Research in Plant Disease (식물병연구)

The Korean Society of Plant Pathology (한국식물병리학회)
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Application of Multiplex RT-PCR for Simultaneous Identification of Tomato Spotted Wilt Virus and Thrips Species in an Individual Thrips on Chrysanthemum

시설재배 국화에서 총채벌레의 종 동정 및 보독 바이러스 동시 검출을 위한 다중 진단법 적용

  • Yoon, Ju-Yeon (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Yoon, Jung-Beom (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Seo, Mi-Hye (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Choi, Seung-Kook (Department of Research Planning and Coordination, Rural Development Administration) ;
  • Cho, In-Sook (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Chung, Bong-Nam (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Yang, Chang Yeol (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration) ;
  • Gangireddygari, Venkata Subba Reddy (Department of Horticultural and Herbal Crop Environment, National Institute of Horticultural and Herbal Science, Rural Development Administration)
  • 윤주연 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • 윤정범 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • 서미혜 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • 최승국 (농촌진흥청 연구정책국 연구운영과) ;
  • 조인숙 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • 정봉남 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • 양창열 (농촌진흥청 국립원예특작과학원 원예특작환경과) ;
  • Received : 2020.11.13
  • Accepted : 2020.12.11
  • Published : 2020.12.31
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Abstract

We have developed a simultaneous diagnostic method that can identify both the species of thrips and tomato spotted wilt virus (TSWV) that are problematic in chrysanthemum plants. This is a method of amplifying DNA by performing reverse transcription-polymerase chain reaction by simultaneously adding primers specific to TSWV coat protein (N) gene and primers specific to the internal transcribed spacer 2 region of Frankliniella occidentalis and F. intonsa using total nucleic acid extracted from one thrips. The sizes of DNA fragments for TSWV, F. occidentalis, and F. intonsa were 777, 287, and 367 bp, respectively. These results showed species identification of thrips and whether thrips carrying TSWV can be simultaneously confirmed. Further usefulness of the simultaneous diagnostic method was made from greenhouse survey at chrysanthemum greenhouses in Taean (Chungcheongnam-do) and Changwon (Gyeongsangnam-do) to investigate the identification of thrips species and the rate of thrips carrying TSWV. Of thrips collected from the greenhouses, 83.7% thrips was F. occidentalis and 72.9% F. occidentalis carried TSWV in Taean. Similarly, the diagnostic method showed that 92.2% thrips was F. occidentalis and 84.0% F. occidentalis carried TSWV in Changwon. These results confirm that F. occidentalis is a dominant thrips species and the thrips species plays a crucial role in the transmission of TSWV in chrysanthemum plants in the greenhouses. Taken together, this study showed a simple diagnostic method for thrips identification and epidemiological studies of the timing and spread of TSWV through thrips in chrysanthemum greenhouses in South Korea.

이번 연구는 국화에서 문제되는 총채벌레의 종 동정 및 보독 바이러스인 토마토반점위조바이러스(Tomato spotted wilt virus, TSWV)를 동시에 확인할 수 있는 진단방법을 개발하였다. 이는 총채벌레 1마리에서 추출한 핵산에 꽃노랑총채벌레 및 대만총채벌레의 ITS2 부분에 특이적인 프라이머와 TSWV 외피단백질(N) 유전자 특이적인 프라이머를 동시에 넣어 reverse transcription-polymerase chain reaction을 수행하여 DNA를 증폭시키는 방법으로 전기영동하여 각각 287, 367, 777 bp의 DNA 단편의 크기를 비교함으로써 총채벌레의 종 동정 및 총채벌레의 TSWV 보독 여부를 동시에 확인할 수 있다. 충청남도 태안 및 경상남도 창원의 국화 시설하우스에서 총채벌레를 포집하여 총채벌레 우점종과 총채벌레의 TSWV 보독율을 조사한 결과, 태안의 국화 시설하우스에서는 꽃노랑총채벌레가 83.7%로 우점하고 있으며 채집된 총채벌레 중 72.9%가 TSWV를 보독하고 있었으며, 창원에서는 꽃노랑총채벌레가 92.2%를 차지하고 있으며 84.0%의 총채벌레에서 TSWV가 진단되었다. 이러한 결과는 Frankliniella occidentalis가 우점종이며 온실의 국화 식물에서 TSWV의 전반에 중요한 역할을 한다는 것을 확인해준다. 이번 연구는 국화 시설하우스에서 총채벌레를 통한 TSWV의 시설하우스내 유입시기 및 확산 경로 등 바이러스의 역학연구를 위한 간편진단법으로 활용 가능함을 예시해준다.

Keywords

References

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