引起葡萄叶斑病的链格孢种类的初步鉴定
|
摘要
为明确北京市大兴区、顺义区和山西省襄汾县葡萄树上叶斑病的病原菌,利用组织分离法对病样进行分离纯化,采用多基因系统发育学分析(ITS,Alt a 1和gpd)和形态学对纯化后菌株进行鉴定,并结合柯赫氏法则对其进行验证。结果表明,共获得37株链格孢属真菌菌株,分属链格孢Alternaria alternata、葡萄链格孢A. viniferae和乔木链格孢A. arborescens,柯赫氏法则检测结果显示接种菌株对所接叶片与果穗均具致病性,说明这3种链格孢种均为葡萄叶斑病的病原菌;葡萄链格孢为主要的病原菌,有28株,故将该类病害命名为葡萄链格孢叶斑病;此外,该类菌株在2～40℃之间均能生长,25℃为最适生长温度。
To determine the causes of the leaf spot symptoms observed in grape vineyards in Daxing and Shunyi Districts of Beijing City and Xiangfen County of Shanxi Province, tissue isolation method,multi-gene phylogenetic analysis(ITS, Alt a 1 and gpd) and morphological study were used to identify the species of pathogens. Koch's postulates were used to verify the results. In total, 37 isolates of Alternaria species, A. altanata, A. viniferae and A. arborescens were isolated. All three Alternaria species were pathogens of grape leaf spot disease. Alternaria viniferae was the dominant species with 28 isolates. Therefore, the disease was named as grape Alternaria leaf spot disease. In addition, the current study showed that the pathogen could grow under a wide range of temperature from 2 to 40℃, and the optimum temperature for the three species was 25℃.
To determine the causes of the leaf spot symptoms observed in grape vineyards in Daxing and Shunyi Districts of Beijing City and Xiangfen County of Shanxi Province, tissue isolation method,multi-gene phylogenetic analysis(ITS, Alt a 1 and gpd) and morphological study were used to identify the species of pathogens. Koch's postulates were used to verify the results. In total, 37 isolates of Alternaria species, A. altanata, A. viniferae and A. arborescens were isolated. All three Alternaria species were pathogens of grape leaf spot disease. Alternaria viniferae was the dominant species with 28 isolates. Therefore, the disease was named as grape Alternaria leaf spot disease. In addition, the current study showed that the pathogen could grow under a wide range of temperature from 2 to 40℃, and the optimum temperature for the three species was 25℃.
引文
Berbee ML,Pirseyedi M,Hubbard S.1999.Cochliobolus phylogenetics and the origin of known,highly virulent pathogens,inferred from ITS and glyceraldehyde-3-phosphate dehydrogenase gene sequences.Mycologia,91(6):964-977
Casieri L,Hofstetter V,Viret O,Gindro K.2009.Fungal communities living in the wood of different cultivars of young Vitis vinifera plants.Phytopathologia Mediterranea,48(1):73-83
Chai ZX.2001.Identification of grape brown spot in Lanzhou Area.Journal of Gansu Agricultural University,36(1):61-64,99(in Chinese)[柴兆祥.2001.兰州地区葡萄褐斑病发生危害及菌种鉴定.甘肃农业大学学报,36(1):61-64,99]
Chen FM,Liu Q,Jiang JH,Huang XH,He DN.2005.Biological properties of endophytic fungus Alternaria alternata var GI009 from Ginkgo biloba.Journal of Northwest Forestry University,20(4):112-114(in Chinese)[陈凤美,刘群,蒋继宏,黄小花,何冬宁.2005.银杏内生链格孢菌GI009生物学特性研究.西北林学院学报,20(4):112-114]
