黄瓜与西芹间作土壤细菌多样性及其对黄瓜枯萎病发生的影响
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摘要
本试验以黄瓜与西芹间作种植模式为处理,黄瓜单作和西芹单作种植模式为对照,利用Illumina公司Miseq平台对上述不同处理土壤进行16S rDNA细菌群落多样性高通量测序分析和田间接种黄瓜枯萎病菌,探讨黄瓜与西芹间作模式土壤细菌的多样性及其对田间黄瓜枯萎病发生的影响。16S rDNA测序结果表明,黄瓜与西芹间作土壤的细菌物种总数最多,群落多样性水平最高,与对照相比显著提高了土壤细菌observed species指数、Shannon指数和Chao1指数(P<0.05);Beta多样性聚类分析表明,黄瓜与西芹间作土壤的环境群落物种与黄瓜单作和西芹单作有一定差异性。在门分类水平上,共检测到45个菌门,其中变形菌门占明显优势,其次为酸杆菌门和放线菌门等;黄瓜与西芹间作土壤细菌种类所占比例最高,达98.63%。在属水平上,共检测到428类菌属,GP6、GP16、GP4、芽单胞菌属、节细菌属5属的丰度值较大;黄瓜与西芹间作土壤的节细菌属分布比例最高,红游动菌属、鞘氨醇单胞菌属和芽球菌属丰度值较大,为黄瓜与西芹间作土壤细菌明显优势菌属。田间接种黄瓜枯萎病菌试验结果表明,采用上述3种不同种植模式土壤种植黄瓜,在黄瓜苗期接种黄瓜枯萎病菌,黄瓜与西芹间作处理的黄瓜枯萎病的田间发病率较西芹单作和黄瓜单作分别降低57.03%~63.54%和66.95%~72.15%。因此,黄瓜与西芹间作增加了土壤细菌群落多样性,降低了黄瓜枯萎病的发病率,对后茬黄瓜土传病害防控具有一定科学指导意义。
Cucumber wilt is a soil disease that is highly prevalent in the production and cultivation of cucumber. The disease is highly difficult to prevent and cure. A number of studies have shown that intercropping was one of the most effective methods of reducing the occurrence of plant soil diseases. Also celery has been proven to have high allelopathy. To explore the application of allelopathic effects of celery on cucumber fusarium wilt control, we conducted cucumber and celery intercropping experiment. In the experiment, three planting patterns were set, which were cucumber-celery intercropping, celery monocropping and cucumber monocropping. The cucumber-celery intercropping pattern was regarded as treatment group and monocropping patterns of celery and cucumber regarded as the control group. The Miseq platform of Illumina Company was used to analyze 16 S rDNA bacterial community diversity through high-throughput sequencing to discuss the influence of cucumber-celery intercropping on cucumber soil bacterial diversity. Cucumber was planted in soil from different planting patterns and inoculated with Fusarium oxysporium f.sp.cucumerinum(Foc) in the pot experiment to investigate the control effect of cucumber-celery intercropping on cucumber fusarium wilt. The 16 S rDNA sequencing results showed that total bacterial species amount and community diversity were highest in soil under cucumber-celery intercropping, which significantly enhanced observed bacteria species index, Shannon index and Chao1 index(P < 0.05). Beta diversity clustering analysis showed there existed a difference in environmental community species between soil from cucumber-celery intercropping and mono-cropped cucumber or celery. Moreover, 15 bacterial phyla were detected. Proteobacteria, which was followed by Acidobacteria and Actinobacteria, had a distinct advantage(35.7%-38.0%). Then the proportion of bacterial species derived from cucumber-celery intercropped soil was highest(98.63%). About 428 bacterial genera were detected with 5 dominant bacterial genera, which were GP6, GP16, GP4, Gemmatimonas and Arthrobacter. Arthrobacter, Rhodoplanes, Sphingomonas and Blastococcus were dominant bacteria genera in cucumber-celery inter-cropped soil. The 4 genera demonstrated that cucumber and celery intercropping enriched the diversity of bacterial communities compared with monocropped celery or cucumber. The results of fusarium wilt inoculation experiment of cucumber suggested that the control efficiency of cucumber-celery intercropping to cucumber fusarium wilt reached 57.03%-63.54% and 66.95%-72.15% relative to monocroped celery and cucumber, respectively. Therefore cucumber-celery intercropping increased the diversity of bacterial communities, and reduced incidence rate of cucumber fusarium wilt. This was of scientific interest for the prevention and control of soil borne diseases.
