acid sequence identity ranged from 87.7 to 98.5%. The N-terminal region of the viral CP gene was highly variable, whereas the C-terminal region was conserved. Genetic algorithm recombination detection (GARD) and single breakpoint recombination (SBP) analyses identified base substitutions between eight ASPV isolates at positions 54 and 57 and position 771, respectively. GABranch analysis was used to determine whether the eight isolates evolved due to positive selection. All values in the GABranch analysis showed a ratio of substitution rates at non-synonymous and synonymous sites (dNS/dS) below 1, suggestive of strong negative selection forces during ASPV CP history. Although negative selection dominated CP evolution in the eight ASPV isolates, SLAC and FEL tests identified four possible positive selection sites at codons 10, 22, 102, and 158. This is the first study of the ASPV genome in Korea.

• Research in Plant Disease
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• v.25 no.1
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• pp.43-47
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• 2019

Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), and Apple scar skin viroid (ASSVd) are economically important viruses that infect pear tree species worldwide. To evaluate the prevalence of these viruses in Korea, we investigated infection degree of three viruses and one viroid for the commercial nursery trees of the pear cultivars, Niitaka, Chuwhang, Wonwhang, and Whasan in 2017 and 2018. The results showed that the infection ratio of ACLSV, ASPV, ASGV, and ASSVd for the scion of pear cultivar Niitaka was 10%, 45%, 77%, and 50%, respectively. From the scion of pear cultivar Chuwhang, infection ratios of ASPV, ASGV, and ASSVd were found to be 70%, 50%, and 60%, respectively. From the scion of pear cultivar Whasan, infection ratios of ACLSV, ASPV, ASGV and ASSVd were found to be 40%, 60%, 93%, and 20%, respectively. From the root stock of pear cultivar Wonwhang, infection ratios of ACLSV, ASPV, ASGV, and ASSVd showed 28%, 57%, 100%, and 14%, respectively. ASGV had the highest recorded infection rate, and ACLSV was characterized by the lowest infection rate. The mixed infection ratio of Niitaka, Chuwhang, Whasan, and Wonwhang was 45%, 60%, 70%, and 85%, respectively.

• The Plant Pathology Journal
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• v.20 no.2
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• pp.147-154
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• 2004

A simple and reliable procedure for RT-PCR detection of Apple stem pitting virus (ASPV), Cherry rasp leaf virus (CRLV), and Cherry necrotic rusty mottle virus (CNRMV) was developed. Two virus specific primer sets for each virus were found to specifically detect each virus among fourteen sets of designed oligonucleotide primers. Total RNAs extracted from healthy and from ASPV-,CRLV- and CNRMV-infected plant tissues were used to synthesize cDNA using oligo dT primer and then amplified by virus-specific primers for each virus. Each primer specifically amplified DNA fragments of 578 bp and 306 bp products for ASPV (prAS CP-C and prAS CP-N primers, respectively); 697 bp and 429 bp products for CRLV (prCR4 and prCR5-JQ3D3 primers, respectively); and 370 bp and 257 bp products for CNRMV (prCN4 and prCN6-NEG 1 primers, respec-tively) by RT-PCR. DNA sequencing of amplified DNA fragments confirmed the nature of each amplified DNA. Altogether, these results suggest that these virus specific primer sets can specifically amplify viral sequences in infected tissues and thus indicate that they can be used for specific detection of each virus.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2018.04a
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• pp.52-52
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• 2018

Apple (Malus domestica) is one of the most economically important fruits in Korea. But virus infection has decreased sustainable production of apple and caused the serious problems such as yield loss and poor fruit quality. Virus or viroid infection including Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), Apple mosaic virus (ApMV) and Apple scar skin viroid (ASSVd) has been also reported in Korea, furthermore, its damages and economic losses have increased constantly. In our research, we tried to survey virus infection for commercial nursery trees of major apple cultivars, especially dwarfing rootstocks 'M.9' and 'M.26' as well as scions. Trees were collected from 11 locations which have produced a great amount of apple nursery stocks in Korea. Infection degree was investigated in apple cultivars, 'Hongro' and 'Fuji' using RT-PCR method. In the scion of cultivar 'Hongro', infection ratio of ACLSV, ASPV and ASGV were 100%, 81.8% and 100% respectively. In the rootstock of cultivar 'Hongro', infection ratio of ACLSV, ASPV, ASGV and ApMV were 90.9%, 81.8%, 100% and 9.1% respectively. In the scion of cultivar 'Fuji', infection ratio of ACLSV, ASPV and ASGV were 81.8%, 90.9% and 100% respectively. In the rootstock of cultivar 'Fuji', infection ratio of ACLSV, ASPV, ASGV and ApMV were 81.8%, 90.9%, 100% and 9.1% respectively. Infection of ASSVd was not detected in both cultivars. From our results, it was found that most of apple rootstocks and scions had multiple infections by apple viruses which have caused economic damage in fruit production.

