枯草芽孢杆菌BC80-6发酵条件的优化及对烟草根黑腐病的控病效果
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摘要
探究了3种枯草芽孢杆菌BC80-6处理液对烟草根黑腐病菌菌丝生长、孢子萌发的抑制作用,对其发酵条件进行了优化,并开展田间控病防效试验。结果表明:10~(-1)倍枯草芽孢杆菌BC80-6的活菌液对烟草根黑腐病菌菌丝生长和孢子萌发的抑制效果最好,抑制率分别为93.2%、95.3%。通过单因素试验及正交试验对菌株BC80-6培养基的配方进行优化,确定培养基的最佳配方为蔗糖1.5%、酵母浸出物0.5%、硫酸镁0.06%(质量体积比)。最佳发酵条件:发酵温度28℃、时间24 h、初始pH 6.5、摇床转速180 r/min、接种量5%、装液量40 mL(250 mL三角瓶)。枯草芽孢杆菌活菌液对烟草根黑腐病的田间防效为88.19%。
To clarify the effects of Bacillus subtilis BC80-6 on Thielaviopsis basicola, the inhibitory effects of three treatment solutions of B. subtilis BC80-6 on the mycelial growth and spore germination of T. basicola were investigated. BC80-6 fermentation conditions were optimized and field control experiments were carried out. The results showed that 10~(-1) times living bacteria solution of B. subtilis BC80-6 had the best inhibitory effects on the mycelial growth and spore germination of B. subtilis with the inhibition rates of 93.2% and 95.3%, respectively. Via a single factor test and an orthogonal experiment, the optimum BC80-6 medium was 1.5% sucrose, 0.5% yeast extract and 0.06% magnesium sulfate(mass/volume ratio). The best fermentation condition was fermentation temperature 28 ℃, time 24 h, initial pH 6.5, rotation speed 180 r/min, inoculation amount 5%(v/v) and medium volume 40 mL in 250 mL triangular bottle. The field control effect of living bacteria solution of B. subtilis on tobacco root black rot was 88.19%.
To clarify the effects of Bacillus subtilis BC80-6 on Thielaviopsis basicola, the inhibitory effects of three treatment solutions of B. subtilis BC80-6 on the mycelial growth and spore germination of T. basicola were investigated. BC80-6 fermentation conditions were optimized and field control experiments were carried out. The results showed that 10~(-1) times living bacteria solution of B. subtilis BC80-6 had the best inhibitory effects on the mycelial growth and spore germination of B. subtilis with the inhibition rates of 93.2% and 95.3%, respectively. Via a single factor test and an orthogonal experiment, the optimum BC80-6 medium was 1.5% sucrose, 0.5% yeast extract and 0.06% magnesium sulfate(mass/volume ratio). The best fermentation condition was fermentation temperature 28 ℃, time 24 h, initial pH 6.5, rotation speed 180 r/min, inoculation amount 5%(v/v) and medium volume 40 mL in 250 mL triangular bottle. The field control effect of living bacteria solution of B. subtilis on tobacco root black rot was 88.19%.
引文
[1]Debois D,Hamze K,Guérineau V,et al.In situ localisation and quantification of surfactins in a Bacillus subtilis swarming community by imaging mass spectrometry[J].Proteomics,2008,8(18):3682-3691.
[2]Ahimou F,Jacques P,Deleu M.Surfactin and iturin Aeffects on Bacillus subtilis surface hydrophobicity[J].Enzyme and Microbial Technology,2000,27(10):749-754.
[3]Stein T.Bacillus subtilis antibiotics:structures,syntheses and specific functions[J].Molecular Microbiology,2005,56(4):845-857.
[4]赵白鸽,孔建,王文夕,等.枯草芽孢杆菌B-903对苹果轮纹菌的抑菌作用及其对病害的控制效果[J].植物病理学报,1997,27(3):213-214.ZHAO Baige,KONG Jian,WANG Wenxi,et al.Suppressions of apple ring rot pathogen(Physalospora piricola)by Bacillus subtilis and ITS effect in field control[J].Acta Phytopathologica Sinica,1997,27(3):213-214.
[5]刘雪,穆常青,蒋细良,等.枯草芽孢杆菌代谢物质的研究进展及其在植病生防中的应用[J].中国生物防治,2006,22(增刊1):179-184.LIU Xue,MU Changqing,JIANG Xiliang,et al.Research progress of the metabolic substances produced by Bacillus subtillis and their application on biocontrol of plant disease[J].Chinese Journal of Biological Control,2006,22(S1):179-184.
[6]李永刚,宋兴舜,赵雪莹,等.生防枯草芽孢杆菌L1特性的初步研究[J].植物保护,2008,34(1):57-61.LI Yonggang,SONG Xingshun,ZHAO Xueying,et al.Preliminary characterization of Bacillus subtilis strain L1[J].Plant Protection,2008,34(1):57-61.
