基于FUNGuild的镰刀菌根腐病发病烟株根际真菌群落研究
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摘要
为了解镰刀菌根腐病发病烟株与未发病烟株在根际真菌群落结构与功能上的差异,联合应用高通量测序和FUNGuild技术研究了镰刀菌根腐病发病烟株与未发病烟株根际土壤中的真菌群落,结果表明:(1)镰刀菌根腐病发病烟株根际真菌群落的Shannon指数为4.14,而未发病烟株则高达4.71,未发病烟株根际真菌群落多样性显著高于发病烟株,镰刀菌根腐病发病烟株与未发病烟株根际土壤中的真菌群落在组成上有显著差异;(2)镰刀菌根腐病发病烟株根际真菌以病理营养型为主,占54.64%,而未发病烟株根际真菌则以腐生营养型为主,占46.10%;(3)镰刀菌根腐病发病烟株根际的病原菌相对丰度为30.73%,显著高于未发病烟株的1.12%,镰刀菌在病原菌群落中占绝对优势地位。高通量测序与FUNGuild联用可对土传病害感病烟株的根际优势病原真菌群落进行准确鉴定,可为烟草病害的防控提供科学参考。
In this study, high-through sequencing and FUNGuild were jointly used to investigate the difference in structure and functions of rhizosphere fungi communities between unaffected and Fusarium root rot diseased tobaccos. The results showed: 1) the structure of rhizosphere fungi communities between unaffected and Fusarium root rot diseased tobacco was significantly different, and the alpha diversity of rhizosphere fungi communities of unaffected tobacco was significantly higher than that of diseased tobacco; 2) pathotroph predominated the rhizosphere fungi communities of Fusarium root rot diseased tobacco(54.64%) while saprotroph dominated the rhizosphere fungi communities of unaffected tobacco(46.10%); 3) The relative abundance of rhizospheric pathogen was 30.73% in rhizosphere fungi communities of Fusarium root rot diseased tobacco, which was significantly higher than that of unaffected tobacco(1.12%). In addition, Fusarium species was dominant in the pathogen communities. The co-application of high-through sequencing and FUNGuild is useful in making a comprehensive identification of rhizogenic dominated pathogenic fungi communities of soil-borne diseased tobacco, which provides a scientific reference for tobacco disease control.
In this study, high-through sequencing and FUNGuild were jointly used to investigate the difference in structure and functions of rhizosphere fungi communities between unaffected and Fusarium root rot diseased tobaccos. The results showed: 1) the structure of rhizosphere fungi communities between unaffected and Fusarium root rot diseased tobacco was significantly different, and the alpha diversity of rhizosphere fungi communities of unaffected tobacco was significantly higher than that of diseased tobacco; 2) pathotroph predominated the rhizosphere fungi communities of Fusarium root rot diseased tobacco(54.64%) while saprotroph dominated the rhizosphere fungi communities of unaffected tobacco(46.10%); 3) The relative abundance of rhizospheric pathogen was 30.73% in rhizosphere fungi communities of Fusarium root rot diseased tobacco, which was significantly higher than that of unaffected tobacco(1.12%). In addition, Fusarium species was dominant in the pathogen communities. The co-application of high-through sequencing and FUNGuild is useful in making a comprehensive identification of rhizogenic dominated pathogenic fungi communities of soil-borne diseased tobacco, which provides a scientific reference for tobacco disease control.
引文
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[5]张笑宇,段宏群,芦阿虔,等.健康烟田与易感黑胫病烟田烟株不同生长时期根际土壤微生物区系变化规律[J].河南农业科学,2018, 47(3):63-69.ZHANG Xiaoyu, DUAN Hongqun, LU Aqian, et al. Variation of R hizosphere Soil Microbial Flora in Healthy and Black ShankSusceptible Tobacco Fields at Different Growth Stages[J]. Journal of Henan Agricultural Sciences, 2018, 47(3):63-69.
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[7]万川,蒋珍茂,赵秀兰,等.深翻和施用土壤改良剂对烟草青枯病发生的影响[J].烟草科技, 2015(2):11-26.WAN Chuan, JIANG Zhenmao, ZHAO Xiulan, et al. Effects of Deep-ploughing and Soil Amendment Application on Incidence of obacco Bacterial Wilt[J]. Tobacco Science&Technology, 2015(2):11-26.
