大叶黄杨灰斑病病原菌鉴定、生物学特性及五种杀菌剂对其抑制作用
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摘要
为明确大叶黄杨灰斑病病原菌种类、生物学特性和对杀菌剂的敏感性,采用组织分离法获得大叶黄杨灰斑病病原菌,利用形态特征和分子生物学对其进行鉴定,系统研究该病原菌的生物学特性,并用生长速率法测定了80%代森锰锌、80%代森锌、70%百菌清、50%多菌灵和430 g/L戊唑醇5种杀菌剂对该病原菌生长的抑制效果。致病性测定结果表明,从感病植株分离所得菌株能在大叶黄杨叶片上成功定殖并扩展,且从发病部位分离得到的菌株与原接种菌株相同。通过病原菌的宏观形态、显微结构和ITS序列分析,确定该病原菌为斑污拟盘多毛孢Pestalotiopsis maculans;当葡萄糖为碳源、蛋白胨为氮源时,菌丝日生长速度最快,分别为4.08、4.90 mm/d;当温度为21～28℃、pH为7时,菌丝日生长速度最快,分别为5.07～5.16 mm/d和5.21 mm/d;50%多菌灵对该菌的抑制效果最好,抑制中浓度EC50最小,为0.06μg/mL。
In order to identify the species, biological characteristics and sensitivity to fungicides of the pathogen causing the gray leaf mold spot of Euonymus japonicus, the pathogen was obtained by tissue isolation from the infected E. japonicus leaves; it was identified according to its morphological and molecular characters, and its biological characteristics were systematically studied; moreover, the inhibitory effect of five fungicides(80% tiezene, 80% mancozeb, 70% chlorothalonil, 50% carbendazim and 430 g/L tebuconazole) on the pathogen was tested by using growth rate method. Based on pathogenicity test, the mycelium was successfully colonized and expanded on E. japonicus leaves, and the same species strain was isolated from the discolored part. According to the pathogen's morphology, microstructure and ITS sequence, the pathogen was identified as Pestalotiopsis maculans. When the carbon source was glucose and the nitrogen source was peptone, the hyphal growth rates were fastest(4.08, 4.90 mm/d,respectively). When temperature ranged from 21℃ to 28℃ and pH was 7, the hyphal growth rates were fastest(5.07-5.16 and 5.21 mm/d, respectively). 50% carbendazimwettable powder had the best inhibition effect on P. maculans, with a smallest EC50 value of 0.06 μg/mL.
In order to identify the species, biological characteristics and sensitivity to fungicides of the pathogen causing the gray leaf mold spot of Euonymus japonicus, the pathogen was obtained by tissue isolation from the infected E. japonicus leaves; it was identified according to its morphological and molecular characters, and its biological characteristics were systematically studied; moreover, the inhibitory effect of five fungicides(80% tiezene, 80% mancozeb, 70% chlorothalonil, 50% carbendazim and 430 g/L tebuconazole) on the pathogen was tested by using growth rate method. Based on pathogenicity test, the mycelium was successfully colonized and expanded on E. japonicus leaves, and the same species strain was isolated from the discolored part. According to the pathogen's morphology, microstructure and ITS sequence, the pathogen was identified as Pestalotiopsis maculans. When the carbon source was glucose and the nitrogen source was peptone, the hyphal growth rates were fastest(4.08, 4.90 mm/d,respectively). When temperature ranged from 21℃ to 28℃ and pH was 7, the hyphal growth rates were fastest(5.07-5.16 and 5.21 mm/d, respectively). 50% carbendazimwettable powder had the best inhibition effect on P. maculans, with a smallest EC50 value of 0.06 μg/mL.
