湖北省稻瘟病菌群体遗传多样性及水稻品种（系）抗瘟性研究
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摘要
稻瘟病是我国水稻的“三大病害”之一,该病在湖北主要水稻产区均有不同程度发生危害,尤其是在山区、丘陵地带发生频率较高,危害程度较大,造成的经济损失也最为严重。稻瘟病的发生与流行,除受稻瘟菌生理小种的组成及其致病力强弱的影响外,水稻品种的抗性、地理环境与气候条件等也是影响病情的重要因素。因此,明确稻瘟菌生理小种类群、致病性和地区分布,对进一步研究病菌的群体遗传多样性具有重要的意义。鉴定分析水稻品种(系)的抗瘟性,对抗病品种的合理利用和抗病良种的选育,以及有效控制稻瘟病的发生与危害具有重要实践应用价值。
本研究采用水稻苗期叶片人工离体接种的方法,鉴定了湖北省主要水稻种植区稻瘟菌生理小种类群及其种群分布,并应用分子生物学方法对病菌的群体遗传结构进行分析,从分子角度明确了湖北省稻瘟菌的遗传多样性。在此基础上,对湖北省稻瘟菌主要菌株的致病性进行鉴定,并对部分水稻主栽品种(系)进行抗病性分析。为湖北省稻瘟病害防治和水稻品种布局以及新品种培育等方面提供科学依据。取得的主要研究结果如下:
1.稻瘟病菌生理小种组成与地理分布。对采自湖北省13个县市稻瘟病标样分离得到的89个稻瘟菌单孢分离物,采用七个鉴别品种进行人工接种,共鉴定出6群20个生理小种,除ZD种群外,ZA、ZB、ZC和ZG、ZE、ZF都有出现,其中ZB群为优势种群,ZB_(31)为优势生理小种。ZG种群的ZG_1小种出现频率仅次于ZB_(31)生理小种。其它小种出现的频率均低于9%。山区与平原交界处和丘陵种植区生理小种分布相对较丰富,有ZA、ZB、ZC和ZG四个种群的13个生理小种,且多为籼型小种群。分析其原因,小种的分布与不同地区种植水稻品种类型、耕作制度、地理环境与气候条件等密切相关。
2.指纹图谱分析。应用指纹图谱方法从分子角度分析了湖北省稻瘟病菌109个菌株的群体遗传结构,明确了在64.872%相似水平下湖北省稻瘟病菌由以A5为优势谱系的13类谱系组成,A5的构成最复杂。稻瘟病菌的山区生态系遗传结构比丘陵和平原生态系更具遗传多样性,且寄主品种组成多样化程度越高,病原菌群体遗传多样性越丰富。指纹图谱分析测定结果与生理小种鉴定结果二者不存在明显对应关系,但指纹鉴定结果更稳定。
3.稻瘟菌致病力分析。根据稻瘟病菌生理小种和聚类分析结果选择出具有代表性的51个菌株,进行致病力分析。结果显示,各代表菌株致病频率差异较大,其范围分布在19.61%～70.59%之间。总的分布基本都能进行致病力归类,高致病菌株在各类中呈特异的分布。鄂西北菌株致病力强且致病类型多,高、强、中、弱4个致擦ψ榫蟹植?鄂北和鄂东北到鄂东南菌株致病类型次之,各组也基本都有分布;鄂中部菌株致病力相对较弱,主要为中弱型菌株。各致病菌类型的分布,可能与当地的地理环境等条件有一定的关系。
4.水稻品种(系)抗瘟性分析。对选用的49个水稻品种(系)抗瘟性鉴定结果分析显示,水稻品种(系)对稻瘟菌的抗感性差异较大,抗病频率分布在37.74%～77.36%之间。同时聚类分析表明,水稻品种(系)的抗病能力的表型反应是其对稻瘟菌抗性机理的具体表现。新品种中VE6208、华413、矮川香29B三个品种抗性属于高抗亚类,可应用于实践生产。
Rice blast is one of the three mian diseases in China. It's vary occurrence in rice producing areas in Hubei Province, especially has a high frequency, greater damage and causing economic losses in mountainous and hilly areas. The occurrence of the epidemic is impacted by the important factor such as resistance of rice varieties, geographical environment and climatic conditions, besides the inherent strength of Race composition and pathogenicity. Therefore, it's a great significance to definitude the physiological races, pathogenicity and geographical distribution of the rice blast fungus for the further study of the genetic diversity. It's very important to identify the anti-blast of rice varieties (lines) for rational use of resistant varieties, breeding disease-resistant varieties, and diseases control.
