台灣感染萵苣之Bidens mottle virus：病毒鑑定及全長度基因體解序分析

Bidens mottle virus Infecting Lettuce in Taiwan: Identification and Sequence Analysis of Full-Length Viral Genome

陳金枝(Chin-Chih Chen) ； 鄧汀欽(Ting-Chin Deng) ； 黃春惠(Chun-Huei Huang) ； 林碧雲(Bi-Yun Lin) ； 廖吉彥(Jye-Yann Liao)

《台灣農業研究》 61卷3期 (2012/09) Pp. 209-221

https://doi.org/10.6156/JTAR/2012.06103.05

摘要

從田間栽培萵苣之葉片呈現黃化嵌紋病徵組織，獲得一個病毒分離株，命名為WF，其與Potyvirus專一性單株抗體產生正反應，並與Potyvirus屬病毒之簡併式引子對HRP-5/Oligo-dT（14）進行反轉錄-聚合酶鏈鎖反應（Reverse transcription-polymerase chain reaction, RT-PCR）可得到一預期之1.3 kb DNA片段，RT-PCR產物經選殖及定序後，證實其鞘蛋白胺基酸序列與Bidens mottle virus（BiMoV）向日葵分離株（SF-1, GenBank Accession number NC_014325）相同度最高，達99.3%。以WF分離株之純化病毒顆粒所製備完成之多元抗血清（As161），與萵苣分離株WF及由美國ATCC購得之BiMoV標準對照株，於SDS-免疫擴散反應中均產生免疫沈澱反應，且彼此之沈澱帶互相融合呈現同源反應。由分子生物學及血清學之證據顯示萵苣WF分離株屬於BiMoV病毒。本研究亦由花蓮栽培之萵苣獲得一病毒分離株HL及咸豐草上分離得三個病毒分離株B3、B4、及B12，此等分離株與WF抗血清反應後均與WF分離株產生血清同源反應，其大蛋白（polyprotein）胺基酸序列均與BiMoV病毒之相同度達97%以上，顯示在台灣之萵苣及咸豐草已受到BiMoV之感染。本研究針對BiMoV-WF、HL、B3、B4、及B12等病毒分離株均已完成其全長度基因體之核甘酸定序，登錄於GenBank取得登錄序號分別為EU250213、EU250214、EU250212、EU250211、及EU25021。由病毒全長度鞘蛋白胺基酸序列分析分離株間的類緣關係，結果顯示萵苣分離株與咸豐草分離株個別形成分群現象，而向日葵分離株與萵苣分離株類緣關係較相近。本研究為BiMoV的萵苣及咸豐草分離株基因體全長度定序及其分子特性之首次報告。

關鍵字

咸豐草斑紋病毒 ； 萵苣 ； 全長度基因體

並列摘要

A virus isolate-WF was isolated from lettuce leaves that exhibit yellowing mosaic. WF isolate was positively reacted with potyvirus-specific monoclonal antibody. A 1.3-kb DNA fragment was amplified from RNA of WF isolate with the potyvirus-degenerate primer pair (HRP-5/Oligo-dT(14)) by reverse transcription- polymerase chain reaction (RT-PCR). The identity of deduced amino acid sequence of coat proteins of WF isolate to that of Bidens mottle virus sunflower isolate SF-1 (GenBank Acc. NC_014325) was 99.3%. When the antiserum against purified virions of WF isolate was used in SDS-immunodiffusion, identical immunoprecipitation reaction was generated between lettuce isolates WF and the typical isolate of BiMoV (ATCC PV165). WF isolate could be recognized as a strain of BiMoV based on the results of the molecular and serological characterizations. Another lettuce isolate (HL) from Hualien and three isolates from Bidens pilosa (B3, B4, B12) were also serologically identical to isolate of BiMoV-PV165 and the identity in amino acid sequences of viral coat proteins were 97%. Full-length of genomic RNAs of BiMoV-WF, HL, B3, B4, and B12 were sequenced depossited into GenBank (accession numbers EU250213, EU250214, EU250212, EU250211, and EU25021, respectively). The phylogenetic analyses based on the deduced amino acid sequences of the full polyproteins with those of different BiMoV isolates revealed that isolates from the same host was classified into the same group. This is the first report of the full-length genomic sequence and molecular characterization of BiMoV infecting lettuce and bidens.

並列關鍵字

Bidens mottle virus ； Lettuce ； Full-length genomic sequences