橡膠樹多主棒孢病菌Cassiicolin基因條形碼數據庫構建及分子檢測技術

李博勛 劉先寶 馮艷麗 解惠婷 黃貴修

摘 ?要??多主棒孢（Corynespora cassiicola）是為害我國主要熱帶作物的植物病原真菌，其寄主范圍廣、形態差異顯著、癥狀類型多樣，且含有一種寄主專化性毒素Cassiicolin，存在6種毒素類型。本研究利用已公布的Cassiicolin毒素基因（Cas1～Cas6型），構建了含287個菌株的國內橡膠樹和部分熱帶作物多主棒孢Cassiicolin基因條形碼數據庫，建立了一套特異性強、靈敏度高的分子檢測技術，可檢測100 pg/μL的目標基因組DNA，系統分析了我國橡膠樹、木薯、番木瓜、瓜菜等主要熱帶作物的919株多主棒孢的毒素類型，發現國內僅存在Cas2和Cas5這2種毒素類型，其中Cas5型的菌株占94.8%，為橡膠樹多主棒孢的優勢種群和特有毒素類型。而Cas2型是橡膠樹和其他作物多主棒孢共有的毒素型。系統發育分析發現，不同毒素類型的多主棒孢菌株與寄主來源密切相關，但與地理來源沒有明顯的相關性，且Cas5型多主棒孢具有明顯的寄主專化性。通過構建多主棒孢Cassiicolin基因條形碼數據庫，為明確我國主要熱帶作物多主棒孢病菌的種群結構和優勢種群情況，發掘、保存多主棒孢菌種資源以及制定病害的防治策略上具有重要的指導意義。

關鍵詞 ?多主棒孢;Cassiicolin毒素基因;DNA條形碼;分子檢測中圖分類號??S763.7??????文獻標識碼??A

Construction of Cassiicolin Genes Barcode Database and Molecular Detection Technology of Corynespora cassiicola?from Hevea brasiliensis?in China

LI Boxun， LIU Xianbao， FENG Yanli， XIE Huiting， HUANG Guixiu^*

Environment and Plant Protection Institute， Chinese Academy of Tropical Agricultural Sciences / Key Laboratory of Integrated Pest Management on Tropical Crops， Ministry of Agriculture and Rural Affairs / Hainan Key Laboratory for Monitoring and Control of Tropical Agricultural Pests， Haikou， Hainan 571101， China

Abstract ?Corynespora cassiicolais a plant pathogenic fungus， which is harmful to major tropical crops in China. It has a wide range of hosts， significant morphological differences， diverse symptoms and contains a host-specific toxin， which encoding up to six distinct protein isoforms. In the study， we used public database to construct Cassiicolin genes barcode database ofC. cassiicolaform major tropical crops in China.?At the same time， we established a set of molecular detection techniques with high specificity and sensitivity， which could detect 100 pg/μL target genomic DNA. Then， we systematically detected the isoforms cassiicolin ofC. cassiicola， which were isolated from rubber tree， cassava， papaya， melon and vegetable in China. There were only two isoforms Cassiicolin in China， among which the Cas5 isoform Cassiicolin accounted for 94.8%， and had obvious host specificity and was specific and advantage population on rubber tree in China. Phylogenetic tree analysis showed that distinct isoforms Cassiicolin ofC. cassiicolawas closely related to host origin， but not to geographical origin. The results of the research would significantly contribute to the population genetic structure， dominant population， preservation ofC. cassiicolaand control strategies.

Keywords ?Corynespora cassiicola; Cassiicolin genes; DNA barcoding; molecular detection

DOI10.3969/j.issn.1000-2561.2019.09.015

多主棒孢（Corynespora cassiicola）是一種重要的植物病原真菌，其寄主范圍十分廣泛，可侵染為害熱帶地區主要經濟作物（橡膠樹、木薯）、瓜菜、果樹和花卉等，生理小種復雜，侵染破壞力極強，病斑類型多樣，具有豐富的遺傳多種樣性^[1-4]。該菌含有特別的寄主專化性毒素Cassiicolin，是一種新型結構的毒素蛋白，這也是該菌最主要的致病因子，存在6種毒素類型^[5]。……