盐碱地白刺不同部位微生物群落高通量分析

doi:10.6048/j.issn.1001-4330.2022.10.026

摘要/Abstract

摘要：

【目的】研究新疆盐碱环境中耐盐植物白刺的根际土壤和叶片内生微生物群落的结构丰富度和多样性。【方法】运用Novaseq测序平台对白刺根际土壤和叶片进行16SrDNA-V4区和ITS1区测序。经FLASH进行拼接,经过Qiim过滤叶绿体和线粒体序列,得到的序列在97%的序列相似性水平上聚类成为操作分类单元(OTUs)。对序列数据进行生物信息学分析,评价物种的Chao1指数、shannon指数以及Alpha多样性。【结果】白刺根际土壤细菌优势菌群为放线菌门(Actinobacteria)、变形菌门(Proteobacteria)、芽单胞菌门(Gemmatimonadetes);真菌优势菌群为子囊菌门(Ascomycota)。叶片内生细菌优势菌群为蓝细菌门(Cyanobacteria)和放线菌门(Actinobacteria);真菌优势菌群为子囊菌门(Ascomycota),白刺不同部位的微生物菌群数量和多样性有显著差异(P<0.05),白刺各样地之间微生物群落构成差异明显。【结论】白刺不同部位微生物群落的丰富度和多样性分布。

关键词: 白刺; 高通量测序; 根际微生物; 叶片内生菌; 微生物多样性; 微生物群落结构

Abstract:

【Objective】 To explore the structural richness and diversity of endophytic microbial communities associated with rhizosphere soil and leaves in the saline-alkali environment of Xinjiang.【Methods】 The 16SrDNA-V4 region amplicon and ITS1 region were sequenced in rhizosphere soil and leaves of Nitraria through Novaseq sequencing system.After each sample was amplified and fused by FLASH, and chloroplast genome sequence and mitochondrial sequence were filtered by Qiim, sequences were finally obtained respectively.With 97% sequence similarity, these sequences were clustered into Operational Taxonomic Units (OTUs).Bioinformatics analysis was carried out on the sequence data to evaluate the species Chao1 index, Shannon index and Alpha diversity.【Results】 The results showed that the dominant bacterial microflora of Nitraria in rhizosphere soil were Actinobacteria, Proteobacteria, Gemmatimonadetes; And the dominant microflora of fungi was Ascomycota, the dominant endophytic bacterial microfloras were Cyanobacteria and Actinobacteria; and the dominant microflora of fungi was Ascomycota.There were significant differences in the number and diversity of microflora in different parts of Nitraria (P<0.05).There were obvious differences in the composition of microbial communities among various species.【Conclusion】 The richness and diversity distribution of microbial community in different parts of Nitraria are revealed by high-throughput sequencing technology, which has provided theoretical basis for the selection of PGPR strains, the improvement of saline soil and the development of microbial fertilizers.

Key words: nitraria; high-throughput sequencing; rhizosphere microorganism; endophyte in leaves; microbial diversity; microbial community structure

中图分类号:

S158

艾海白尔·卡斯木, 樊永红, 迪拉热·海米提. 盐碱地白刺不同部位微生物群落高通量分析[J]. 新疆农业科学, 2022, 59(10): 2562-2573.

Akbar Kasim, FAN Yonghong, Dilara Hamit. High Throughput Analysis of the Microbial Community in Different Parts of Nitraria in Saline Alkali Land[J]. Xinjiang Agricultural Sciences, 2022, 59(10): 2562-2573.