Deighton FC.1976.Three fungi on leaves of Vitis.Transactions of the British Mycological Society,67(2):223-232
Fan YM,Hao JZ,Zhang MZ,Zhao ZY,Wang HQ,Wu YH,Zhang J.2007.Grape Scolecotrichum leaf spot disease pathogen identification and symptoms recognition in Xinjiang.Plant Protection,33(3):140-142(in Chinese)[范咏梅,郝敬哲,张明智,赵震宇,王惠卿,吴玉华,张娟.2007.新疆葡萄褐纹病病原鉴定及症状识别.植物保护,33(3):140-142]
Fang ZD.1998.Research methods of plant pathology(3rd edition).Beijing:China Agriculture Press(in Chinese)[方中达.1998.植病研究法(第3版).北京:中国农业出版社]
Farr DF,Rossman AY.2015.Fungal databases.Retrieved From https://nt.ars-grin.gov/fungaldatabases/.2017-12-10
French AM.1989.California plant disease host index.California:California Department of Food and Agriculture Sacramento
González V,Tello ML.2011.The endophytic mycota associated with Vitis vinifera in central Spain.Fungal Diversity,47(1):29-42
Guo QQ,Yang ZH,Cui YJ,Xu J,Zhu JH.2013.A method for molecular detection of Alternaria solani and its applicability to early diagnosis of potato early blight in the field.Journal of Plant Protection,40(4):381-382(in Chinese)[郭倩倩,杨志辉,崔亚婧,徐进,朱杰华.2013.马铃薯早疫病菌的分子鉴定与田间早期诊断.植物保护学报,40(4):381-382]
Guo YL,Liu XJ.2003.Flora fungorum sinicorum.Vol.20,Mycovellosiella,Passalora,Phaeoramularia.Beijing:Science Press(in Chinese)[郭英兰,刘锡琎.2003.中国真菌志.第二十卷,菌绒孢属钉孢属色链隔孢属.北京:科学出版社]
Hall TA.1999.BioEdit:a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT.Nucleic Acids Symposium Series,41:95-98
He PC.2005.Grape science.Beijing:China Agriculture Press(in Chinese)[贺普超.2005.葡萄学.北京:中国农业出版社]
Hong GS,Cramer RA,Lawrence CB,Lawrence CB,Pryor BM.2005.Alt a 1 allergen homologs from Alternaria and related taxa:analysis of phylogenetic content and secondary structure.Fungal Genetics and Biology,42(2):119-129
Hong SK,Wan GK,Yun HK,Choi KJ.2008.Morphological variations,genetic diversity and pathogenicity of Colletotrichum species causing grape ripe rot in Korea.Plant Pathology Journal,24(3):269-278
Jayawardena RS,Liu M,Maharachchikumbura SSN,Zhang W,Xing QK,Hyde KD,Nilthong S,Li XH,Yan JY.2016.Neopestalotiopsis vitis sp.nov.causing grapevine leaf spot in China.Phytotaxa,258(1):63-74
KakalíkováL,Jankura E,?robárováA.2009.First report of Alternaria bunch rot of grapevines in Slovakia.Australasian Plant Disease Notes,4(1):68-69
Katoh K,Toh H.2008.Recent developments in the MAFFT multiple sequence alignment program.Briefings in Bioinformatics,9(4):286-298
Liang CH,Jayawardena RS,Zhang W,Wang XG,Liu M,Liu L,Zang CQ,Xu XR,Hyde KD,Yan JY,et al.2016.Identification and characterization of Pseudocercospora species causing grapevine leaf spot in China.Journal of Phytopathology,164(2):75-85
