百合灰霉病病原菌鉴定及其部分生物学特性测定
|
摘要
【目的】鉴定北京昌平地区百合灰霉病病原菌,并明确其生物学特性,为该病害的预测、防治及百合抗病育种提供理论依据。【方法】利用柯赫氏法则从东方百合索邦灰霉病病株中分离纯化获得致病菌,结合形态学及分子生物学方法鉴定病原菌种类;并利用菌落生长法和血球计数板孢子计量法研究病原菌的生物学特性。【结果】从索邦灰霉病叶片病斑上分离获得灰霉病病原菌(标记为CX1),利用形态学和分子生物学方法鉴定CX1菌株为灰葡萄孢菌(Botrytis cinerea Pers.)。CX1菌株菌丝生长和产孢的最适培养基为PSA培养基,最适温度为20℃,最适光照环境为光暗交替(12 h光照/12 h黑暗),菌丝生长的最适p H为5～6,产孢的最适pH为6,菌丝生长和产孢的最适碳源分别为可溶性淀粉和蔗糖,最适氮源分别为牛肉膏和硫酸铵。【结论】引起北京昌平地区百合灰霉病的病原菌为灰葡萄孢菌。依据百合灰葡萄孢菌的生物学特性,生产中除使用化学和生物防治外,还可通过改变栽培环境、合理施肥、合理灌溉、合理密植、轮作等农业措施来抑制灰葡萄孢菌的生长和产孢,从而减轻病害发生。
【Objective】The present study was performed to identify the pathogen causing lily gray mold in Changping District,Beijing City,and to determine biological characteristics of the pathogen. The results would provide a scientific basis for forecasting,control and resistance breeding of lily gray mold.【Method】The gray mold pathogen was isolated and purified from Lilium Oriental hybrid Sorbonne strains according to Koch's rule and identified combining morphological characteristics study and molecular biological analysis. Biological characteristics of the pathogen were determined by measuring the diameter of the colony and counting the number of conidia using hemocytometer measurement.【Result】The gray mold pathogen isolated from the pathogenetic lily leaves was marked as CX1. CX1 was identified as Botrytis cinerea Pers. according to morphological characteristics study and molecular biological analysis. For the mycelial growth and sporulation of CX1,the optimum medium was PSA,the optimum temperature was 20 ℃ and the optimum light condition was 12 h∶12 h alternation of light and darkness. The optimal pH value for mycelial growth and sporulation were5-6 and 6,respectively. Soluble starch was the best carbon source and beef extract was the best nitrogen source for mycelial growth. The optimum carbon source and nitrogen source for sporulation were sucrose and(NH4)2 SO4,respectively.【Conclusion】The pathogen of lily gray mold occurs in Changping,Beijing was B. cinerea Pers. Based on the studies about biological characteristics of B. cinerea Pers.,apart from chemical and biological controls,growth of B. cinerea Pers. and sporulation can be inhibited by changing cultivation environment,reasonable fertilization,irrigation,planting density and rotation,so as to reduce the disease damage.
【Objective】The present study was performed to identify the pathogen causing lily gray mold in Changping District,Beijing City,and to determine biological characteristics of the pathogen. The results would provide a scientific basis for forecasting,control and resistance breeding of lily gray mold.【Method】The gray mold pathogen was isolated and purified from Lilium Oriental hybrid Sorbonne strains according to Koch's rule and identified combining morphological characteristics study and molecular biological analysis. Biological characteristics of the pathogen were determined by measuring the diameter of the colony and counting the number of conidia using hemocytometer measurement.【Result】The gray mold pathogen isolated from the pathogenetic lily leaves was marked as CX1. CX1 was identified as Botrytis cinerea Pers. according to morphological characteristics study and molecular biological analysis. For the mycelial growth and sporulation of CX1,the optimum medium was PSA,the optimum temperature was 20 ℃ and the optimum light condition was 12 h∶12 h alternation of light and darkness. The optimal pH value for mycelial growth and sporulation were5-6 and 6,respectively. Soluble starch was the best carbon source and beef extract was the best nitrogen source for mycelial growth. The optimum carbon source and nitrogen source for sporulation were sucrose and(NH4)2 SO4,respectively.【Conclusion】The pathogen of lily gray mold occurs in Changping,Beijing was B. cinerea Pers. Based on the studies about biological characteristics of B. cinerea Pers.,apart from chemical and biological controls,growth of B. cinerea Pers. and sporulation can be inhibited by changing cultivation environment,reasonable fertilization,irrigation,planting density and rotation,so as to reduce the disease damage.