[7]乔俊卿,陈志谊,梁雪杰,等.枯草芽孢杆菌Bs916在番茄根部的定殖[J].江苏农业学报,2015,31(6):1278-1283.QIAO Junqing,CHEN Zhiyi,LIANG Xuejie,et al.Colonization of Bacillus subtilis BS916 on tomato roots[J].Jiangsu Journal of Agricultural Sciences,2015,31(6):1278-1283.
[8]刘庆丰,熊国如,毛自朝,等.枯草芽孢杆菌XF-1的根围定殖能力分析[J].植物保护学报,2012,39(5):425-430.LIU Qingfeng,XIONG Guoru,MAO Zichao,et al.Analyses for the colonization ability of Bacillus subtilis XF-1 in the rhizosphere[J].Acta Phytophylacica Sinica,2012,39(5):425-430.
[9]顾真荣,马承铸,韩长安.枯草芽孢杆菌G3防治植病盆栽试验[J].上海农业学报,2002,18(1):77-80.GU Zhenrong,MA Chengzhu,HAN Changan.Pot test on plant disease with Bacillus subtilis G3strain[J].Acta Agriculturae Shanghai,2002,18(1):77-80.
[10]黎起秦,罗宽,林纬,等.内生菌B47的定殖能力及其对番茄青枯病的防治作用[J].植物保护学报,2006,33(4):363-368.LI Qiqin,LUO Kuan,LIN Wei,et al.Analysis on the colonization of entophytic bacteria B47 and its control on tomato bacterial wilt[J].Acta Phytophylacica Sinica,2006,33(4):363-368.
[11]易龙,马冠华,肖崇刚.烟草根黑腐病拮抗内生细菌的筛选及其抑菌作用[J].微生物学通报,2012,39(10):1464-1470.YI Long,MA Guanhua,XIAO Chonggang.Inhibition effect and screening of antagonistic endophytic bacteria against Thielaviopsis basicola[J].Microbiology China,2012,39(10):1464-1470.
[12]檀根甲,李增智,刘淑芳,等.枯草芽孢杆菌BS80-6对苹果采后炭疽病的控病效果及作用机制[J].植物保护学报,2008,35(3):227-232.TAN Genjia,LI Zengzhi,LIU Shufang,et al.Mechanisms of action and efficacy of Bacillus subtilis BS80-6 active against postharvest anthracnose pathogen on apples[J].Acta Phytophylacica Sinica,2008,35(3):227-232.
[13]叶云峰,黎起秦,袁高庆,等.枯草芽孢杆菌B47菌株高产抗菌物质的培养基及发酵条件优化[J].微生物学通报,2011,38(9):1339-1346.YE Yunfeng,LI Qiqin,YUAN Gaoqing,et al.Optimization of culture medium and fermentation conditions for high production of antimicrobial substance by Bacillus subtilis strain B47[J].Microbiology China,2011,38(9):1339-1346.
[14]GB/T 23222-2008烟草病虫害分级及调查方法[S].GB/T 23222-2008 Grade and investigation method of tobacco diseases and insect pests[S].
[15]St?ver A G,Driks A.Secretion,localization,and antibacterial activity of TasA,a Bacillus subtilis spore-associated protein[J].Journal of Bacteriology,1999,181(5):1664-1672.
[16]孔建,赵白鸽,王文夕,等.枯草芽孢杆菌(Bacillus subtilis(Cohen))B-903菌株抗菌物质对植物病原真菌的抑制作用[J].植物病理学报,1995,25(1):69-72.KONG Jian,ZHAO Baige,WANG Wenxi,et al.On the inhibitory action of Bacillus subtilis B-903 strain’s antifungal substance on plant pathogenic fungi[J].Acta Phytopathologica Sinica,1995,25(1):69-72.
[17]Wilson C L,Chalutz E.Postharvest biological control of Penicillium rots of citrus with antagonistic yeasts and bacteria[J].Scientia Horticulturae,1989,40(2):105-112.
[18]Gutter Y,Littauer F.Antagonistic action of Bacillus subtilis against citrus fruit pathogens[J].Bulletin of the Research Council of Israel,1953,33(3):192-197.
[19]秦艳,李卫芬,黄琴.枯草芽孢杆菌发酵条件的优化[J].饲料研究,2007(12):70-74.QIN Yan,LI Weifen,HUANG Qin.Optimization of Bacillus subtilis fermentation conditions[J].Feed Research,2007(12):70-74.
[20]刘雪,叶婧,穆长青,等.枯草芽孢杆菌B-332菌株发酵条件的研究[J].中国农学通报,2012,28(10):230-235.LIU Xue,YE Jing,MU Changqing,et al.Research on the fermentation conditions of Bacillus subtillus B-332[J].Chinese Agricultural Science Bulletin,2012,28(10):230-235.
