• The Plant Pathology Journal
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• v.40 no.4
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• pp.408-413
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• 2024

The emergence of rice black-streaked dwarf virus (RBSDV) poses a significant threat to global cereal crop cultivation, necessitating the urgent development of reliable detection and quantification techniques. This study introduces a reliable approach for the precise and sensitive quantification of the RBSDV in cereal crop samples, employing a reverse transcription digital polymerase chain reaction (RT-dPCR) assay. We assessed the specificity and sensitivity of the RT-dPCR assay proposed for precise RBSDV detection and quantification. Our findings demonstrate that RT-dPCR was specific for detection of RBSDV, with no cross-reactivity observed with other viruses infecting cereal crops. The RT-dPCR sensitivity was over 10 times that of RT-quantitative PCR (RT-qPCR). The detection limit of RT-dPCR was 0.096 copies/㎕. In addition, evaluation of RT-dPCR assay with field samples was conducted on 60 different cereal crop samples revealed that RT-dPCR (45/60) exhibited superior accuracy compared with RT-qPCR (23/60). In this study, we present a specific and accurate RT-dPCR assay for the detection and quantification of RBSDV.

• The Plant Pathology Journal
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• v.38 no.2
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• pp.159-166
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• 2022

Barley yellow dwarf virus (BYDV) has been a major viral pathogen causing significant losses of cereal crops including oats worldwide. It spreads naturally through aphids, and a rapid, specific, and reliable diagnostic method is imperative for disease monitoring and management. Here, we established a rapid and reliable method for isothermal reverse transcription recombinase polymerase amplification (RT-RPA) combined with a lateral flow strips (LFS) assay for the detection of BYDV-infected oat samples based on the conserved sequences of the BYDV coat protein gene. Specific primers and a probe for RT-RPA reacted and optimally incubated at 42℃ for 10 min, and the end-labeled amplification products were visualized on LFS within 10 min. The RT-RPA-LFS assay showed no cross-reactivity with other major cereal viruses, including barley mild mosaic virus, barley yellow mosaic virus, and rice black streaked dwarf virus, indicating high specificity of the assay. The sensitivity of the RT-RPA-LFS assay was similar to that of reverse transcription polymerase chain reaction, and it was successfully validated to detect BYDV in oat samples from six different regions and in individual aphids. These results confirm the outstanding potential of the RT-RPA-LFS assay for rapid detection of BYDV.

• Applied Biological Chemistry
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• v.37 no.3
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• pp.148-153
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• 1994

Rice black-streaked dwarf virus (RBSDV), a member of the plant reoviridae fijivirus group, causes a serious damage for rice production in Korea. To characterize the RBSDV genome, virus particles were produced by feeding of planthopper (Laodelphax striatellus F.) carring RBSDV to maize plants for 2 days. In $30{\sim}40$ days after feeding, the viral particles were purified from the infected maize roots by using $10{\sim}40%$ sucrose gradient centrifugation. After treatment of 10% SDS to remove the viral coat proteins, ten viral double-stranded RNAs were resolved in agrose gel electrophoresis. Total dsRNA was then used to synthesize cDNA by reverse transcriptase and a cDNA library was constructed in the ${\lambda}gt11$ vector. The phages that contain RBSDV cDNA fragments were selected by hybridizing with the random-primed probe prepared from RBSDV dsRNAs. After subcloning of several cDNA fragments into the pUC19 plasmid vector, one clone (pRV3) was chosen for sequencing. The pRV3 clone was shown to be located on the RBSDV genome fragment No.3 by RNA gel-blot analysis. Sequence analysis of the clone revealed that the pRV3 contains two partial open reading frames.

• Research in Plant Disease
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• v.14 no.3
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• pp.223-225
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• 2008

This study was carried out to identify the Rice black-streaked dwarf virus that infected maize plants collected from Gochang-gun in Jeollabukdo in 2005. The genomic dsRNA from infected plants was extracted and the genome pattern was analyzed by polyacrylamide gel electrophoresis. Results of the electrophoresis revealed the already known to-segment genome and the difference of mobility was confirmed in isolates by collected areas. The RBSDV was identified from the result of RT-PCR using the template of extracted dsRNA and specific primer. The results of S10 cloned to pGEM-T vector and the conducted in sequence analysis consisted of 1,801nt and 559aa. This was of the same size as the RBSDV S10 identified in rice, and the change was confirmed in 18 base and displayed homology of 99%.

