大豆细菌性斑点病菌生理小种研究
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摘要
本研究在我国大豆的主产区，主要是东北的吉林省、辽宁省和黑龙江省，对大豆细菌性斑点病进行了广泛的田间调查，在大量的标本采集后，进行了大豆细菌性斑点病的病原细菌分离、纯化，用吉林20号大豆品种对所采集的菌株进行了病原菌的致病性测定，其方法是：用培养钵培育大豆的幼苗，在单叶上，采用高压喷雾方法(147kPa=1.5kg／cm~2)于叶背喷雾接种，出现水浸状斑即可，接种后套上塑料袋保湿培养24小时。之后置于培养箱或温室中，在20～25℃下培养10～14天。按照如下标准记载结果。感病反应(S)：接种3天后产生坏死斑点，并有水浸状晕圈；抗病反应(R)：接种3天后产生坏死枯斑，无水浸状晕圈；或无反应。之后，又进行了植物病原细菌的染色反应、形态特征、培养性状以及生理生化特征的测定。根据所有的测定结果，确证得到了42个大豆细菌性斑点病菌菌株。
根据有关资料记载及反复的田间观察，主要从东北三省的各个院校及科研院所，收集了可供试验的62个大豆品种。用所得到的42个菌株对这62个大豆品种进行抗感反应测定，其方法同上文所迷的对病原菌的致病性测定方法一致。
在进行了反复的3～5次的抗感反应测定后，按照如下原则确定鉴定品种：1．抗感反应表现稳定性，即：同一菌株对同品种，在相同环境条件下，几次重复接种的抗感反应表现一致；2．抗病类型和感病类型有代表性，即：在所选定的鉴别品种中，即有抗病性的品种，也有感病性的品种；3．品种来源地有代表性，即：大豆品种来自大豆主产区的各地。按照此原则，确定了大豆品种十胜长叶、长农4号、丹豆4号、早丰3号、吉林28号和晋特1号作为大豆细菌性斑点病菌生理小种的鉴别寄主，构成大豆细菌性斑点病菌生理小种鉴别寄主体系。
用所采集到的42个菌株再对上述6个鉴别寄主进行反复接种3～5次，根据测定结果，将采集到的42个菌株划分为14个生理小种，即C_1、C_2、C_3、C_4、C_5、C_6、C_7、C_8、C_9、C_(10)、C_(11)、C_(12)、C_(13)和C_(14)。根据菌株的来源地，确定了这14个生理小种在我国的大豆主产区(主要是吉林省、辽宁省和黑龙江省)的存在与分布情况。
吉林省存在。J龟种C；龟C。龟C．龟C。龟C；屯C，电C：、C；、C；。电C；；龟C；。、C；玉和C。·，共13个，
只小种C；不存在；辽宁省存在小种C；龟C。龟C，咆Cl。和C；，共5个；黑龙江省存在小种C；龟
C。、C；、C：、C；；、C；；、C；。和C；。，共8个．另外，来源于湖北省的1个菌株PSG547，属于小
种C；。这个菌株是在武汉地区采集到的，可以认为湖北省的武汉地区存在PSG的小种C；。。
根据采集菌株的来源地，可将生理小种的分布情况进一步确定到所在省份的地市区．
在吉林省，长春地区存在小种C。、C。、C。、C；、C，屯C；．、C；；和C；；，共8个小种；吉林
地区存在一种G、c：、八；、o；、凸l和o，共6个小种；四平地区存在小种G龟o、乙、o．
和C；。，共5个小种．
在辽宁省，铁岭地区存在小种C；、C．、C；和C1l，共一个小种；沈阳地区存在小种C；；
和C；；，共2个小种．
在黑龙江省，绥化地区存在小种C；、C。、C；；和C；，共4个小种；哈尔滨地区存在小
种C；、C：、C；；和；；，共4个，J、种．
这燃别寄主与所确定的14个生理小种间的交叉互作反应明显，继别品种的反
应在重复试验中表现稳定，能反应出生理小种的特性，鉴别能力较强．反过来，也可利
用这些生理小种的代表菌株来区分品种的抗感反应类群。这对渊品种的抗病基因，筛
选和鉴别生理小种都是有意义的．
本躯别寄主主要来自于我国大豆的品种（品系）资源（十胜长叶来自于日本）．
同国外的一舱别寄主断比较，结果显示，本套鉴别寄主有鉴别力，能将我国大豆主
产区的大豆细菌性斑点病菌生理小种划腑更为清晰，更为切合实际．固此，用本舱
别寄主来确定我国的生理小种的存在与分布，更有实用性与针对性。
本研究在国内首次建立了大豆细菌性斑点病菌生理小种鉴别寄主体系．并首次按此
体系确定了我国大豆主产区大豆细菌性斑点病菌生理＊、种的存在与分布倩况．此项研究
成果可应用于我国大豆品种对大豆细菌性斑点病菌的投病性鉴定，以及大豆细菌性斑点
病菌生理分化的研究，并为抗病育种工作打下可靠的驯．
After extensive field investigation and considerable samples collection in China, mainly in Jilin, Liaoning and Heilongjiang provinces, and then pathogenic isolation and purification of bacterial blight of soybean were made. And pathogenic features on soybean (cultivar Jilin 20) were tested. Primary leaves of 10- to 14-day-old plants (primary leaves just fully expanded) were inoculated with water suspensions of bacteria adjusted to IX 108CFU/ml. Suspensions were introduced