• The Plant Pathology Journal
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• v.16 no.1
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• pp.48-51
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• 2000

Citrus tristeza virus (CTV) was identified form CTV-infected early satsuma mandarin (Citus unshiu) and yuzu (C.junos) by RT-PCR. The total RNAs were isolated from citrus bark and seaf tissues infected with CTV and reverse transcription was followed with primers designed for amplifying CTV coat protein gene. DNA fragments 738 bp were amplified by RT-PCR and these products were colned for sequence analysis. Based on the sequence analysis, this PCR product has 97% sequence homology to CTV (T-385) CP gene isolated from USA. RT-PCR assay for CTV detection was more sensitivity than ELISA assay which was done with anti-CTV CP antibody. This is the frist report about CTV identification in Cheju island Korea.

• Plant Disease and Agriculture
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• v.5 no.1
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• pp.34-40
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• 1999

The virus infection rates and disease symptoms of three major citrus viruses, citrus tristeza virus (CTV), satsuma dwarf virus (SDV), and citrus tatter leaf virus (CTLV), were investigated at 35 citrus orchards in Cheju Island from 1995. The infection rates of CTV, SDV, and CTLV were 69.8%, 8.6%, and 9.3%, respectively. However, depending on cultivars there were significant differences in the infection rates. The infection rates of CTV were highest in early satsuma mandarin (Citrusunshiu) with 80.9% and lowest in very early satsuma mandarin with 51.9%. In SDV, the highest was in very early stasuma mandarin with 23.1% and the lowest was in early satsuma mandarin with 6.3%. And the highest infection rate in very early satsuma mandarin with 17.9% and the lowest in tangors with 7.3% in CTLV. The symptoms of virus-infected citrus were very diverse; small and abnormal shape of fruits, abnormal leaves such as narrow boat and small spoon shapes of leaves, stem-pitting on the twig, bud-union crease and swelling of the graft part, reduction of the plant vigor and poor yields.

• The Plant Pathology Journal
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• v.38 no.4
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• pp.287-295
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• 2022

Citrus tristeza virus (CTV) is efficiently transmitted in a semi-persistent manner by the brown citrus aphid (Toxoptera citricida (Kirkaldy)). Currently, the most sensitive method for detecting plant viruses in insect vectors is reverse-transcription quantitative polymerase chain reaction (RT-qPCR). In this study, the elongation factor-1 alpha (EF-1α) gene and acidic p0 ribosomal protein (RPAP0) gene were confirmed to be suitable reference genes for RT-qPCR normalization in viruliferous T. citricida aphids using the geNorm, NormFinder, and BestKeeper tools. Then the relative CTV titer in aphids (T. citricida) at different post-acquisition feeding times on healthy plants was quantified by RT-qPCR using EF-1α and RPAP0 as reference genes. The relative CTV titer retained in the aphids gradually decreased with increasing feeding time. During the first 0.5 h of feeding time on healthy plants, the remaining CTV titer in aphids showed about 80% rapid loss for the highly transmissible isolate CT11A and 40% loss for the poorly transmissible isolate CTLJ. The relative CTV titer in aphids during more than 12 h post-acquisition times for CT11A was significantly lower than at the other feeding times, which is similar to the trend found for CTLJ. To our knowledge, this is the first report about the relative titer variation of CTV remaining in T. citricida at different post-acquisition feeding times on healthy plants.

• The Plant Pathology Journal
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• v.15 no.6
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• pp.335-339
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• 1999

Citrus tristeza virus(CTV), an aphid-borne closterovirus, is one of the most destructive pathogens of citrus. It has caused rapid decline in growth, stem pitting and death in citrus trees. A reverse transcriptase polymerase chain reaction (RT-PCR) assay was developed for detection of CTV and investigation of the CTV infection status of citrus and its related cultivars in Cheju island. For RT-PCR based CTV detection, primers were designed to amplify 670bp of coat protein gene. A screening test for CTV in citrus cultivars was conducted from March to July in 1999. Seventy individual citrus trees representing 9 species of 3 genera were tested. The infection rates of CTV for leaves from the years or older trees of late maturing citrus varieties such as Yuzu (C. junos Sieb. ex Tanaka), Navel orange (C.sinensis Osbeck), Kiyomitanger (C. unshiu x C. sinensis), and Shiranuhi ((C. unshiu x C. sinensis) x C. reticulata) were 100%, 80%, 60%, and 60% respectively. The CTV infection rates in Early satsuma mandarins such as 'Miyagawa Early' Satsuma mandarins (C. unshiu Marc. var. Miyagawa) and 'Okitsu Early' Satsuma mandarins (C. unshiu Marc. var. Okitsu) were 100%, and 60%, respectively. CTV was not detected in Cheju native Dangyooja (C. unshiu Marc. var. Osbeck), Trifoliate orange (Poncirus trifoliata) and Kumquat (Fortunella margarita Swingle). In conclusion, RT-PCR assay can be successfully applied to the detection of CTV in citrus trees.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1999.10a
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• pp.85.1-85
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• 1999

• The Plant Pathology Journal
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• v.16 no.1
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• pp.43-47
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• 2000

Citrus tristeza virus (CTV) is the causal agent of one of the most important diseases of citrus. Recently, CTV has been detected in Cheju Island by ELISA. The coat protein (CP) gene of CTV isolated form Cheju Island was cloned by RT-PCR and the nucleotide was analyzed in this study. Citrus leaves were collected from trees showing decline symptoms from various region of Cheju Island in the summer of 1998 and 1999. The CP gene open reading frame is composed of 670 nucleotides and encodes a polypeptide of 223 amono acids. Sequence analysis the CP gene revealed that two CTV strains present in Cheju Island. Viruses collected form Sogwipo area and Cheju City area in 1999 ahowed 91-93% nucleotide sequence homology with CTV T36 strain. Viruses collected form Cheju City area in 1999 and Sogwipo City in 1998 showed 94-98% nucleotide sequence homology with CTV SY568 strain. A efficient viral RNA extraction methods was developed by modifying procedure for animal virus RNA purification methods and PCR product was detected form one tenth of RNA purified from as small as 45 mg fresh or frozen tissue.

