桑树中拮抗内生细菌的分离筛选及其抑菌机制
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摘要
以小麦赤霉病菌为指示菌,进行桑树拮抗内生细菌的分离筛选,开展拮抗内生细菌的对峙培养和盆栽控病试验,并采用分光光度法检测SSY26拮抗菌引入小麦穗部,小麦穗组织中的多酚氧化酶(PPO)、过氧化物酶(POD)和苯丙氨酸解氨酶(PAL)的酶活性。结果显示,SSY26菌株对小麦赤霉病菌具有较好的抑制作用,抑菌率达到58.65%,显微观察显示SSY26菌株可引起小麦赤霉病菌菌丝膨大、畸形,影响其正常生长;分子鉴定SSY26菌株为芽孢杆菌属微生物。小麦盆栽试验结果表明,第18天时SSY26菌悬液对小麦赤霉病的防治效果为27.30%,与化学药剂处理组没有显著差异,且桑树内生细菌SSY26引入到小麦穗部,可提高穗部组织中PPO、PAL及POD等防御酶活性。
The antimicrobial activity of endophytic bacterium from Morus alba L. was examined against wheat head blight using dual culture test and disease-control pot experiment, and the related enzymes activities(PPO, PAL and POD) in wheat spike treated with antagonistic endophytic bacteria SSY26 were detected by spectrophotometric method. The results showed that the SSY26 strain had strong inhibit effect against wheat head blight, with inhibitory rate of 58.65%, and the SSY26 strain, which was identified as Bacillus by molecular identification, could cause the expansion and deformity of phytopathogen mycelia of wheat head blight. The pot experimental results showed that the inhibit effect was 27.30% in the eighteenth day after treated with suspensions of SSY26 strain, without cant difference between SSY26 strain suspensions and chemical treatment, and the enzymes activities of PPO, PAL and POD in wheat spike increased treated with the suspensions of SSY26 strain.
The antimicrobial activity of endophytic bacterium from Morus alba L. was examined against wheat head blight using dual culture test and disease-control pot experiment, and the related enzymes activities(PPO, PAL and POD) in wheat spike treated with antagonistic endophytic bacteria SSY26 were detected by spectrophotometric method. The results showed that the SSY26 strain had strong inhibit effect against wheat head blight, with inhibitory rate of 58.65%, and the SSY26 strain, which was identified as Bacillus by molecular identification, could cause the expansion and deformity of phytopathogen mycelia of wheat head blight. The pot experimental results showed that the inhibit effect was 27.30% in the eighteenth day after treated with suspensions of SSY26 strain, without cant difference between SSY26 strain suspensions and chemical treatment, and the enzymes activities of PPO, PAL and POD in wheat spike increased treated with the suspensions of SSY26 strain.
引文
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[2]罗茂,彭华,高健,等.玉米纹枯病抗性相关miRNA的鉴定与功能分析[J].中国生物化学与分子生物学报,2012,28(12):1122-1132.
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[5]徐剑宏,王建伟,胡晓丹,等.小麦赤霉病菌拮抗菌AF0907的分离鉴定及其拮抗特性[J].江苏农业学报,2013,29(3):517-522.
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[7]苏博,江胡彪,乔建礼,等.玉米纹枯病生防菌DZSY21的抗病机制[J].浙江农业学报,2017,29(3):450-459.
[8]成静,孙璇,丁婷,等.杜仲内生细菌DZBG07对小麦全蚀病的生防作用[J].生物学杂志,2015,32(6):105-108,112.
[9]GHIMIRE S R,CHARLTON N D,CRAVEN K D.The mycorrhizal fungus,Sebacina vermifera,enhances seed germination and biomass production in switch grass(Panicum virgatum L.)[J].Bioenerg Res,2009,2(1/2):51-58.
[10]蔡学清,陈炜,林娜,等.内生细菌在荔枝体内的定殖及其防病保鲜功能[J].应用生态学报,2011,22(8):2140-2146.