• The Plant Pathology Journal
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• v.29 no.1
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• pp.99-104
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• 2013

To detect five plant viruses (Beet black scorch virus, Beet necrotic yellow vein virus, Eggplant mottled dwarf virus, Pelargonium zonate spot virus, and Rice yellow mottle virus) for quarantine purposes, we designed 15 RT-PCR primer sets. Primer design was based on the nucleotide sequence of the coat protein gene, which is highly conserved within species. All but one primer set successfully amplified the targets, and gradient PCRs indicated that the optimal temperature for the 14 useful primer sets was $51.9^{\circ}C$. Some primer sets worked well regardless of annealing temperature while others required a very specific annealing temperature. A primer specificity test using plant total RNAs and cDNAs of other plant virus-infected samples demonstrated that the designed primer sets were highly specific and generated reproducible results. The newly developed RT-PCR primer sets would be useful for quarantine inspections aimed at preventing the entry of exotic plant viruses into Korea.

• Applied Biological Chemistry
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• v.38 no.6
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• pp.522-527
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• 1995

To develop an antiviral agent for the rice black-streaked dwarf virus (RBSDV), a hammerhead type ribozyme, which has a potential target site on the genome segment 3, was designed. Oligonucleotides for the ribozyme and its substrate were synthesized, annealed, and cloned into a plasmid pBluescript II KS(+). Ribozyme and substrate RNAs were then synthesized by in vitro transcription with $T_3$ RNA polymerase, obtaining RNAs in expected size, 193 and 182 nucleotides, respectively. The substrate RNA was efficiently cleaved into two fragments when incubated with the ribozyme at $55^{\circ}C$, while the cleavage was not detected at $37^{\circ}C$. In addition, the segment 3 RNA of RBSDV was also cleaved into two fragments by the same ribozyme at $55^{\circ}C$. Taken together, our results demonstrated that the hammerhead ribozyme has an in vitro endonucleolytic activity and may be used as an antiviral agent in transgenic plants.

• Korean Journal Plant Pathology
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• v.1 no.2
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• pp.109-114
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• 1985

The experiments were conducted to clarify the influence of transplanting date on the occurrence of rice virus in field condition of 1984. The rate of RBSDV (rice black-slreaked dwarf virus) viruliferous vector, smaller brown planthopper (Laodelphax striatellus Fallen), was shown to be $13.6\%$ at the 2nd adult and that of rice stripe virus (RSV) viruliferous was $6.7\%$ at the 2nd adult. The vector in the field was begun to come from May 29, the maximum densities were 19.6 insects per hill on June 13 in cultivar Chucheongbyeo, 19.3 in Nagdongbyeo, 7.4 in Cheongcheongbyeo and 4.9 in Samgangoyes. The number of vectors per hill was inclined to increase by early transplanting. Although the infection of rice virus in nursery bed was not recognized until May 30 transplanting, the nursery infection could be seen from June 10 transplanting. The highest rate of nursery infection with RSV was $4.1\%$ at June 10 transplanting plot, and that of RBSDV was $14.2\%$at June 20 trans planting plot. The infection of rice virus in paddy field was the highest at May 20 transplanting plot, the lowest at July 10 plot. The earlier transplanting, the more severe occurrence of rice viruses. Occurrence of infected plants with RBSDV was shown to increase more rapidly at May 20 and May 30 planting plot than May 10 plots. However, the occurrence of infected plant with RSV was more rapid at May 10 transplanting plot than May 20 and 30.

• Proceedings of the Korean Society of Crop Science Conference
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• 2005.08a
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• pp.120-121
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• 2005

• Korean journal of applied entomology
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• v.22 no.3 s.56
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• pp.193-197
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• 1983

In 1981 Rice Black-Streaked Dwarf Virus (RBSDV) was severely occurred in Yeongnam area of Korea. The influence of RBSDV to rice plant was studied with two susceptible cultivars, Nagdongbyeo and Cheongcheongbyeo. The stunting rate was determined by the percentage of plant height of infected plants vs. healthy plants. When the rice plants were severely stunted by RBSDV, the yield components and yield were greatly reduced. The stunting of rice plants infected with RBSDV was caused mostly by the shortening of internodes in upper parts of the culm. The relationship between stunting rate of rice plants and yield was shown to have a negative exponential correlation. The regression equations of the relationship are experssed as follows: In Cheongcheongbyeo $Y=46.6lxe^{-0.0624_\chi}$, and in Nagdongbyeo $Y=54.82xe^{-0.067_\chi}$.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2000.10a
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• pp.62.1-62
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• 2000