Liang CH,Zhao KH,Liu CY,Wang L.2010.Advances in study on grape leaf rot.Liaoning Agricultural Sciences,2010(S):117-119(in Chinese)[梁春浩,赵奎华,刘长远,王璐.2010.葡萄褐斑病研究进展.辽宁农业科学,2010(S):117-119]
Liu T,Ma L,Du NS.2002.The survey of the grape’s biological action.Chinese Journal of Nature,24(2):81-87(in Chinese)[刘涛,马龙,堵年生.2002.葡萄的生物学作用研究进展.自然杂志,24(2):81-87]
Peever TL,Su G,Carpenter-Boggs L,Timmer LW.2004.Molecular systematics of citrus-associated Alternaria species.Mycologia,96(1):119-134
Peregrine WTH,Ahmad KB.1982.Brunei:a first annotated list of plant diseases and associated organisms.Phytopathological Papers,27:1-87
Qu WW,Liu X,Yang KQ,Liu HX,Hou LQ.2012.Identification and phylogenetic analysis of walnut-associated Alternaria spp.in Shandong Province,China.Journal of Plant Protection,39(2):121-128(in Chinese)[曲文文,刘霞,杨克强,刘会香,侯立群.2012.山东省危害核桃的链格孢属真菌鉴定及其系统发育.植物保护学报,39(2):121-128]
Simmons EG,Roberts RG.1993.Alternaria themes and variations(73).Mycotaxon,46(1):109-140
Song WC,Guo QY,Wang HK,Wang DY,Lei D,Liu Y,Alaiyi.2014.Grape common diseases and their occurrence status quo in Turpan and some other areas in northern Xinjiang.Xinjiang Agricultural Sciences,51(5):893-901(in Chinese)[宋卫翠,郭庆元,王洪凯,王登元,雷丹,刘瑶,阿来依.2014.吐鲁番地区及北疆部分地区葡萄常见病害种类及发生现状.新疆农业科学,51(5):893-901]
Swofford DL.2002.PAUP*4.0:phylogenetic analysis using parsimony.Sunderland:Sinauer Associates
Tao WC,Zhang W,Yan JY,Hyde KD,McKenzie EHC,Li XH,Wang Y.2014.A new Alternaria species from grapevine in China.Mycological Progress,13:999
Wang B.2015.Influence of temperature and the p H on the colony growth of Alternaria alternata(Fr.)Keissler of Trifolium repens Linn.Gansu Agriculture Science and Technology,(8):59-61(in Chinese)[王斌.2015.温度和pH对白三叶草链格孢菌菌落生长的影响.甘肃农业科技,(8):59-61]
Wang GZ,Fan ZQ,Ru QH,An HW.2002.Survey of grapevine diseases in the eastern foot of Helan Mountain,Ningxia.Northern Horticulture,(6):64-65(in Chinese)[王国珍,樊仲庆,茹庆华,安宏伟.2002.宁夏贺兰山东麓葡萄病害调查.北方园艺,(6):64-65]
Wang YY,Liu B,Zhou YY,Zhu XF,Gao ZG,Chen LJ.2015.Biological characteristics and screening of fungicides of maize Alternaria tenuis Nees.Journal of Shenyang Agricultural University,46(5):538-542(in Chinese)[王媛媛,刘彬,周园园,朱晓峰,高增贵,陈立杰.2015.玉米链格孢病菌生物学特性及防治药剂离体活性筛选.沈阳农业大学学报,46(5):538-542]
Wang ZY.2009.Chinese grape diseases and insect pests and integrated control techniques.Beijing:China Agriculture Press(in Chinese)[王忠跃.2009.中国葡萄病虫害与综合防控技术.北京:中国农业出版社]
Weir TL,Huff DR,Christ BJ,Romaine CP.1998.RAPD-PCR analysis of genetic variation among isolates of Alternaria solani and Alternaria alternata from potato and tomato.Mycologia,90(5):813-821
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
Wilcox WF,Gubler WD,Uyemoto JK.2015.Compendium of grape diseases,disorders,and pests.Saint Paul:The American Phytopathological Society Press,pp.1-220