Cucumber wilt is a soil disease that is highly prevalent in the production and cultivation of cucumber. The disease is highly difficult to prevent and cure. A number of studies have shown that intercropping was one of the most effective methods of reducing the occurrence of plant soil diseases. Also celery has been proven to have high allelopathy. To explore the application of allelopathic effects of celery on cucumber fusarium wilt control, we conducted cucumber and celery intercropping experiment. In the experiment, three planting patterns were set, which were cucumber-celery intercropping, celery monocropping and cucumber monocropping. The cucumber-celery intercropping pattern was regarded as treatment group and monocropping patterns of celery and cucumber regarded as the control group. The Miseq platform of Illumina Company was used to analyze 16 S rDNA bacterial community diversity through high-throughput sequencing to discuss the influence of cucumber-celery intercropping on cucumber soil bacterial diversity. Cucumber was planted in soil from different planting patterns and inoculated with Fusarium oxysporium f.sp.cucumerinum(Foc) in the pot experiment to investigate the control effect of cucumber-celery intercropping on cucumber fusarium wilt. The 16 S rDNA sequencing results showed that total bacterial species amount and community diversity were highest in soil under cucumber-celery intercropping, which significantly enhanced observed bacteria species index, Shannon index and Chao1 index(P < 0.05). Beta diversity clustering analysis showed there existed a difference in environmental community species between soil from cucumber-celery intercropping and mono-cropped cucumber or celery. Moreover, 15 bacterial phyla were detected. Proteobacteria, which was followed by Acidobacteria and Actinobacteria, had a distinct advantage(35.7%-38.0%). Then the proportion of bacterial species derived from cucumber-celery intercropped soil was highest(98.63%). About 428 bacterial genera were detected with 5 dominant bacterial genera, which were GP6, GP16, GP4, Gemmatimonas and Arthrobacter. Arthrobacter, Rhodoplanes, Sphingomonas and Blastococcus were dominant bacteria genera in cucumber-celery inter-cropped soil. The 4 genera demonstrated that cucumber and celery intercropping enriched the diversity of bacterial communities compared with monocropped celery or cucumber. The results of fusarium wilt inoculation experiment of cucumber suggested that the control efficiency of cucumber-celery intercropping to cucumber fusarium wilt reached 57.03%-63.54% and 66.95%-72.15% relative to monocroped celery and cucumber, respectively. Therefore cucumber-celery intercropping increased the diversity of bacterial communities, and reduced incidence rate of cucumber fusarium wilt. This was of scientific interest for the prevention and control of soil borne diseases.
引文
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[8]GóMEZ-RODRíGUEZ O,ZAVALETA-MEJíA E,GONZá1EZHEMáNDEZ V A,et al.Allelopathy and microclimatic modification of intercropping with marigold on tomato early blight disease development[J].Field Crops Research,2003,83(1):27–34