into the leaves at a pressure of 147kPa. Norm of the susceptible (S) and resistant (R) reactions is : S= compatible combination, typified by water-soaking after 3 days followed by necrosis; R= incompatible hypersensitive resistant reaction with no water-soaking or no reaction. Then the physiological and biochemical characters of the strains were also tested. Finally, 42 strains of Pseudomonas syringae pv. glycinea were defined.
According to relative data and field survey, 62 soybean cultivars from colleges and institutes in Northeast China were collected, reactions of 62 soybean cultivars to the 42 strains were tested in the same way pathogenic features on soybean (cultivar Jilin 20 ) were done.
After the reaction were repeatedly tested for 3 ~ 5 times, 6 differential hosts, Shishengchangye, Changnong 4, Dandou 4, Zaofeng 3, Jilin 28, and Jinte, were screened. Norm as the differential hosts is: 1. Stability of susceptibility and resistance; 2. Combination of susceptible and resistant cultivars; 3: different cultivars from different areas. Finally the 6 differential hosts consist of identification system.
With the system, 42 strains were fallen into 14 physiological races (C1, C2 C3, C4, C5, C6, C7, C8, Q9, C10, C11, C12, C13 & C14). In the same time, distributions of all of the identified races were described in the China, especially in Jilin, Liaoning and Heilongjiang provinces.
There are race C2,C3,C4,C5,C6,C7,C8,C9,C10,C11,C12,C13 & C14 in Jilin province; there are race C1, C4, C9, C13 & C14 in Liaoning province; there are race C1, C3, C7, C8, C11, C12, C13 & C14 in Heilongjiang province. Besides the strain PSG547 from Hubei province belongs to race
m
. Furthermore according to the origin of strains, the race distribution situation in some districts was also confirmed.
In Jilin province, there are race C2, C3, C5, C6, C9, C10, C11 & C12 at Changchun district; race C5, C8, C11, C12, C13 & C14 at Jilin district; race C2, C4, C7, C10 & C12 at Siping district. In Liaoning province, there are race C1. C4, C9 & C13 at Tieling district; race C13 & C14 at Shenyang district. In Heilongjiang province, there are race C1,C2,C13& C14 at Suihua district; race C7,C8, C11 & C12 at Haerbin district. The strain PSG547 is from Wuhan district in Hubei province, therefore there is race Cio at this district.
