生防放线菌CY-14的鉴定及其发酵液对香蕉炭疽病的防治效果
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摘要
香蕉炭疽病是由香蕉炭疽菌Colletotrichum musae侵染引起的,是香蕉采后贮藏、运输和销售过程中发生的主要病害之一,严重影响中国香蕉的北运和出口。从土壤中分离到1株对香蕉炭疽菌具有拮抗作用的放线菌CY-14,通过形态特征、培养特性、生理生化特性和16S rDNA序列分析对该菌株进行了初步鉴定,并对其发酵液进行了活性评价。结果表明:经初步鉴定,该放线菌为链霉菌属Streptomyces sp.;其发酵液稀释10倍对香蕉炭疽菌的抑制率达92.5%,同时对其他8种植物病原真菌均具有较强的抑菌活性,抗菌谱较广;其发酵液热稳定性较好,经100℃处理2 h后,体积分数为10%的发酵液对香蕉炭疽菌的抑制率为55.7%,发酵液稀释1 000倍后仍可完全抑制香蕉炭疽菌的孢子萌发;CY-14纯发酵液对香蕉炭疽病具有一定的防效,与对照药剂1 000 mg/L百菌清的防效相当。菌株CY-14作为香蕉炭疽病的生防放线菌,具有发现新型抗生素或者活性化合物的潜能,可对其进行深入的研究。
Banana anthracnose is caused by Colletotrichum musae. In China, it is one of the main diseases that happens during the post harvest storage, including transportation and sale, which seriously affects the north transportation and export of the banana. A actinomycete strain of CY-14, which had antagonistic effect on C. musae, was isolated from the soil. The strain was preliminarily identified by morphology, physiological and biochemical characteristics, and 16 S rDNA sequence analysis. The bioactivity of its fermentation broth was also evaluated. The results showed that strain CY-14 was preliminarily identified as Streptomyces sp. The inhibition rate of 10 times diluted fermentation broth to C. musae was 92.5%. The fermentation broth also had a broad-spectrum antifungal activity to other 8 kinds of plant pathogenic fungi. In addition, the fermentation broth had good thermal stability. After heated up at 100 ℃ for 2 hours, the inhibition rate of 10% fermentation broth was 55.7%. The 1 000 times diluted fermentation broth could completely inhibited the spore germination of C. musae. While the pure fermentation broth had the same control effect as 1 000 mg/L chlorothalonil. Strain CY-14, as a biocontrol actinomycetes of banana anthracnose, has a potential application in the discovery of new antibiotics or active compounds, and is well worthy of further study.
Banana anthracnose is caused by Colletotrichum musae. In China, it is one of the main diseases that happens during the post harvest storage, including transportation and sale, which seriously affects the north transportation and export of the banana. A actinomycete strain of CY-14, which had antagonistic effect on C. musae, was isolated from the soil. The strain was preliminarily identified by morphology, physiological and biochemical characteristics, and 16 S rDNA sequence analysis. The bioactivity of its fermentation broth was also evaluated. The results showed that strain CY-14 was preliminarily identified as Streptomyces sp. The inhibition rate of 10 times diluted fermentation broth to C. musae was 92.5%. The fermentation broth also had a broad-spectrum antifungal activity to other 8 kinds of plant pathogenic fungi. In addition, the fermentation broth had good thermal stability. After heated up at 100 ℃ for 2 hours, the inhibition rate of 10% fermentation broth was 55.7%. The 1 000 times diluted fermentation broth could completely inhibited the spore germination of C. musae. While the pure fermentation broth had the same control effect as 1 000 mg/L chlorothalonil. Strain CY-14, as a biocontrol actinomycetes of banana anthracnose, has a potential application in the discovery of new antibiotics or active compounds, and is well worthy of further study.
引文
[1]雷照鸣,梁艳琼,贺春萍,等.20%咪鲜胺水乳剂防治香蕉炭疽病效果评价[J].安徽农业科学,2013,41(11):4830-4831.LEI Z M,LIANG Y Q,HE C P,et al.Evaluation on 20%prochoraz emulsion water on controlling banana anthracnos[J].J Anhui Agric Sci,2013,41(11):4830-4831.
