基于高通量测序技术的酿酒葡萄产区土壤微生物多样性

doi:10.6048/j.issn.1001-4330.2020.05.009

摘要/Abstract

摘要： 【目的】研究新疆4大酿酒葡萄产区土壤中的微生物群落结构及多样性,为构建新疆酿酒葡萄主产区的微生物菌库奠定基础。【方法】分别从新疆酿酒葡萄主产区采集土壤样品,提取总DNA,通过Illumina MiSeq高通量测序结合生物信息学分析,得到新疆酿酒葡萄主产区的土壤微生物群落结构及多样性。【结果】在12份土壤样品中,伊犁河谷土壤中细菌的群落多样性最高,天山北麓土壤中细菌群落多样性最低;吐哈盆地中真菌群落多样性最低,焉耆盆地中真菌的群落多样性最高。节杆菌属细菌和耐冷酵母属真菌是吐哈盆地和天山北麓葡萄园土壤中的优势菌属;芽单胞菌属细菌和刚毛四枝孢属真菌是伊犁河谷葡萄园土壤中的优势菌属;节杆菌属细菌和赤霉菌属真菌是焉耆盆地葡萄园土壤中的优势菌属。土壤pH值对细菌群落组成的影响最显著,Mn元素含量对真菌群落组成的影响最显著。【结论】揭示了新疆酿酒葡萄主产区土壤中细菌和真菌群落组成,新疆酿酒葡萄产区的微生物多样性存在差异。

关键词: Illumina MiSeq高通量测序; 酿酒葡萄; 土壤; 微生物群落; 多样性

Abstract: 【Objective】 To find out the microbial community structure and diversity in the soil of the four major wine grape producing areas in Xinjiang and to lay a foundation for constructing the microbial bank in the main wine grape producing areas in the region. 【Methods】 After total DNA was extracted, Illumina MiSeq high-throughput sequencing system and bioinformatics analysis were used for the investigation of microbial communities and species diversity of main wine grape producing area in Xinjiang. 【Results】 In the twelve soil samples, the results showed that the highest microbial diversity of bacteria was the soil in Yili river-valley and the lowest one was in the soil of northern slope of Tianshan Mountain, and the highest microbial diversity of fungus was in the soil of Yanqi basin and the lowest microbial diversity of fungus was the Tuha basin soil. The dominant bacteria and fungus were Arthrobacter and Guehomyces in Tuha basin and the northern slope of Tianshan Mountain wine grape producing area. The dominant bacteria and fungus were Gemmatimonas and Tetracladium in Yili river-valley wine grape producing area. The dominant bacteria and fungus were Arthrobacter and Gibberella in Yanqi basin wine grape producing area. Soil pH value had the most significant effect on the composition of bacterial community, and the content of Mn had the most significant effect on the composition of fungal community. 【Conclusion】 This study revealed the composition of bacterial and fungal communities in the soil in main wine grape producing areas in Xinjiang, which indicated that there were differences in microbial community diversity.

Key words: Illumina MiSeq high-throughput sequencing system; wine grapes; soil; microbial community; diversity

中图分类号:

S188

王伟, 布丽根·加冷别克, 胡晓东, 夏俊芳, 张志东, 顾美英, 武运. 基于高通量测序技术的酿酒葡萄产区土壤微生物多样性[J]. 新疆农业科学, 2020, 57(5): 859-868.

WANG Wei, Buligen Jialengbieke, HU Xiaodong, XIA Junfang, ZHANG Zhidong, GU Meiying, WU Yun. Analysis of the Microbial Community Diversity of Soil from Wine Grape Productiing Area in Xinjiang Based on High-throughput Sequencing[J]. Xinjiang Agricultural Sciences, 2020, 57(5): 859-868.

