Characterization of Pitaya Mosaic Virus ans Analysis of Its Full-length Sequence

• Main Authors: Minq-Ru Liou, 劉命如
• Other Authors: Ruey-Fen Liou
• Format: Others
• Language: zh-TW
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/43692234880863395579

碩士 === 國立臺灣大學 === 植物病理學研究所 === 88 === Pitaya (Hylocereus undatus Britt. & Rose), which belongs to Cactaceae, has become an important tropic fruit in Taiwan in recent years. Pitaya showing mosaic symptoms on the pads was collected from fields in the Kawnshi area in 1999. Indicator plant tests indicated that the causal agent of the mosaic symptoms was mechanically transmissible, and local lesions developed on the leaves of Chenopodium amaranticolor 5 days after inoculation. After three successive single lesion transfers on the same plants, a virus isolate was obtained. In addition to C. amaranticolor, this virus is capable of infecting C. quinoa, Celosia argentea, and Gomphrena globosa. A back inoculation assay was performed to ensure that the virus, named Pitaya mosaic virus (PtMV) was indeed the causal agent of the mosaic symptom observed on pitaya. Electron microscopic examination of negatively stained virus particles from pad extracts of the diseased plants revealed that the virus has a flexuous rod-shaped morphology with a length of 483 to 517 nm. Purified viruses had an A260/A280 ratio of 1.20. Antiserum raised against PtMV reacted positively with infected plants and was used for detection of PtMV by ELISA. The molecular mass of the coat protein (CP) was estimated to be 26 kDa by SDS-polyacylamide gel electrophoresis. Analysis of viral RNA by formaldehyde agarose gel electrophoresis revealed the presence of a single species of RNA approximately 6-7 kb in length. Molecular cloning of the viral genome was subsequently carried out by rapid amplification of cDNA ends (RACE). Sequences of PtMV analyzed so far contained 6605 nucleotides (excluding 3’ poly(A) tail), which included a total of 7 putative open reading frames (ORF). Of the 7 ORFs being identified, ORF1 encodes RNA dependent RNA polymerase (RdRp), ORF2, ORF3, and ORF4 encode proteins which are involved in virus movement, ORF5 encodes coat protein. Each of these proteins displays significant homology to the corresponding ORFs of other potexviruses, indicating that genome organization of PtMV is basically similar as those of other potexviruses. Phylogenetic analysis based on multiple sequence alignments of RdRp and CP reveals a closest relationship of PtMV to Papaya mosaic virus.