引文
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Deng YC,Deng Y.2008.Prevention and control of diseases and insect pests of Euonymus japonicas.China Flowers&Horticulture,(10):44-45(in Chinese)[邓运川,邓越.2008.大叶黄杨的病虫害防治.中国花卉园艺,(10):44-45]
Ding R,Wang YL,Li BX,Xie CP.2010.Identification and characterization of mango leaf blight disease pathogen Pestalotiopsis mangiferae.South China Fruits,39(4):20-24(in Chinese)[丁榕,王延丽,李博勋,谢昌平.2010.杧果拟盘多毛孢叶枯病菌鉴定及其生物学特性研究.中国南方果树,39(4):20-24]
Fang ZD.1998.Research method of plant disease(3rd edition).Beijing:China Agriculture Press,pp.123-125(in Chinese)[方中达.1998.植病研究方法(第3版).北京:中国农业出版社,pp.123-125]
Gao GW,Zhou XL,Yue HC,Chi MY,Huang JG.2017.Effect of environmental factors on conidial germination of Euonymus japonicas powdery mildew.Shandong Agricultural Sciences,49(12):90-92(in Chinese)[高国伟,周小林,岳弘辰,迟梦宇,黄金光.2017.环境因子对冬青卫矛白粉病菌分生孢子萌发的影响.山东农业科学,49(12):90-92]
Gao J.2017.Control effect of five fungicides including chlorothalonil on pathogen causing powdery mildew on Euonymus japonicas.Agriculture and Technology,37(22):61(in Chinese)[高军.2017.百菌清等5种杀菌剂对大叶黄杨白粉病的防效试验.农业与技术,37(22):61]
Ge QX,Chen YX,Xu T.2009.Flora Fungorum Sinicorum(Vol.38):Pestalotiopsis.Beijing:Science Press(in Chinese)[葛起新,陈育新,徐同.2009.中国真菌志(第38卷):拟盘多毛孢属.北京:科学出版社]
Jian JG,Zhang WY,Bai T,Chen JY,Chen GX.2009.Experiment against Cercospora destructiva Rav on Euonymus japonicus with ten fungicides in the field.Forest Pest and Disease,(4):30-32(in Chinese)[江建国,张文颖,白涛,陈京元,陈国新.2009.10种杀菌剂对大叶黄杨褐斑病田间防治试验.中国森林病虫,(4):30-32]
Kirk PM,Cannon PF,David JC,Stalpers JA.2001.Ainsworth&Bisby’s dictionary of the fungi(9th edition).Wallingford:CABIPublishing
Li JB,Deng K.2015.Studies on the occurrence and control of Euonymus japonicus.Beijing Agriculture,(30):84-85(in Chinese)[李建波,邓珂.2015.大叶黄杨白粉病发生规律及其防治研究.北京农业,(30):84-85]
Liu QL,Zhang JX,Xu RF.2006.Preliminary study on the bionomics of pathogen of leaf spot of Euonymus japonicus in the northern Henan.Journal of Anhui Agricultural Sciences,34(20):5279-5280(in Chinese)[刘起丽,张建新,徐瑞富.2006.豫北地区大叶黄杨褐斑病病原菌生物学特性的研究.安徽农业科学,34(20):5279-5280]
Liu XY,Xu YM,Wang CH.2006.Study with suggestion and analysis of current status on disease for Euonymus japonicas Thunb.Hunan Forestry Science and Technology,(6):42-45(in Chinese)[刘兴元,徐有明,王长辉.2006.大叶黄杨病害的研究现状分析与建议.湖北林业科技,(6):42-45]
Nag Raj TR.1993.Coelomycetous anamorphs with appendage-bearing conidia.Waterloo,Canada:Mycolgue Publication,pp.231-241
Trapero A,Romero MA,Varo R,Sánchez ME.2003.First report of Pestalotiopsis maculans causing necrotic leaf spots in nursery plants of Arbutus unedo and Ceratonia siliqua in Spain.Plant Disease,87(10):1263
Wang JN,Zhao HH,Wei YY,Li XD,Liang C,Li BD.2016.The pathogen causing Lasiodiplodia twig blight of blueberry.Mycosystema,35(6):657-665(in Chinese)[王佳宁,赵洪海,尉莹莹,李晓东,梁晨,李宝笃.2016.蓝莓毛色二胞枝枯病的病原菌.菌物学报,35(6):657-665]