[9]REN L X,SU S M,YANG X M,et al.Intercropping with aerobic rice suppressed Fusarium wilt in watermelon[J].Soil Biology and Biochemistry,2008,40(3):834–844
[10]董艳,董坤,郑毅,等.不同品种小麦与蚕豆间作对蚕豆枯萎病的防治及其机理[J].应用生态学报,2014,25(7):1979-1987DONG Y,DONG K,ZHENG Y,et al.Faba bean fusarium wilt(Fusarium oxysporum)control and its mechanism in different wheat varieties and faba bean intercropping system[J].Chinese Journal of Applied Ecology,2014,25(7):1979-1987
[11]苏世鸣.西瓜与旱作水稻间作抑制西瓜连作枯萎病及其机理[D].南京:南京农业大学,2007SU S M.Decreasing Fusarium wilt of watermelon in continuing cropping system by intercropping with aerobic rice[D].Nanjing:Nanjing Agricultural University,2007
[12]高晓敏,王琚钢,李杰,等.西芹鲜根丙酮浸提物层析流分对黄瓜枯萎病菌的化感作用以及化感物质鉴定[J].中国生态农业学报,2014,22(11):1364–1371GAO X M,WANG J G,LI J,et al.Allelopathic effects of fresh parsley root acetone extracts on Fusarium oxysporum f.sp.cucumberinum and allelochemicals identification[J].Chinese Journal of Eco-Agriculture,2014,22(11):1364–1371
[13]钱程,云兴福,高晓敏,等.西芹鲜根浸提液作用后黄瓜枯萎病菌弱毒菌株的筛选[J].中国生态农业学报,2013,21(5):606–614QIAN C,YUN X F,GAO X M,et al.Screening of virulenceattenuated strains for Fusarium oxysporum f.sp.cucumerinum after treatment with parsley fresh root extract[J].Chinese Journal of Eco-Agriculture,2013,21(5):606–614
[14]包妍妍,云兴福,张东东,等.西芹根物质浸提液处理后黄瓜叶片内几种氮代谢物质含量的变化[J].中国生态农业学报,2012,20(11):1508–1513BAO Y Y,YUN X F,ZHANG D D,et al.Nitrogen metabolites in cucumber leaves treated with parsley root extracts[J].Chinese Journal of Eco-Agriculture,2012,20(11):1508–1513
[15]MACDONALD C A,THOMAS N,ROBINSON L,et al.Physiological,biochemical and molecular responses of the soil microbial community after afforestation of pastures with Pinus radiata[J].Soil Biology Biochemistry,2009,41(8):1642–1651
[16]WHITNEY D,ROSSMAN A,HAYDEN N.Evaluating an existing subsurface flow constructed wetland in Akumal,Mexico[J].Ecological Engineering,2003,20(1):105–111
[17]王玉彦,吴凤芝,周新刚.不同间作模式对设施黄瓜生长及土壤环境的影响[J].中国蔬菜,2009,(16):8–13WANG Y Y,WU F Z,ZHOU X G.Effects of different intercropping patterns on the growth of cucumber in greenhouse and soil environment[J].China Vegetables,2009,(16):8–13
[18]吴凤芝,周新刚.不同作物间作对黄瓜病害及土壤微生物群落多样性的影响[J].土壤学报,2009,46(5):899–906WU F Z,ZHOU X G.Effect of intercropping of cucumber with different crops on cucumber diseases and soil microbial community diversity[J].Acta Pedologica Sinica,2009,46(5):899–906
[19]高晓敏,王琚钢,云兴福,等.西芹醇层物对黄瓜枯萎病菌的化感作用及化感物质鉴定[J].中国生物防治学报,2017,33(2):281–288GAO X M,WANG J G,YUN X F,et al.Allelopathic effects of fresh parsley root ethanol extracts on Fusarium oxysporum f.sp.cucumberinum and allelochemicals identification[J].Chinese Journal of Biological Control,2017,33(2):281–288
[20]焦中志,李相昆,张立成,等.反硝化聚磷菌菌种筛分与除磷特性分析[J].沈阳建筑大学学报:自然科学版,2009,25(3):535–540JIAO Z Z,LI X K,ZHANG L C,et al.Isolation and characteristic of denitrifying phosphorus accumulating organism[J].Journal of Shenyang Architecture University:Natural Science,2009,25(3):535–540
[21]单娜娜,赖波.风沙土成土演变过程中土壤微生物生物学特性研究进展与展望[J].新疆环境保护,2004,26(S1):79–82SHAN N N,LAI B.Study progresses and prospect on the ecological characteristics of soil-inhabiting microorganism in soil forming process of Aeolian sand soil[J].Environmental Protection of Xinjiang,2004,26(S1):79–82
[22]孙红敏,余利岩,张玉琴.地嗜皮菌科放线菌的研究进展[J].微生物学报,2015,55(12):1521–1529SUN H M,YU L Y,ZHANG Y Q.Recent advance in Geodermatophilaceae—A review[J].Acta Microbiologica Sinica,2015,55(12):1521–1529