引文
白滨,杨花莲,何苏琴,于安芬.2013.兰州百合叶枯病病原菌形态特征及生物学特性研究[J].中国蔬菜,(16):78-84.[Bai B,Yang H L,He S Q,Yu A F.2013.Studies on morphology and biological characteristics of Botrytis elliptica(Berk.)Cooke causing foliage blight of Lilium davidii Duch.var.unicolor(Hoog)Cotton[J].China Vegetables,(16):78-84.]
包满珠.2011.花卉学[M].第3版.北京:中国农业出版社:253-268.[Bao M Z.2011.Floriculture[M].The 3rdEdition.Beijing:China Agriculture Press:253-268.]
陈长卿,张博,杨丽娜,高洁.2012.越橘灰霉病病原菌鉴定及其生物学特性研究[J].吉林农业大学学报,34(5):511-516.[Chen C Q,Zhang B,Yang L N,Gao J.2012.Identification and biological characteristics of pathogenic bacteria of blueberry gray mold[J].Journal of Jilin Agricultural University,34(5):511-516.]
杜运鹏.2014.我国百合属植物资源评价及抗病基因同源序列(RGA)的研究[D].北京:北京林业大学.[Du Y P.2014.Collection and evaluation of Lilium spp.in China(Liliaceae)and the research on resistance gene analog(RGA)[D].Beijing:Beijing Forestry University.]
方中达.2007.植病研究方法[M].第3版.北京:中国农业出版社.[Fang Z D.2007.Plant Pathology Research Methods[M].The 3rdEdition.Beijing:China Agriculture Press.]
高智谋,李艳梅,李喜玲,章战华,营金凤.2009.源自不同寄主的灰葡萄孢生物学特性的比较研究[J].菌物学报,28(3):370-377.[Gao Z M,Li Y M,Li X L,Zhang Z H,Ying J F.2009.Comparison of the biological characteristics of Botrytis cinerea isolates from different hosts[J].Mycosystema,28(3):370-377.]
巩慧玲,孙爱洁,李茜,汤国纲,袁慧君,冯再平,李善家.2015.兰州百合鳞茎贮藏腐烂病原菌的鉴定及生物学特性[J].食品工业科技,36(20):97-101.[Gong H L,Sun A J,Li X,Tang G G,Yuan H J,Feng Z P,Li S J.2015.Identification of the pathogen from stored Lanzhou lily bulb rot disease and biological characteristics[J].Science and Technology of Food Industry,36(20):97-101.]
韩青,杨野,陈瑞,葛峰,陈朝银,刘迪秋.2015.超表达岷江百合类萌发素蛋白基因LrGLP2增强烟草对几种病原真菌的抗性[J].植物生理学报,51(12):2223-2230.[Han Q,Yang Y,Chen R,Ge F,Chen C Y,Liu D Q.2015.Overexpression of germin-like protein gene from Lilium regale enhance resistance to several pathogenic fungi in transgenic tobacco plants[J].Plant Physiology Journal,51(12):2223-2230.]
李聪丽,阳亚婕,傅本重,张志林,付应林,李国元.2017.香葱灰霉病病原鉴定及其生物学特性研究[J].河南农业科学,46(9):73-78.[Li C L,Yang Y J,Fu B Z,Zhang ZL,Fu Y L,Li G Y.2017.Identification and biological characterization of the pathogen of Allium ascalonicum Gray Mould[J].Journal of Henan Agricultural Sciences,46(9):73-78.]
梁巧兰,张娜,魏列新,徐秉良.2017.深绿木霉蛋白质TraT2A诱导兰州百合抗灰霉病的作用[J].中国生物防治学报,33(4):545-551.[Liang Q L,Zhang N,Wei LX,Xu B L.2017.Effect of Trichoderma atroviride proteinaceous TraT2A induced Lanzhou lily resistant to gray mold caused by Botrytis cinerea[J].Chinese Journal of Biological Control,33(4):545-551.]
刘春元,刘建华,刘文原.2003.海棠灰霉病菌生物学特性的研究[J].河南农业大学学报,37(1):61-64.[Liu C Y,Liu J H,Liu W Y.2003.A study on the biological characteristics of Botrytis cinerea in Begonia semperflorens[J].Journal of Henan Agricultural University,37(1):61-64.]