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[4]周亚奎,陈旭玉,郑服丛.香蕉炭疽病生物防治研究进展[J].中国农学通报,2008,24(4):328-331.ZHOU Y K,CHEN X Y,ZHENG F C.Advance in the study on biological control of banana anthracnose[J].Chin Agric Sci Bull,2008,24(4):328-331.
[5]李其利,宁平,郑露,等.不同培养条件对球孢链霉菌JK-1生长及其产生抗菌物质的影响[J].南方农业学报,2012,43(11):1682-1687.LI Q L,NING P,ZHENG L,et al.Different culture conditions'effects on growth and antifungal substances production of Streptomyces globisporus JK-1[J].J Southern Agric,2012,43(11):1682-1687.
[6]李立梅,李鑫,沈佳龙,等.杨树烂皮病生防链霉菌的筛选及鉴定[J].植物保护学报,2017,44(1):137-144.LI L M,LI X,SHEN J L,et al.Screening and indentification of an actinomycete Streptomyces for the biocontrol of poplar canker disease[J].J Plant Prot,2017,44(1):137-144.
[7]梁春浩.葡萄霜霉病生防放线菌PY-1鉴定及抑菌活性物质结构解析[D].沈阳:沈阳农业大学,2016.LIANG C H.Identification of actinomycetes PY-1 against Plasmopara viticola and structure determination of antimicrobial substances[D].Shenyang:Shenyang Agricultural University,2016.
[8]罗建军,李辉,耿鹏,等.拮抗放线菌MY-4的分离、鉴定及其对果蔬病原菌的抑制作用[J].果树学报,2012,29(4):644-649.LUO J J,LI H,GENG P,et al.Isolation and identification of actinomyce strain MY-4 and its antagonistic activity against pathogenic fungi of fruits and vegetables[J].J Fruit Sci,2012,29(4):644-649.
[9]刘重喜.大豆根部内生放线菌的筛选、鉴定及其活性代谢产物研究[D].哈尔滨:东北农业大学,2013.LIU C X.Sreening and indentification of endophytic actinomycetes from soybean roots and study of their metabolites with bioactivity[D].Harbin:Northeast Agricultural University,2013.
[10]韩秀芳.土壤放线菌分离菌株的分类鉴定与拮抗性筛选[D].北京:北京理工大学,2015.HAN X F.Taxonomic identification and resistance screening of actinomycetes[D].Beijing:Beijing Institute of Technology,2015.
[11]李小俊,成丽霞,吴彦彬,等.拮抗菌抗菌谱及发酵液拮抗能力测定的新方法[J].生物技术,2007,17(1):55-58.LI X J,CHENG L X,WU Y B,et al.New methods of testing fermentation liquid antagonistic effect of antagonistic bacteria and antimicrobial spectrum[J].Biotechnology,2007,17(1):55-58.
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[13]阎逊初.放线菌的分类和鉴定[M].北京:科学出版社,1992.YAN X C.Classification and identification of actinomycetes[M].Beijing:Science Press,1992.
[14]MEHLING A,WEHMEIER U F,PIEPERSBERG W.Nucleotide sequences of Streptomycete 16S ribosomal DNA:towards a specific identification system for streptomycetes using PCR[J].Microbiology,1995,141(9):2139-2147.
[15]CAI Z Y,LI G H,LIN C H,et al.Identifying pathogenicity genes in the rubber tree anthracnose fungus Colletotrichum gloeosporioides through random insertional mutagenesis[J].Microbiol Res,2013,168(6):340-350.
[16]徐大勇,李志栋,李峰,等.番茄灰霉病菌拮抗放线菌的筛选、鉴定及其活性评价[J].中国生物防治学报,2012,28(2):298-302.XU D Y,LI Z D,LI F,et al.Screening,identification and inhibitory activity of an actinomycete strain and against Botrytis cinerea[J].Chin J Biol Control,2012,28(2):298-302.
