灰葡萄孢蚕豆分离物的遗传多样性

doi:10.3864/j.issn.0578-1752.2014.12.006

摘要/Abstract

摘要： 【目的】灰葡萄孢是引起蚕豆赤斑病、花腐病和荚腐病的重要病原菌，其为包含两个称作类群I和类群II的复合种，具有高度的遗传变异。然而，危害蚕豆的灰葡萄孢复合种地位及遗传特征尚未被研究。论文旨在对中国6个不同地理来源蚕豆灰葡萄孢复合种的地位进行鉴定，研究其遗传多样性及群体间的遗传分化，为探明病害流行规律及制定防治策略提供依据。【方法】在PDA培养基上观察分离物形态特征。用Flipper和Boty特异性引物检测分离物的转座子基因型。葡萄孢复合种所属类群通过Bc-hch基因的PCR扩增酶切多态性鉴定，SSR标记分析解析其遗传多样性和遗传分化。【结果】100个灰葡萄孢分离物均为灰葡萄孢类群II，共产生菌核型、菌丝型和分生孢子型3种类型菌落形态，其中76个分离物为菌核型。92个分离物为转座子基因型，分别含有Boty、Flipper和Boty+Flipper，其中仅含Boty分离物最多，占检测分离物的61.0%。江苏分离物只含有单一Flipper，重庆、四川、甘肃和河北分离物只含有单一Boty。基于SSR分析，100个分离物被鉴定为92个多位点基因型，6个群体的平均基因多样性指数和平均基因型多样性指数分别为0.5140和0.9982；分子方差分析（AMOVA）发现群体内遗传变异占总变异的86.70%。标准关联指数rD值范围为0.0312—0.1261，平均0.0491；遗传固定指数（FST）在0.0085—0.2650，平均为0.1330。UPGMA聚类将100个分离物划分为10个遗传组，大部分遗传组包含不同地理来源的分离物。贝叶斯（Bayesian）推理分析将92个多位点基因型分为两个遗传类群，其中重庆和四川群体聚为一类，青海群体单独聚为一类，而江苏、河北和甘肃群体由两个不同遗传类群组成。【结论】源于蚕豆的灰葡萄孢群体由灰葡萄孢类群II组成，分离物的菌落形态以菌核型为主，绝大多数分离物为转座子基因型，但转座子基因型分布明显存在地域差异。调查的6个群体都具有高水平的遗传多样性，遗传变异主要来源于群体内，生殖方式以有性繁殖为主，大部分群体间存在中等到大的遗传分化。

关键词: 蚕豆 , 赤斑病 , 灰葡萄孢 , 转座子 , 遗传多样性

Abstract: 【Objective】Botrytis cinerea is one of the most important pathogens causing chocolate spot, blossom blight and pod rot in broad bean. B. cinerea is a species complex that comprises two cryptic species, Group I and Group II, and exhibits a great morphological and genetic diversity. However, the status of B. cinerea species complex from broad bean as well as their genetic characteristics have been poorly understood. Therefore, this study aims to determine the status and investigate the genetic diversity and genetic differentiation of isolates of B. cinerea complex from broad bean obtained from six different geographic regions. 【Method】The B. cinerea isolates were observed for cultural characteristics on potato dextrose agar. Transposons types of the isolates were detected using Flipper- and Boty-specific primers. Group determination of the isolates was analyzed by PCR-RFLP of Bc-hch gene, and their genetic characteristics was analyzed using SSR markers. 【Result】 All the 100 isolates were identified as B. cinerea group II. Three colony types, including mycelial type, sclerotial type and conidial type, were produced, and 76 isolates belonged to sclerotial type. Three transposon genotypes were identified in 92 isolates, which contained Boty-only, Fliper-only and Boty + Fliper genotypes. The most prevalent transposon genotype was Boty-only which was detected in isolates from Chongqing, Sichuan, Gansu and Hebei, while genotype Fliper-only was only found in isolates from Jiangsu. SSR analysis detected 92 multilocus genotypes (MLGs) among the 100 isolates with averaged gene and genotypic diversity of 0.5140 and 0.9982, respectively. Analysis of AMOVA revealed that genetic variation within populations accounted for 86.70% of the total genetic variation. The standardized index of association (rD) was estimated to range from 0.0312 to 0.1261 with the average value of 0.0491. In all comparsions, the values of fixation index (FST) ranged from 0.0085-0.2650 with the average value of 0.1330. All 100 isolates were divided into 10 genetic groups by UPGMA clustering analysis, and most of genetic groups included isolates from different geographic regions. In addition, two genetic groups were inferred in 92 MLGs by Bayesian analysis, and both populations from Chongqing and Sichuan belonged to one group, the population from Qinghai belonged to the other group, while popultions from Jiangsu, Hebei and Gansu were admixture of two genetic groups.【Conclusion】The populations of B. cinerea complex from broad bean made up of only B. cinerea group II. The sclerotial type was main colony morphology. Most of the isolates belonged to transposon genotypes whose distribution existed distinct geographic difference. Great genetic diversity was found in all six populations, and genetic variation mainly presented within populations. Sexual reproduction was the main mode of reproduction. Moderate to large genetic differentiation was detected among most populations.

Key words: Vicia faba , chocolate spot , Botrytis cinerea , transposon , genetic diversity

黄燕1, 2, 朱振东1, 段灿星1, 武小菲1, 东方阳2. 灰葡萄孢蚕豆分离物的遗传多样性[J]. 中国农业科学, 2014, 47(12): 2335-2347.

HUANG Yan-1, 2 , ZHU Zhen-Dong-1, DUAN Can-Xing-1, WU Xiao-Fei-1, DONG Fang-Yang-2. Genetic Diversity of Botrytis cinerea Isolates from Broad Bean[J]. Scientia Agricultura Sinica, 2014, 47(12): 2335-2347.

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