[11]YáNEZ-MENDIZáBAL V,ZERIOUH H,VI?AS I,et al.Biological control of peach brown rot(Monilinia spp.)by Bacillus subtilis CPA-8 is based on production of fengycin-like lipopeptides[J].Eur J Plant Pathol,2012,132(4):609-619.
[12]赫荣乔.植物内生菌成为我国当前微生物研究领域的热点[J].微生物学通报,2009,36(1):1.
[13]李峰,徐大勇,王光利,等.一株拮抗赤霉病的小麦内生细菌的筛选和抑菌活性[J].生态学杂志,2011,30(8):1738-1743.
[14]辜运富,张云飞,张小平.一株抗玉米纹枯病内生细菌的分离鉴定及其抗病促生作用[J].微生物学通报,2008,35(8):1240-1245.
[15]张淑梅,沙长青,赵晓宇,等.一株抗真菌内生多粘芽孢杆菌的分离鉴定及对水稻恶苗病菌的抑制作用[J].中国生物工程杂志,2010,30(2):84-88.
[16]杨悦,高绘菊,路国兵,等.桑树内生真菌Macrophomina phaseolina MOD-1产油脂培养基及摇瓶发酵条件的优化试验[J].蚕业科学,2010,36(1):12-19.
[17]路国兵,冀宪领,张瑶,等.桑树内生细菌的分离及生防益菌的筛选[J].蚕业科学,2007,33(3):350-354.
[18]方翔,徐伟芳,牛娜,等.一株桑树内生拮抗菌的分离、鉴定及发酵条件优化[J].微生物学报,2018,58(12):2147-2160.
[19]谭广秀.具有抑菌活性的桑树内生菌的分离鉴定及其抑菌物质研究[D].镇江:江苏科技大学,2012.
[20]HAN Y,WANG Y,BI J L,et al.Constitutive and induced activities of defense-related enzymes in aphid-resistant and aphid-susceptible cultivars of wheat[J].J Chem Ecol,2009,35(2):176-182.
[21]CHEN F,WANG M,ZHENG Y,et al.Quantitative changes of plant defense enzymes and phytohormone in biocontrol of cucumber Fusarium wilt by Bacillus subtilis B579[J].World J Microbiol Biotechnol,2010,26(4):675-684.
[22]MATARESE F S,SARROCCO S,GRUBER S,et al.Biocontrol of Fusarium head blight:interactions between Trichoderma and mycotoxigenic Fusarium[J].Microbiology,2012,158(1):98-106.
[23]慕立义.植物化学保护研究方法[M].北京:中国农业出版社,1994.
[24]丁婷,江腾,江海洋,等.喜树内生真菌的分离、鉴定以及抗植物病原菌活性的初步研究[J].激光生物学报,2010,19(4):501-506.
[25]杜军凯,余桂红,王秀娥,等.赤霉病主效抗性QTL区域SSCP标记的发掘与验证[J].麦类作物学报,2010,30(5):829-834.
[26]李洪连,徐敬友.农业植物病理学实验实习指导[M].2版.北京:中国农业出版社,2007.
[27]AQUINO-BOLA?OS E N,MERCADO-SILVA E.Effects of polyphenol oxidase and peroxidase activity,phenolics and lignin content on the browning of cut jicama[J].Postharvest Biol Tec,2004,33(3):275-283.
[28]梁军锋,薛泉宏,牛小磊,等.7株放线菌在辣椒根部定殖及对辣椒叶片PAL与PPO活性的影响[J].西北植物学报,2005,25(10):2118-2123.
[29]YOSHIDA S,OHBA A,LIANG Y M,et al.Specificity of Pseudomonas isolates on healthy and Fusarium head blight-infected spikelets of wheat heads[J].Microb Ecol,2012,64(1):214-225.
[30]HE J,BOLAND G J,ZHOU T.Concurrent selection for microbial suppression of Fusarium graminearum,Fusarium head blight and deoxynivalenol in wheat[J].J Appl Microbio,2009,106(6):1805-1817.