刘树芳,李艳琼,曾莉,李铭刚,杨佩文,李一青,尚慧,崔晓龙,李家瑞,文孟良.2006.放线菌次生代谢产物对百合灰霉病菌的抑菌活性筛选[J].西南农业学报,19(4):627-630.[Liu S F,Li Y Q,Zeng L,Li M G,Yang P W,Li Y Q,Shang H,Cui X L,Li J R,Wen M L.2006.Primary study on inhibition activity of actinomycetes seconday metabolites to Botrytis elliptica(Berk.)Cooke.[J].Southwest China Journal of Agricultural Sciences,19(4):627-630.]
罗长维,陈友.2018.凤丹灰霉病病原鉴定与发病条件研究[J].河南农业科学,47(8):78-82.[Luo C W,Chen Y.2018.Pathogen identification and incidence conditions of gray mould on Paeonia ostii[J].Journal of Henan Agricultural Sciences,47(8):78-82.]
仇硕,夏科,李秀娟,周浩,唐凤鸾,赵志国,赵健.2018.龙牙百合两种病害致病菌的分离、鉴定及抑制效应研究[J].广西植物,38(1):109-118.[Qiu S,Xia K,Li X J,Zhou H,Tang F L,Zhao Z G,Zhao J.2018.Isolation,identification and inhibiting effects of pathogens that caused two kinds of diseases in Lilium brownii var.viridulum[J].Guihaia,38(1):109-118.]
瞿友均,徐源辉.2010.百合灰霉病的发生特点和综合防治方法[J].中国植保导刊,30(4):26-27.[Qu Y J,Xu Y H.2010.Lily gray mould occurrence and control[J].China Plant Protection,30(4):26-27.]
宋翔宇.2016.椭圆灰葡萄孢(Botrytis cinerea)胁迫下亚洲百合的生理响应及调节基因MAPK的克隆[D].沈阳:沈阳农业大学.[Song X Y.2016.Physiological responses of Lilium asiatic hybrid to Botrytis cinerea stress and cloning of regulatory gene-MAPK[D].Shenyang:Shenyang Agriculture Uuniversity.]
唐祥宁,肖爱萍,游春平,曾宜杰,彭桂云,刘文越,张德贵.1998.百合灰霉病病菌生物学特性研究[J].江西农业大学学报,20(4):485-489.[Tang X N,Xiao A P,You C P,Zeng Y J,Peng G Y,Liu W Y,Zhang D G.1998.Astudy on the biological properties of Botrytis elliptica causing lily gray mould[J].Acta Agriculturae Universitatis Jiangxiensis,20(4):485-489.]
王丹.2008.不同作物灰霉病菌生物学特性的比较研究和番茄灰霉病的生防研究[D].雅安:四川农业大学.[Wang D.2008.Study on biological characteristics of different strains of Botrytis cinerea and study on biocontrol to tomato Botrytis cinera[D].Ya’an:Sichuan Agricultural University.]
袁月,周如军,傅俊范,卢占慧,史晓琪,李自博.2016.不同人参灰霉病的灰葡萄孢菌生物学特性及致病性比较[J].沈阳农业大学学报,47(3):271-277.[Yuan Y,Zhou RJ,Fu J F,Lu Z H,Shi X Q,Li Z B.2016.Comparison of the biological characteristics and pathogenicity of Botrytis cinerea isolates[J].Journal of Shenyang Agricultural University,47(3):271-277.]
张悦,李丹,陈子牛,黄晓雯.2015.不同环境因素对昆明本地灰葡萄孢、黄曲霉菌株生长的影响[J].江苏农业科学,43(7):118-120.[Zhang Y,Li D,Chen Z N,Huang X W.2015.Effects of different environmental factors on the growth of Botrytis cinerea and Aspergillus flavus in Kunming[J].Journal of Jiangsu Agricultural Sciences,43(7):118-120.]
朱丽梅,胡凤荣,罗凤霞.2010.不同百合品种对百合灰霉病的抗病性鉴定[J].植物保护,36(3):148-151.[Zhu LM,Hu F R,Luo F X.2010.Identification of disease resistance of Lilium species to lily gray mould[J].Plant Protection,36(3):148-151.]
朱丽梅,张长青,陈浩,陆慧.2011.百合灰霉病高效杀菌剂的筛选和联合作用研究[J].金陵科技学院学报,27(1):51-54.[Zhu L M,Zhang C Q,Chen H,Lu H.2011.Screening and combined action of efficient fungicides for controlling lily Botrytis cinerea[J].Journal of Jinling Institute of Technology,27(1):51-54.]
Cao X,Shi S,Zhang Z.2018.First report of Botrytis leaf blight on lily(Lilium longiflorum)caused by Botrytis cinerea in Beijing,China[J].Plant Disease,102(5):1033-1034.