[23]GARBEVA P,VAN VEEN J A,VAN ELSAS J D.Microbial diversity in soil:Selection of microbial populations by plant and soil type and implications for disease suppressiveness[J].Annual Review of Phytopathology,2004,42:243–270
[24]GORISSEN A,VAN OVERBEEK L S,VAN ELSAS J D.Pig slurry reduces the survival of Ralstonia solanacearum biovar2 in soil[J].Canadian Journal of Microbiology,2004,50(8):587–593
[25]BENIZRI E,PIUTTI S,VERGER S,et al.Replant diseases:bacterial community structure and diversity in peach rhizosphere as determined by metabolic and genetic fingerprinting[J].Soil Biology and Biochemistry,2005,37(9):1738–1746
[26]邓晓,李勤奋,侯宪文,等.香蕉枯萎病不同感病级别植株根际与非根际土壤微生物物种多样性研究[J].中国农学通报,2012,28(30):239–248DENG X,LI Q F,HOU X W,et al.Study on the microbial species diversity of rhizosphere and non-rhizosphere soils from different grades infected by banana fusarium wilt[J].Chinese Agricultural Science Bulletin,2012,28(30):239–248
[27]GARBEVA P,VAN VEEN J A,VAN ELSAS J D.Assessment of the diversity,and antagonism towards Rhizoctonia solani AG3,of Pseudomonas species in soil from different agricultural regimes[J].FEMS Microbiology Ecology,2004,47(1):51–64
[2]苏阿德,谢关林,李斌,等.芽孢杆菌在促进番茄生长和控制青枯病上的比较优势[J].浙江大学学报:农业与生命科学版,2004,30(6):603–610SU A D,XIE G L,LI B,et al.Comparative performance of Bacillus spp.in growth promotion and suppression of tomato bacterial wilt caused by Ralstonia solanacearum[J].Journal of Zhejiang University:Agriculture and Life Sciences,2004,30(6):603–610
[3]高芬,马利平,乔雄梧,等.蜡质芽孢杆菌BC98-Ⅰ发酵液与抑菌粗提物对黄瓜枯萎病菌的抑菌特性研究[J].中国生态农业学报,2006,14(1):189–192GAO F,MA L P,QIAO X W,et al.The antimicrobial characters of the fermenting liquor of BC98-Ⅰ(B.cereus)and its antifungal crude extraction on Fusarium oxysporum(Schl.)f.sp.cucumerinum Owen[J].Chinese Journal of Eco-Agriculture,2006,14(1):189–192
[4]徐美娜,王光华,靳学慧.土传病害生物防治研究进展[J].吉林农业科学,2005,30(2):39–42XU M N,WANG G H,JIN X H.Progress in studies on biological control of soil mediated diseases[J].Journal of Jilin Agricultural Sciences,2005,30(2):39–42
[5]吴敏娜,张惠文,李新宇,等.土壤抑真菌作用与细菌群落结构的关系[J].应用生态学报,2008,19(7):1574–1578WU M N,ZHANG H W,LI X Y,et al.Relationship between fungistasis of soil and bacterial community structure[J].Chinese Journal of Applied Ecology,2008,19(7):1574–1578
[6]YANG C H,CROWLEY D E,MENGE J A.16S r DNA fingerprinting of rhizosphere bacterial communities associated with healthy and phytophthora infected avocado roots[J].FEMS Microbiology Ecology,2001,35(2):129–136
[7]ZHU Y Y,CHEN H R,FAN J H,et al.Genetic diversity and disease control in rice[J].Nature,2000,406(6797):718–722
[8]GóMEZ-RODRíGUEZ O,ZAVALETA-MEJíA E,GONZá1EZHEMáNDEZ V A,et al.Allelopathy and microclimatic modification of intercropping with marigold on tomato early blight disease development[J].Field Crops Research,2003,83(1):27–34
[9]REN L X,SU S M,YANG X M,et al.Intercropping with aerobic rice suppressed Fusarium wilt in watermelon[J].Soil Biology and Biochemistry,2008,40(3):834–844
[10]董艳,董坤,郑毅,等.不同品种小麦与蚕豆间作对蚕豆枯萎病的防治及其机理[J].应用生态学报,2014,25(7):1979-1987DONG Y,DONG K,ZHENG Y,et al.Faba bean fusarium wilt(Fusarium oxysporum)control and its mechanism in different wheat varieties and faba bean intercropping system[J].Chinese Journal of Applied Ecology,2014,25(7):1979-1987