[21]章四平,刘圣明,王建新,等.枯草芽孢杆菌生防菌株NJ-18的发酵条件优化[J].南京农业大学学报,2010,33(2):58-62.ZHANG Siping,LIU Shengming,WANG Jianxin,et al.Optimizing fermentation condition for the antagonistic Bacillus subtilis NJ-18 strain[J].Journal of Nanjing Agricultural University,2010,33(2):58-62.
[22]周芳,谈静惠,王美琴,等.枯草芽孢杆菌D-29菌株发酵条件的优化[J].山西农业科学,2013,41(12):1333-1335.ZHOU Fang,TAN Jinghui,WANG Meiqin,et al.Optimization of fermentation conditions of Bacillus subtilis strain D-29[J].Journal of Shanxi Agricultural Sciences,2013,41(12):1333-1335.
[23]孙沙沙,夏振远,吴德喜.枯草芽孢杆菌CG24发酵条件优化[J].云南农业大学学报,2013,28(1):36-43.SUN Shasha,XIA Zhenyuan,WU Dexi.Optimizing fermentation condition for the Bacillus subtilis[J].Journal of Yunnan Agricultural University,2013,28(1):36-43.
[24]易龙,肖崇刚,马冠华.防治烟草根黑腐病拮抗芽孢菌株的筛选[J].植物病理学报,2011,41(3):333-336.YI Long,XIAO Chonggang,MA Guanhua.Screening of antagonistic Bacillus spp.against Thielaviopsis basicola[J].Acta Phytopathologica Sinica,2011,41(3):333-336.
[2]Ahimou F,Jacques P,Deleu M.Surfactin and iturin Aeffects on Bacillus subtilis surface hydrophobicity[J].Enzyme and Microbial Technology,2000,27(10):749-754.
[3]Stein T.Bacillus subtilis antibiotics:structures,syntheses and specific functions[J].Molecular Microbiology,2005,56(4):845-857.
[4]赵白鸽,孔建,王文夕,等.枯草芽孢杆菌B-903对苹果轮纹菌的抑菌作用及其对病害的控制效果[J].植物病理学报,1997,27(3):213-214.ZHAO Baige,KONG Jian,WANG Wenxi,et al.Suppressions of apple ring rot pathogen(Physalospora piricola)by Bacillus subtilis and ITS effect in field control[J].Acta Phytopathologica Sinica,1997,27(3):213-214.
[5]刘雪,穆常青,蒋细良,等.枯草芽孢杆菌代谢物质的研究进展及其在植病生防中的应用[J].中国生物防治,2006,22(增刊1):179-184.LIU Xue,MU Changqing,JIANG Xiliang,et al.Research progress of the metabolic substances produced by Bacillus subtillis and their application on biocontrol of plant disease[J].Chinese Journal of Biological Control,2006,22(S1):179-184.
[6]李永刚,宋兴舜,赵雪莹,等.生防枯草芽孢杆菌L1特性的初步研究[J].植物保护,2008,34(1):57-61.LI Yonggang,SONG Xingshun,ZHAO Xueying,et al.Preliminary characterization of Bacillus subtilis strain L1[J].Plant Protection,2008,34(1):57-61.
[7]乔俊卿,陈志谊,梁雪杰,等.枯草芽孢杆菌Bs916在番茄根部的定殖[J].江苏农业学报,2015,31(6):1278-1283.QIAO Junqing,CHEN Zhiyi,LIANG Xuejie,et al.Colonization of Bacillus subtilis BS916 on tomato roots[J].Jiangsu Journal of Agricultural Sciences,2015,31(6):1278-1283.
[8]刘庆丰,熊国如,毛自朝,等.枯草芽孢杆菌XF-1的根围定殖能力分析[J].植物保护学报,2012,39(5):425-430.LIU Qingfeng,XIONG Guoru,MAO Zichao,et al.Analyses for the colonization ability of Bacillus subtilis XF-1 in the rhizosphere[J].Acta Phytophylacica Sinica,2012,39(5):425-430.
[9]顾真荣,马承铸,韩长安.枯草芽孢杆菌G3防治植病盆栽试验[J].上海农业学报,2002,18(1):77-80.GU Zhenrong,MA Chengzhu,HAN Changan.Pot test on plant disease with Bacillus subtilis G3strain[J].Acta Agriculturae Shanghai,2002,18(1):77-80.
[10]黎起秦,罗宽,林纬,等.内生菌B47的定殖能力及其对番茄青枯病的防治作用[J].植物保护学报,2006,33(4):363-368.LI Qiqin,LUO Kuan,LIN Wei,et al.Analysis on the colonization of entophytic bacteria B47 and its control on tomato bacterial wilt[J].Acta Phytophylacica Sinica,2006,33(4):363-368.
