一株桑树内生拮抗菌的分离、鉴定及发酵条件优化
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摘要
【目的】利用植物内生拮抗菌防治植物病害是一种有效的生物防治手段。本研究从健康桑树中分离筛选桑椹菌核病拮抗性内生细菌,为桑椹菌核病生物防治提供优良菌种。【方法】釆用组织分离培养法及抑菌圈法分离、筛选桑椹菌核病拮抗性内生细菌;根据菌体形态、培养特征、生理生化特性及基于16S rDNA序列的系统发育分析,对抑菌活性显著且稳定的菌株进行菌种鉴定;进而利用菌丝生长速率法检测活性发酵液的抑菌谱与热稳定性,并通过单因素及正交试验优化该菌株产生抑菌活性物质的发酵条件。【结果】从健康桑树中共分离获得55株内生细菌,其中XP-27菌株对核盘菌PZ-2的抑菌活性稳定且拮抗效果明显;XP-27菌株形态、培养特征、生理生化特性与芽孢杆菌属相符,基于16S rDNA序列的系统发育分析结果显示该菌株与多株甲基营养型芽孢杆菌(Bacillus methylotrophicus)的亲缘关系最近,且处于系统发育树的同一分枝,故将XP-27菌株鉴定为甲基营养型芽孢杆菌,命名为B.methylotrophicusXP-27;抑菌谱与热稳定性实验结果表明XP-27菌株对灰霉菌SWU5、腐霉菌SWU3等10种常见植物病原菌具不同程度的抑制作用,且其发酵滤液热稳定性强;发酵条件优化结果表明该菌株最佳培养基配方与培养条件为:牛肉膏1.00%,淀粉1.50%,K_2HPO_4 0.05%,MgSO_4·7H_2O 0.10%,初始pH值为8.0,培养温度为30°C,接种量为7%,发酵时间为120 h。【结论】筛选获得的桑树内生细菌B. methylotrophicus XP-27对桑椹菌核病病原菌核盘菌PZ-2具有显著拮抗作用,可作为开发桑椹菌核病生防制剂的候选菌株。
[Objective] Application of antagonistic endophyte is a potential strategy for biological control of plant diseases. In this study, the antagonistic endophytic bacteria were isolated and screened from healthy mulberry, to provide candidate strains for the biological control of mulberry fruit sclerotiniose. [Methods] Tissue isolation method was used to isolate the endophytic bacteria from healthy mulberry tissues. The endophytic bacterium with strong and stable antagonistic activity on the pathogens of mulberry fruit sclerotiniose was screened by inhibition zone method. The antagonistic bacterium was identified through morphological features, cultural, physiological and biochemical characteristics and 16 S rDNA phylogenetic analysis. Antimicrobial spectrum and thermal stability of cell-free fermentation supernatant were assayed by mycelial growth rate. The fermentation conditions and medium composition were optimized through single factor and orthogonal experiment. [Results] In total 55 endophytic bacteria were isolated from healthy mulberry. Among them, strain XP-27 was found to have strong and stable antagonistic activity against Sclerotinia sclerotiorum PZ-2(the pathogen of mulberry fruit sclerotiniose). Morphological features, cultural, physiological and biochemical characteristics indicated that XP-27 belongs to genus of Bacillus. Based on 16 S rDNA phylogenetic analysis, XP-27 was close to the Bacillus methylotrophicus and within the same clade with several strains of Bacillus methylotrophicus. Therefore, strain XP-27 was identified as Bacillus methylotrophicus, and named to be B. methylotrophicus XP-27. Antimicrobial spectrum assayed results showed that strain XP-27 had variously antagonistic activity on 10 plant pathogens and had excellent heat stability. The results of fermentation optimization showed that the optimal medium was composed of 1.00% beef extract, 1.50% starch, 0.05% K_2HPO_4, 0.10% MgSO_4·7 H_2O and the optimal culture conditions were inoculation size of 7% for 120 h at 30 °C with initial pH 8.0. [Conclusion] The above results indicated that mulberry endophytic bacterium B. methylotrophicus XP-27 has strong antagonistic activity on S. sclerotiorum PZ-2, being a candidate strain for developing the biological control agent against mulberry fruit sclerotiniose.