Woudenberg JHC.2015.Restyling Alternaria.Ph.D Thesis.Wageningen:Wageningen University
Yang C,Zhang GL,Ren YZ,Zhong YQ,Gao XL,Li Z,Zhang YP,Li GY.2017.Identification of the pathogens of grape spike-stalk brown along the Tianshan Mountains.Plant Protection,43(3):129-135(in Chinese)[杨超,张国丽,任毓忠,钟杨琼,高祥雷,李卓,张亚平,李国英.2017.北疆沿天山北坡一带葡萄穗轴褐枯病病原菌的鉴定.植物保护,43(3):129-135]
Yao M.2015.Identification and control of two leaf diseases in grape.Journal of Agricultural Catastrophology,5(9):5-6,10(in Chinese)[姚敏.2015.葡萄2种常见叶部病害的识别和防治.农业灾害研究,5(9):5-6,10]
Zhang R,Yue HM,Sun GY.2009.Sequence and phylogenetic analysis of OPA2-1 region from Alternaria small-spored species.Mycosystema,28(4):553-557(in Chinese)[张荣,岳海梅,孙广宇.2009.小孢子链格孢OPA2-1核苷酸序列分析及系统发育研究.菌物学报,28(4):553-557]
Zhang TY.2003.Flora fungorum sinicorum.Vol.16,Alternaria.Beijing:Science Press,pp.19-20(in Chinese)[张天宇.2003.中国真菌志.第十六卷,链格孢属.北京:科学出版社,pp.19-20]
Zhao YQ,Yu HR,Shi K,Zhang LJ,Zhang DM,Zhu PT,Chai C,Jia AM.2017.Identification of the pathogen causing leaf spot on sorghum.Acta Phytopathologica Sinica,47(2):282-285(in Chinese)[赵艳琴,于华荣,石凯,张丽娟,张冬梅,朱鹏涛,柴畅,贾爱民.2017.高粱链格孢叶斑病病原鉴定.植物病理学报,47(2):282-285]
Casieri L,Hofstetter V,Viret O,Gindro K.2009.Fungal communities living in the wood of different cultivars of young Vitis vinifera plants.Phytopathologia Mediterranea,48(1):73-83
Chai ZX.2001.Identification of grape brown spot in Lanzhou Area.Journal of Gansu Agricultural University,36(1):61-64,99(in Chinese)[柴兆祥.2001.兰州地区葡萄褐斑病发生危害及菌种鉴定.甘肃农业大学学报,36(1):61-64,99]
Chen FM,Liu Q,Jiang JH,Huang XH,He DN.2005.Biological properties of endophytic fungus Alternaria alternata var GI009 from Ginkgo biloba.Journal of Northwest Forestry University,20(4):112-114(in Chinese)[陈凤美,刘群,蒋继宏,黄小花,何冬宁.2005.银杏内生链格孢菌GI009生物学特性研究.西北林学院学报,20(4):112-114]
Deighton FC.1976.Three fungi on leaves of Vitis.Transactions of the British Mycological Society,67(2):223-232
Fan YM,Hao JZ,Zhang MZ,Zhao ZY,Wang HQ,Wu YH,Zhang J.2007.Grape Scolecotrichum leaf spot disease pathogen identification and symptoms recognition in Xinjiang.Plant Protection,33(3):140-142(in Chinese)[范咏梅,郝敬哲,张明智,赵震宇,王惠卿,吴玉华,张娟.2007.新疆葡萄褐纹病病原鉴定及症状识别.植物保护,33(3):140-142]
Fang ZD.1998.Research methods of plant pathology(3rd edition).Beijing:China Agriculture Press(in Chinese)[方中达.1998.植病研究法(第3版).北京:中国农业出版社]
Farr DF,Rossman AY.2015.Fungal databases.Retrieved From https://nt.ars-grin.gov/fungaldatabases/.2017-12-10
French AM.1989.California plant disease host index.California:California Department of Food and Agriculture Sacramento
González V,Tello ML.2011.The endophytic mycota associated with Vitis vinifera in central Spain.Fungal Diversity,47(1):29-42