[17]柳志强,郑媛,吴亚腾,等.阴香内生细菌YX6的鉴定及对香蕉炭疽病的防效[J].西南农业学报,2014,27(6):2402-2404.LIU Z Q,ZHENG Y,WU Y T,et al.Study on identification of Cinnamomum burmannii endophytic bacterial strain YX6 and its control effect on Colletotrichum musae[J].Southwest China J Agric Sci,2014,27(6):2402-2404.
[18]张斯颖,骆焱平,胡坚,等.阴香内生菌YXG2-3的抑菌活性与生长条件优化[J].西北农林科技大学学报(自然科学版),2018,46(7):87-94.ZHANG S Y,LUO Y P,HU J,et al.Optimal growth conditions and antibacterial activities of endophyte YXG2-3 from Cinnamoumum burmanii[J].J Northwest A&.F Univ(Nat.Sci.Ed.),2018,46(7):87-94.
[19]欧雄常,柳凤,何红,等.红树内生细菌AmS2防治香蕉采后炭疽病的研究[J].保鲜与加工,2013,13(5):15-19.OU X C,LIU F,HE H,et al.Biological control banana anthracnose by endophytic bacterial strain AmS2 from mangrove[J].Storage and Process,2013,13(5):15-19.
[20]李振华,凌金锋,曾会才.6株土壤链霉菌的抑真菌活性及其发酵液对采后香蕉果实炭疽病的防效[J].热带农业科学,2006,26(3):35-37,55.LI Z H,LING J F,ZENG H C.Antifungal activity of six soil Streptomyces strains and their control effects against banana postharvest anthracnose[J].Chin J Trop Agric,2006,26(3):35-37,55.
[21]郝永丽,宗兆锋.4株放线菌的防病促生作用研究[J].西北农业学报,2007,16(3):257-259.HAO Y L,ZONG Z F.Biocontrol and growth promotion effects of four actinomycetes strains[J].Acta Agriculturae Boreali-Occidentalis Sinica,2007,16(3):257-259.
[22]付岗,黄思良,史国英,等.香蕉叶斑病的药剂防治研究[J].植物保护,2010,36(1):143-147.FU G,HUANG S L,SHI G Y,et al.Studies on the control of the banana leaf spot diseases with pesticides[J].Plant Prot,2010,36(1):143-147.
[23]罗萍,区建发,杨炜钊,等.香蕉炭疽菌拮抗生防菌株的筛选及鉴定[J].现代食品科技,2011,27(3):275-278.LUO P,OU J F,YANG W Z,et al.Screening and identification of antagonistic organisms against banana anthracnose fungus[J].Mod Food Sci Technol,2011,27(3):275-278.
[24]静玮,张鲁斌,朱德明,等.茶树油控制香蕉采后炭疽病害的研究[J].热带作物学报,2011,32(3):500-503.JING W,ZHANG L B,ZHU D M,et al.Evaluation of tea tree oil to control postharvest banana disease of Colletotrichum musae[J].Chin JTrop Crops,2011,32(3):500-503.
[25]BAZIE S,AYALEW A,WOLDETSADIK K.Integrated management of postharvest banana anthracnose(Colletotrichum musae)through plant extracts and hot water treatment[J].Crop Prot,2014,66:14-18.
[26]MAQBOOL M,ALI A,RAMACHANDRAN S,et al.Control of postharvest anthracnose of banana using a new edible composite coating[J].Crop Prot,2010,29(10):1136-1141.
[2]付岗,黄思良,谢玲,等.45%咪鲜胺水乳剂对香蕉采后炭疽病的控制效果[J].安徽农业科学,2009,37(23):11056-11057,11063.FU G,HUANG S L,XIE L,et al.Control effect of 45%prochloraz EW against postharvest banana anthracnose[J].J Anhui Agric Sci,2009,37(23):11056-11057,11063.