[31]毛腾霄.枯草芽孢杆菌BS-8D防治玉米纹枯病(Rhizoctonia solani)的研究[D].雅安:四川农业大学,2006.
[32]GOND S K,BERGEN M S,TORRES M S,et al.Endophytic Bacillus spp.produce antifungal lipopeptides and induce host defence gene expression in maize[J].Microbiol Res,2015,172:79-87.
[33]SHRESTHA B K,KARKI H S,GROTH D E,et al.Biological control activities of rice-associated Bacillus sp.strains against sheath blight and bacterial panicle blight of rice[J].PLoS One,2016,11(1):e0146764.
[34]REN J H,LI H,WANG Y F,et al.Biocontrol potential of an endophytic Bacillus pumilus JK-SX001 against poplar canker[J].Biol Control,2013,67(3):421-430.
[35]王光达,黄初女,吴委林,等.不同玉米品种对大斑病的抗性与相关防御酶活性的关系研究[J].玉米科学,2014,22(5):146-152.
[36]冯远娇,王建武,苏贻娟,等.茉莉酸在玉米地上部与地下部系统性诱导防御反应中的作用[J].中国农业科学,2009,42(8):2726-2735.
[2]罗茂,彭华,高健,等.玉米纹枯病抗性相关miRNA的鉴定与功能分析[J].中国生物化学与分子生物学报,2012,28(12):1122-1132.
[3]赵兰飞.小麦赤霉病Ⅱ型抗性机理研究及相关基因的功能鉴定[D].泰安:山东农业大学,2016.
[4]姚金保,马鸿翔,张平平,等.小麦优良亲本宁麦9号的研究与利用[J].核农学报,2012,26(1):17-21.
[5]徐剑宏,王建伟,胡晓丹,等.小麦赤霉病菌拮抗菌AF0907的分离鉴定及其拮抗特性[J].江苏农业学报,2013,29(3):517-522.
[6]周苗苗,刘振林,唐丽霖,等.小麦赤霉病菌拮抗内生菌的抑菌活性及鉴定[J].核农学报,2016,30(3):460-467.
[7]苏博,江胡彪,乔建礼,等.玉米纹枯病生防菌DZSY21的抗病机制[J].浙江农业学报,2017,29(3):450-459.
[8]成静,孙璇,丁婷,等.杜仲内生细菌DZBG07对小麦全蚀病的生防作用[J].生物学杂志,2015,32(6):105-108,112.
[9]GHIMIRE S R,CHARLTON N D,CRAVEN K D.The mycorrhizal fungus,Sebacina vermifera,enhances seed germination and biomass production in switch grass(Panicum virgatum L.)[J].Bioenerg Res,2009,2(1/2):51-58.
[10]蔡学清,陈炜,林娜,等.内生细菌在荔枝体内的定殖及其防病保鲜功能[J].应用生态学报,2011,22(8):2140-2146.
[11]YáNEZ-MENDIZáBAL V,ZERIOUH H,VI?AS I,et al.Biological control of peach brown rot(Monilinia spp.)by Bacillus subtilis CPA-8 is based on production of fengycin-like lipopeptides[J].Eur J Plant Pathol,2012,132(4):609-619.
[12]赫荣乔.植物内生菌成为我国当前微生物研究领域的热点[J].微生物学通报,2009,36(1):1.
[13]李峰,徐大勇,王光利,等.一株拮抗赤霉病的小麦内生细菌的筛选和抑菌活性[J].生态学杂志,2011,30(8):1738-1743.
[14]辜运富,张云飞,张小平.一株抗玉米纹枯病内生细菌的分离鉴定及其抗病促生作用[J].微生物学通报,2008,35(8):1240-1245.
[15]张淑梅,沙长青,赵晓宇,等.一株抗真菌内生多粘芽孢杆菌的分离鉴定及对水稻恶苗病菌的抑制作用[J].中国生物工程杂志,2010,30(2):84-88.
[16]杨悦,高绘菊,路国兵,等.桑树内生真菌Macrophomina phaseolina MOD-1产油脂培养基及摇瓶发酵条件的优化试验[J].蚕业科学,2010,36(1):12-19.