[Objective] Application of antagonistic endophyte is a potential strategy for biological control of plant diseases. In this study, the antagonistic endophytic bacteria were isolated and screened from healthy mulberry, to provide candidate strains for the biological control of mulberry fruit sclerotiniose. [Methods] Tissue isolation method was used to isolate the endophytic bacteria from healthy mulberry tissues. The endophytic bacterium with strong and stable antagonistic activity on the pathogens of mulberry fruit sclerotiniose was screened by inhibition zone method. The antagonistic bacterium was identified through morphological features, cultural, physiological and biochemical characteristics and 16 S rDNA phylogenetic analysis. Antimicrobial spectrum and thermal stability of cell-free fermentation supernatant were assayed by mycelial growth rate. The fermentation conditions and medium composition were optimized through single factor and orthogonal experiment. [Results] In total 55 endophytic bacteria were isolated from healthy mulberry. Among them, strain XP-27 was found to have strong and stable antagonistic activity against Sclerotinia sclerotiorum PZ-2(the pathogen of mulberry fruit sclerotiniose). Morphological features, cultural, physiological and biochemical characteristics indicated that XP-27 belongs to genus of Bacillus. Based on 16 S rDNA phylogenetic analysis, XP-27 was close to the Bacillus methylotrophicus and within the same clade with several strains of Bacillus methylotrophicus. Therefore, strain XP-27 was identified as Bacillus methylotrophicus, and named to be B. methylotrophicus XP-27. Antimicrobial spectrum assayed results showed that strain XP-27 had variously antagonistic activity on 10 plant pathogens and had excellent heat stability. The results of fermentation optimization showed that the optimal medium was composed of 1.00% beef extract, 1.50% starch, 0.05% K_2HPO_4, 0.10% MgSO_4·7 H_2O and the optimal culture conditions were inoculation size of 7% for 120 h at 30 °C with initial pH 8.0. [Conclusion] The above results indicated that mulberry endophytic bacterium B. methylotrophicus XP-27 has strong antagonistic activity on S. sclerotiorum PZ-2, being a candidate strain for developing the biological control agent against mulberry fruit sclerotiniose.
引文
[1]Liang GQ,Wu JJ,Shen W,Lu CX,He J.Nutritional analysis and evaluation of mulberry fresh fruit.Modern Agricultural Science and Technology,2011,(16):320-321,325.(in Chinese)梁贵秋,吴婧婧,沈蔚,陆春霞,何骥.桑椹鲜果的营养分析与评价.现代农业科技,2011,(16):320-321,325.
[2]He L,Hu JH,Xu L,Yao TS,Ran C,Li HJ,Liu HQ,Lei HD.Identification and biological characteristics of pathogen of mulberry disease.Southwest China Journal of Agricultural Sciences,2010,23(3):760-763.(in Chinese)贺磊,胡军华,徐立,姚廷山,冉春,李鸿筠,刘浩强,雷慧德.一株桑椹致病菌的鉴定及其生物学特性研究.西南农业学报,2010,23(3):760-763.
[3]Kuai YZ,Wu FA.A review on pathogens of mulberry fruit sclerotiniosis and its control technology.Science of Sericulture,2012,38(6):1099-1104.(in Chinese)蒯元璋,吴福安.桑椹菌核病病原及病害防治技术综述.蚕业科学,2012,38(6):1099-1104.
[4]LüRH,Zhao AC,Yu J,Wang CH,Liu CY,Cai YX,Yu MD.Biological and epidemiological characteristics of the pathogen of hypertrophy sorosis scleroteniosis,Ciboria shiraiana.Acta Microbiologica Sinica,2017,57(3):388-398.(in Chinese)吕蕊花,赵爱春,余建,王传宏,刘长英,蔡雨翔,余茂德.桑椹肥大性菌核病病原菌生物学特性及流行性.微生物学报,2017,57(3):388-398.
[5]Hu JH,Cai YX,Zhou SJ,Zhang JC,Zhang HL,Chen YB,Li PM,Ying GM.Diversity of mulberry sclerotiniose pathogen and ITS analysis.Journal of Ningbo University(Natural Science&Engineering Edition),2011,24(3):20-23.(in Chinese)胡君欢,蔡岳兴,周书军,张建成,张慧丽,陈银宝,李培民,应国铭.宁波桑果基地菌核病菌的多样性与ITS初步分析.宁波大学学报(理工版),2011,24(3):20-23.
[6]Hong SK,Kim WG,Sung GB,Nam SH.Identification and distribution of two fungal species causing sclerotial disease on mulberry fruits in Korea.Mycobiology,2007,35(2):87-90.