参考文献

[1] 龙妍, 惠竹梅, 程建梅, 等. 生草葡萄园土壤微生物分布及土壤酶活性研究 [J]. 西北农林科技大学学报, 2007, 35(6): 99-103.
LONG Yan, HUI Zhumei, CHENG Jianmei, et al. Ecological distributing of soil microorganisms and activity of soil enzymes in vineyard green covering [J]. Journal of Northwest A & F University, 2007, 35(6): 99-103.
[2] Renouf, V, Claisse, O, & Lonvaud-Funel, A. Inventory and monitoring of wine microbial consortia [J]. Applied Microbiology and Biotechnology, , 2007,75(1): 149-164.
[3] Vincent, R, Olivier, C, & Aline, L. Understanding the microbial ecosystem on the grape herry surfur through numeration and identification of yeast and bacteria [J]. Australian Journal of Grape and Wine Research, 2010, 11(3): 316-327.
[4] Hirano, S. S, & Upper, C. D. Bacteria in the leaf ecosystem with emphasis on Pseudomonas syringae-a pathogen, ice nucleus, and epiphyte [J]. Microbiology and Molecular Biology Reviews, 2000, 64(3): 624-653.
[5] Park, J. Y, Han, S. H, & Lee, J. H. Draft genome sequence of the biocontrol bacterium Pseudomonas putida B001, an oligotrophic bacterium that induces systemic resistance to plant diseases [J]. Journal of Bacteriology, 2011, 193(23): 6795-6796.
[6] Sun, H. H, Ma, H. Q, & Hao, M. L. Identification of yeast population dynamics of spontaneous fermentation in Beijing wine region, China [J]. Annals of Microbiology, 2009, 59(1): 69-76.
[7] Li, E., Liu, A., & Xue, B. Yeast species associated with spontaneous wine fermentation of Cabernet Sauvignon from Ningxia, China [J]. World Journal of Microbiology & Biotechnology, 2011, 27(10): 2475-2482.
[8] 柴丽红, 崔晓龙, 彭谦, 等. 青海两盐湖细菌多样性研究 [J]. 微生物学报, 2004, 44(3): 271-275.
CHAI Lihong, CUI Xiaolong, PENG Qian, et al. Bacterial diversity of two salt lakes in Qinghai [J]. Acta Microbiologica Sinica, 2004, 44(3): 271-275.
[9] 曹露芬, 殷婷婷, 袁振亚, 等. 基于高通量测序的陕西花马池和苟池微生物群落多样性分析[J]. 生物技术通讯, 2016, 27(3): 374-380.
CAO Lufeng, YIN Tingting, YUAN Zhenya, et al. Analysis of the microbial community composition of Huama lake and Gou lake in Shanxi province based on high-throughput sequencing [J]. Letters in Biotechnology, 2016, 27(3): 374-380.
[10] Wu, L. Y, Wen, C. Q, & Qin, Y. J. Phasing amplicon sequencing on Illumina Miseq for robust environmental microbial community analysis [J]. BMC Microbiol, 2015(15): 125.
[11] You, J, Wu, G, & Ren, F. P. Microbial community dynamics in Baolige oilfield during MEOR treatment, revealed by Illumina MiSeq sequencing. Applied Microbiology & Biotechnology, 2015, 100(3): 1469-1478.
[12] 于荣, 徐明岗, 王伯仁. 土壤活性有机质测定方法的比较 [J]. 中国土壤肥料, 2005, (2): 49-52.
YU Rong, XU Mingang, WANG Boren. (2005). Study on methods for determining labile orange matter of soils [J]. Soils and Fertilizers, 2005 (2): 49-52.
[13] 沈文, 陈均志, 代春吉. 微波消解-凯式定氮法测定牛奶中蛋白质含量 [J]. 食品研究与开发, 2009, 30(5): 96-97.
SHEN Wen, CHEN Junzhi, DAI Chunji. Determination of protein content in milk by microwave digestion-kjeldahl [J]. Food Research and Development, 2009, 30(5): 96-97.
[14] 于翠, 胡兴明, 李欣, 等. 3种土壤类型桑园根际与非根际微生物种群结构及代谢功能多样性分析[J]. 蚕业科学, 2015, 41(1): 28-36.
YU Cui, HU Xingmin, LI Xin, et al. Microbial community structure and metabolic function diversity between rhizospheric and non-rhizospheric soil in three types of mulberry fields [J]. Science of Sericulture, 2015, 41(1): 28-36.
[15] 丁雷, 李俊华, 赵思峰, 等. 生物有机肥和拮抗菌对土壤有效养分和土壤酶活性的影响[J]. 新疆农业科学, 2011, 48(3): 504-510.