[11]苏世鸣.西瓜与旱作水稻间作抑制西瓜连作枯萎病及其机理[D].南京:南京农业大学,2007SU S M.Decreasing Fusarium wilt of watermelon in continuing cropping system by intercropping with aerobic rice[D].Nanjing:Nanjing Agricultural University,2007
[12]高晓敏,王琚钢,李杰,等.西芹鲜根丙酮浸提物层析流分对黄瓜枯萎病菌的化感作用以及化感物质鉴定[J].中国生态农业学报,2014,22(11):1364–1371GAO X M,WANG J G,LI J,et al.Allelopathic effects of fresh parsley root acetone extracts on Fusarium oxysporum f.sp.cucumberinum and allelochemicals identification[J].Chinese Journal of Eco-Agriculture,2014,22(11):1364–1371
[13]钱程,云兴福,高晓敏,等.西芹鲜根浸提液作用后黄瓜枯萎病菌弱毒菌株的筛选[J].中国生态农业学报,2013,21(5):606–614QIAN C,YUN X F,GAO X M,et al.Screening of virulenceattenuated strains for Fusarium oxysporum f.sp.cucumerinum after treatment with parsley fresh root extract[J].Chinese Journal of Eco-Agriculture,2013,21(5):606–614
[14]包妍妍,云兴福,张东东,等.西芹根物质浸提液处理后黄瓜叶片内几种氮代谢物质含量的变化[J].中国生态农业学报,2012,20(11):1508–1513BAO Y Y,YUN X F,ZHANG D D,et al.Nitrogen metabolites in cucumber leaves treated with parsley root extracts[J].Chinese Journal of Eco-Agriculture,2012,20(11):1508–1513
[15]MACDONALD C A,THOMAS N,ROBINSON L,et al.Physiological,biochemical and molecular responses of the soil microbial community after afforestation of pastures with Pinus radiata[J].Soil Biology Biochemistry,2009,41(8):1642–1651
[16]WHITNEY D,ROSSMAN A,HAYDEN N.Evaluating an existing subsurface flow constructed wetland in Akumal,Mexico[J].Ecological Engineering,2003,20(1):105–111
[17]王玉彦,吴凤芝,周新刚.不同间作模式对设施黄瓜生长及土壤环境的影响[J].中国蔬菜,2009,(16):8–13WANG Y Y,WU F Z,ZHOU X G.Effects of different intercropping patterns on the growth of cucumber in greenhouse and soil environment[J].China Vegetables,2009,(16):8–13
[18]吴凤芝,周新刚.不同作物间作对黄瓜病害及土壤微生物群落多样性的影响[J].土壤学报,2009,46(5):899–906WU F Z,ZHOU X G.Effect of intercropping of cucumber with different crops on cucumber diseases and soil microbial community diversity[J].Acta Pedologica Sinica,2009,46(5):899–906
[19]高晓敏,王琚钢,云兴福,等.西芹醇层物对黄瓜枯萎病菌的化感作用及化感物质鉴定[J].中国生物防治学报,2017,33(2):281–288GAO X M,WANG J G,YUN X F,et al.Allelopathic effects of fresh parsley root ethanol extracts on Fusarium oxysporum f.sp.cucumberinum and allelochemicals identification[J].Chinese Journal of Biological Control,2017,33(2):281–288
[20]焦中志,李相昆,张立成,等.反硝化聚磷菌菌种筛分与除磷特性分析[J].沈阳建筑大学学报:自然科学版,2009,25(3):535–540JIAO Z Z,LI X K,ZHANG L C,et al.Isolation and characteristic of denitrifying phosphorus accumulating organism[J].Journal of Shenyang Architecture University:Natural Science,2009,25(3):535–540
[21]单娜娜,赖波.风沙土成土演变过程中土壤微生物生物学特性研究进展与展望[J].新疆环境保护,2004,26(S1):79–82SHAN N N,LAI B.Study progresses and prospect on the ecological characteristics of soil-inhabiting microorganism in soil forming process of Aeolian sand soil[J].Environmental Protection of Xinjiang,2004,26(S1):79–82
[22]孙红敏,余利岩,张玉琴.地嗜皮菌科放线菌的研究进展[J].微生物学报,2015,55(12):1521–1529SUN H M,YU L Y,ZHANG Y Q.Recent advance in Geodermatophilaceae—A review[J].Acta Microbiologica Sinica,2015,55(12):1521–1529
[23]GARBEVA P,VAN VEEN J A,VAN ELSAS J D.Microbial diversity in soil:Selection of microbial populations by plant and soil type and implications for disease suppressiveness[J].Annual Review of Phytopathology,2004,42:243–270
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