[7]LüRH,Zhao AC,Li J,Wang XL,Yu YS,Lu C,Yu MD.Biological study of hypertrophy sorosis scleroteniosis and its molecular characterization based on LSU rRNA.African Journal of Microbiology Research,2013,7(26):3405-3411.
[8]Sultana R,Ju HJ,Chae JC,Kim K,Lee KJ.Identification of Ciboria carunculoides RS103V,a fungus causing popcorn disease on mulberry fruits in Korea.Research in Plant Disease,2013,19(4):308-312.
[9]Ren JQ,Zeng XL,Chen L,Qiu SC,Yang Y,Zhang MH,Zheng ZY.Advances in integrated control of mulberry fruit sclerotiniosis.Science of Sericulture,2017,43(4):699-703.(in Chinese)任杰群,曾秀丽,陈力,邱诗春,杨义,张明海,郑章云.桑椹菌核病综合防治技术研究新进展.蚕业科学,2017,43(4):699-703.
[10]Abudu Rosuli·Kurban,Shi ZM,Mamat Imin·Ayoufu,Shi CY,Lv HR,Liu YZ.The impact of plastic film mclching on the white mulberry fruit disease and fruit quality.Hubei Agricultural Sciences,2016,55(22):5846-5848,5857.(in Chinese)阿布都肉苏力?库尔班,师志敏,麦麦提依明?阿尤甫,石彩云,吕化荣,刘永忠.果桑地表覆膜对桑白果病发病及果实品质的影响.湖北农业科学,2016,55(22):5846-5848,5857.
[11]Ye MQ,Kuang ZS,Zhao XJ,Yang Q,Li QR,Xiao Y,Wang ZJ,Dai FW,Luo GQ.Screening of fungicide to control Ciboria carunculoides under laboratory conditions.Pakistan Journal of Zoology,2014,46(1):53-59.
[12]Ju WT,Kim HB,Sung GB,Park KY,Kim YS.Mulberry popcorn disease occurrence in Korea region and development of integrative control method.International Journal of Industrial Entomology,2016,33(1):36-40.
[13]Sultana R,Kim K.Bacillus thuringiensis C25 suppresses popcorn disease caused by Ciboria shiraiana in mulberry(Morus australis L.).Biocontrol Science and Technology,2016,26(2):145-162.
[14]Ye MQ,Yue HL,Luo GQ,Yang Q,Kuang ZS.Effect of a fungal pathogen,Trichoderma hamatum,on growth and germination of Ciboria carunculoides under laboratory conditions.Pakistan Journal of Zoology,2014,46(5):1377-1384.
[15]Bacon CW,White JF.Microbial endophytes.New York:Marcel Dekker,2000.
[16]Xie J,Strobel GA,Feng T,Ren HS,Mends MT,Zhou ZY,Geary B.An endophytic Coniochaeta velutina producing broad spectrum antimycotics.Journal of Microbiology,2015,53(6):390-397.
[17]Amaresan N,Jayakumar V,Kumar K,Thajuddin N.Isolation and characterization of plant growth promoting endophytic bacteria and their effect on tomato(Lycopersicon esculentum)and chilli(Capsicum annuum)seedling growth.Annals of Microbiology,2012,62(2):805-810.
[18]Santhanam R,Baldwin IT,Groten K.In wild tobacco,Nicotiana attenuata,variation among bacterial communities of isogenic plants is mainly shaped by the local soil microbiota independently of the plants'capacity to produce jasmonic acid.Communicative&Integrative Biology,2015,8(2):e1017160.
[19]Santhanam R,Luu VT,Weinhold A,Goldberg J,Oh Y,Baldwin IT.Native root-associated bacteria rescue a plant from a sudden-wilt disease that emerged during continuous cropping.Proceedings of the National Academy of Sciences of the United States of America,2015,112(36):E5013-E5020.
[20]Mu ZM,Lu GB,Ji XL,Gai YP,Wang YW,Gao HJ,Zha CY.Identification and colonization of an antagonistic endophytic Burkholderia cepacia Lu10-1 isolated from mulberry.Acta Microbiologica Sinica,2008,48(5):623-630.(in Chinese)牟志美,路国兵,冀宪领,盖英萍,王彦文,高绘菊,查传勇.桑树内生拮抗细菌Burkholderia cepacia Lu10-1的分离鉴定及其内生定殖.微生物学报,2008,48(5):623-630.
[21]Tan GX,Wu QY,Jia JQ,Gui ZZ,Du JJ,Chen L.Isolation,identification and antibacterial activity of an endophytic bacterial strain in mulberry.Science of Sericulture,2012,38(3):412-417.(in Chinese)谭广秀,吴琼英,贾俊强,桂仲争,杜金娟,陈炼.一株桑树内生拮抗细菌的分离鉴定与抑菌活性.蚕业科学,2012,38(3):412-417.
[22]Xie J,Xia T,Lin LP,Zuo WD,Zhou ZY.Isolation and identification of an antagonistic endophytic bacterial strain from mulberry.Science of Sericulture,2009,35(1):121-125.(in Chinese)谢洁,夏天,林立鹏,左伟东,周泽扬.一株桑树内生拮抗菌的分离鉴定.蚕业科学,2009,35(1):121-125.
[23]Xie J,Ren HS,Tang CM,Zuo WD,Chen J,Huang CS,Wang ZJ,Dai FW,Zhou ZY.Identification of a mulberry endophytic bacterium and its antagonistic activity on Scleromitula shiraiana.Science of Sericulture,2015,41(5):815-824.(in Chinese)谢洁,任慧爽,唐翠明,左伟东,陈洁,黄传书,王振江,戴凡炜,周泽扬.一株桑树内生细菌的鉴定和对桑椹核地杖菌的拮抗作用.蚕业科学,2015,41(5):815-824.
[24]Ren HS,Xu WF,Wang AY,Zuo WD,Xie J.Research on biodiversity of endophytic bacteria and the antagonistic endophytes in mulberry.Journal of Southwest University(Natural Science Edition),2017,39(1):36-45.(in Chinese)任慧爽,徐伟芳,王爱印,左伟东,谢洁.桑树内生细菌多样性及内生拮抗活性菌群的研究.西南大学学报(自然科学版),2017,39(1):36-45.
[25]布坎南RE,吉本斯NE.伯杰细菌鉴定手册.中国科学院微生物研究所《伯杰细菌鉴定手册》翻译组,译.第8版.北京:科学出版社,1984.
[26]东秀珠,蔡妙英.常见细菌系统鉴定手册.北京:科学出版社,2001.
[27]沈萍,陈向东.微生物学实验.第4版.北京:高等教育出版社,2007.
[28]Weisburg WG,Barns SM,Pelletier DA,Lane DJ.16Sribosomal DNA amplification for phylogenetic study.Journal of Bacteriology,1991,173(2):697-703.
[29]方中达.植病研究方法.第3版.北京:中国农业出版社,1998.
[30]Sun GZ,Yao T,Feng CJ,Chen L,Li JH,Wang LD.Identification and biocontrol potential of antagonistic bacteria strains against Sclerotinia sclerotiorum and their growthpromoting effects on Brassica napus.Biological Control,2017,104:35-43.
[31]Dunlap CA,Kim SJ,Kwon SW,Rooney AP.Bacillus velezensis is not a later heterotypic synonym of Bacillus amyloliquefaciens;Bacillus methylotrophicus,Bacillus amyloliquefaciens subsp.plantarum and‘Bacillus oryzicola’are later heterotypic synonyms of Bacillus velezensis based on phylogenomics.International Journal of Systematic and Evolutionary Microbiology,2016,66(3):1212-1217.
[32]Gond SK,Bergen MS,Torres MS,White Jr JF.Endophytic Bacillus spp.produce antifungal lipopeptides and induce host defence gene expression in maize.Microbiological Research,2015,172:79-87.
[33]Shrestha BK,Karki HS,Groth DE,Jungkhun N,Ham JH.Biological control activities of rice-associated Bacillus sp.strains against sheath blight and bacterial panicle blight of rice.PLoS One,2016,11(1):e0146764.
[34]Stein T.Bacillus subtilis antibiotics:structures,syntheses and specific functions.Molecular Microbiology,2005,56(4):845-857.
[35]Pérez-García A,Romero D,de Vicente A.Plant protection and growth stimulation by microorganisms:biotechnological applications of Bacilli in agriculture.Current Opinion in Biotechnology,2011,22(2):187-193.
[36]Wu LQ,Gu HK,Wang Q,Shang HZ,Liu GJ,Bao F.Antagonistic efficacy and growth-promoting effect of Bacillus methylotrophicus isolated from Dendrobium huoshanense.Biotechnology Bulletin,2016,32(8):200-206.(in Chinese)武利勤,顾海科,王青,尚宏忠,刘桂君,包放.石斛内生甲基营养芽胞杆菌的拮抗和促生作用研究.生物技术通报,2016,32(8):200-206.
[37]Jiang Y,Yin W,Chen CQ,Tian L,Xu P.Identification and optimized fermentation condition of an endophyte antagonistic bacteria NJ13.Agrochemicals,2013,52(2):97-101.(in Chinese)姜云,尹望,陈长卿,田磊,许朋.人参内生拮抗细菌NJ13的鉴定及发酵条件.农药,2013,52(2):97-101.
[38]Cai L,Ou YQ,Hou XZ,Wang JL,Fu L.Isolation and identification of endophytic bacteria from Fenghuang Dancong tea.Science and Technology of Food Industry,2016,37(10):246-250.(in Chinese)蔡丽,欧燕清,侯小桢,王金良,傅力.凤凰单丛茶内生拮抗细菌的筛选与鉴定.食品工业科技,2016,37(10):246-250.
[2]He L,Hu JH,Xu L,Yao TS,Ran C,Li HJ,Liu HQ,Lei HD.Identification and biological characteristics of pathogen of mulberry disease.Southwest China Journal of Agricultural Sciences,2010,23(3):760-763.(in Chinese)贺磊,胡军华,徐立,姚廷山,冉春,李鸿筠,刘浩强,雷慧德.一株桑椹致病菌的鉴定及其生物学特性研究.西南农业学报,2010,23(3):760-763.
[3]Kuai YZ,Wu FA.A review on pathogens of mulberry fruit sclerotiniosis and its control technology.Science of Sericulture,2012,38(6):1099-1104.(in Chinese)蒯元璋,吴福安.桑椹菌核病病原及病害防治技术综述.蚕业科学,2012,38(6):1099-1104.
[4]LüRH,Zhao AC,Yu J,Wang CH,Liu CY,Cai YX,Yu MD.Biological and epidemiological characteristics of the pathogen of hypertrophy sorosis scleroteniosis,Ciboria shiraiana.Acta Microbiologica Sinica,2017,57(3):388-398.(in Chinese)吕蕊花,赵爱春,余建,王传宏,刘长英,蔡雨翔,余茂德.桑椹肥大性菌核病病原菌生物学特性及流行性.微生物学报,2017,57(3):388-398.
[5]Hu JH,Cai YX,Zhou SJ,Zhang JC,Zhang HL,Chen YB,Li PM,Ying GM.Diversity of mulberry sclerotiniose pathogen and ITS analysis.Journal of Ningbo University(Natural Science&Engineering Edition),2011,24(3):20-23.(in Chinese)胡君欢,蔡岳兴,周书军,张建成,张慧丽,陈银宝,李培民,应国铭.宁波桑果基地菌核病菌的多样性与ITS初步分析.宁波大学学报(理工版),2011,24(3):20-23.
[6]Hong SK,Kim WG,Sung GB,Nam SH.Identification and distribution of two fungal species causing sclerotial disease on mulberry fruits in Korea.Mycobiology,2007,35(2):87-90.
[7]LüRH,Zhao AC,Li J,Wang XL,Yu YS,Lu C,Yu MD.Biological study of hypertrophy sorosis scleroteniosis and its molecular characterization based on LSU rRNA.African Journal of Microbiology Research,2013,7(26):3405-3411.
[8]Sultana R,Ju HJ,Chae JC,Kim K,Lee KJ.Identification of Ciboria carunculoides RS103V,a fungus causing popcorn disease on mulberry fruits in Korea.Research in Plant Disease,2013,19(4):308-312.
[9]Ren JQ,Zeng XL,Chen L,Qiu SC,Yang Y,Zhang MH,Zheng ZY.Advances in integrated control of mulberry fruit sclerotiniosis.Science of Sericulture,2017,43(4):699-703.(in Chinese)任杰群,曾秀丽,陈力,邱诗春,杨义,张明海,郑章云.桑椹菌核病综合防治技术研究新进展.蚕业科学,2017,43(4):699-703.