DING Lei, LI Junhua, ZHAO Sifeng, et al. Effects of application of biological organic fertilizer and antagonistic bacteria on soil available nutrients and soil enzyme activities [J]. Xinjiang Agricultural Sciences, 2011, 48(3): 504-510.
[16] 吴湘琳, 严晶, 窦晓静,等. 新疆温宿县果园土壤微量元素含量及有效性评价 [J]. 新疆农业科学, 2017, 54(3): 520-527.
WU Xianglin, YAN Jing, DOU Xiaojin, et al. Contents and availability of trace elements in orchard soil in Wensu county of Xinjiang [J]. Xinjiang Agricultural Sciences, 2017, 54(3): 520-527.
[17] Zhan, B, Fadista, J. T, & homsen, B. Global assessment of genomic variation in cattle by genome resequencing and high-throughput genotyping [J]. BMC Genomics, 2011, 12(2): 557.
[18] 许宇静, 张煜坤, 沈雪梅, 等. 采用Illumina MiSeq测序技术分析断奶幼兔盲肠微生物群落的多样性 [J]. 动物营养学报, 2015, 27(9): 2793-2802.
XU Yujin, ZHANG Yukun, SHEN Xuemei, et al. Caecal microbiota diversity analysis of weaning rabbits using Illumina Miseq sequencing technology [J]. Journal of Animal Nutrition, 2015, 27(9): 2793-2802.
[19] Magoc, T, & Salzberg, S. L. FLASH: fast length adjustment of short reads to improve genome assemblies [J]. Bioinformatics, 2001,27(21):2957-2963.
[20] Schloss, P. D, Westcott, S. L, & Ryabin, T. Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities [J]. Applied and Environmental Microbiology, 2009, 75(23): 7537-7541.
[21] Edgar, R.C. Search and clustering orders of magnitude faster than blast [J]. Bioinformatics, 2010, 26(19): 2460-2461.
[22] Altschul, S.F, Gish, W, & Miller, W. Basic local alignment search tool [J]. Journal of Molecular Biology,1990, 215(3): 403-410.
[23] 王茹华, 张启发, 周宝利, 等. 浅析植物根分泌物与根际微生物的相互作用关系 [J]. 土壤通报, 2007, 38(1): 167-172.
WANG Ruhua, ZHANG Qifa, ZHOU Baoli, et al. Analysis on the interaction between root exudates and rhizosphere microbes [J]. Journal of Soil Science, 2007, 38(1): 167-172.
[24] Burns, K. N, Kluepfel, D. A., & Strauss, S. L. Vineyard soil bacterial diversity and composition revealed by 16S rRNA genes: differentiation by geographic features [J]. Soil Biology & Biochemistry, 2015, 91: 232-247.
[25] 何芝, 赵天涛, 邢志林, 等. 典型生活垃圾填埋场覆盖土微生物群落分析 [J]. 中国环境科学, 2015, 35(12): 3744-3753.
HE Zhi, ZHAO Tiantao, XING Zhilin, et al. Analysis of bacterial community composition in landfill cover soil [J]. China Environmental Science, 2015, 35(12): 3744-3753.
[26] 李翠霞, 李华, 金刚, 等. 新疆葡萄酒产区优良酒类酒球菌的分离、鉴定 [J]. 食品科学, 2012, 33(1): 141-146.
LI Cuixia, LI Hua, JIN Gang, et al. Isolation and identification of excellent Oenococcus oeni from Xinjiang wines [J]. Food Science, 2012, 33(1): 141-146.
[27] BLATTEL. V, WIRTH. K, & CLAUS, H. A lytic enzyme cocktail from Streptomyces sp. B578 for the control of lactic and acetic acid bacteria in wine [J]. Applied Microbiology and Biotechnology, 2009, 83(5): 839-848.
[28] Lijuan H, Kai W, Hui M, et al. Identification and Characterization of Erysiphe necator-Responsive MicroRNAs in Chinese Wild Vitis pseudoreticulata by High-Throughput Sequencing[J]. Frontiers in Plant Science, 2016,(7): 621.
[29] Ferrari, V, Dellacassa, E, & Coniberti, A. Role of grapevine vegetative expression on Aspergillus spp. incidence and OTA accumulation in wines produced in a temperate humid climate [J]. Food Additives & Contaminants: Part A: Chemistry, Analysis, Control, Exposure & Risk Assessment, 2017, 34(2): 299-306.
[30] