一株小麦赤霉病拮抗菌的筛选鉴定及防治效果分析
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摘要
自小麦赤霉病发病田块土壤中,分离得到1株小麦赤霉病高效拮抗菌株。通过形态特征、生理生化特性和16S rDNA序列比对分析,将这株拮抗菌株鉴定为奇异变形杆菌(proteus mirabilis)。P. mirabilis DY05发酵液和无细胞上清均可显著抑制禾谷镰刀菌菌丝体生长(抑制率分别为79.50%和51.25%)和分生孢子萌发(抑制率均为100%),减少呕吐毒素产生(分别减少84.32%和82.82%)。田间赤霉病防治试验结果显示,接种DY05,可降低发病率52.13%,同时病情指数降低48.74%,显示出较好的防治效果。促生生理活性评价试验结果显示,菌株DY05可以产生铁载体和IAA,并具有溶磷作用和ACC脱氨酶活性,具有很好的促生长潜力。盆栽试验结果表明,菌株DY05对小麦植株生长具有显著的促进作用。与对照组相比,菌株DY05处理可以显著增加小麦幼苗茎高、根长、鲜重和干重,其中茎高、根长、鲜重和干重分别提高了22.21%、26.41%、44.77%和26.53%。分离得到的拮抗菌株DY05具有拮抗病原真菌和促进植物生长的双重功能,为开发禾谷镰刀菌生物防治制剂提供了菌种材料。
One bacterial strain against Fusarium graminearum was isolated from a cultivated soil and identified as Proteus mirabilis basing on morphologic observation, biochemical characteristic analysis and 16 S rDNA sequences alignment. Both the fermentation broth and the cell-free supernatant of P. mirabilis DY05 could significantly inhibit the hyphal growth of F.graminearum(with the inhibition rate 79.50% and 51.25% respectively) and conidiospore germination of F. graminearum(both inhibition rates were 100%), as well as reduce DON production in wheat(decreased by 84.32% and 82.82%respectively). Field assays showed a significant reduction of Fusarium head blight(FHB) diseases on wheat when DY05 suspension was sprayed on wheat spikes after inoculation with F. graminearum(incidence rate decreased by 52.13%, while the disease index decreased by 48.74%). The modified CAS assay, the Salkowski colorimetry, molybdenum blue method and ACC deaminase assay were used to measure production of siderophores, HCN, IAA, phosphate solubilizing capacity and ACC deaminase in the strain DY05. The results indicated that strain DY05 could not only produce siderophores, IAA and ACC deaminase, but also solubilize phosphate. Moreover, in pot plant experiments, the strain DY05 showed significant growth promoting effects on the stem height, root length, fresh weight and dry weight of wheat seedlings(increased by22.21%, 26.41%, 44.77% and 26.53% respectively). Overall, the antagonistic strain DY05 was found to antagonize F.graminearum and showed a high plant growth promotion, and were therefore could offer available resource for development of new biocontrol agents.
One bacterial strain against Fusarium graminearum was isolated from a cultivated soil and identified as Proteus mirabilis basing on morphologic observation, biochemical characteristic analysis and 16 S rDNA sequences alignment. Both the fermentation broth and the cell-free supernatant of P. mirabilis DY05 could significantly inhibit the hyphal growth of F.graminearum(with the inhibition rate 79.50% and 51.25% respectively) and conidiospore germination of F. graminearum(both inhibition rates were 100%), as well as reduce DON production in wheat(decreased by 84.32% and 82.82%respectively). Field assays showed a significant reduction of Fusarium head blight(FHB) diseases on wheat when DY05 suspension was sprayed on wheat spikes after inoculation with F. graminearum(incidence rate decreased by 52.13%, while the disease index decreased by 48.74%). The modified CAS assay, the Salkowski colorimetry, molybdenum blue method and ACC deaminase assay were used to measure production of siderophores, HCN, IAA, phosphate solubilizing capacity and ACC deaminase in the strain DY05. The results indicated that strain DY05 could not only produce siderophores, IAA and ACC deaminase, but also solubilize phosphate. Moreover, in pot plant experiments, the strain DY05 showed significant growth promoting effects on the stem height, root length, fresh weight and dry weight of wheat seedlings(increased by22.21%, 26.41%, 44.77% and 26.53% respectively). Overall, the antagonistic strain DY05 was found to antagonize F.graminearum and showed a high plant growth promotion, and were therefore could offer available resource for development of new biocontrol agents.