Guo QQ,Yang ZH,Cui YJ,Xu J,Zhu JH.2013.A method for molecular detection of Alternaria solani and its applicability to early diagnosis of potato early blight in the field.Journal of Plant Protection,40(4):381-382(in Chinese)[郭倩倩,杨志辉,崔亚婧,徐进,朱杰华.2013.马铃薯早疫病菌的分子鉴定与田间早期诊断.植物保护学报,40(4):381-382]
Guo YL,Liu XJ.2003.Flora fungorum sinicorum.Vol.20,Mycovellosiella,Passalora,Phaeoramularia.Beijing:Science Press(in Chinese)[郭英兰,刘锡琎.2003.中国真菌志.第二十卷,菌绒孢属钉孢属色链隔孢属.北京:科学出版社]
Hall TA.1999.BioEdit:a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT.Nucleic Acids Symposium Series,41:95-98
He PC.2005.Grape science.Beijing:China Agriculture Press(in Chinese)[贺普超.2005.葡萄学.北京:中国农业出版社]
Hong GS,Cramer RA,Lawrence CB,Lawrence CB,Pryor BM.2005.Alt a 1 allergen homologs from Alternaria and related taxa:analysis of phylogenetic content and secondary structure.Fungal Genetics and Biology,42(2):119-129
Hong SK,Wan GK,Yun HK,Choi KJ.2008.Morphological variations,genetic diversity and pathogenicity of Colletotrichum species causing grape ripe rot in Korea.Plant Pathology Journal,24(3):269-278
Jayawardena RS,Liu M,Maharachchikumbura SSN,Zhang W,Xing QK,Hyde KD,Nilthong S,Li XH,Yan JY.2016.Neopestalotiopsis vitis sp.nov.causing grapevine leaf spot in China.Phytotaxa,258(1):63-74
KakalíkováL,Jankura E,?robárováA.2009.First report of Alternaria bunch rot of grapevines in Slovakia.Australasian Plant Disease Notes,4(1):68-69
Katoh K,Toh H.2008.Recent developments in the MAFFT multiple sequence alignment program.Briefings in Bioinformatics,9(4):286-298
Liang CH,Jayawardena RS,Zhang W,Wang XG,Liu M,Liu L,Zang CQ,Xu XR,Hyde KD,Yan JY,et al.2016.Identification and characterization of Pseudocercospora species causing grapevine leaf spot in China.Journal of Phytopathology,164(2):75-85
Liang CH,Zhao KH,Liu CY,Wang L.2010.Advances in study on grape leaf rot.Liaoning Agricultural Sciences,2010(S):117-119(in Chinese)[梁春浩,赵奎华,刘长远,王璐.2010.葡萄褐斑病研究进展.辽宁农业科学,2010(S):117-119]
Liu T,Ma L,Du NS.2002.The survey of the grape’s biological action.Chinese Journal of Nature,24(2):81-87(in Chinese)[刘涛,马龙,堵年生.2002.葡萄的生物学作用研究进展.自然杂志,24(2):81-87]
Peever TL,Su G,Carpenter-Boggs L,Timmer LW.2004.Molecular systematics of citrus-associated Alternaria species.Mycologia,96(1):119-134
Peregrine WTH,Ahmad KB.1982.Brunei:a first annotated list of plant diseases and associated organisms.Phytopathological Papers,27:1-87
Qu WW,Liu X,Yang KQ,Liu HX,Hou LQ.2012.Identification and phylogenetic analysis of walnut-associated Alternaria spp.in Shandong Province,China.Journal of Plant Protection,39(2):121-128(in Chinese)[曲文文,刘霞,杨克强,刘会香,侯立群.2012.山东省危害核桃的链格孢属真菌鉴定及其系统发育.植物保护学报,39(2):121-128]
Simmons EG,Roberts RG.1993.Alternaria themes and variations(73).Mycotaxon,46(1):109-140
Song WC,Guo QY,Wang HK,Wang DY,Lei D,Liu Y,Alaiyi.2014.Grape common diseases and their occurrence status quo in Turpan and some other areas in northern Xinjiang.Xinjiang Agricultural Sciences,51(5):893-901(in Chinese)[宋卫翠,郭庆元,王洪凯,王登元,雷丹,刘瑶,阿来依.2014.吐鲁番地区及北疆部分地区葡萄常见病害种类及发生现状.新疆农业科学,51(5):893-901]
