臺灣博碩士論文加值系統

中文摘要
羅勒(Ocimum basilicum L.) 俗稱九層塔，為台灣地區常見之香辛蔬菜，本文記載在南台灣高屏地區田間，發現一種病毒性的羅勒新病害，受感染的植株葉片呈現嵌紋及皺縮之病徵。將單斑分離所得之病毒機械接種於5科21種植物，結果計有藜科植物3種及茄科植物1種受感染，而且接種之羅勒幼苗呈現與田間相類似的病徵。棉蚜(Aphis gossypii)及桃蚜(Myzus persicae)等兩種蚜蟲可以非持續型方式媒介傳播此病毒。以電顯觀察部分純化病毒可見到直徑約30 nm之球型粒子。病毒之物理性質包括熱不活化溫度介於60-70℃、耐稀釋度為10-3-10-4、室溫下耐保存性為8天、而-70℃下其活性則可維持達10週以上。部分純化病毒經十二烷基硫酸鈉聚丙烯胺膠體電泳(sodium dodecyl sulfate polyacrylamide gel electrophoresis, SDS-PAGE)可測得分子量分別為42和26 kDa之兩種蛋白質。凝膠雙向免疫擴散反應或酵素免疫分析法(ELISA)試驗結果顯示本病毒可與Broad bean wilt virus 抗體發生特異性反應但卻不與Cucumber mosaic virus 抗體發生反應。依據病毒粒子型態及大小、蚜蟲傳播模式、血清學及鞘蛋白特性等分類特徵，初步判定此病毒應屬Comoviridae科Fabavirus屬之Broad bean wilt virus；此為台灣地區田間羅勒病毒病害之首度報導。

Abstract
An unusual disease occurred in bush basil (Ocimum basilicum L.) fields mainly in Pingtung County and to a lesser extent in Kaohsiung County in southern Taiwan. Foliar symptoms of affected plants included mosaic and leaf distortion. A virus isolate was obtained from the symptomatic basil and subsequently tested for its infectivity on 21 plant species from 5 families by mechanical inoculation. Three species in Chenopodiaceae and one of the plants from Solanaceae were infected. Inoculation of the isolated virus on basil seedling induced symptoms similar to those observed from the fields. Two aphid species, Aphis gossypii Glover and Myzus persicae Sulzer, were found to transmit the virus in a non-persistent manner to healthy basil seedlings. The virus was partially purified. By electron microscopy, isometric virus particles, approximately 30 nm in diameter, were visible. The thermal inactivation point of the virus isolate was 60-70C, dilution end-point was 10-3 to 10-4 and the longevity in vitro of infected sap was 8 days at 24C and more than 10 weeks at -70C. Two protein species with molecular weights of about 42 and 26 kDa were detected after partially purified virus was denatured with sodium dodecyl sulfate (SDS) and analyzed by polyacrylamide gel electrophoresis. The partially purified virus and crude sap from infected Chenopodium quinoa leaves reacted specifically with Broad bean wilt virus (BBWV) antiserum in SDS-agar immunodiffusion test but not with Cucumber mosaic virus (CMV) antiserum. In indirect enzyme linked immunosorbent assay (ELISA), CMV polyclonal antibody did not show correlation with partially purified preparations. The virus was identified as Broad bean wilt virus a member of the Fabavirus genus belonging to the family Comoviridae on the basis of its particle morphology and size, manner of aphid transmission, serology and capsid protein. This was the first report on the natural occurrence of a plant virus of the genus Fabavirus to cause a disease on bush basil in Taiwan.

Table of Contents
Abstract in English……………………………………………...…vi
Abstract in Chinese………………………………………...….....viii
Acknowledgements………………………………………….….…ix
Table of Contents………………………………………….…...…xii
List of Tables…………………………………………………..…xiv
List of Illustrations…………………………………………..……xv
I.Introduction……………………………………………….…1
II.Materials and Methods………………………………………7
A. Virus source and isolation…………………….…..….7
B. Plants and growing conditions………………………..8
C. Aphid colonies, source and test plants………………..8
1. Cotton aphid transmission characteristics
of BsMV…………………………...…………..9
1) Preacquisition fasting time………………..10
2) Acquisition access time………………...…11
3) Inoculation access time……………...……11
4) Retention time………………………….....12
5) Number of aphids………………………....12
2. Green peach aphid transmission efficiency of BsMV………………………………...……....13
D. Test of host range………………………………...…13
E. Physical properties of BsMV…………...……….…..14
1. Dilution end-point…………………………………15
2. Thermal inactivation point………..……………….15
3. Longevity in vitro…………………………………16
F. Electron microscopy…………………………...……17
1. Negative staining………………………………….17
G. Virus purification……………………………………17
H. Determination of the molecular weight of viral coat
protein by polyacrylamide gel electrophoresis…...…19
I. Serological assays……..……….……………………20
1. Indirect ELISA……………...…………………….20
2. Ouchterlony test………………………………...…21
III. Results………………………………………………...…....23
IV. Discussion……………………………………………...…..51
V. References…………………………………...………….….58
VI. Biographical sketch……………………………………...…64

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