引文
[1]李正辉,向晶晶,陈婧鸿,等.小麦赤霉病拮抗菌的分离与鉴定[J].麦类作物学报,2007,27(1):149-152.
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[4]陆道训,秦华宝,赵立军,等.70%甲基硫菌灵WP防治小麦赤霉病的药效研究[J].安徽农业科学,2005,33(11):2014-2014.
[5]姬勇,陈太虎,付佑胜.30%多酮可湿性粉剂防治小麦赤霉病及白粉病田间试验[J].安徽农学通报,2008,14(7):168-169.
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[10]杨慧勇,李飞凤,陆琼娴,等.拮抗菌株AFR0406对小麦赤霉病菌和纹枯病菌的生物活性测定[J].江苏农业科学,2006(6):142-144.
[11]管章玲,辛海峰,李建宏,等.小麦赤霉病拮抗菌的筛选及应用[J].江苏农业学报,2012,28(2):309-313.
[12]东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社,2001.
[13]YOON S H,HA S M,KWON S,et al.Introducing Ez Bio Cloud:a taxonomically united database of 16S r RNAgene sequences and whole-genome assemblies[J].International Journal of Systematic and Evolutionary Microbiology,2017,67(5):1613-1617.
[14]KUMAR S,STECHER G,TAMURA K.MEGA7:Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets[J].Molecular Biology and Evolution,2016,33(7),1870-1874.
[15]王瑶,赵月菊,邢福国,等.禾谷镰刀菌拮抗菌株的筛选及鉴定[J].核农学报,2017,31(6):1128-1136.
[16]RODRIGUES E P,RODRIGUES L S,OLIVEIRA A L MD,et al.Azospirillum amazonense inoculation:effects on growth,yield and N2 fixation of rice(Oryza sativa L.)[J].Plant&Soil,2009,316(1/2):323-323.
[17]王平,董飚,李阜棣,等.小麦根圈细菌铁载体的检测[J].微生物学通报,1994,21(6):323-326.
[18]GLICKMANN E,DESSAUX Y.A Critical Examination of the Specificity of the Salkowski Reagent for Indolic Compounds Produced by Phytopathogenic Bacteria[J].Applied and Environmental Microbiology,1995,61(2):793.
[19]周仲强.植物根际促生细菌功能菌株的筛选[D].长春:吉林大学,2007:19-20
[20]宗英,赵月菊,刘阳,等.一株贝莱斯芽孢杆菌抑制禾谷镰刀菌的研究[J].核农学报,2018,32(2):310-317.
[21]周丽娜,王莉莉,张永娜,等.2株放线菌的抗菌活性及分类学地位[J].中国农学通报,2015,31(11):182-189.
[22]刘伟成,潘洪玉,席景会,等.小麦赤霉病拮抗性芽孢杆菌生防作用的研究[J].麦类作学报,2005,25(4):95-100.
[23]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.
[24]蔺国强,廖玉才,宫安东,等.禾谷镰刀菌拮抗菌的筛选与鉴定[J].华中农业大学学报:自然科学版,2013,32(3):28-32.
[25]黄婷婷.多菌灵、氰烯菌酯对禾谷镰孢菌DON毒素合成的影响[D].南京:南京农业大学,2012:21-23.
[26]孟丽媛,李冰,孙乐妮,等.水稻根际促生细菌促生生理活性评价[J].中国土壤与肥料,2014(1):90-94.
[2]张雪娇,石晶晶,常娜,等.河北省冬小麦赤霉病拮抗细菌的分离与鉴定[J].作物杂志,2017(2):157-162.
[3]张洁,伊艳杰,王金水,等.小麦赤霉病的防治技术研究进展[J].中国植保导刊,2014,34(1):24-28.
