| Summary: | 碩士 === 國立中興大學 === 植物病理學系所 === 107 === Lisianthus (Eustoma rusellianum) is one of the major cut-flower crops for exportation in Taiwan. Plants with symptoms of stunting, cup-shaped leaf curl and enations on petals and dorsal veins of leaves were found and collected from Fang-Yuen (FY) and Bei-Gang (BG) areas in central Taiwan in 2015. Results of biological, molecular and phylogenetic analyses indicated that the causal agents are two begomoviruses of the family Geminiviridae. Some isolates, that are FY0 (LC089013), FY2 (LC089014), and BG2 (LC089766), shared 93.3% nucleotide sequence identity to that of papaya leaf curl Guangdong virus (PaLCuGdV) (KP876482) while some isolates, namely BG1 (LC091538) and BG9 (LC091539), were most similar to that of DNA-A of tomato leaf curl Thailand virus (TYLCTHV) (KT322144) with 87.2% similarity. Based on the International Committee on Taxonomy of Viruses (ICTV) demarcation criteria of 91% nucleotide sequence identity for Begomovirus species, the FY0 and related isolates should be lisianthus-infecting isolates of PaLCuGdV and the isolates BG1 and BG9 is considered as a novel species of Begomovirus and temporarily designated as lisianthus enation leaf curl virus (LisELCV). We used PaLCuGdV-FY0 and LisELCV-BG9 as models for further analyses. Both viruses contained typical genome organization of begomovirus DNA-A with 6 open reading frames (ORFs) but different in genome size with PaLCuGdV-FY0 2732 nucleotides (nts) and LisELCV-BG9 2759 nts. In silico analyses revealed that LisELCV may be a recombinant with origins derived from PaLCuGdV-W1 (KF446659) and DNA-A component of TYLCTHV-NT35 (GU723735). The LisELCV-BG9 consists of whole C1 and C4 genes, part of C2 (89%) and C3 (53%) genes of PaLCuGdV-W1 (KF446659) and whole V1 and V2 as well as part of C3 (47%) and 1 C2 (11%) genes of DNA-A component of TYLCTHV-NT35 (GU723735). Infectious clones of both viruses, i.e. pCAMBIA1304-FY0 and pCAMBIA1304-BG9, were generated by rolling circle amplification and were successfully introduced to lisianthus and Nicotiana benthamiana plants by agro-infiltration. The inoculated lisianthus plants showed symptoms similar to those of diseased samples originally collected in the fields and the inoculated N. benthamiana plants also displayed symptoms of leaf curl and enation indicating the pathogenicity of both viruses. However, the pCAMBIA1304-BG9 was less virulent than pCAMBIA1304-FY0 with milder symptoms and longer incubation period after agro-infiltration. Nevertheless, synergistic effects of shorter incubation period and severer symptoms associated with co-inoculation by less virulent LisELCV-BG9 and lisianthus-infecting PaLCuGdV-FY0 as well as ageratum yellow vein virus (AYVV) are observed. Specific primers for detecting lisianthus begomoviruses in multiplex polymerase chain reaction have been developed and can be used effectively in field surveys. The etiology of two emerging lisianthus viral diseases and the characteristics of their causal agents have been studied and the detecting tools and techniques for lisianthus-infecting begomoviruses have been developed in this study.
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