• Journal of Plant Biotechnology
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• v.32 no.1
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• pp.45-50
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• 2005

Virus-free stocks was produced by the combination of the heat treatment of virus infected plant and shoot-tip grafting (STS). To produce virus-free stocks, the plants infected with citrus viruses were used for virus-free stock production using the modified method of STG in thermotherapy at $40^{\circ}C$ for 16 hours in the light, and at $30^{\circ}C$ for 8 hours of darkness for 4 weeks. Trifoliate orange (P. trifoliata) were used as rootstock seedling for STG. Percentages of virus-free stocks against citrus tristeza virus (CTV), satsuma dwarf virus (SDV) and citrus tatter leaf virus (CTLV) were $75.7\%,\;100.0,\%\;82.6\%$ respectively. Shoot tip size for successful STG were as small as possible. Less than $0.3\;\cal{mm}$ of shoot tips gave the hight efficiency of virus free plants but survival rates were low. And, survival rate after shoot-tip culture was analyzed and the rates were dependant on the cultivars; Yuzu cultivar showed the hight survival rate ($74.6\%$) and early satsuma mandarin (Iwasagi) was $13.3\%$ as the lowest cultivar. But citrus trees were not succeed to grown, turned brown, and died.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1999.10a
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• pp.18.2-18
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• 1999

• The Plant Pathology Journal
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• v.40 no.2
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• pp.125-138
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• 2024

Citrus yellow vein clearing virus (CYVCV) is a member of the Alphaflexiviridae family that causes yellow vein clearing symptoms on citrus leaves. A total of 118 leaf samples from nine regions of six provinces in Korea were collected from various citrus species in 2020 and 2021. Viral diagnosis using next-generation sequencing and reverse transcription polymerase chain reaction (RT-PCR) identified four viruses: citrus tristeza virus, citrus leaf blotch virus, citrus vein enation virus, and CYVCV. A CYVCV incidence of 9.3% was observed in six host plants, including calamansi, kumquat, Persian lime, and Eureka lemon. Among the citrus infected by CYVCV, only three samples showed a single infection; the other showed a mixed infection with other viruses. Eureka lemon and Persian lime exhibited yellow vein clearing, leaf distortion, and water-soak symptom underside of the leaves, while the other hosts showed only yellowing symptoms on the leaves. The complete genome sequences were obtained from five CYVCV isolates. Comparison of the isolates reported from the different geographical regions and hosts revealed the high sequence identity (95.2% to 98.8%). Phylogenetic analysis indicated that all the five isolates from Korea were clustered into same clade but were not distinctly apart from isolates from China, Pakistan, India, and Türkiye. To develop an efficient diagnosis system for the four viruses, a simultaneous detection method was constructed using multiplex RT-PCR. Sensitivity evaluation, simplex RT-PCR, and stability testing were conducted to verify the multiplex RT-PCR system developed in this study. This information will be useful for developing effective disease management strategies for citrus growers in Korea.

• The Plant Pathology Journal
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• v.29 no.1
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• pp.31-41
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• 2013

The presence of Citrus tristeza virus (CTV) has previously been reported in citrus growing regions of Turkey. All serologically and biologically characterized isolates including I$\breve{g}$d${\i}$r, which was the first identified CTV isolates from Turkey, were considered mild isolates. In this study, molecular characteristics of the I d r isolate were determined by different methods. Analysis of the I$\breve{g}$d${\i}$r isolate by western blot and BD-RT-PCR assays showed the presence of MCA13 epitope, predominantly found in severe isolates, in the I$\breve{g}$d${\i}$r isolate revealing that it contains a severe component. For further characterization, the coat protein (CP) and the RNA-depen-dent RNA polymerase (RdRp) genes representing the 3' and 5' half of CTV genome, respectively, were amplified from dsRNA by RT-PCR. Both genes were cloned separately and two clones for each gene were sequenced. Comparisons of nucleotide and deduced amino acid sequences showed that while two CP gene sequences were identical, two RdRp clones showed only 90% and 91% sequence identity in their nucleotide and amino acid sequences, respectively, suggesting a mixed infection with different strains. Phylogenetic analyses of the CP and RdRp genes of I$\breve{g}$d${\i}$r isolate with previously characterized CTV isolates from different citrus growing regions showed that the CP gene was clustered with NZRB-TH30, a resistance breaking isolate from New Zealand, clearly showing the presence of severe component. Furthermore, two different clones of the RdRp gene were clustered separately with different CTV isolates with a diverse biological activity. While the RdRp-1 was clustered with T30 and T385, two well-characterized mild isolates from Florida and Spain, respectively, the RdRp-2 was most closely related to NZRB-G90 and NZRB-TH30, two well-characterized resistance breaking and stem pitting (SP) isolates from New Zealand confirming the mixed infection. These results clearly demonstrated that the I$\breve{g}$d${\i}$r isolate, which was previously described as biologically a mild isolate, actually contains a mixture of mild and severe strains.