The identification of Magnaporthe grisea Physiological Races and populations distribution in the major rice growing areas and rice blast often hair zone of Huber province based on the rice seedling stage in vivo and in vitro vaccination experiments, and identified population genetic structure of rice blast fungus by molecular biology Ways in these area, Defined Magnaporthe grisea genetic diversity From the molecular perspective. Identified the pathogenic of main rice blast fungus strains and analyzed disease resistance of parts of rice varieties (lines) in Huber Province for blast victims on this basis.
It is providing a scientific basis to control rice blast and layout varieties areas as well as cultivate new varieties of the main research results are as follows:
1. Identification and analysis of the Physiological Races of Rice Blast Fungus
From the identification of 89 isolates in 2007 and 2008, there were 6 populations 20 races in addition to ZD group in the counties of Huber province by testing 89 isolates and using distinguish host of China. Their dominant group is ZB, frequency is 55.1%, the frequency of ZA comes to 20.2% following it. Their dominant races are ZB31, Frequency is 20.22%, the frequency of ZG1 and ZB15 also follow it, come to 13.48% and 8.99%, they belong to the group of indica races. Structures of the rice blast fungus populations in the different area were obviously dissimilar. The highest frequency of other races does not exceed 6% at the same time. The distributions of races are relatively abundant in the junction of mountains and plains also called hills, there are thirteen race in four groups. And most of them are the indicate hills environment is also suitable for rice blast happened. The distribution of races in regions is closely related to the types of rice varieties, farming systems, geography and climate.
2. Fingerprint analysis
Genetic structures of 109 isolates of Magnaporthe grisea of Huber Province were analyzed by fingerprinting method from the molecular perspective, Divided them into 13 lineages, that the dominant is A5 at the level of 66.07%, and the composition of lineage A5 is the most complex one. The composition of dominant Lineage is more complex than the rare. The genetic structure of Magnaporthe grisea of mountain ecosystems is more diversity than the hills and plains, and there is a regular pattern that the change of population genetic diversity is the same with the composition of host variation. The results of fingerprint Determination is very different with the results of the identification of physiological race, but the fingerprint Determination is more stable.
3. Pathogenicity analysis of Magnaporthe grisea.
The pathogenicity of 53 isolates of Magnaporthe grisea mainly from combination of physiological races and the results of cluster analysis was analyzed by inoculation onto 49 rice varieties. Their pathogenic frequency (FP) varied greatly, from 19.6 1% to 70.59%. Their pathogenicity is related to the origins at same extent. Virulence of the isolates from Enshi area was the strongest with 50% PF, The general distribution can be classified by pathogenicity basically, and high pathogenic isolates distributed in the every lineage distinctively. The pathogenic of isolates from northwest of Huber are highly and various, distributed from high to low lingages of pathogenic. From the north to east and southeast are followed, but they also distribute in every lingages. The middle and weak isolates mainly distribute in the central, plain and lake area. Distribution of various types of pathogens may have a certain relationship with local geographical conditions and environment.