Wang RJ,Zhao SL,Zhang SF.2015 Main diseases and insect pests of Euonymus japonicus and their control in Longnan City.Journal of Agricultural Catastrophology,5(7):2-4(in Chinese)[王让军,赵淑玲,张少飞.2015.陇南市大叶黄杨主要病虫害及其防治.农业灾害研究,5(7):2-4]
Wei JG,Xu T,Pan XH,Liao WJ.2006.Progress of research on taxonomy of Pestalotiopsis.Journal of Guangxi Agricultural and Biological Science,25(1):78-85(in Chinese)[韦继光,徐同,潘秀湖,廖旺姣.2006.拟盘多毛孢属的分类学研究进展.广西农业生物科学,25(1):78-85]
White TJ,Bruns T,Lee S,Taylor J.1990.Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics.//Innis MA,Gelfand DH,Sninsky JJ,White TJ.PCR protocols:a guide to methods and applications.New York:Academic Press,pp.315-322
Xi ZG,You JM,Zheng L,Liu H,Huang JB.2018.Identification and biological characteristics of the pathogen causing leaf spot on Coptis chinensis in Hubei Province.Journal of Plant Protection,45(2):315-321(in Chinese)[席中刚,游景茂,郑露,刘浩,黄俊斌.2018.湖北省黄连叶斑病病原鉴定及其生物学特性.植物保护学报,45(2):315-321]
Xing GB,Li LH.2016.Cultivation and management technology of Euonymus japonicas.Agriculture of Henan,(14):48-49(in Chinese)[邢贵彬,李连海.2016.大叶黄杨栽培与管护技术.河南农业,(14):48-49]
Xiong YQ,Li CT,Liu XH.2004.A study of the cold resistance of Euonymus japonicus and its application.Chinese Landscape Architecture,(4):36-38(in Chinese)[熊佑清,李崇涛,刘晓辉.2004.大叶黄杨的抗寒性及其应用研究.中国园林,(4):36-38]
Ye H,Wei W,Wu B,Su MS,Wang DX,Jiang ZP.2014.Isolation and primary identification of the pathogen of anthracnose in Camellia oleifera.Acta Agriculturae Universitatis Jiangxiensis,36(2):314-318(in Chinese)[叶航,韦维,吴波,苏明声,王东雪,江泽鹏.2014.油茶炭疽病病原菌的分离和初步鉴定.江西农业大学学报,36(2):314-318]
Yu JY,Dong LK,Wang ZH,Shen J,Xu H,Nie CR,Duan QM.2017.Identification of pathogen for leaf spot disease of Photinia serrulata Lind.and determination of its sensitivity to fungicides.Acta Phytopathologica Sinica,47(4):440-447(in Chinese)[于静亚,董立坤,王志华,沈锦,徐慧,聂超仁,段庆明.2017.石楠叶斑病病原鉴定及对药物敏感性测定.植物病理学报,47(4):440-447]
Zhang LX,Wu ZR,Lin B,Wang Q,Tan GJ.2018.Sensitivity of Alternaria alternata in Trichosanthes kirilowii to four fungicides in Anhui Province.Plant Protection,44(3):203-206(in Chinese)[张立新,吴治然,林彬,王琦,檀根甲.2018.安徽省栝蒌叶枯病菌对4种杀菌剂的敏感性.植物保护,44(3):203-206]
Zhang MY,Wen XL,Zhao MJ,Li XY,Qi HX.2018.Biological characteristics of grape leaf spot pathogen of Pestalotiopsis uvicola(Speg.)Bissett.Journal of Hebei Normal University of Science&Technology,32(1):39-42(in Chinese)[张梦影,温晓蕾,赵美键,李昕颖,齐慧霞.2018.葡萄尤韦可拟盘多毛孢叶斑病病菌的生物学特性.河北科技师范学院学报,32(1):39-42]
Zhao HH,Yue QH,Liang C.2014.The pathogen causing Pestalotiopsis twig dieback of blueberry.Mycosystema,33(3):577-583(in Chinese)[赵洪海,岳清华,梁晨.2014.蓝莓拟盘多毛孢枝枯病的病原菌.菌物学报,33(3):577-583]
Zhao JN,Ma Z,Liu ZP,Shang QX,Zhao XY,Wei YM.2016.Pestalotiopsis clavispora causing leaf spot on strawberry.Mycosystema,35(1):114-120(in Chinese)[赵景楠,马喆,刘正坪,尚巧霞,赵晓燕,魏艳敏.2016.草莓拟盘多毛孢叶斑病的病原菌.菌物学报,35(1):114-120]