图/表 10

参考文献 42

[1]	赵晨光, 孟和巴雅尔, 丁积禄. 阿拉善白刺生长季叶片尺度耗水特性[J]. 防护林科技, 2017,(10):46-50.
	ZHAO Chenguang, Menghebayaer, DING Jilu. The characteristics of water consumption of blade scale for nitraria tangutorun in alxa league[J]. Protection Forest Science and Technology, 2017,(10):46-50.
[2]	席军强, 杨自辉, 郭树江, 等. 不同类型白刺沙丘土壤理化性状与微生物相关性研究[J]. 草业学报, 2015,(6):64-74.
	XI Junqiang, YANG Zihui, GUO Shujiang. The correlation between soil physical and chemical properties and soil microbes in different types of Nitraria dune[J]. Acta Prataculturae Sinica, 2015,(6):64-74.
[3]	张芝萍, 安富博, 赵艳丽, 等. 青土湖间断性水淹干扰对白刺沙堆土壤特性与生物量的影响[J]. 安徽农业科学, 2020, 48(1):70-72,82.
	ZHANG Zhiping, AN Fubo, ZHAO Yanli, et al. Effects of Intermittent Flooding Interference on Soil Properties and Biomass of Nitraria[J]. Journal of Anhui Agricultural Sciences, 2020, 48(1):70-72,82.
[4]	梁晓霞. 浅谈温室效应及其影响[J]. 中国高新区, 2019,(7):203.
	LIANG Xiaoxia. On Greenhouse Effect and Its Impact[J]. Science & Technology Industry Parks, 2019,(7):203.
[5]	Misra S, Dixit V K, Khan M H, et al. Exploitation of agro-climatic environment for selection of 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase producing salt tolerant indigenous plant growth promoting rhizobacteria.[J]. Microbiological Research, 2017: 25-34.
[6]	李娥, 胡华冉, 李蛟男, 等. 内生真菌提高植物抵御盐胁迫的研究进展[J]. 生物技术通报, 2019, 35(11):169-178. DOI
	LI E, HU Huaran, LI Jiaonan. Research Progress on Endophytic Fungi Improving Plant Resistance to Salt Stress[J]. Biotechnology Bulletin, 2019, 35(11):169-178. DOI
[7]	梁晋刚, 刘鹏程, 张秀杰. 基于16S rDNA高通量测序技术研究转基因作物对根际细菌群落结构的影响[J]. 江苏农业科学, 2018, 46(6):5-8.
	LING Jingang, LIU Pengcheng, ZHANG Xiujie. Effects S rDNA transgenic crops on rhizosphere bacterial community structure based on 16 high-throughput sequencing techniques[J]. Jiangsu Agricultural Sciences, 2018, 46(6):5-8.
[8]	楼骏, 柳勇, 李延. 高通量测序技术在土壤微生物多样性研究中的研究进展[J]. 中国农学通报, 2014, 30(15):256-260.
	LOU Jun, LIU Yong, LI Yan. Review of High-throughput Sequencing Techniques in Studies of Soil Microbial Diversity[J]. Chinese Agricultural Science Bulletin, 2014, 30(15):256-260.
[9]	Sarkar A, Ghosh P K, Pramanik K, et al. A halotolerant Enterobacter sp.displaying ACC deaminase activity promotes rice seedling growth under salt stress[J]. Research in Microbiology, 2018, 169(1): 20-32. DOI URL
[10]	叶文雨, 谢序泽, 许钰滢, 等. 基于高通量测序技术分析2种菌草根际土壤真菌群落多样性[J]. 热带作物学报, 2020, 41(3):556-563.
	YE Wenyu, XIE Xuze, XU Yuying, et al. High-throughput Sequencing Analysis of Fungal Community in Rhizosphere Soils of Two Plants for Mushroom Cultivation (JUNCAO)[J]. Chinese Journal of Tropical Crops, 2020, 41(3):556-563.
[11]	方静, 赵小庆, 史功赋, 等. 农作物根际微生物的研究进展[J]. 北方农业学报, 2019, 47(4):102-107. DOI
	FANG Jing, ZHAO Xiaoqing, SHI Gongfu. Research progress on rhizosphere microorganisms of crops[J]. Journal of Northern Agriculture, 2019, 47(4):102-107.
[12]	Qin S, Feng W, Wang T, et al. Plant growth-promoting effect and genomic analysis of the beneficial endophyte Streptomyces sp.KLBMP 5084 isolated from halophyte Limonium sinense[J]. Plant and Soil, 2017, 416(1): 117-132. DOI URL
[13]	杨美玲, 张霞, 王绍明, 等. 基于高通量测序的裕民红花根际土壤细菌群落特征分析[J]. 微生物学通报, 2018, 45(11):2429-2438.
	YANG Meiling, ZHANG Xia, WANG Shaoming, et al. High throughput sequencing analysis of bacterial communities in Yumin safflower[J]. Microbiology China, 2018, 45(11):2429-2438.
[14]	艾铄, 张丽杰, 肖芃颖, 等. 高通量测序技术在环境微生物领域的应用与进展[J]. 重庆理工大学学报(自然科学), 2018, 32(9):111-121.
	AI Shuo, ZHANG Lijie, XIAO Pengyi, et al. Application and Progress of High-Throughput Sequencing Technology in the Field of Environmental Microorganisms[J]. Journal of Chongqing University of Technology (Natural Science Ed.), 2018, 32(9):111-121.