Chiou A L,Wu W S.2003.Formulation of Bacillus amyloliquefaciens B190 for control of lily grey mould(Botrytis elliptica)[J].Journal of Phytopathology,151(1):13-18.
包满珠.2011.花卉学[M].第3版.北京:中国农业出版社:253-268.[Bao M Z.2011.Floriculture[M].The 3rdEdition.Beijing:China Agriculture Press:253-268.]
陈长卿,张博,杨丽娜,高洁.2012.越橘灰霉病病原菌鉴定及其生物学特性研究[J].吉林农业大学学报,34(5):511-516.[Chen C Q,Zhang B,Yang L N,Gao J.2012.Identification and biological characteristics of pathogenic bacteria of blueberry gray mold[J].Journal of Jilin Agricultural University,34(5):511-516.]
杜运鹏.2014.我国百合属植物资源评价及抗病基因同源序列(RGA)的研究[D].北京:北京林业大学.[Du Y P.2014.Collection and evaluation of Lilium spp.in China(Liliaceae)and the research on resistance gene analog(RGA)[D].Beijing:Beijing Forestry University.]
方中达.2007.植病研究方法[M].第3版.北京:中国农业出版社.[Fang Z D.2007.Plant Pathology Research Methods[M].The 3rdEdition.Beijing:China Agriculture Press.]
高智谋,李艳梅,李喜玲,章战华,营金凤.2009.源自不同寄主的灰葡萄孢生物学特性的比较研究[J].菌物学报,28(3):370-377.[Gao Z M,Li Y M,Li X L,Zhang Z H,Ying J F.2009.Comparison of the biological characteristics of Botrytis cinerea isolates from different hosts[J].Mycosystema,28(3):370-377.]
巩慧玲,孙爱洁,李茜,汤国纲,袁慧君,冯再平,李善家.2015.兰州百合鳞茎贮藏腐烂病原菌的鉴定及生物学特性[J].食品工业科技,36(20):97-101.[Gong H L,Sun A J,Li X,Tang G G,Yuan H J,Feng Z P,Li S J.2015.Identification of the pathogen from stored Lanzhou lily bulb rot disease and biological characteristics[J].Science and Technology of Food Industry,36(20):97-101.]
韩青,杨野,陈瑞,葛峰,陈朝银,刘迪秋.2015.超表达岷江百合类萌发素蛋白基因LrGLP2增强烟草对几种病原真菌的抗性[J].植物生理学报,51(12):2223-2230.[Han Q,Yang Y,Chen R,Ge F,Chen C Y,Liu D Q.2015.Overexpression of germin-like protein gene from Lilium regale enhance resistance to several pathogenic fungi in transgenic tobacco plants[J].Plant Physiology Journal,51(12):2223-2230.]
李聪丽,阳亚婕,傅本重,张志林,付应林,李国元.2017.香葱灰霉病病原鉴定及其生物学特性研究[J].河南农业科学,46(9):73-78.[Li C L,Yang Y J,Fu B Z,Zhang ZL,Fu Y L,Li G Y.2017.Identification and biological characterization of the pathogen of Allium ascalonicum Gray Mould[J].Journal of Henan Agricultural Sciences,46(9):73-78.]
梁巧兰,张娜,魏列新,徐秉良.2017.深绿木霉蛋白质TraT2A诱导兰州百合抗灰霉病的作用[J].中国生物防治学报,33(4):545-551.[Liang Q L,Zhang N,Wei LX,Xu B L.2017.Effect of Trichoderma atroviride proteinaceous TraT2A induced Lanzhou lily resistant to gray mold caused by Botrytis cinerea[J].Chinese Journal of Biological Control,33(4):545-551.]
刘春元,刘建华,刘文原.2003.海棠灰霉病菌生物学特性的研究[J].河南农业大学学报,37(1):61-64.[Liu C Y,Liu J H,Liu W Y.2003.A study on the biological characteristics of Botrytis cinerea in Begonia semperflorens[J].Journal of Henan Agricultural University,37(1):61-64.]
刘树芳,李艳琼,曾莉,李铭刚,杨佩文,李一青,尚慧,崔晓龙,李家瑞,文孟良.2006.放线菌次生代谢产物对百合灰霉病菌的抑菌活性筛选[J].西南农业学报,19(4):627-630.[Liu S F,Li Y Q,Zeng L,Li M G,Yang P W,Li Y Q,Shang H,Cui X L,Li J R,Wen M L.2006.Primary study on inhibition activity of actinomycetes seconday metabolites to Botrytis elliptica(Berk.)Cooke.[J].Southwest China Journal of Agricultural Sciences,19(4):627-630.]