Deng YC,Deng Y.2008.Prevention and control of diseases and insect pests of Euonymus japonicas.China Flowers&Horticulture,(10):44-45(in Chinese)[邓运川,邓越.2008.大叶黄杨的病虫害防治.中国花卉园艺,(10):44-45]
Ding R,Wang YL,Li BX,Xie CP.2010.Identification and characterization of mango leaf blight disease pathogen Pestalotiopsis mangiferae.South China Fruits,39(4):20-24(in Chinese)[丁榕,王延丽,李博勋,谢昌平.2010.杧果拟盘多毛孢叶枯病菌鉴定及其生物学特性研究.中国南方果树,39(4):20-24]
Fang ZD.1998.Research method of plant disease(3rd edition).Beijing:China Agriculture Press,pp.123-125(in Chinese)[方中达.1998.植病研究方法(第3版).北京:中国农业出版社,pp.123-125]
Gao GW,Zhou XL,Yue HC,Chi MY,Huang JG.2017.Effect of environmental factors on conidial germination of Euonymus japonicas powdery mildew.Shandong Agricultural Sciences,49(12):90-92(in Chinese)[高国伟,周小林,岳弘辰,迟梦宇,黄金光.2017.环境因子对冬青卫矛白粉病菌分生孢子萌发的影响.山东农业科学,49(12):90-92]
Gao J.2017.Control effect of five fungicides including chlorothalonil on pathogen causing powdery mildew on Euonymus japonicas.Agriculture and Technology,37(22):61(in Chinese)[高军.2017.百菌清等5种杀菌剂对大叶黄杨白粉病的防效试验.农业与技术,37(22):61]
Ge QX,Chen YX,Xu T.2009.Flora Fungorum Sinicorum(Vol.38):Pestalotiopsis.Beijing:Science Press(in Chinese)[葛起新,陈育新,徐同.2009.中国真菌志(第38卷):拟盘多毛孢属.北京:科学出版社]
Jian JG,Zhang WY,Bai T,Chen JY,Chen GX.2009.Experiment against Cercospora destructiva Rav on Euonymus japonicus with ten fungicides in the field.Forest Pest and Disease,(4):30-32(in Chinese)[江建国,张文颖,白涛,陈京元,陈国新.2009.10种杀菌剂对大叶黄杨褐斑病田间防治试验.中国森林病虫,(4):30-32]
Kirk PM,Cannon PF,David JC,Stalpers JA.2001.Ainsworth&Bisby’s dictionary of the fungi(9th edition).Wallingford:CABIPublishing
Li JB,Deng K.2015.Studies on the occurrence and control of Euonymus japonicus.Beijing Agriculture,(30):84-85(in Chinese)[李建波,邓珂.2015.大叶黄杨白粉病发生规律及其防治研究.北京农业,(30):84-85]
Liu QL,Zhang JX,Xu RF.2006.Preliminary study on the bionomics of pathogen of leaf spot of Euonymus japonicus in the northern Henan.Journal of Anhui Agricultural Sciences,34(20):5279-5280(in Chinese)[刘起丽,张建新,徐瑞富.2006.豫北地区大叶黄杨褐斑病病原菌生物学特性的研究.安徽农业科学,34(20):5279-5280]
Liu XY,Xu YM,Wang CH.2006.Study with suggestion and analysis of current status on disease for Euonymus japonicas Thunb.Hunan Forestry Science and Technology,(6):42-45(in Chinese)[刘兴元,徐有明,王长辉.2006.大叶黄杨病害的研究现状分析与建议.湖北林业科技,(6):42-45]
Nag Raj TR.1993.Coelomycetous anamorphs with appendage-bearing conidia.Waterloo,Canada:Mycolgue Publication,pp.231-241
Trapero A,Romero MA,Varo R,Sánchez ME.2003.First report of Pestalotiopsis maculans causing necrotic leaf spots in nursery plants of Arbutus unedo and Ceratonia siliqua in Spain.Plant Disease,87(10):1263
Wang JN,Zhao HH,Wei YY,Li XD,Liang C,Li BD.2016.The pathogen causing Lasiodiplodia twig blight of blueberry.Mycosystema,35(6):657-665(in Chinese)[王佳宁,赵洪海,尉莹莹,李晓东,梁晨,李宝笃.2016.蓝莓毛色二胞枝枯病的病原菌.菌物学报,35(6):657-665]