[15]	葛艺, 徐绍辉, 徐艳. 根际微生物组构建的影响因素研究进展[J]. 浙江农业学报, 2019, 31(12):2120-2130. DOI
	GE Yi, XU Shaohui, XU Yan. Review on influencing factors of rhizosphere microbiome assemblage[J]. Acta Agriculturae Zhejiangensis, 2019, 31(12):2120-2130. DOI
[16]	姬文秀, 李虎林, 冷雪, 等. 产ACC脱氨酶人参内生细菌的分类和促生特性分析[J]. 吉林农业大学学报, 2019, 41(2):168-174.
	JI Wenxiu, LI Hulin, LENG Xue. Isolation and Promoting Growth Properties of Endophytic Bacteria Producing ACC Deaminase of Panax ginseng[J]. Journal of Jilin Agricultural University, 2019, 41(2):168-174.
[17]	陈泽斌, 李冰, 高熹, 等. 铁皮石斛不同部位内生细菌群落高通量分析[J]. 河南农业科学, 2017, 46(11):98-102,112.
	CHEN Zebin, LI Bing, GAO Xi, et al. High Throughput Analysis of the Endophytic Bacterial Community in Different Parts of Dendrobium candidum[J]. Journal of Henan Agricultural Sciences, 2017, 46(11):98-102,112.
[18]	沙月霞. 不同水稻组织内生细菌的群落多样性[J]. 微生物学报, 2018, 58(12):2216-2228.
	SHA Yuexia. Diversity o f bacterial endophytic community in different rice tissues[J]. Acta microbiologica Sinica, 2018, 58(12):2216-2228.
[19]	周密密, 周欣, 孙民琴, 等. 高通量测序技术分析进境高粱种子的真菌多样性[J]. 南京农业大学学报, 2019, 42(4):665-671.
	ZHOU MiMi, ZHOU Xin, SUN Mingqin, et al. Fungal diversity in imported sorghum by high-throughput sequencing technology[J]. Journal of Nanjing AgriculturalUniversity, 2019, 42(4):665-671.
[20]	刘开辉, 丁小维, 张波, 等. 高通量测序分析云南腾冲热海热泉真菌多样性[J]. 微生物学报, 2017, 57(9):1314-1322.
	LIU Kaihui, DING Xiaowei, ZHANG Bo, et al. High -throughput sequencing to reveal fungal diversity in hot springs of Rehai at Tengchong in Yunnan[J]. Acta Microbiologica Sinica, 2017, 57(9):1314-1322.
[21]	林丽, 陈泽斌, 何群香, 等. 烟草不同部位内生细菌的多样性[J]. 江苏农业科学, 2017, 45(22):274-278.
	LIN Li, CHEN Zebin, HE Qunxiang, et al. Diversity of endophytic bacteria in different parts of tobacco[J]. Jiangsu Agricultural Sciences, 2017, 45(22):274-278.
[22]	刘蓬蓬, 陈江宁, 贾天柱. 基于Illumina MiSeq高通量测序分析黄芪内生真菌的多样性[J]. 中国实验方剂学杂志, 2018, 24(12):34-38.
	LIU Pengpeng, CHEN Jiangning, JIA Tianzhu. Analysis of Diversity of Endophytic Fungi in Astragali Radix by Illumina MiSeq High-throughput Sequencing Technology[J]. Chinese Journal of Experimental Traditional Medical Formulae, 2018, 24(12):34-38.
[23]	丁新景, 敬如岩, 黄雅丽, 等. 基于高通量测序的4种不同树种人工林根际土壤细菌结构及多样性[J]. 林业科学, 2018, 54(1):81-89.
	DING Xinjing, JING Ruyan, HUANG Yali, et al. Bacterial Structure and Diversity of Rhizosphere Soil of Four Tree Species in Yellow River Delta Based on High-Throughput[J]. Sequencing Scientia Silvae Sinicae, 2018, 54(1):81-89.
[24]	Jia Y, Liao Z, Chew H, et al. Effect of Pennisetum giganteum z.x.lin mixed nitrogen-fixing bacterial fertilizer On The Growth, Quality, Soil Fertility And Bacterial Community Of Pakchoi (Brassica Chinensis L.)[J]. Plos One, 2020, 15(2).
[25]	赵帅, 周娜, 赵振勇, 等. 新疆北部主要盐生植物根部内生细菌群落结构的高通量分析[J]. 微生物学报, 2016, 56(10):1583-1594. PMID
	ZHAO Shuai, ZHOU Na, ZHAO Zhenyong, et al. Estimation of endophytic b cterial diversity in root of halophytes in Northern Xinjiang by high throughput sequencing[J]. Acta microbiologica Sinica, 2016, 56(10):1583-1594. PMID
[26]	Zhou Y, Bao J, Zhang D, et al. Effect of heterocystous nitrogen-fixing cyanobacteria against rice sheath blight and the underlying mechanism[J]. Applied Soil Ecology, 2020
[27]	李发虎, 李明, 刘金泉, 等. 生物炭对温室黄瓜根际土壤真菌丰度和根系生长的影响[J]. 农业机械学报. 2017, 48(4):265-270,341.
	LI Fahu, LI Ming, LIU Jinquan, et al. Effect of Biochar on Fungal Abundance of Rhizosphere Soil and Cucumber Root Growth in Greenhouse[J]. Transactions of The Chinese Society of Agricultural Machinery, 2017, 48(4):265-270,341.
[28]	杨潇湘, 张蕾, 黄小琴, 等. 基于高通量测序分析大豆和油菜根际微生物群落结构的差异[J]. 应用生态学报, 2019, 30(7):2345-2351. DOI