The mutual reactions between 6 hosts and 14 races are obvious and stable, and the reactions of 6 hosts can clearly feature 14 races. Meanwhile the strains belonged to these races can be used to divide cultivars into different reaction groups. That is helpful to analyze cultivars' resistant genes, screen and identify races.
The 6 hosts are mainly from native cultivars ( Shishengchangye from Japan). Compared with foreign differential hosts (the hosts confirmed by Cross) , they have stronger differential ability. With the 6 hosts, the races from native areas can be more clearly and practically defined, and distribution of races can more conform with practical situations.
In this thesis, identification hosts system of Pseudomotias syringae pv. glycinea was first established hi China, and distributions of races were also first confirmed in the varieties of districts. The findings of the research can be used to identify soybean cultivars' resistance and pathogenic physiological differentiation, and they are to be of value in breeding field.
根据有关资料记载及反复的田间观察，主要从东北三省的各个院校及科研院所，收集了可供试验的62个大豆品种。用所得到的42个菌株对这62个大豆品种进行抗感反应测定，其方法同上文所迷的对病原菌的致病性测定方法一致。
在进行了反复的3～5次的抗感反应测定后，按照如下原则确定鉴定品种：1．抗感反应表现稳定性，即：同一菌株对同品种，在相同环境条件下，几次重复接种的抗感反应表现一致；2．抗病类型和感病类型有代表性，即：在所选定的鉴别品种中，即有抗病性的品种，也有感病性的品种；3．品种来源地有代表性，即：大豆品种来自大豆主产区的各地。按照此原则，确定了大豆品种十胜长叶、长农4号、丹豆4号、早丰3号、吉林28号和晋特1号作为大豆细菌性斑点病菌生理小种的鉴别寄主，构成大豆细菌性斑点病菌生理小种鉴别寄主体系。
用所采集到的42个菌株再对上述6个鉴别寄主进行反复接种3～5次，根据测定结果，将采集到的42个菌株划分为14个生理小种，即C_1、C_2、C_3、C_4、C_5、C_6、C_7、C_8、C_9、C_(10)、C_(11)、C_(12)、C_(13)和C_(14)。根据菌株的来源地，确定了这14个生理小种在我国的大豆主产区(主要是吉林省、辽宁省和黑龙江省)的存在与分布情况。
吉林省存在。J龟种C；龟C。龟C．龟C。龟C；屯C，电C：、C；、C；。电C；；龟C；。、C；玉和C。·，共13个，
只小种C；不存在；辽宁省存在小种C；龟C。龟C，咆Cl。和C；，共5个；黑龙江省存在小种C；龟
C。、C；、C：、C；；、C；；、C；。和C；。，共8个．另外，来源于湖北省的1个菌株PSG547，属于小
种C；。这个菌株是在武汉地区采集到的，可以认为湖北省的武汉地区存在PSG的小种C；。。
根据采集菌株的来源地，可将生理小种的分布情况进一步确定到所在省份的地市区．
在吉林省，长春地区存在小种C。、C。、C。、C；、C，屯C；．、C；；和C；；，共8个小种；吉林
地区存在一种G、c：、八；、o；、凸l和o，共6个小种；四平地区存在小种G龟o、乙、o．
和C；。，共5个小种．
在辽宁省，铁岭地区存在小种C；、C．、C；和C1l，共一个小种；沈阳地区存在小种C；；
和C；；，共2个小种．
在黑龙江省，绥化地区存在小种C；、C。、C；；和C；，共4个小种；哈尔滨地区存在小
种C；、C：、C；；和；；，共4个，J、种．
这燃别寄主与所确定的14个生理小种间的交叉互作反应明显，继别品种的反
应在重复试验中表现稳定，能反应出生理小种的特性，鉴别能力较强．反过来，也可利
用这些生理小种的代表菌株来区分品种的抗感反应类群。这对渊品种的抗病基因，筛
选和鉴别生理小种都是有意义的．
本躯别寄主主要来自于我国大豆的品种（品系）资源（十胜长叶来自于日本）．
同国外的一舱别寄主断比较，结果显示，本套鉴别寄主有鉴别力，能将我国大豆主
产区的大豆细菌性斑点病菌生理小种划腑更为清晰，更为切合实际．固此，用本舱
别寄主来确定我国的生理小种的存在与分布，更有实用性与针对性。
本研究在国内首次建立了大豆细菌性斑点病菌生理小种鉴别寄主体系．并首次按此
体系确定了我国大豆主产区大豆细菌性斑点病菌生理＊、种的存在与分布倩况．此项研究
成果可应用于我国大豆品种对大豆细菌性斑点病菌的投病性鉴定，以及大豆细菌性斑点
病菌生理分化的研究，并为抗病育种工作打下可靠的驯．