[8]夏艳,徐茜,董瑜,等.烟草青枯病菌拮抗菌的筛选、鉴定及生防特性研究[J].中国生态农业学报, 2014, 22(2):201-207.XIA Yan, XU Qian, DONG Yu, et al. Screening, identification and characterization of antagonistic bacteria against Ralstonia solanacearum[J]. Chinese Journal of Eco-Agriculture, 2014, 22(2):201-207.
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[10]王婷婷,王仁刚,李咏梅,等.贵州烟草根围AM真菌多样性的初步研究[J].菌物学报, 2014, 33(1):143-151.WANG Tingting, WANG Rengang, LI Yongmei, et al. A primary study on diversity of arbuscular mycorrhizal fungi associated with tobacco roots from Guizhou Province[J]. Mycosystema, 2014,33(1):143-151.
[11]王娜,吕国忠,孙晓东,等.烟草根际土壤真菌多样性的研究[J].菌物学报, 2012, 31(6):827-836.WANGNa,LVGuozhong,SUNXiaodong,etal.Fungal diversity in tobacco rhizosphere soil[J]. Mycosystema, 2012,31(6):827-836.
[12]FUNGuild:An open annotation tool for parsing fungal community datasets by ecological guild[J]. Fungal Ecology, 2016, 20(1):241-248.
[13]Leff J W, Lynch R C, Kane N C, et al. Plant domestication and the assembly of bacterial and fungal communities associated with strains of the common sunflower, Helianthus annuus[J]. New Phytologist, 2016, 214(1).
[14]Kola?íkováZ, Kohout P, Krüger C, et al. Root-associated fungal communities along a primary succession on a mine spoil:Distinct ecological guilds assemble differently[J]. Soil Biology&Biochemistry, 2017, 113:143-152.
[15]Gardes M, Bruns TD. ITS primers with enhanced specificity for basidiomycetes--application to the identification of mycorrhizae and rusts. Molecular Ecology 1993; 2:113-118.
[16]White T, Bruns T, Lee S, Taylor FJRM, White T, Lee SH, et al.Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR protocols:a guide to methods and applications. 1990.
[17]Li X, Zhang Y, Ding C, et al. Declined soil suppressiveness to Fusarium oxysporum, by rhizosphere microflora of cotton in soil sickness[J]. Biology&Fertility of Soils, 2015, 51(8):935-946.
[18]Mago T, Salzberg S L. FLASH:fast length adjustment of short reads to improve genome assemblies[J]. Bioinformatics, 2011,27(21):2957-2963.
[19]Bolger A M, Lohse M, Usadel B. Trimmomatic:a flexible trimmer for Illumina sequence data[J]. Bioinformatics, 2014, 30(15):2114-2120.
[20]Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics 2010; 26:2460.
[21]Koljalg U, Larsson K K, Nilsson R H, et al. UNITE:a database providing web‐based methods for the molecular identification of ectomycorrhizal fungi[J]. New Phytologist, 2005, 166(3):1063–1068.Nguyen N H, Song Z, Bates S T, et al.
[22]Berendsen R L, Pieterse C M J, Bakker P A H M. The rhizosphere microbiome and plant health[J]. Trends in Plant Science, 2012,17(8):478-86.
[23]李小龙,李红丽,曾强,等.钙镁磷肥对青枯病发病烟株根际土壤微生物区系的影响[J].中国烟草学报, 2016, 22(1):71-75.LI Xiaolong, LI Hongli, ZENG Qiang, et al. Effects of calciummagnesium phosphates on microbial flora in rhizosphere soil of tobacco plant effected with Ralstonia solanacearum[J]. Acta Tabacaria Sinica, 2016, 22(1):71-75.
[24]LiXG,DingCF,Zhang TL, WangXX.Fungalpathogen accumulation at the expense of plant-beneficial fungi as a consequence of consecutive peanut monoculturing. Soil Biology&Biochemistry 2014; 72:11-18.
[25]Tedersoo L, Bahram M, P?lme S, et al. Global diversity and geography of soil fungi[J]. Science, 2014, 346(6213):1-11.