4. Identification of blast resistance among Rice varieties
The resistance of 49 varieties (lines) mainly from Hubei province was analyzed by inoculated with 53 magnaporthe grisea, their resistance frequencies (RF) are also varied greatly, between 37.74% and 77.36%. Fingerprint analysis showed that the phenotype response of Rice varieties(lines) to rice blast fungus is a concrete manifestation of the resistance mechanism, and the resistance mechanism of rice varieties(lines) may have a certain commonality. The Resistant frequencies of five New varieties, VE6208, Hua413, Aichuanxiang29B,T1007 and VE6223 are in the highest resistance sub-lineages, accoding to the RF, VE6208, Hua413, Aichuanxiang29B are the best varieties that can be applied to actual production.
本研究采用水稻苗期叶片人工离体接种的方法,鉴定了湖北省主要水稻种植区稻瘟菌生理小种类群及其种群分布,并应用分子生物学方法对病菌的群体遗传结构进行分析,从分子角度明确了湖北省稻瘟菌的遗传多样性。在此基础上,对湖北省稻瘟菌主要菌株的致病性进行鉴定,并对部分水稻主栽品种(系)进行抗病性分析。为湖北省稻瘟病害防治和水稻品种布局以及新品种培育等方面提供科学依据。取得的主要研究结果如下:
1.稻瘟病菌生理小种组成与地理分布。对采自湖北省13个县市稻瘟病标样分离得到的89个稻瘟菌单孢分离物,采用七个鉴别品种进行人工接种,共鉴定出6群20个生理小种,除ZD种群外,ZA、ZB、ZC和ZG、ZE、ZF都有出现,其中ZB群为优势种群,ZB_(31)为优势生理小种。ZG种群的ZG_1小种出现频率仅次于ZB_(31)生理小种。其它小种出现的频率均低于9%。山区与平原交界处和丘陵种植区生理小种分布相对较丰富,有ZA、ZB、ZC和ZG四个种群的13个生理小种,且多为籼型小种群。分析其原因,小种的分布与不同地区种植水稻品种类型、耕作制度、地理环境与气候条件等密切相关。
2.指纹图谱分析。应用指纹图谱方法从分子角度分析了湖北省稻瘟病菌109个菌株的群体遗传结构,明确了在64.872%相似水平下湖北省稻瘟病菌由以A5为优势谱系的13类谱系组成,A5的构成最复杂。稻瘟病菌的山区生态系遗传结构比丘陵和平原生态系更具遗传多样性,且寄主品种组成多样化程度越高,病原菌群体遗传多样性越丰富。指纹图谱分析测定结果与生理小种鉴定结果二者不存在明显对应关系,但指纹鉴定结果更稳定。
3.稻瘟菌致病力分析。根据稻瘟病菌生理小种和聚类分析结果选择出具有代表性的51个菌株,进行致病力分析。结果显示,各代表菌株致病频率差异较大,其范围分布在19.61%～70.59%之间。总的分布基本都能进行致病力归类,高致病菌株在各类中呈特异的分布。鄂西北菌株致病力强且致病类型多,高、强、中、弱4个致擦ψ榫蟹植?鄂北和鄂东北到鄂东南菌株致病类型次之,各组也基本都有分布;鄂中部菌株致病力相对较弱,主要为中弱型菌株。各致病菌类型的分布,可能与当地的地理环境等条件有一定的关系。
4.水稻品种(系)抗瘟性分析。对选用的49个水稻品种(系)抗瘟性鉴定结果分析显示,水稻品种(系)对稻瘟菌的抗感性差异较大,抗病频率分布在37.74%～77.36%之间。同时聚类分析表明,水稻品种(系)的抗病能力的表型反应是其对稻瘟菌抗性机理的具体表现。新品种中VE6208、华413、矮川香29B三个品种抗性属于高抗亚类,可应用于实践生产。
Rice blast is one of the three mian diseases in China. It's vary occurrence in rice producing areas in Hubei Province, especially has a high frequency, greater damage and causing economic losses in mountainous and hilly areas. The occurrence of the epidemic is impacted by the important factor such as resistance of rice varieties, geographical environment and climatic conditions, besides the inherent strength of Race composition and pathogenicity. Therefore, it's a great significance to definitude the physiological races, pathogenicity and geographical distribution of the rice blast fungus for the further study of the genetic diversity. It's very important to identify the anti-blast of rice varieties (lines) for rational use of resistant varieties, breeding disease-resistant varieties, and diseases control.