• Journal of Plant Biotechnology
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• v.44 no.4
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• pp.379-387
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• 2017

Various sizes (0.2 ~ 1.2 mm) and developmental stages (referred to as Stage 1 ~ 3) of apical and lateral meristems were excised, together or separately, directly from dormant buds of apple 'Hongro'. They were mixed infected by Apple scar skin viroid (ASSVd), Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV) and Apple stem grooving virus (ASGV), which are major viruses attacking apples. A total of 31 callus lines (> 10 mm in diameter) were obtained by culturing the explants on Murashige and Skoog (MS) medium supplemented with 3% sucrose, 3.0 mg/L benzyladenine (BA) and 0.1 mg/L indole-3-butyric acid (IBA), and they were subjected to RT-PCR analysis for virus detection. A high rate of virus elimination (expressed as the percentage of calli that did not amplify during RT-PCR, i.e., RT-PCR negative calli per total number of calli obtained) was achieved for ACLSV (100%), ASSVd (93.7%), and ASPV (93.7%), whereas it was only 25.8% for ASGV. ASPV was detected in the presence of 2 ~ 3 bracts. Simultaneous virus elimination of ASSVd, ASPV, ACLSV, and ASGV occurred during the meristem culture, in which the early stages of the dormant buds (Stage 1) were used, because ASGV was mostly eliminated during that stage. The results of the present study will be valuable for the production of virus-free apple trees.

• Research in Plant Disease
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• v.26 no.2
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• pp.95-102
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• 2020

The investigation of the infection rate of domestic apple orchards by four types of apple viruses (Apple chlorotic leaf spot virus [ACLSV], Apple stem pitting virus [ASPV], Apple stem grooving virus [ASGV], Apple mosaic virus [ApMV]) and one type of viroid (Apple scar skin viroid, ASSVd) found that most apple trees were infected with viruses and viroid at the rate of 97.3%. By region, the infection rate in Jeongseon stood at 98.8%, Danyang at 100%, Yesan at 100%, Jangsu at 89.1%, and Muju at 98.1%. By each virus and viroid, the infection rate of ASGV was the highest at 93.4%, followed by ASPV at 85.7%, ACLSV at 59.0%, ASSVd at 6.7%, and ApMV at the lowest 3.6%. In addition, 84.8% of the cases were infected with two or more types of viruses and viroid, nearly seven times the single type infection rate of 12.4%, and the cases infected with three viruses, ASPV, ACLSV, and ASGV accounted for 56.2%, more than the half the total number of trees investigated.

• The Plant Pathology Journal
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• v.32 no.5
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• pp.441-451
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• 2016

Deep sequencing has generated 52 contigs derived from five viruses; Apple chlorotic leaf spot virus (ACLSV), Apple stem grooving virus (ASGV), Apple stem pitting virus (ASPV), Apple green crinkle associated virus (AGCaV), and Apricot latent virus (ApLV) were identified from eight apple samples showing small leaves and/or growth retardation. Nucleotide (nt) sequence identity of the assembled contigs was from 68% to 99% compared to the reference sequences of the five respective viral genomes. Sequences of ASPV and ASGV were the most abundantly represented by the 52 contigs assembled. The presence of the five viruses in the samples was confirmed by RT-PCR using specific primers based on the sequences of each assembled contig. All five viruses were detected in three of the samples, whereas all samples had mixed infections with at least two viruses. The most frequently detected virus was ASPV, followed by ASGV, ApLV, ACLSV, and AGCaV which were withal found in mixed infections in the tested samples. AGCaV was identified in assembled contigs ID 1012480 and 93549, which showed 82% and 78% nt sequence identity with ORF1 of AGCaV isolate Aurora-1. ApLV was identified in three assembled contigs, ID 65587, 1802365, and 116777, which showed 77%, 78%, and 76% nt sequence identity respectively with ORF1 of ApLV isolate LA2. Deep sequencing assay was shown to be a valuable and powerful tool for detection and identification of known and unknown virome in infected apple trees, here identifying ApLV and AGCaV in commercial orchards in Korea for the first time.

• Journal of Plant Biotechnology
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• v.46 no.3
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• pp.228-235
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• 2019

Apple (Malus pumila) is one of the most economically important fruits in Korea. but virus infection has decreased the sustainable production of apples and caused serious problems such as yield loss and poor fruit quality. Virus or viroid infection including apple chlorotic leaf spot virus (ACLSV), apple stem pitting virus (ASPV), apple stem grooving virus (ASGV), apple mosaic virus (ApMV) and apple scar skin viroid (ASSVd) have been also reported in Korea. In many cases, as apple gets infected with virus and viroid with no specific symptoms, the damage and symptoms caused by the viruses are not detected. In our research, viruses in the rootstock were eliminated for a virus-free apple dwarfing rootstock of M.9 and M.26. The virus elimination methods were apical meristem culture, thermotherapy ($37^{\circ}C$, 6 weeks) and chemotherapy($Ribavirin^{(R)}$). The detection of apple viruses was accomplished by Enzyme-linked Immuno-Sorbent Assay (ELlSA) and reverse transcription-polymerase chain reaction (RT-PCR). RT- PCR method was 10 ~ 30% more sensitive than the ELISA method. The efficiency of virus elimination was enhanced in apical meristem culture method. The acquisition rate of virus-free apple dwarfing rootstocks was 30 ~ 40% higher in apical meristem culture. After the meristem culturing of M.9, the infection ratio of ACLSV, ASPV and ASGV was 45%, 60% and 50%, respectively. In the apple dwarfing rootstock of M.26, the infection ratio of ACLSV, ASPV and ASGV was 40%, 55% and 55%, respectively. Based on this study, the best method for the production of virus-free apple dwarfing rootstocks was the apical meristem culture.