	YANG Xiaoxiang, ZHANG Lei, HUANG Xiaoqin, et al. Difference of the microbial community structure in the rhizosphere of soybean and oilseed rape based on high-throughput pyrosequencing analysis[J]. Chinese Journal of Applied Ecology, 2019, 30(7):2345-2351. DOI
[29]	费诗萱, 张敏, 王迎, 等. 具有ACC脱氨酶活性的红枣根际促生菌株的分类筛选及其促生效果研究[J]. 西北林学院学报, 2019, 34(6):140-146.
	FEI Shixuan, ZHANG Min, WANG Ying, et al. Isolation, Screening and Promoting Effects of Plant Growth-promoting Rhizobacteria (PGPR)Containing ACC Deaminase from Jujube[J]. Journal of Northwest Forestry University, 2019, 34(6):140-146.
[30]	马欣, 成妍, 马蓉丽. 植物根围促生细菌促生机制研究进展[J]. 山东农业科学, 2019, 51(5):148-154
	MA Xin, CHENG Yan, MA Rongli. Research Progress of Growth-Promoting Mechanisms of Plant Growth-Promoting rhizobacteria[J]. Shandong Agricultural Sciences, 2019, 51(5):148-154..
[31]	刘丹丹, 李敏, 刘润进. 我国植物根围促生细菌研究进展[J]. 生态学杂志, 2016, 35(3):815-824.
	LIU Dandan, LI Min, LIU Runjin. Recent advances in the study of plant growth -promoting rhizobacteria in China[J]. Chinese Journal of Ecology, 2016, 35(3):815-824.
[32]	刘少芳, 王若愚. 植物根际促生细菌提高植物耐盐性研究进展[J]. 中国沙漠, 2019, 39(2):1-12.
	LIU Shaofang, WANG Ruoyu. Advance in Research on Plant Salt Tolerance Improved by Plant-growth-promoting Rhizobacteria[J]. Journal of Desert Research, 2019, 39(2):1-12.
[33]	段娜, 李清河, 陈晓娜, 等. 模拟干旱和氮沉降对唐古特白刺根系生长特征的影响[J]. 草地学报, 2019, 27(4):956-961. DOI
	DUAN Na, LI Qinghe, CHEN Xiaona, et al. Combined effects of simulated drought stress and nitrogen deposition on root growth characteristics of nitraria tangutorum.[J]. Acta Agrestila Sinica, 2019, 27(4):956-961.
[34]	Gowtham H G, Singh S B, Murali M, et al. Induction of drought tolerance in tomato upon the application of ACC deaminase producing plant growth promoting rhizobacterium Bacillus subtilis Rhizo SF 48[J]. Microbiological Research, 2020.
[35]	魏伟, 吴小芹, 乔欢. 马尾松根际高效解磷真菌的筛选鉴定及其促生效应[J]. 林业科学, 2014, 50(9):82-88.
	WEI Wei, WU Xiaoqin, QIAO Huan. Screening and identification of phosphate-solubilizing fungi of pinus massoniana rhizosphere and its application[J]. Scientia Silvae Sinicae, 2014, 50(9):82-88.
[36]	桂许维, 张扬, 宋庆妮, 等. 毛竹林钾矿物分解细菌的分类与鉴定[J]. 森林与环境学报, 2018, 38(4):499-505.
	GUI Xuwei, ZHANG Yang, SONG Qinni, et al. Isolation and identification of the mineral potassium-solubilizing bacteria in Phyllostachys edulis forest[J]. Journal of Forest and Environment, 2018, 38(4):499-505.
[37]	Mustafa S, Kabir S, Shabbir U, et al. Plant growth promoting rhizobacteria in sustainable agriculture: from theoretical to pragmatic approach[J]. Symbiosis, 2019, 78:115-123. DOI
[38]	Pereira L B, Andrade G S, Meneghin S P, et al. Prospecting Plant Growth-Promoting Bacteria Isolated from the Rhizosphere of Sugarcane Under Drought Stress.[J]. Current Microbiology, 2019, 76(11): 1345-1354. DOI PMID
[39]	Rana K L, Kour D, Kaur T, et al. Endophytic microbes: Biodiversity, plant growth-promoting mechanisms and potential applications for agricultural sustainability[J]. Antonie van Leeuwenhoek, 2020, 113(8):1075-1107. DOI PMID
[40]	陆奇丰, 黄至欢, 唐文秀. 细胞分裂素和赤霉素对3种山茶科种子萌发特性的影响[J]. 种子, 2019, 38(2):25-30.
	LU Qifeng, HUANG Zhihuan, TANG Wenxiu. Effect of 6-benzylaminopurine and gibberellin on seed germination of three theaceae[J]. Seed, 2019, 38(2):25-30.
[41]	孙萌. 铁载体高产菌株的ARTP选育及其铁载体产量提高机理的初步分析[D]. 无锡: 江南大学, 2017.
	SUN Meng. ARTP selection of high siderophore productions trains and preliminary analysis on mechanism of the siderophore improving[D]. Wuxi: Jiangnan University, 2017.
[42]	路璐. CARD-FISH技术及其在微生物生态学研究中的应用[J]. 微生物学杂志, 2017, 37(6):87-97.
	LU Lu. Catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) and its application in microbial ecology study[J]. Journal of Microbiology, 2017, 37(6):87-97.