Swofford DL.2002.PAUP*4.0:phylogenetic analysis using parsimony.Sunderland:Sinauer Associates
Tao WC,Zhang W,Yan JY,Hyde KD,McKenzie EHC,Li XH,Wang Y.2014.A new Alternaria species from grapevine in China.Mycological Progress,13:999
Wang B.2015.Influence of temperature and the p H on the colony growth of Alternaria alternata(Fr.)Keissler of Trifolium repens Linn.Gansu Agriculture Science and Technology,(8):59-61(in Chinese)[王斌.2015.温度和pH对白三叶草链格孢菌菌落生长的影响.甘肃农业科技,(8):59-61]
Wang GZ,Fan ZQ,Ru QH,An HW.2002.Survey of grapevine diseases in the eastern foot of Helan Mountain,Ningxia.Northern Horticulture,(6):64-65(in Chinese)[王国珍,樊仲庆,茹庆华,安宏伟.2002.宁夏贺兰山东麓葡萄病害调查.北方园艺,(6):64-65]
Wang YY,Liu B,Zhou YY,Zhu XF,Gao ZG,Chen LJ.2015.Biological characteristics and screening of fungicides of maize Alternaria tenuis Nees.Journal of Shenyang Agricultural University,46(5):538-542(in Chinese)[王媛媛,刘彬,周园园,朱晓峰,高增贵,陈立杰.2015.玉米链格孢病菌生物学特性及防治药剂离体活性筛选.沈阳农业大学学报,46(5):538-542]
Wang ZY.2009.Chinese grape diseases and insect pests and integrated control techniques.Beijing:China Agriculture Press(in Chinese)[王忠跃.2009.中国葡萄病虫害与综合防控技术.北京:中国农业出版社]
Weir TL,Huff DR,Christ BJ,Romaine CP.1998.RAPD-PCR analysis of genetic variation among isolates of Alternaria solani and Alternaria alternata from potato and tomato.Mycologia,90(5):813-821
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
Wilcox WF,Gubler WD,Uyemoto JK.2015.Compendium of grape diseases,disorders,and pests.Saint Paul:The American Phytopathological Society Press,pp.1-220
Woudenberg JHC.2015.Restyling Alternaria.Ph.D Thesis.Wageningen:Wageningen University
Yang C,Zhang GL,Ren YZ,Zhong YQ,Gao XL,Li Z,Zhang YP,Li GY.2017.Identification of the pathogens of grape spike-stalk brown along the Tianshan Mountains.Plant Protection,43(3):129-135(in Chinese)[杨超,张国丽,任毓忠,钟杨琼,高祥雷,李卓,张亚平,李国英.2017.北疆沿天山北坡一带葡萄穗轴褐枯病病原菌的鉴定.植物保护,43(3):129-135]
Yao M.2015.Identification and control of two leaf diseases in grape.Journal of Agricultural Catastrophology,5(9):5-6,10(in Chinese)[姚敏.2015.葡萄2种常见叶部病害的识别和防治.农业灾害研究,5(9):5-6,10]
Zhang R,Yue HM,Sun GY.2009.Sequence and phylogenetic analysis of OPA2-1 region from Alternaria small-spored species.Mycosystema,28(4):553-557(in Chinese)[张荣,岳海梅,孙广宇.2009.小孢子链格孢OPA2-1核苷酸序列分析及系统发育研究.菌物学报,28(4):553-557]
Zhang TY.2003.Flora fungorum sinicorum.Vol.16,Alternaria.Beijing:Science Press,pp.19-20(in Chinese)[张天宇.2003.中国真菌志.第十六卷,链格孢属.北京:科学出版社,pp.19-20]
Zhao YQ,Yu HR,Shi K,Zhang LJ,Zhang DM,Zhu PT,Chai C,Jia AM.2017.Identification of the pathogen causing leaf spot on sorghum.Acta Phytopathologica Sinica,47(2):282-285(in Chinese)[赵艳琴,于华荣,石凯,张丽娟,张冬梅,朱鹏涛,柴畅,贾爱民.2017.高粱链格孢叶斑病病原鉴定.植物病理学报,47(2):282-285]