After extensive field investigation and considerable samples collection in China, mainly in Jilin, Liaoning and Heilongjiang provinces, and then pathogenic isolation and purification of bacterial blight of soybean were made. And pathogenic features on soybean (cultivar Jilin 20) were tested. Primary leaves of 10- to 14-day-old plants (primary leaves just fully expanded) were inoculated with water suspensions of bacteria adjusted to IX 108CFU/ml. Suspensions were introduced into the leaves at a pressure of 147kPa. Norm of the susceptible (S) and resistant (R) reactions is : S= compatible combination, typified by water-soaking after 3 days followed by necrosis; R= incompatible hypersensitive resistant reaction with no water-soaking or no reaction. Then the physiological and biochemical characters of the strains were also tested. Finally, 42 strains of Pseudomonas syringae pv. glycinea were defined.
According to relative data and field survey, 62 soybean cultivars from colleges and institutes in Northeast China were collected, reactions of 62 soybean cultivars to the 42 strains were tested in the same way pathogenic features on soybean (cultivar Jilin 20 ) were done.
After the reaction were repeatedly tested for 3 ~ 5 times, 6 differential hosts, Shishengchangye, Changnong 4, Dandou 4, Zaofeng 3, Jilin 28, and Jinte, were screened. Norm as the differential hosts is: 1. Stability of susceptibility and resistance; 2. Combination of susceptible and resistant cultivars; 3: different cultivars from different areas. Finally the 6 differential hosts consist of identification system.
With the system, 42 strains were fallen into 14 physiological races (C1, C2 C3, C4, C5, C6, C7, C8, Q9, C10, C11, C12, C13 & C14). In the same time, distributions of all of the identified races were described in the China, especially in Jilin, Liaoning and Heilongjiang provinces.
There are race C2,C3,C4,C5,C6,C7,C8,C9,C10,C11,C12,C13 & C14 in Jilin province; there are race C1, C4, C9, C13 & C14 in Liaoning province; there are race C1, C3, C7, C8, C11, C12, C13 & C14 in Heilongjiang province. Besides the strain PSG547 from Hubei province belongs to race
m
. Furthermore according to the origin of strains, the race distribution situation in some districts was also confirmed.
In Jilin province, there are race C2, C3, C5, C6, C9, C10, C11 & C12 at Changchun district; race C5, C8, C11, C12, C13 & C14 at Jilin district; race C2, C4, C7, C10 & C12 at Siping district. In Liaoning province, there are race C1. C4, C9 & C13 at Tieling district; race C13 & C14 at Shenyang district. In Heilongjiang province, there are race C1,C2,C13& C14 at Suihua district; race C7,C8, C11 & C12 at Haerbin district. The strain PSG547 is from Wuhan district in Hubei province, therefore there is race Cio at this district.
The mutual reactions between 6 hosts and 14 races are obvious and stable, and the reactions of 6 hosts can clearly feature 14 races. Meanwhile the strains belonged to these races can be used to divide cultivars into different reaction groups. That is helpful to analyze cultivars' resistant genes, screen and identify races.