[3]KHAN S H,AKED J,MAGAN N.Control of the anthracnose pathogen of banana(Colletotrichum musae)using antioxidants alone and in combination with thiabendazole or imazalil[J].Plant Pathol,2001,50(5):601-608.
[4]周亚奎,陈旭玉,郑服丛.香蕉炭疽病生物防治研究进展[J].中国农学通报,2008,24(4):328-331.ZHOU Y K,CHEN X Y,ZHENG F C.Advance in the study on biological control of banana anthracnose[J].Chin Agric Sci Bull,2008,24(4):328-331.
[5]李其利,宁平,郑露,等.不同培养条件对球孢链霉菌JK-1生长及其产生抗菌物质的影响[J].南方农业学报,2012,43(11):1682-1687.LI Q L,NING P,ZHENG L,et al.Different culture conditions'effects on growth and antifungal substances production of Streptomyces globisporus JK-1[J].J Southern Agric,2012,43(11):1682-1687.
[6]李立梅,李鑫,沈佳龙,等.杨树烂皮病生防链霉菌的筛选及鉴定[J].植物保护学报,2017,44(1):137-144.LI L M,LI X,SHEN J L,et al.Screening and indentification of an actinomycete Streptomyces for the biocontrol of poplar canker disease[J].J Plant Prot,2017,44(1):137-144.
[7]梁春浩.葡萄霜霉病生防放线菌PY-1鉴定及抑菌活性物质结构解析[D].沈阳:沈阳农业大学,2016.LIANG C H.Identification of actinomycetes PY-1 against Plasmopara viticola and structure determination of antimicrobial substances[D].Shenyang:Shenyang Agricultural University,2016.
[8]罗建军,李辉,耿鹏,等.拮抗放线菌MY-4的分离、鉴定及其对果蔬病原菌的抑制作用[J].果树学报,2012,29(4):644-649.LUO J J,LI H,GENG P,et al.Isolation and identification of actinomyce strain MY-4 and its antagonistic activity against pathogenic fungi of fruits and vegetables[J].J Fruit Sci,2012,29(4):644-649.
[9]刘重喜.大豆根部内生放线菌的筛选、鉴定及其活性代谢产物研究[D].哈尔滨:东北农业大学,2013.LIU C X.Sreening and indentification of endophytic actinomycetes from soybean roots and study of their metabolites with bioactivity[D].Harbin:Northeast Agricultural University,2013.
[10]韩秀芳.土壤放线菌分离菌株的分类鉴定与拮抗性筛选[D].北京:北京理工大学,2015.HAN X F.Taxonomic identification and resistance screening of actinomycetes[D].Beijing:Beijing Institute of Technology,2015.
[11]李小俊,成丽霞,吴彦彬,等.拮抗菌抗菌谱及发酵液拮抗能力测定的新方法[J].生物技术,2007,17(1):55-58.LI X J,CHENG L X,WU Y B,et al.New methods of testing fermentation liquid antagonistic effect of antagonistic bacteria and antimicrobial spectrum[J].Biotechnology,2007,17(1):55-58.
[12]中国科学院微生物研究所放线菌分类组.链霉菌鉴定手册[M].北京:科学出版社,1975:13-15.Taxonomy of Actinomycetes from the Institute of Microbiology,Academy of Sciences of China Academy of Sciences.Streptomyces identification manual[M].Beijing:Science Press,1975,13-15.
[13]阎逊初.放线菌的分类和鉴定[M].北京:科学出版社,1992.YAN X C.Classification and identification of actinomycetes[M].Beijing:Science Press,1992.
[14]MEHLING A,WEHMEIER U F,PIEPERSBERG W.Nucleotide sequences of Streptomycete 16S ribosomal DNA:towards a specific identification system for streptomycetes using PCR[J].Microbiology,1995,141(9):2139-2147.
[15]CAI Z Y,LI G H,LIN C H,et al.Identifying pathogenicity genes in the rubber tree anthracnose fungus Colletotrichum gloeosporioides through random insertional mutagenesis[J].Microbiol Res,2013,168(6):340-350.