[10]Abudu Rosuli·Kurban,Shi ZM,Mamat Imin·Ayoufu,Shi CY,Lv HR,Liu YZ.The impact of plastic film mclching on the white mulberry fruit disease and fruit quality.Hubei Agricultural Sciences,2016,55(22):5846-5848,5857.(in Chinese)阿布都肉苏力?库尔班,师志敏,麦麦提依明?阿尤甫,石彩云,吕化荣,刘永忠.果桑地表覆膜对桑白果病发病及果实品质的影响.湖北农业科学,2016,55(22):5846-5848,5857.
[11]Ye MQ,Kuang ZS,Zhao XJ,Yang Q,Li QR,Xiao Y,Wang ZJ,Dai FW,Luo GQ.Screening of fungicide to control Ciboria carunculoides under laboratory conditions.Pakistan Journal of Zoology,2014,46(1):53-59.
[12]Ju WT,Kim HB,Sung GB,Park KY,Kim YS.Mulberry popcorn disease occurrence in Korea region and development of integrative control method.International Journal of Industrial Entomology,2016,33(1):36-40.
[13]Sultana R,Kim K.Bacillus thuringiensis C25 suppresses popcorn disease caused by Ciboria shiraiana in mulberry(Morus australis L.).Biocontrol Science and Technology,2016,26(2):145-162.
[14]Ye MQ,Yue HL,Luo GQ,Yang Q,Kuang ZS.Effect of a fungal pathogen,Trichoderma hamatum,on growth and germination of Ciboria carunculoides under laboratory conditions.Pakistan Journal of Zoology,2014,46(5):1377-1384.
[15]Bacon CW,White JF.Microbial endophytes.New York:Marcel Dekker,2000.
[16]Xie J,Strobel GA,Feng T,Ren HS,Mends MT,Zhou ZY,Geary B.An endophytic Coniochaeta velutina producing broad spectrum antimycotics.Journal of Microbiology,2015,53(6):390-397.
[17]Amaresan N,Jayakumar V,Kumar K,Thajuddin N.Isolation and characterization of plant growth promoting endophytic bacteria and their effect on tomato(Lycopersicon esculentum)and chilli(Capsicum annuum)seedling growth.Annals of Microbiology,2012,62(2):805-810.
[18]Santhanam R,Baldwin IT,Groten K.In wild tobacco,Nicotiana attenuata,variation among bacterial communities of isogenic plants is mainly shaped by the local soil microbiota independently of the plants'capacity to produce jasmonic acid.Communicative&Integrative Biology,2015,8(2):e1017160.
[19]Santhanam R,Luu VT,Weinhold A,Goldberg J,Oh Y,Baldwin IT.Native root-associated bacteria rescue a plant from a sudden-wilt disease that emerged during continuous cropping.Proceedings of the National Academy of Sciences of the United States of America,2015,112(36):E5013-E5020.
[20]Mu ZM,Lu GB,Ji XL,Gai YP,Wang YW,Gao HJ,Zha CY.Identification and colonization of an antagonistic endophytic Burkholderia cepacia Lu10-1 isolated from mulberry.Acta Microbiologica Sinica,2008,48(5):623-630.(in Chinese)牟志美,路国兵,冀宪领,盖英萍,王彦文,高绘菊,查传勇.桑树内生拮抗细菌Burkholderia cepacia Lu10-1的分离鉴定及其内生定殖.微生物学报,2008,48(5):623-630.
[21]Tan GX,Wu QY,Jia JQ,Gui ZZ,Du JJ,Chen L.Isolation,identification and antibacterial activity of an endophytic bacterial strain in mulberry.Science of Sericulture,2012,38(3):412-417.(in Chinese)谭广秀,吴琼英,贾俊强,桂仲争,杜金娟,陈炼.一株桑树内生拮抗细菌的分离鉴定与抑菌活性.蚕业科学,2012,38(3):412-417.
[22]Xie J,Xia T,Lin LP,Zuo WD,Zhou ZY.Isolation and identification of an antagonistic endophytic bacterial strain from mulberry.Science of Sericulture,2009,35(1):121-125.(in Chinese)谢洁,夏天,林立鹏,左伟东,周泽扬.一株桑树内生拮抗菌的分离鉴定.蚕业科学,2009,35(1):121-125.