[26]桑维钧,祝明亮,吴兴禄,等.烟草镰刀菌根腐病研究初报[J].山地农业生物学报, 1998, 42(3):140-141.SANG Weijun, ZHU Mingliang, WU Xinglu, et al. A Preliminary report of tobacco root rot caused by Fusarium sp.[J]. Journal of Mountain Agriculture and Biology, 1998, 42(3):140-141.
[27]吴安忠,程崖芝,巫升鑫,等.烟草镰刀菌根腐病的病原鉴定[J].中国烟草学报, 2018, 24(2):135-140.WU Anzhong, CHENG Yazhi, WU Shengxin, et al. Identification of tobacco Fusarium root rot pathogen[J]. Acta Tabacaria Sinica,2018, 24(2):135-140.
[2]何永宏,曾乙心,刘林,等.温度和品种抗性对烟草青枯病潜育期的影响[J].烟草科技, 2017, 50(6):16-32HE Yonghong, ZENG Yixin, LIU Lin, et al. Effects of temperature and varietal resistance on latent period of tobacco bacterial wilt[J].Tobacco Science&Technology, 2017, 50(6):16-32
[3]潘建菁,纪成灿,刘冬霞,等.青枯菌侵染后烟株体内过氧化物酶活性的变化及其与抗病性的关系[J].中国烟草科学, 2004,25(3):28-30.PANG Jianjing, JI Chengcan, LIU Dongxia, et al. Changes in activities of peroxidase in flue-cured tobacco infected with Ralstonia solanacearum E.F.Smith and their relations to the resistance[J]. Chinese Tobacco Science, 2004, 25(3):28-30.
[4]高正良,周本国,雷艳丽.黑胫病菌在烟草品种间的侵染差异及I-S关系研究[J].中国烟草学报, 2004, 10(4):31-35.GAO Zhengliang, ZHOU Benguo, LEI Yanli. Study on the Infection difference of tobacco variety to Phytophthora nicotianae andtherelationshipbetweenincidenceandseverity[J]. Acta Tabacaria Sinica, 2004, 10(4):31-35.
[5]张笑宇,段宏群,芦阿虔,等.健康烟田与易感黑胫病烟田烟株不同生长时期根际土壤微生物区系变化规律[J].河南农业科学,2018, 47(3):63-69.ZHANG Xiaoyu, DUAN Hongqun, LU Aqian, et al. Variation of R hizosphere Soil Microbial Flora in Healthy and Black ShankSusceptible Tobacco Fields at Different Growth Stages[J]. Journal of Henan Agricultural Sciences, 2018, 47(3):63-69.
[6]李远远.连作烟田土壤微生物多样性及微生物制剂应用研究[D].郑州大学, 2017.LI Yuanyuan. Microbal Diversity in Continuous Cropped Tobacco Field and Application of Microbial Agents[D]. Zhengzhou University, 2017.
[7]万川,蒋珍茂,赵秀兰,等.深翻和施用土壤改良剂对烟草青枯病发生的影响[J].烟草科技, 2015(2):11-26.WAN Chuan, JIANG Zhenmao, ZHAO Xiulan, et al. Effects of Deep-ploughing and Soil Amendment Application on Incidence of obacco Bacterial Wilt[J]. Tobacco Science&Technology, 2015(2):11-26.
[8]夏艳,徐茜,董瑜,等.烟草青枯病菌拮抗菌的筛选、鉴定及生防特性研究[J].中国生态农业学报, 2014, 22(2):201-207.XIA Yan, XU Qian, DONG Yu, et al. Screening, identification and characterization of antagonistic bacteria against Ralstonia solanacearum[J]. Chinese Journal of Eco-Agriculture, 2014, 22(2):201-207.
[9]王晶晶,蒋士君,常淑娴,等.两株生防菌对烟草黑胫病的抑制活性及其鉴定[J].中国烟草学报, 2011, 17(6):89-93.WANG Jingjing, JIANG Shijun, CHANG Shuxian, et al. Antifungal activity of two biocontrol strains against tobacco black shank and their identification[J]. Acta Tabacaria Sinica, 2011, 17(6):89-93.
