大花杓兰根际与非根际土壤真菌多样性的高通量测序分析
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摘要
采用Illumina Miseq高通量测序技术对大花杓兰根际与非根际土壤真菌群落组成进行对比分析。结果表明,大花杓兰根际土壤获得63 780条Clean tags,非根际土壤获得48 094条Clean tags。基于≥97%的相似度水平,通过聚类共获得1 223个OTUs,根际和非根际土壤真菌OTUs数量分别为1 004和945,二者共有726个OTUs。Chao1和Shannon指数揭示根际土壤真菌群落在丰富度和多样性上均高于其非根际土壤。与非根际土壤相比,在门、纲、目、科和属分类水平上丰度显著增加的根际土壤真菌类群分别为担子菌门(Basidiomycota)、伞菌纲(Agaricomycetes)、伞菌目(Agaricales)和鸡油菌目(Cantharellales)、小菇科(Mycen-aceae)和角担菌科(Ceratobasidiaceae)、小菇属(Mycena)和角担菌属(Ceratobasidium)。首次从分子水平上揭示大花杓兰根际与非根际土壤真菌群落的多样性对大花杓兰的保育具有重要意义。
The composition of fungal communities in the rhizosphere and non-rhizosphere soils from Cypripedium macranthum were comparatively analyzed,combinated the Illumina Miseq platform with bioinformatics.The results showed that a total of 63 780 and 48 094 clean tags were obtained from rhizospheric and non-rhizospheric soil samples,respectively.At the 97% sequence similarity levels,1 223 operational taxonomic units(OTUs)were identified based on UCLUST algorithm.1 004 OTUs were obtained from rhizospheric soil and 945 OTUs from non-rhizospheric soil,726 OTUs were shared by two groups of samples.The Chao1 and Shannon index indicated that the abundance and diversity of rhizosphere fungi were higher than that of non-rhizosphere fungi.Compared with the nonrhizosphere soil at the classification levels of phylum,class,order,family and genera,the dominant groups of C.macranthumrhizosphere fungi were Basidiomycota,Agaricomycetes,Agaricales and Cantharellales,Mycenaceae and Ceratobasidiaceae,Mycena and Ceratobasidium,respectively.It was firstly revealed that the diversity of fungal communities in rhizosphere and non-rhizosphere soils from C.macranthumis of great significance for its conservation.
The composition of fungal communities in the rhizosphere and non-rhizosphere soils from Cypripedium macranthum were comparatively analyzed,combinated the Illumina Miseq platform with bioinformatics.The results showed that a total of 63 780 and 48 094 clean tags were obtained from rhizospheric and non-rhizospheric soil samples,respectively.At the 97% sequence similarity levels,1 223 operational taxonomic units(OTUs)were identified based on UCLUST algorithm.1 004 OTUs were obtained from rhizospheric soil and 945 OTUs from non-rhizospheric soil,726 OTUs were shared by two groups of samples.The Chao1 and Shannon index indicated that the abundance and diversity of rhizosphere fungi were higher than that of non-rhizosphere fungi.Compared with the nonrhizosphere soil at the classification levels of phylum,class,order,family and genera,the dominant groups of C.macranthumrhizosphere fungi were Basidiomycota,Agaricomycetes,Agaricales and Cantharellales,Mycenaceae and Ceratobasidiaceae,Mycena and Ceratobasidium,respectively.It was firstly revealed that the diversity of fungal communities in rhizosphere and non-rhizosphere soils from C.macranthumis of great significance for its conservation.
引文
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[2]MCCORMICK M K,JACQUEMYN H.What constrains the distribution of orchid populations[J].New Phytologist,2014,202(2):392-400.
[3]张毓,赵世伟.一种促进大花杓兰种子共生萌发的真菌及其应用[P].中国:CN201110402377.6,2011-11-06.ZHANG Y,ZHAO SH W.Application and Identification of Mycorrhizal Fungi Promoting Symbiotic Seed Germination of Cypripedium macranthos[P].China:CN201110402377.6,2011-11-06.
[4]SHIMURA H,SADAMOTO M,MATSUURA M,et al.Characterization of mycorrhizal fungi isolated from the threatened Cypripedium macranthos,in a northern island of Japan:two phylogenetically distinct fungi associated with the orchid[J].Mycorrhiza,2009,19(8):525-534.
[5]张亚平.中国北方三种杓兰内生真菌多样性及其对原球茎生长的效应[D].四川雅安:四川农业大学,2013.ZHANG Y P.Diversity of endophytic fungi in three Cypripediumspecies in northern China and their effects on protocorm development[D].Ya’an Sichuan:Sichuan Agricultural University,2013.
[6]COATS V C,RUMPHO M E.The rhizosphere microbiota of plant invaders:an overview of recent advances in the microbiomics of invasive plants[J].Frontiers in Microbiology,2014,5(5):368.