The 6 hosts are mainly from native cultivars ( Shishengchangye from Japan). Compared with foreign differential hosts (the hosts confirmed by Cross) , they have stronger differential ability. With the 6 hosts, the races from native areas can be more clearly and practically defined, and distribution of races can more conform with practical situations.
In this thesis, identification hosts system of Pseudomotias syringae pv. glycinea was first established hi China, and distributions of races were also first confirmed in the varieties of districts. The findings of the research can be used to identify soybean cultivars' resistance and pathogenic physiological differentiation, and they are to be of value in breeding field.
引文
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[30] 高洁等．我国大豆细菌性斑点病菌生理分化的初步研究．吉林农业大学学报．1998，20(1)：10—12．
[31] Thomas, M.D. and Leary, J.V.. A New Race of Pseudomonas glycinea. Phytopathology, 1980, 70:310-312.
[32] Fett, W.F., and L. Sequeira.. further characterization of physiologic race of Pseudomonas glycinea. Can. J. Bot., 1981, 59:283-287.
[33] Kueharek, T.,stall, B., A bacterial leaf spot of soybean caused by a new race of Pseudomonas syringae pv. glycinea. Review of Plant Pathology, 1986,65(9):509.
[34] Staskawicz, B., K.D. Dahlbeck, H. Keen, and C. Napoli. Molecular characterization of cloned avirulence genes from race 0 and race 1 of Pseudomonas syringae pv. glycinea. J. Bacterol. 1987, 169:5789-5794.
[35] 张克勤译(N．W．Schaad主编)．植物病原细菌鉴定实验指南．贵州人民出版社，1986，1：3．
[36] 方中达．植病研究方法．农业出版社，1979，1：174—175．
[37] Kucharek, T.,stall, B., A bacterial leaf spot of soybean caused by a new race of Pseudomonas syringae pv. glycinea. Review of Plant Pathology, 1986,65(9):509. Surico, G.,Kennedy, B,W.,and Ereolani, G.L.. Multiplication of on soybean primary leaves exposed to aerosolized inoeulum. Phytopatology, 1981,71:532-536.
[38] Matthias Ullerch, Stefan Bereswill, Beate Volksch, Wolfgang Fritsche. Molecular characterization of field isolates of Pseudomonas syringae pv. glycinea differing in coronatine production. Journa of General Microbiology, 1993, 139:1927-1937.
[39] Bruegger B.B. and Keen N.T.. Specific elicitors of glyceollin accumulation in the Pseudomonas glycinea-soybean host-parasite system. Physiological Plant Pathology, 1979,15:43-51.
[40] Gulya, Jr. And dunleavy, J.M.. Inhibition of chlorophyll synthesis by Pseudomonas glycinea. Crop Science, 1979, 19(March-April):261-263.
[41] Robert M. Zacharius and E, win Kailan. Isoflavonoid changes in soybean cell suspensions when challenged with intact bacteria or fungal elicitors. Plant physiology, 1990,135:732-736.
[42] Thomas Hoffman, J. scott Schmidt, Xiangyang Zheng ,and Andrew F. Bent. Isolation of ethylene-lnsensitive soybea mutants that are altered in pathogen susceptibility and gene-for-gene disease resistance. Plant Physiology, 1999, 119:935-949.
[43] Jacyn Baker, C. Gary L. Harmon, Judith a. Glazener, and elizabeth W. Orlandi. A noninvasive technique for monitoring peroxidative and H_2O_2-scavenging activities during interactions between bacterial plant pathogens and suspension cells. Plant Physiology, 1995,108:353-359.
[44] Tamaki, S.J. Kobayashi, D.Y. and Keen, N.T. Sequence domains required for the activity of avirulence genes avrB and avrC from Pseudomonas syringae pv. glycinea. Journal of Bacteriology, 1991,1:301-307.
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