Wang RJ,Zhao SL,Zhang SF.2015 Main diseases and insect pests of Euonymus japonicus and their control in Longnan City.Journal of Agricultural Catastrophology,5(7):2-4(in Chinese)[王让军,赵淑玲,张少飞.2015.陇南市大叶黄杨主要病虫害及其防治.农业灾害研究,5(7):2-4]
Wei JG,Xu T,Pan XH,Liao WJ.2006.Progress of research on taxonomy of Pestalotiopsis.Journal of Guangxi Agricultural and Biological Science,25(1):78-85(in Chinese)[韦继光,徐同,潘秀湖,廖旺姣.2006.拟盘多毛孢属的分类学研究进展.广西农业生物科学,25(1):78-85]
White TJ,Bruns T,Lee S,Taylor J.1990.Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics.//Innis MA,Gelfand DH,Sninsky JJ,White TJ.PCR protocols:a guide to methods and applications.New York:Academic Press,pp.315-322
Xi ZG,You JM,Zheng L,Liu H,Huang JB.2018.Identification and biological characteristics of the pathogen causing leaf spot on Coptis chinensis in Hubei Province.Journal of Plant Protection,45(2):315-321(in Chinese)[席中刚,游景茂,郑露,刘浩,黄俊斌.2018.湖北省黄连叶斑病病原鉴定及其生物学特性.植物保护学报,45(2):315-321]
Xing GB,Li LH.2016.Cultivation and management technology of Euonymus japonicas.Agriculture of Henan,(14):48-49(in Chinese)[邢贵彬,李连海.2016.大叶黄杨栽培与管护技术.河南农业,(14):48-49]
Xiong YQ,Li CT,Liu XH.2004.A study of the cold resistance of Euonymus japonicus and its application.Chinese Landscape Architecture,(4):36-38(in Chinese)[熊佑清,李崇涛,刘晓辉.2004.大叶黄杨的抗寒性及其应用研究.中国园林,(4):36-38]
Ye H,Wei W,Wu B,Su MS,Wang DX,Jiang ZP.2014.Isolation and primary identification of the pathogen of anthracnose in Camellia oleifera.Acta Agriculturae Universitatis Jiangxiensis,36(2):314-318(in Chinese)[叶航,韦维,吴波,苏明声,王东雪,江泽鹏.2014.油茶炭疽病病原菌的分离和初步鉴定.江西农业大学学报,36(2):314-318]
Yu JY,Dong LK,Wang ZH,Shen J,Xu H,Nie CR,Duan QM.2017.Identification of pathogen for leaf spot disease of Photinia serrulata Lind.and determination of its sensitivity to fungicides.Acta Phytopathologica Sinica,47(4):440-447(in Chinese)[于静亚,董立坤,王志华,沈锦,徐慧,聂超仁,段庆明.2017.石楠叶斑病病原鉴定及对药物敏感性测定.植物病理学报,47(4):440-447]
Zhang LX,Wu ZR,Lin B,Wang Q,Tan GJ.2018.Sensitivity of Alternaria alternata in Trichosanthes kirilowii to four fungicides in Anhui Province.Plant Protection,44(3):203-206(in Chinese)[张立新,吴治然,林彬,王琦,檀根甲.2018.安徽省栝蒌叶枯病菌对4种杀菌剂的敏感性.植物保护,44(3):203-206]
Zhang MY,Wen XL,Zhao MJ,Li XY,Qi HX.2018.Biological characteristics of grape leaf spot pathogen of Pestalotiopsis uvicola(Speg.)Bissett.Journal of Hebei Normal University of Science&Technology,32(1):39-42(in Chinese)[张梦影,温晓蕾,赵美键,李昕颖,齐慧霞.2018.葡萄尤韦可拟盘多毛孢叶斑病病菌的生物学特性.河北科技师范学院学报,32(1):39-42]
Zhao HH,Yue QH,Liang C.2014.The pathogen causing Pestalotiopsis twig dieback of blueberry.Mycosystema,33(3):577-583(in Chinese)[赵洪海,岳清华,梁晨.2014.蓝莓拟盘多毛孢枝枯病的病原菌.菌物学报,33(3):577-583]
Zhao JN,Ma Z,Liu ZP,Shang QX,Zhao XY,Wei YM.2016.Pestalotiopsis clavispora causing leaf spot on strawberry.Mycosystema,35(1):114-120(in Chinese)[赵景楠,马喆,刘正坪,尚巧霞,赵晓燕,魏艳敏.2016.草莓拟盘多毛孢叶斑病的病原菌.菌物学报,35(1):114-120]