[7]MCCORMICK M K,TAYLOR D L,WHIGHAM D F,et al.Germination patterns in three terrestrial orchids relate to abundance of mycorrhizal fungi[J].Journal of Ecology,2016,104(3):744-754.
[8]WAUD M,WIEGAND T,BRYS R,et al.Nonrandom seedling establishment corresponds with distance-dependent decline in mycorrhizal abundance in two terrestrial orchids[J].New Phytologist,2016,211(1):255-264.
[9]MAGOCT,SALZBERG S L.FLASH:fast length adjustment of short reads to improve genome assemblies[J].Bioinformatics,2011,27(21):2957-2963.
[10]BOLGER A M,LOHSE M,USADEL B.Trimmomatic:a flexible trimmer for Illumina sequence data[J].Bioinformatics,2014,30(15):2114-2120.
[11]EDGAR R C,HAAS B J,CLEMENTE J C,et al.UCH-IME improves sensitivity and speed of chimera detection[J].Bioinformatics,2011,27(16):2194-2200.
[12]EDGAR R C.Search and clustering orders of magnitude faster than BLAST[J].Bioinformatics,2010,26(19):2460-2461.
[13]SCHLOSS P D,WESTCOTT S L,RYABIN T,et al.Introducing mothur:open-source,platform-independent,community-supported software for describing and comparing microbial communities[J].Applied Environment Microbiology,2009,75(23):7537-7541.
[14]MENDES R,GARBEVA P,RAAIJMAKERS J M.The rhizosphere microbiome:significance of plant beneficial,plant pathogenic,and human pathogenic microorganisms[J].FEMS Microbiology Reviews,2013,37(5):634-663.
[15]徐红梅,倪方方,宋腾蛟,等.高通量测序揭示杭白芍根际真菌群落年际变化[J].浙江中医药大学学报,2016,40(12):942-947.XU H M,NI F F,SONG T J,et al.Fungal interannual changes in the rhizosphere soil of Paeonia lactiflora by454Pyrosequencing[J].Journal of Zhejiang Chinese Medical University,2016,40(12):942-947.
[16]李倩,杨水平,崔广林,等.不同种植年限条件下黄花蒿根际土壤微生物生物量、酶活性及真菌群落组成[J].草业学报,2017,26(1):34-42.LI Q,YANG SH P,CUI G L,et al.Microbial biomass,enzyme activity and composition of the fungal community in rhizospheric soil cropped with Artenisia annua for several years[J].Acta Prataculturas Sinica,2017,26(1):34-42.
[17]李越鲲,孙燕飞,雷勇辉,等.枸杞根际土壤真菌群落多样性的高通量测序[J].微生物学报,2017,57(7):1049-1059.LI Y K,SUN Y F,LEI Y H,et al.Fungal community diversity in rhizosphere soil of Lycium barbarum L.based on high-throughput sequencing[J].Acta Microbiologica Sinica,2017,57(7):1049-1059.
[18]AL-SADI A M,AL-MAZROUI S S,PHILLIPS A J.Evaluation of culture-based techniques and 454pyrosequencing for the analysis 7of fungal diversity in potting media and organic fertilizers[J].Journal of Applied Microbiology,2015,119(2):500-509.
[19]OJA J,VAHTRA J,BAHRAM M.Local-scale spatial structure and community composition of orchid mycorrhizal fungi in semi-natural grasslands[J].Mycorrhiza,2017,27(4):355-367.
[20]VOYRON S,ERCOLE E,GHIGNONE S,et al.Fine-scale spatial distribution of orchid mycorrhizal fungi in the soil of host-rich grasslands[J].New Phytologist,2017,213:1428-1439.
[21]黄运峰,杨小波.兰科菌根研究综述[J].热带亚热带植物学报,2008,16(3):283-288.HUANG Y F,YANG X B.Summary research on orchidaceous mycorrhizae[J].Journal of Tropical and Subtropical Botany,2008,16(3):283-288.
[22]赵文静,周明,孙海,等.额尔古纳国家级自然保护区内4种林型土壤真菌的多样性[J].东北林业大学学报,2014,42(5):105-109.ZHAO W J,ZHOU M,SUN H,et al.Soil fungi diversity in four forest types of Erguna National Nature[J].Reserve Journal of Northeast Forestry University,2014,42(5):105-109.
[23]王殿东,谭永忠,田雪亮,等.茎瘤芥(榨菜)根肿病不同发病时期根际土壤真菌群落变化特征的高通量分析[J].中国蔬菜,2016(5):33-37.WANG D D,TAN Y ZH,TIAN X L,et al.High throughput analysis on change characteristics of rhizosphere soil fungal community during different occurance times of tuber mustard clubroot disease[J].China Vegetables,2016(5):33-37.
[24]MIAO C P,MI Q L,QIAO X G,et al.Rhizospheric fungi of Panax notoginseng:diversity and antagonism to host phytopathogens[J].Journal of Ginseng Research,2016,40(2):127-134.