罗长维,陈友.2018.凤丹灰霉病病原鉴定与发病条件研究[J].河南农业科学,47(8):78-82.[Luo C W,Chen Y.2018.Pathogen identification and incidence conditions of gray mould on Paeonia ostii[J].Journal of Henan Agricultural Sciences,47(8):78-82.]
仇硕,夏科,李秀娟,周浩,唐凤鸾,赵志国,赵健.2018.龙牙百合两种病害致病菌的分离、鉴定及抑制效应研究[J].广西植物,38(1):109-118.[Qiu S,Xia K,Li X J,Zhou H,Tang F L,Zhao Z G,Zhao J.2018.Isolation,identification and inhibiting effects of pathogens that caused two kinds of diseases in Lilium brownii var.viridulum[J].Guihaia,38(1):109-118.]
瞿友均,徐源辉.2010.百合灰霉病的发生特点和综合防治方法[J].中国植保导刊,30(4):26-27.[Qu Y J,Xu Y H.2010.Lily gray mould occurrence and control[J].China Plant Protection,30(4):26-27.]
宋翔宇.2016.椭圆灰葡萄孢(Botrytis cinerea)胁迫下亚洲百合的生理响应及调节基因MAPK的克隆[D].沈阳:沈阳农业大学.[Song X Y.2016.Physiological responses of Lilium asiatic hybrid to Botrytis cinerea stress and cloning of regulatory gene-MAPK[D].Shenyang:Shenyang Agriculture Uuniversity.]
唐祥宁,肖爱萍,游春平,曾宜杰,彭桂云,刘文越,张德贵.1998.百合灰霉病病菌生物学特性研究[J].江西农业大学学报,20(4):485-489.[Tang X N,Xiao A P,You C P,Zeng Y J,Peng G Y,Liu W Y,Zhang D G.1998.Astudy on the biological properties of Botrytis elliptica causing lily gray mould[J].Acta Agriculturae Universitatis Jiangxiensis,20(4):485-489.]
王丹.2008.不同作物灰霉病菌生物学特性的比较研究和番茄灰霉病的生防研究[D].雅安:四川农业大学.[Wang D.2008.Study on biological characteristics of different strains of Botrytis cinerea and study on biocontrol to tomato Botrytis cinera[D].Ya’an:Sichuan Agricultural University.]
袁月,周如军,傅俊范,卢占慧,史晓琪,李自博.2016.不同人参灰霉病的灰葡萄孢菌生物学特性及致病性比较[J].沈阳农业大学学报,47(3):271-277.[Yuan Y,Zhou RJ,Fu J F,Lu Z H,Shi X Q,Li Z B.2016.Comparison of the biological characteristics and pathogenicity of Botrytis cinerea isolates[J].Journal of Shenyang Agricultural University,47(3):271-277.]
张悦,李丹,陈子牛,黄晓雯.2015.不同环境因素对昆明本地灰葡萄孢、黄曲霉菌株生长的影响[J].江苏农业科学,43(7):118-120.[Zhang Y,Li D,Chen Z N,Huang X W.2015.Effects of different environmental factors on the growth of Botrytis cinerea and Aspergillus flavus in Kunming[J].Journal of Jiangsu Agricultural Sciences,43(7):118-120.]
朱丽梅,胡凤荣,罗凤霞.2010.不同百合品种对百合灰霉病的抗病性鉴定[J].植物保护,36(3):148-151.[Zhu LM,Hu F R,Luo F X.2010.Identification of disease resistance of Lilium species to lily gray mould[J].Plant Protection,36(3):148-151.]
朱丽梅,张长青,陈浩,陆慧.2011.百合灰霉病高效杀菌剂的筛选和联合作用研究[J].金陵科技学院学报,27(1):51-54.[Zhu L M,Zhang C Q,Chen H,Lu H.2011.Screening and combined action of efficient fungicides for controlling lily Botrytis cinerea[J].Journal of Jinling Institute of Technology,27(1):51-54.]
Cao X,Shi S,Zhang Z.2018.First report of Botrytis leaf blight on lily(Lilium longiflorum)caused by Botrytis cinerea in Beijing,China[J].Plant Disease,102(5):1033-1034.
Chiou A L,Wu W S.2003.Formulation of Bacillus amyloliquefaciens B190 for control of lily grey mould(Botrytis elliptica)[J].Journal of Phytopathology,151(1):13-18.