美人蕉嵌紋病相關之菜豆黃化嵌紋病毒特性之研究

Characterization of an isolate of Bean yellow mosaic virus associated with canna mosaic in Taiwan

"美人蕉（Canna spp.）為一年或多年生之美人蕉科草本觀賞植物。屏東田間美人蕉嵌紋病葉以奎藜（Chenopodium quinoa）進行單斑分離，獲得一種病毒顆粒長度約為750-830 nm的絲狀病毒，暫稱為CA-FT分離株。該病毒熱不活化溫度為55℃，耐稀釋度為10-4，24℃下活性可維持1天，-80℃下則可保存6個月。機械接種17科68種供試植物，僅藜科之奎藜、紅藜（C. amaranticolor）、綠藜（C. murale）可被感染，於接種葉出現黃斑病徵。電子顯微鏡觀察美人蕉嵌紋病葉及奎藜單斑病組織超薄切片，可在細胞質中觀察到風車狀（pinwheel）及薄板狀（laminated aggregates）內含體。病毒鞘蛋白分子量經電泳及西方轉漬反應分析估算約為32kDa。病毒鞘蛋白經免疫注射白兔後，可製備得力價達稀釋1024倍之抗血清；瓊脂免疫雙向擴散反應（SDS-agar gel double diffusion test）除能與同源抗原反應外，另可與菜豆黃化嵌紋病毒（Bean yellow mosaic virus, BYMV）發生專一性反應。以針對Potyvirus屬病毒鞘蛋白基因之引子對進行反轉錄聚合酶連鎖反應（RT-PCR），美人蕉田間嵌紋病葉及奎藜接種病葉皆可增幅出長度約1263個核苷酸（nucleotide, nt）之核酸產物，其與NCBI GenBank中親緣較近之前12個BYMV序列比對後，發現與中國報導在美人蕉所分離之菜豆黃化嵌紋病毒（BYMV）核苷酸序列之相同度達到93%，胺基酸序列之相似度亦達93%，3'端非轉譯區（3'-NCR）則有95%之相同度。綜合以上結果顯示本研究自美人蕉所分離之病毒應為BYMV之美人蕉病毒株（canna isolate），此亦為菜豆黃化嵌紋病毒在台灣美人蕉上發生的首度報導。"

"Canna (Canna spp.) is an annual ornamental plant. A filamentous virus of about 750-830 nm in particle size, was successfully isolated from mosaic canna plant by mechanical inoculation in Pingtung, and was temporarily named as CA-FT. The thermal inactivation point was 55℃, dilution end point was 10-4 and the longevity in vitro was 1 day at 24℃ and more than 6 month at -800C .The host range test indicated that CA-FT is a virus with very narrow host range and only infected 3 out of 68 species of plants tested. Inclusion bodies of pinwheel and Laminated aggregates, an indication of typical potyvirus infection, were found in cells of infected Chenopodium quinoa and canna. SDS-PAGE showed the virus coat protein contains a subunit with relative molecular weight of about 32 kDa. An antiserum with a titer of 1024 was prepared by immunizing a white rabbit with the purified virions. The antiserum produced can be effectively applied to detect studied virus . The antiserum produced reacted strongly with homologous antigen and with Bean yellow mosaic virus (BYMV), indicating the serological relatedness between the studied canna virus and BYMV. A cDNA fragment, consists of partial NIb, full CP, and 3'NCR of the virus, was amplified with RT-PCR and was cloned. The sequence of the 1263-nt cDNA fragment was found to be corresponding to a cannainfecting BYMV isolate in China. The sequence identities of the CP gene and the 3'-non coding regions (3'-NCR) of both viruses were 93％ and 95％, respectively, while that of CP amino acid sequence was 93％. Based on these results, the virus CA-FT isolated from canna with mosaic symptom should be a strain of BYMV. This is a new record of BYMV infecting canna in Taiwan."