[4]陆道训,秦华宝,赵立军,等.70%甲基硫菌灵WP防治小麦赤霉病的药效研究[J].安徽农业科学,2005,33(11):2014-2014.
[5]姬勇,陈太虎,付佑胜.30%多酮可湿性粉剂防治小麦赤霉病及白粉病田间试验[J].安徽农学通报,2008,14(7):168-169.
[6]叶华智,余桂容,严吉明,等.小麦赤霉病的生物防治研究[J].拮抗芽孢杆菌B4、B6菌株的防病机制.四川农业大学学报,2003,21(1):18-22.
[7]DUNLAP C A,SCHISLER D A,PRICE N P,et al.Cyclic lipopeptide profile of three Bacillus subtilis strains:antagonists of Fusarium head blight[J].Journal of Microbiology,2011,49(4):603-609.
[8]CRANE J M,GIBSON D M,VAUGHAN R H,et al.Iturin levels on wheat spikes linked to biological control of Fusarium head blight by Bacillus amyloliquefaciens[J].Phytopathology,2013,103(2):146-155.
[9]陈亮,李瑞静,秦素雅,等.小麦赤霉病拮抗菌株筛选及其抑制作用研究[J].中国植保导刊,2017,37(5):12-17.
[10]杨慧勇,李飞凤,陆琼娴,等.拮抗菌株AFR0406对小麦赤霉病菌和纹枯病菌的生物活性测定[J].江苏农业科学,2006(6):142-144.
[11]管章玲,辛海峰,李建宏,等.小麦赤霉病拮抗菌的筛选及应用[J].江苏农业学报,2012,28(2):309-313.
[12]东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社,2001.
[13]YOON S H,HA S M,KWON S,et al.Introducing Ez Bio Cloud:a taxonomically united database of 16S r RNAgene sequences and whole-genome assemblies[J].International Journal of Systematic and Evolutionary Microbiology,2017,67(5):1613-1617.
[14]KUMAR S,STECHER G,TAMURA K.MEGA7:Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets[J].Molecular Biology and Evolution,2016,33(7),1870-1874.
[15]王瑶,赵月菊,邢福国,等.禾谷镰刀菌拮抗菌株的筛选及鉴定[J].核农学报,2017,31(6):1128-1136.
[16]RODRIGUES E P,RODRIGUES L S,OLIVEIRA A L MD,et al.Azospirillum amazonense inoculation:effects on growth,yield and N2 fixation of rice(Oryza sativa L.)[J].Plant&Soil,2009,316(1/2):323-323.
[17]王平,董飚,李阜棣,等.小麦根圈细菌铁载体的检测[J].微生物学通报,1994,21(6):323-326.
[18]GLICKMANN E,DESSAUX Y.A Critical Examination of the Specificity of the Salkowski Reagent for Indolic Compounds Produced by Phytopathogenic Bacteria[J].Applied and Environmental Microbiology,1995,61(2):793.
[19]周仲强.植物根际促生细菌功能菌株的筛选[D].长春:吉林大学,2007:19-20
[20]宗英,赵月菊,刘阳,等.一株贝莱斯芽孢杆菌抑制禾谷镰刀菌的研究[J].核农学报,2018,32(2):310-317.
[21]周丽娜,王莉莉,张永娜,等.2株放线菌的抗菌活性及分类学地位[J].中国农学通报,2015,31(11):182-189.
[22]刘伟成,潘洪玉,席景会,等.小麦赤霉病拮抗性芽孢杆菌生防作用的研究[J].麦类作学报,2005,25(4):95-100.
[23]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.
[24]蔺国强,廖玉才,宫安东,等.禾谷镰刀菌拮抗菌的筛选与鉴定[J].华中农业大学学报:自然科学版,2013,32(3):28-32.
[25]黄婷婷.多菌灵、氰烯菌酯对禾谷镰孢菌DON毒素合成的影响[D].南京:南京农业大学,2012:21-23.
[26]孟丽媛,李冰,孙乐妮,等.水稻根际促生细菌促生生理活性评价[J].中国土壤与肥料,2014(1):90-94.