[10]王婷婷,王仁刚,李咏梅,等.贵州烟草根围AM真菌多样性的初步研究[J].菌物学报, 2014, 33(1):143-151.WANG Tingting, WANG Rengang, LI Yongmei, et al. A primary study on diversity of arbuscular mycorrhizal fungi associated with tobacco roots from Guizhou Province[J]. Mycosystema, 2014,33(1):143-151.
[11]王娜,吕国忠,孙晓东,等.烟草根际土壤真菌多样性的研究[J].菌物学报, 2012, 31(6):827-836.WANGNa,LVGuozhong,SUNXiaodong,etal.Fungal diversity in tobacco rhizosphere soil[J]. Mycosystema, 2012,31(6):827-836.
[12]FUNGuild:An open annotation tool for parsing fungal community datasets by ecological guild[J]. Fungal Ecology, 2016, 20(1):241-248.
[13]Leff J W, Lynch R C, Kane N C, et al. Plant domestication and the assembly of bacterial and fungal communities associated with strains of the common sunflower, Helianthus annuus[J]. New Phytologist, 2016, 214(1).
[14]Kola?íkováZ, Kohout P, Krüger C, et al. Root-associated fungal communities along a primary succession on a mine spoil:Distinct ecological guilds assemble differently[J]. Soil Biology&Biochemistry, 2017, 113:143-152.
[15]Gardes M, Bruns TD. ITS primers with enhanced specificity for basidiomycetes--application to the identification of mycorrhizae and rusts. Molecular Ecology 1993; 2:113-118.
[16]White T, Bruns T, Lee S, Taylor FJRM, White T, Lee SH, et al.Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. PCR protocols:a guide to methods and applications. 1990.
[17]Li X, Zhang Y, Ding C, et al. Declined soil suppressiveness to Fusarium oxysporum, by rhizosphere microflora of cotton in soil sickness[J]. Biology&Fertility of Soils, 2015, 51(8):935-946.
[18]Mago T, Salzberg S L. FLASH:fast length adjustment of short reads to improve genome assemblies[J]. Bioinformatics, 2011,27(21):2957-2963.
[19]Bolger A M, Lohse M, Usadel B. Trimmomatic:a flexible trimmer for Illumina sequence data[J]. Bioinformatics, 2014, 30(15):2114-2120.
[20]Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics 2010; 26:2460.
[21]Koljalg U, Larsson K K, Nilsson R H, et al. UNITE:a database providing web‐based methods for the molecular identification of ectomycorrhizal fungi[J]. New Phytologist, 2005, 166(3):1063–1068.Nguyen N H, Song Z, Bates S T, et al.
[22]Berendsen R L, Pieterse C M J, Bakker P A H M. The rhizosphere microbiome and plant health[J]. Trends in Plant Science, 2012,17(8):478-86.
[23]李小龙,李红丽,曾强,等.钙镁磷肥对青枯病发病烟株根际土壤微生物区系的影响[J].中国烟草学报, 2016, 22(1):71-75.LI Xiaolong, LI Hongli, ZENG Qiang, et al. Effects of calciummagnesium phosphates on microbial flora in rhizosphere soil of tobacco plant effected with Ralstonia solanacearum[J]. Acta Tabacaria Sinica, 2016, 22(1):71-75.
[24]LiXG,DingCF,Zhang TL, WangXX.Fungalpathogen accumulation at the expense of plant-beneficial fungi as a consequence of consecutive peanut monoculturing. Soil Biology&Biochemistry 2014; 72:11-18.
[25]Tedersoo L, Bahram M, P?lme S, et al. Global diversity and geography of soil fungi[J]. Science, 2014, 346(6213):1-11.
[26]桑维钧,祝明亮,吴兴禄,等.烟草镰刀菌根腐病研究初报[J].山地农业生物学报, 1998, 42(3):140-141.SANG Weijun, ZHU Mingliang, WU Xinglu, et al. A Preliminary report of tobacco root rot caused by Fusarium sp.[J]. Journal of Mountain Agriculture and Biology, 1998, 42(3):140-141.
[27]吴安忠,程崖芝,巫升鑫,等.烟草镰刀菌根腐病的病原鉴定[J].中国烟草学报, 2018, 24(2):135-140.WU Anzhong, CHENG Yazhi, WU Shengxin, et al. Identification of tobacco Fusarium root rot pathogen[J]. Acta Tabacaria Sinica,2018, 24(2):135-140.
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