样本Sample group	细菌Bacteria			真菌Fungus
样本Sample group	丰富度指数 Chao1	多样性指数 Shannon	覆盖率 Coverage (%)	丰富度指数 Chao1	多样性指数 Shannon	覆盖率 Coverage(%)
BS₁	1 355.13						7.205	99.6	376.65	2.280	99.9
BS₂	1457.25						7.878	99.7	597.40	4.453	99.9
BS₃	1 503.46						7.884	99.6	445.21	3.256	99.9
BP₁	375.98						2.251	99.8	59.67	1.540	100
BP₂	289.47						0.991	99.9	21.00	1.336	100
BP₃	323.50						0.864	99.8	21.00	1.563	100

样本Sample group	细菌Bacteria			真菌Fungus
样本Sample group	丰富度指数 Chao1	多样性指数 Shannon	覆盖率 Coverage (%)	丰富度指数 Chao1	多样性指数 Shannon	覆盖率 Coverage(%)
BS₁	1 355.13						7.205	99.6	376.65	2.280	99.9
BS₂	1457.25						7.878	99.7	597.40	4.453	99.9
BS₃	1 503.46						7.884	99.6	445.21	3.256	99.9
BP₁	375.98						2.251	99.8	59.67	1.540	100
BP₂	289.47						0.991	99.9	21.00	1.336	100
BP₃	323.50						0.864	99.8	21.00	1.563	100

