广西南方水稻黑条矮缩病毒及水稻齿叶矮缩病毒的分子检测

Molecular detection of southern rice black-streaked dwarf virus and rice ragged stunt virus in Guangxi

【目的】探索南方水稻黑条矮缩病毒(SRBSDV)和水稻齿叶矮缩病毒(RRSV)的分子检测技术,了解两种病毒在广西的分布,为生产上开展两种病害的防治奠定基础。【方法】用RT-PCR技术检测采自广西各地的水稻、杂草及稻飞虱样品;用CF-11纤维素粉提取水稻茎叶组织中的dsRNA,1.5%琼脂糖凝胶电泳观察dsRNA图谱。【结果】从水稻、稗草、五节芒、李氏禾和白背飞虱样品中获得850bp左右的SRBSDV序列,从水稻和褐飞虱样品中获得700bp左右的RRSV序列;首次从五节芒和李氏禾中检测到SRBSDV。SRBSDV单独侵染、SRBSDV和RRSV复合侵染可见8条dsRNA条带,但带谱不一致;RRSV单独侵染可见5条dsRNA带。在供检测的181株水稻样品中,除了来自北海、贵港和梧州市的水稻样品外,其余样品均检测到SRBSDV;除了来自梧州、贺州、玉林、贵港和来宾的水稻样品外,其余样品均检测出RRSV;其中带SRBSDV的样品数占44.8%,带RRSV的样品占21.5%,两种病毒复合感染占11.0%。【结论】研究设计的特异性引物能用于RT-PCR检测SRBSDV和RRSV、水稻样品提取dsRNA能快速直观地鉴定SRBSDV、RRSV单独感染及混合感染水稻。用设计的引物首次检测出SRBSDV的两种新寄主五节芒和李氏禾。

[Objective] The current research was conducted to explore molecular biological methods for detecting southern rice black-streaked dwarf virus （SRBSDV） and rice ragged stunt virus （RRSV）, understand the distribution of two viruses in Guangxi, and lay the foundation for controlling the two viruses in production. [Method]RT-PCR technique was used to detect the two viruses from rice, weeds and vector insects samples gathered from different regions of Guangxi. Fibrous cellulose powder （CF-11） was used to extract dsRNA from rice leaves and stems. dsRNA electrophoregram was observed on 1.5% agarose gel under ultraviolet light. [Result]A 850 bp sequence of SRBSDV was obtained from rice, Echinochloa crusgalli, Miscanthus floridulus, Leersia hexandra and Sogatella furcifera by RT-PCR, and a 700 bp sequence of RRSV from rice and Nilaparvata lugens. SRBSDV was detected from Miscanthus floridulus and Leersia hexandra for the first time. Eight bands of dsRNA were detected from rice infected by SRBSDV alone and by both SRBSDV and RRSV, but the sizes of dsRNA bands were different. Five bands of dsRNA were detected from rice infected by RRSV. Among the detected 181 rice samples from different regions of Guangxi, except rice samples from Beihai, Guigang and Wuzhou, SRBSDV were detected and reached 44.8 percent of the total samples. Except rice samples from Hezhou, Guigang, Yulin, Laibin and Wuzhou, RRSV were detected and reached 21.5 percent of the total samples, in which 11.0% of rice samples were infected by both SRBSDV and RRSV. [Conclusion]The special primer designed could be used in RT-PCR to detect SRBSDV and RRSV. dsRNA extracted from rice samples could rapidly and visually identify rice samples infected by SRBSDV, RRSV or both. Thanks to the designed special primer, two new hosts to SRBSDV, Miscanthus floridulu （Labnll.） Warb and Leersia hexandra, were discovered for the very first time.