[17]路国兵,冀宪领,张瑶,等.桑树内生细菌的分离及生防益菌的筛选[J].蚕业科学,2007,33(3):350-354.
[18]方翔,徐伟芳,牛娜,等.一株桑树内生拮抗菌的分离、鉴定及发酵条件优化[J].微生物学报,2018,58(12):2147-2160.
[19]谭广秀.具有抑菌活性的桑树内生菌的分离鉴定及其抑菌物质研究[D].镇江:江苏科技大学,2012.
[20]HAN Y,WANG Y,BI J L,et al.Constitutive and induced activities of defense-related enzymes in aphid-resistant and aphid-susceptible cultivars of wheat[J].J Chem Ecol,2009,35(2):176-182.
[21]CHEN F,WANG M,ZHENG Y,et al.Quantitative changes of plant defense enzymes and phytohormone in biocontrol of cucumber Fusarium wilt by Bacillus subtilis B579[J].World J Microbiol Biotechnol,2010,26(4):675-684.
[22]MATARESE F S,SARROCCO S,GRUBER S,et al.Biocontrol of Fusarium head blight:interactions between Trichoderma and mycotoxigenic Fusarium[J].Microbiology,2012,158(1):98-106.
[23]慕立义.植物化学保护研究方法[M].北京:中国农业出版社,1994.
[24]丁婷,江腾,江海洋,等.喜树内生真菌的分离、鉴定以及抗植物病原菌活性的初步研究[J].激光生物学报,2010,19(4):501-506.
[25]杜军凯,余桂红,王秀娥,等.赤霉病主效抗性QTL区域SSCP标记的发掘与验证[J].麦类作物学报,2010,30(5):829-834.
[26]李洪连,徐敬友.农业植物病理学实验实习指导[M].2版.北京:中国农业出版社,2007.
[27]AQUINO-BOLA?OS E N,MERCADO-SILVA E.Effects of polyphenol oxidase and peroxidase activity,phenolics and lignin content on the browning of cut jicama[J].Postharvest Biol Tec,2004,33(3):275-283.
[28]梁军锋,薛泉宏,牛小磊,等.7株放线菌在辣椒根部定殖及对辣椒叶片PAL与PPO活性的影响[J].西北植物学报,2005,25(10):2118-2123.
[29]YOSHIDA S,OHBA A,LIANG Y M,et al.Specificity of Pseudomonas isolates on healthy and Fusarium head blight-infected spikelets of wheat heads[J].Microb Ecol,2012,64(1):214-225.
[30]HE J,BOLAND G J,ZHOU T.Concurrent selection for microbial suppression of Fusarium graminearum,Fusarium head blight and deoxynivalenol in wheat[J].J Appl Microbio,2009,106(6):1805-1817.
[31]毛腾霄.枯草芽孢杆菌BS-8D防治玉米纹枯病(Rhizoctonia solani)的研究[D].雅安:四川农业大学,2006.
[32]GOND S K,BERGEN M S,TORRES M S,et al.Endophytic Bacillus spp.produce antifungal lipopeptides and induce host defence gene expression in maize[J].Microbiol Res,2015,172:79-87.
[33]SHRESTHA B K,KARKI H S,GROTH D E,et al.Biological control activities of rice-associated Bacillus sp.strains against sheath blight and bacterial panicle blight of rice[J].PLoS One,2016,11(1):e0146764.
[34]REN J H,LI H,WANG Y F,et al.Biocontrol potential of an endophytic Bacillus pumilus JK-SX001 against poplar canker[J].Biol Control,2013,67(3):421-430.
[35]王光达,黄初女,吴委林,等.不同玉米品种对大斑病的抗性与相关防御酶活性的关系研究[J].玉米科学,2014,22(5):146-152.
[36]冯远娇,王建武,苏贻娟,等.茉莉酸在玉米地上部与地下部系统性诱导防御反应中的作用[J].中国农业科学,2009,42(8):2726-2735.