分类 Taxon	土壤Soil		叶片Leaves
	bacteria	fungus	bacteria	fungus
	OTUs数	1 985	878	593	252
界Kingdom	2	2	1	2
门Phylum	29	9	17	6
纲Class	44	28	31	12
目Order	93	60	66	19
科Family	167	114	123	25
属Genus	305	167	212	28
种Species	103	159	97	25

分类 Taxon	土壤Soil		叶片Leaves
	bacteria	fungus	bacteria	fungus
	OTUs数	1 985	878	593	252
界Kingdom	2	2	1	2
门Phylum	29	9	17	6
纲Class	44	28	31	12
目Order	93	60	66	19
科Family	167	114	123	25
属Genus	305	167	212	28
种Species	103	159	97	25

细菌门 phylum of bacteria	占比 proportion(%)			细菌门 phylum of bacteria	占比 proportion(%)
细菌门 phylum of bacteria	BS₁	BS₂	BS₃	细菌门 phylum of bacteria	BP₁	BP₂	BP₃
放线菌门(Actinobacteria)	26.17	24.48	48.16	蓝细菌门(Cyanobacteria)	69.86	93.17	93.05
变形菌门(Proteobacteria)	23.91	22.63	19.40	放线菌门(Actinobacteria)	26.28	5.14	5.41
芽单胞菌门(Gemmatimonadetes)	23.62	19.18	9.46	变形菌门(Proteobacteria)	2.58	0.93	0.99
拟杆菌门(Bacteroidetes)	7.48	17.60	8.00	厚壁菌门(Firmicutes)	0.66	0.45	0.33
浮霉菌门(Planctomycetes)	4.65	6.07	5.36	拟杆菌门(Bacteroidetes)	0.29	0.13	0.09
绿弯菌门(Chloroflexi)	2.96	1.62	1.48	芽单胞菌门(Gemmatimonadetes)	0.22	0.09	0.06
酸杆菌门(Acidobacteria)	3.21	3.15	4.86	栖热菌门(Deinococcus-thermus)	0.02	0.03	0.01
庞微菌门(Verrucomicrobia)	0.65	1.46	0.85	酸杆菌门(Acidobacteria)	0.007	-	0.002
厚壁菌门(Firmicutes)	0.79	1.53	0.77	绿弯菌门(Chloroflexi)	0.016	-	0.002