Interactions between Cymbidium mosaic virus and Odontoglossum ringspot virus in Nicotiana benthamiana

• Main Authors: Shang-Ming Guo, 郭尚明
• Other Authors: Ya-Chun Chang
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/e3x9c3

碩士 === 國立臺灣大學 === 植物病理與微生物學研究所 === 105 === Mixed infection of plant viruses usually leads to intrahost virus-virus interactions. Odontoglossum ringspot virus (ORSV) and Cymbidium mosaic virus (CymMV) commonly co-infect orchid plants and cause more severe symptoms, which is defined as synergistic effect. Recently, we found that the synergistic effect between ORSV and CymMV did exist on Nicotiana benthamiana protoplasts. This interaction seems to be regulated by the silencing suppression activity of ORSV p126. In this study, we continued to explore the interactions between ORSV and CymMV on N. benthamiana. In addition to p126, transiently expressed ORSV capsid protein (CP) facilitated CymMV accumulation on the inoculated leaves of N. benthamiana, but ORSV movement protein did not. The mechanism under this phenomenon remains unknown. Individual domains of ORSV p126 were proved without RNA silencing suppression ability and could not improve CymMV accumulation. In this study, we constructed five different domain combination of p126 and found that all four domains are necessary for RNA silencing suppression. Surprisingly, viral RNA and CP accumulation of both ORSV and CymMV had no significant difference between singly and doubly inoculated leaves of N. benthamiana plants through agroinoculation. However, by means of sap inoculation, more severe symptoms on both inoculated and systemic leaves of doubly infected plants were observed compared to singly infected ones. Next, we detected the viruses in systemic leaves of ORSV and CymMV doubly infected plants by indirect-ELISA, and found that the systemic movement-deficient CymMV could systemically infect N. benthamiana. These results suggested that although mixed infection of ORSV and CymMV did not exhibit synergistic interaction on inoculated leaves, ORSV still facilitated CymMV in other mechanism, probably on movement. Interestingly, facilitation on CymMV systemic movement disappeared when CymMV was co-inoculated with systemic movement-deficient ORSV (ORSVE100), which suggested the systemic movement of CymMV may rely on ORSV CP or ORSV infection processes. To understand the specificity of ORSV-CymMV synergism, we co-expressed CymMV with some well-known RNA silencing suppressors (RSSs), e.g. Turnip mosaic virus (TuMV) HCPro, Cucumber mosaic virus 2b, Potato virus X (PVX) p25 and Tomato bushy stunt virus p19. Except for PVX p25, all RSSs could significantly increase the accumulation of CymMV, which indicated that p126-mediated enhancement of CymMV accumulation probably can be replaced by other RSSs. Furthermore, we were curious about whether CymMV can systemically infect N. benthamiana with the aid of other ORSV-related or ORSV-unrelated viruses. For mixed infection of TuMV+CymMV, and PVX+CymMV, about 57% and 50% infected plants showed systemic CymMV infection. Co-infection of CymMV and Tomato mild green mosaic virus (TMGMV), a tobamovirus, facilitated systemic movement of CymMV on all tested plants. Finally, we constructed three eGFP-expressing CymMV clones and used one of them to confirm the experimental results.