The identification of Magnaporthe grisea Physiological Races and populations distribution in the major rice growing areas and rice blast often hair zone of Huber province based on the rice seedling stage in vivo and in vitro vaccination experiments, and identified population genetic structure of rice blast fungus by molecular biology Ways in these area, Defined Magnaporthe grisea genetic diversity From the molecular perspective. Identified the pathogenic of main rice blast fungus strains and analyzed disease resistance of parts of rice varieties (lines) in Huber Province for blast victims on this basis.
It is providing a scientific basis to control rice blast and layout varieties areas as well as cultivate new varieties of the main research results are as follows:
1. Identification and analysis of the Physiological Races of Rice Blast Fungus
From the identification of 89 isolates in 2007 and 2008, there were 6 populations 20 races in addition to ZD group in the counties of Huber province by testing 89 isolates and using distinguish host of China. Their dominant group is ZB, frequency is 55.1%, the frequency of ZA comes to 20.2% following it. Their dominant races are ZB31, Frequency is 20.22%, the frequency of ZG1 and ZB15 also follow it, come to 13.48% and 8.99%, they belong to the group of indica races. Structures of the rice blast fungus populations in the different area were obviously dissimilar. The highest frequency of other races does not exceed 6% at the same time. The distributions of races are relatively abundant in the junction of mountains and plains also called hills, there are thirteen race in four groups. And most of them are the indicate hills environment is also suitable for rice blast happened. The distribution of races in regions is closely related to the types of rice varieties, farming systems, geography and climate.
2. Fingerprint analysis
Genetic structures of 109 isolates of Magnaporthe grisea of Huber Province were analyzed by fingerprinting method from the molecular perspective, Divided them into 13 lineages, that the dominant is A5 at the level of 66.07%, and the composition of lineage A5 is the most complex one. The composition of dominant Lineage is more complex than the rare. The genetic structure of Magnaporthe grisea of mountain ecosystems is more diversity than the hills and plains, and there is a regular pattern that the change of population genetic diversity is the same with the composition of host variation. The results of fingerprint Determination is very different with the results of the identification of physiological race, but the fingerprint Determination is more stable.
3. Pathogenicity analysis of Magnaporthe grisea.
The pathogenicity of 53 isolates of Magnaporthe grisea mainly from combination of physiological races and the results of cluster analysis was analyzed by inoculation onto 49 rice varieties. Their pathogenic frequency (FP) varied greatly, from 19.6 1% to 70.59%. Their pathogenicity is related to the origins at same extent. Virulence of the isolates from Enshi area was the strongest with 50% PF, The general distribution can be classified by pathogenicity basically, and high pathogenic isolates distributed in the every lineage distinctively. The pathogenic of isolates from northwest of Huber are highly and various, distributed from high to low lingages of pathogenic. From the north to east and southeast are followed, but they also distribute in every lingages. The middle and weak isolates mainly distribute in the central, plain and lake area. Distribution of various types of pathogens may have a certain relationship with local geographical conditions and environment.
4. Identification of blast resistance among Rice varieties
The resistance of 49 varieties (lines) mainly from Hubei province was analyzed by inoculated with 53 magnaporthe grisea, their resistance frequencies (RF) are also varied greatly, between 37.74% and 77.36%. Fingerprint analysis showed that the phenotype response of Rice varieties(lines) to rice blast fungus is a concrete manifestation of the resistance mechanism, and the resistance mechanism of rice varieties(lines) may have a certain commonality. The Resistant frequencies of five New varieties, VE6208, Hua413, Aichuanxiang29B,T1007 and VE6223 are in the highest resistance sub-lineages, accoding to the RF, VE6208, Hua413, Aichuanxiang29B are the best varieties that can be applied to actual production.
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