[23]Xie J,Ren HS,Tang CM,Zuo WD,Chen J,Huang CS,Wang ZJ,Dai FW,Zhou ZY.Identification of a mulberry endophytic bacterium and its antagonistic activity on Scleromitula shiraiana.Science of Sericulture,2015,41(5):815-824.(in Chinese)谢洁,任慧爽,唐翠明,左伟东,陈洁,黄传书,王振江,戴凡炜,周泽扬.一株桑树内生细菌的鉴定和对桑椹核地杖菌的拮抗作用.蚕业科学,2015,41(5):815-824.
[24]Ren HS,Xu WF,Wang AY,Zuo WD,Xie J.Research on biodiversity of endophytic bacteria and the antagonistic endophytes in mulberry.Journal of Southwest University(Natural Science Edition),2017,39(1):36-45.(in Chinese)任慧爽,徐伟芳,王爱印,左伟东,谢洁.桑树内生细菌多样性及内生拮抗活性菌群的研究.西南大学学报(自然科学版),2017,39(1):36-45.
[25]布坎南RE,吉本斯NE.伯杰细菌鉴定手册.中国科学院微生物研究所《伯杰细菌鉴定手册》翻译组,译.第8版.北京:科学出版社,1984.
[26]东秀珠,蔡妙英.常见细菌系统鉴定手册.北京:科学出版社,2001.
[27]沈萍,陈向东.微生物学实验.第4版.北京:高等教育出版社,2007.
[28]Weisburg WG,Barns SM,Pelletier DA,Lane DJ.16Sribosomal DNA amplification for phylogenetic study.Journal of Bacteriology,1991,173(2):697-703.
[29]方中达.植病研究方法.第3版.北京:中国农业出版社,1998.
[30]Sun GZ,Yao T,Feng CJ,Chen L,Li JH,Wang LD.Identification and biocontrol potential of antagonistic bacteria strains against Sclerotinia sclerotiorum and their growthpromoting effects on Brassica napus.Biological Control,2017,104:35-43.
[31]Dunlap CA,Kim SJ,Kwon SW,Rooney AP.Bacillus velezensis is not a later heterotypic synonym of Bacillus amyloliquefaciens;Bacillus methylotrophicus,Bacillus amyloliquefaciens subsp.plantarum and‘Bacillus oryzicola’are later heterotypic synonyms of Bacillus velezensis based on phylogenomics.International Journal of Systematic and Evolutionary Microbiology,2016,66(3):1212-1217.
[32]Gond SK,Bergen MS,Torres MS,White Jr JF.Endophytic Bacillus spp.produce antifungal lipopeptides and induce host defence gene expression in maize.Microbiological Research,2015,172:79-87.
[33]Shrestha BK,Karki HS,Groth DE,Jungkhun N,Ham JH.Biological control activities of rice-associated Bacillus sp.strains against sheath blight and bacterial panicle blight of rice.PLoS One,2016,11(1):e0146764.
[34]Stein T.Bacillus subtilis antibiotics:structures,syntheses and specific functions.Molecular Microbiology,2005,56(4):845-857.
[35]Pérez-García A,Romero D,de Vicente A.Plant protection and growth stimulation by microorganisms:biotechnological applications of Bacilli in agriculture.Current Opinion in Biotechnology,2011,22(2):187-193.
[36]Wu LQ,Gu HK,Wang Q,Shang HZ,Liu GJ,Bao F.Antagonistic efficacy and growth-promoting effect of Bacillus methylotrophicus isolated from Dendrobium huoshanense.Biotechnology Bulletin,2016,32(8):200-206.(in Chinese)武利勤,顾海科,王青,尚宏忠,刘桂君,包放.石斛内生甲基营养芽胞杆菌的拮抗和促生作用研究.生物技术通报,2016,32(8):200-206.
[37]Jiang Y,Yin W,Chen CQ,Tian L,Xu P.Identification and optimized fermentation condition of an endophyte antagonistic bacteria NJ13.Agrochemicals,2013,52(2):97-101.(in Chinese)姜云,尹望,陈长卿,田磊,许朋.人参内生拮抗细菌NJ13的鉴定及发酵条件.农药,2013,52(2):97-101.
[38]Cai L,Ou YQ,Hou XZ,Wang JL,Fu L.Isolation and identification of endophytic bacteria from Fenghuang Dancong tea.Science and Technology of Food Industry,2016,37(10):246-250.(in Chinese)蔡丽,欧燕清,侯小桢,王金良,傅力.凤凰单丛茶内生拮抗细菌的筛选与鉴定.食品工业科技,2016,37(10):246-250.