台灣梨衰弱病菌質體檢測技術之研發與應用

Development and Application of PCR-based Strategies for the Detection of the Phytoplasma Causing Pear Decline in Taiwan

劉秀玲(H. L. Liu) ； 林長平(C. P. Lin)

《植物病理學會刊》 16卷1期 (2007/03) Pp. 1-10

https://doi.org/10.6649/PPB.200703_16(1).0001

摘要

從1994年六月起，台灣中部梨樹栽培區發生台灣梨衰弱病(pear decline in Taiwan, PDTW)，其病徵表現與國外發表之梨衰弱病(pear decline)極為相似，皆會引起罹病植株出現紅葉、衰弱萎凋等病徵，嚴重時造成植株死亡。經由針對該病原之核醣體RNA基因序列(rDNA)進行研究，本研究室已證實此病害係由屬於apple proliferation group (group 16SrX)之台灣梨衰弱病菌質體(PDTW phytoplasma)所引起。本研究利用南方雜配實驗，證實PDTW phytoplasma之核醣體RNA基因於基因體中應具二重複套組，適合做為檢測用之標的序列。研究中亦針對此PDTW phytoplasma之rDNA陸續開發出多項聚合酵素連鎖反應(polymerase chain reaction, PCR)之檢測技術，其中包括專一性放大式聚合酵素連鎖反應(specific booster PCR)、多引子聚合酵素連鎖反應(multiplex PCR)、巢式聚合酵素連鎖反應(nested PCR)與PCR-RFLP (PCR-restriction fragment length polymorphism)等，可供田間罹病梨株、媒介昆蟲及梨接穗之病原菌檢測。本研究並由2002年至2004年完成歷時3年之各月份罹病梨樹中PDTW phytoplasma之PCR偵測調查，以瞭解罹病梨樹中PDTW phytoplasma含量之季節性變化，結果顯示罹病梨樹中PDTW phytoplasma之檢出率於春季三至五月間開始上升，夏季六至九月份間為病原菌檢出率最高，而冬季落葉期之檢出率則降為零。

關鍵字

台灣梨衰弱病菌質體 ； 聚合酵素連鎖反應 ； 核醣體RNA基因 ； 季節性變化

並列摘要

In 1994, pears exhibited a slow decline symptom were observed in orchards of central Taiwan. According to the molecular evidences of the ribosomal RNA gene (rDNA), the pear decline in Taiwan (PDTW) has been proven to be caused by PDTW phytoplasma closely related to the members of the apple proliferation group (16SrX) in our previous study. According to the rDNA sequence of PDTW phytoplasma, various polymerase chain reaction (PCR)-based strategies including specific booster PCR, multiplex PCR, nested PCR and PCR-RFLP were developed and applied in this study for the detection of the causative agent in pear trees. The study of the seasonal variation in the detection of PDTW phytoplasma was conducted in four pear orchards in Dungshr and Heping. Samples collected from 7-20 infected trees were detected by booster PCR monthly in three consecutive years from 2002 to 2004, Unless there is no leaf sample can be collected from the pear trees in the winter, PDTW phytoplasma in the pear trees can be detected readily. The maximum detection rates of PDTW phytoplasma were obtained in summer.

並列關鍵字

PDTW phytoplasma ； polymerase chain reaction ； ribosomal RNA gene ； seasonal variation