[16]徐大勇,李志栋,李峰,等.番茄灰霉病菌拮抗放线菌的筛选、鉴定及其活性评价[J].中国生物防治学报,2012,28(2):298-302.XU D Y,LI Z D,LI F,et al.Screening,identification and inhibitory activity of an actinomycete strain and against Botrytis cinerea[J].Chin J Biol Control,2012,28(2):298-302.
[17]柳志强,郑媛,吴亚腾,等.阴香内生细菌YX6的鉴定及对香蕉炭疽病的防效[J].西南农业学报,2014,27(6):2402-2404.LIU Z Q,ZHENG Y,WU Y T,et al.Study on identification of Cinnamomum burmannii endophytic bacterial strain YX6 and its control effect on Colletotrichum musae[J].Southwest China J Agric Sci,2014,27(6):2402-2404.
[18]张斯颖,骆焱平,胡坚,等.阴香内生菌YXG2-3的抑菌活性与生长条件优化[J].西北农林科技大学学报(自然科学版),2018,46(7):87-94.ZHANG S Y,LUO Y P,HU J,et al.Optimal growth conditions and antibacterial activities of endophyte YXG2-3 from Cinnamoumum burmanii[J].J Northwest A&.F Univ(Nat.Sci.Ed.),2018,46(7):87-94.
[19]欧雄常,柳凤,何红,等.红树内生细菌AmS2防治香蕉采后炭疽病的研究[J].保鲜与加工,2013,13(5):15-19.OU X C,LIU F,HE H,et al.Biological control banana anthracnose by endophytic bacterial strain AmS2 from mangrove[J].Storage and Process,2013,13(5):15-19.
[20]李振华,凌金锋,曾会才.6株土壤链霉菌的抑真菌活性及其发酵液对采后香蕉果实炭疽病的防效[J].热带农业科学,2006,26(3):35-37,55.LI Z H,LING J F,ZENG H C.Antifungal activity of six soil Streptomyces strains and their control effects against banana postharvest anthracnose[J].Chin J Trop Agric,2006,26(3):35-37,55.
[21]郝永丽,宗兆锋.4株放线菌的防病促生作用研究[J].西北农业学报,2007,16(3):257-259.HAO Y L,ZONG Z F.Biocontrol and growth promotion effects of four actinomycetes strains[J].Acta Agriculturae Boreali-Occidentalis Sinica,2007,16(3):257-259.
[22]付岗,黄思良,史国英,等.香蕉叶斑病的药剂防治研究[J].植物保护,2010,36(1):143-147.FU G,HUANG S L,SHI G Y,et al.Studies on the control of the banana leaf spot diseases with pesticides[J].Plant Prot,2010,36(1):143-147.
[23]罗萍,区建发,杨炜钊,等.香蕉炭疽菌拮抗生防菌株的筛选及鉴定[J].现代食品科技,2011,27(3):275-278.LUO P,OU J F,YANG W Z,et al.Screening and identification of antagonistic organisms against banana anthracnose fungus[J].Mod Food Sci Technol,2011,27(3):275-278.
[24]静玮,张鲁斌,朱德明,等.茶树油控制香蕉采后炭疽病害的研究[J].热带作物学报,2011,32(3):500-503.JING W,ZHANG L B,ZHU D M,et al.Evaluation of tea tree oil to control postharvest banana disease of Colletotrichum musae[J].Chin JTrop Crops,2011,32(3):500-503.
[25]BAZIE S,AYALEW A,WOLDETSADIK K.Integrated management of postharvest banana anthracnose(Colletotrichum musae)through plant extracts and hot water treatment[J].Crop Prot,2014,66:14-18.
[26]MAQBOOL M,ALI A,RAMACHANDRAN S,et al.Control of postharvest anthracnose of banana using a new edible composite coating[J].Crop Prot,2010,29(10):1136-1141.