[11]易龙,马冠华,肖崇刚.烟草根黑腐病拮抗内生细菌的筛选及其抑菌作用[J].微生物学通报,2012,39(10):1464-1470.YI Long,MA Guanhua,XIAO Chonggang.Inhibition effect and screening of antagonistic endophytic bacteria against Thielaviopsis basicola[J].Microbiology China,2012,39(10):1464-1470.
[12]檀根甲,李增智,刘淑芳,等.枯草芽孢杆菌BS80-6对苹果采后炭疽病的控病效果及作用机制[J].植物保护学报,2008,35(3):227-232.TAN Genjia,LI Zengzhi,LIU Shufang,et al.Mechanisms of action and efficacy of Bacillus subtilis BS80-6 active against postharvest anthracnose pathogen on apples[J].Acta Phytophylacica Sinica,2008,35(3):227-232.
[13]叶云峰,黎起秦,袁高庆,等.枯草芽孢杆菌B47菌株高产抗菌物质的培养基及发酵条件优化[J].微生物学通报,2011,38(9):1339-1346.YE Yunfeng,LI Qiqin,YUAN Gaoqing,et al.Optimization of culture medium and fermentation conditions for high production of antimicrobial substance by Bacillus subtilis strain B47[J].Microbiology China,2011,38(9):1339-1346.
[14]GB/T 23222-2008烟草病虫害分级及调查方法[S].GB/T 23222-2008 Grade and investigation method of tobacco diseases and insect pests[S].
[15]St?ver A G,Driks A.Secretion,localization,and antibacterial activity of TasA,a Bacillus subtilis spore-associated protein[J].Journal of Bacteriology,1999,181(5):1664-1672.
[16]孔建,赵白鸽,王文夕,等.枯草芽孢杆菌(Bacillus subtilis(Cohen))B-903菌株抗菌物质对植物病原真菌的抑制作用[J].植物病理学报,1995,25(1):69-72.KONG Jian,ZHAO Baige,WANG Wenxi,et al.On the inhibitory action of Bacillus subtilis B-903 strain’s antifungal substance on plant pathogenic fungi[J].Acta Phytopathologica Sinica,1995,25(1):69-72.
[17]Wilson C L,Chalutz E.Postharvest biological control of Penicillium rots of citrus with antagonistic yeasts and bacteria[J].Scientia Horticulturae,1989,40(2):105-112.
[18]Gutter Y,Littauer F.Antagonistic action of Bacillus subtilis against citrus fruit pathogens[J].Bulletin of the Research Council of Israel,1953,33(3):192-197.
[19]秦艳,李卫芬,黄琴.枯草芽孢杆菌发酵条件的优化[J].饲料研究,2007(12):70-74.QIN Yan,LI Weifen,HUANG Qin.Optimization of Bacillus subtilis fermentation conditions[J].Feed Research,2007(12):70-74.
[20]刘雪,叶婧,穆长青,等.枯草芽孢杆菌B-332菌株发酵条件的研究[J].中国农学通报,2012,28(10):230-235.LIU Xue,YE Jing,MU Changqing,et al.Research on the fermentation conditions of Bacillus subtillus B-332[J].Chinese Agricultural Science Bulletin,2012,28(10):230-235.
[21]章四平,刘圣明,王建新,等.枯草芽孢杆菌生防菌株NJ-18的发酵条件优化[J].南京农业大学学报,2010,33(2):58-62.ZHANG Siping,LIU Shengming,WANG Jianxin,et al.Optimizing fermentation condition for the antagonistic Bacillus subtilis NJ-18 strain[J].Journal of Nanjing Agricultural University,2010,33(2):58-62.
[22]周芳,谈静惠,王美琴,等.枯草芽孢杆菌D-29菌株发酵条件的优化[J].山西农业科学,2013,41(12):1333-1335.ZHOU Fang,TAN Jinghui,WANG Meiqin,et al.Optimization of fermentation conditions of Bacillus subtilis strain D-29[J].Journal of Shanxi Agricultural Sciences,2013,41(12):1333-1335.
[23]孙沙沙,夏振远,吴德喜.枯草芽孢杆菌CG24发酵条件优化[J].云南农业大学学报,2013,28(1):36-43.SUN Shasha,XIA Zhenyuan,WU Dexi.Optimizing fermentation condition for the Bacillus subtilis[J].Journal of Yunnan Agricultural University,2013,28(1):36-43.
[24]易龙,肖崇刚,马冠华.防治烟草根黑腐病拮抗芽孢菌株的筛选[J].植物病理学报,2011,41(3):333-336.YI Long,XIAO Chonggang,MA Guanhua.Screening of antagonistic Bacillus spp.against Thielaviopsis basicola[J].Acta Phytopathologica Sinica,2011,41(3):333-336.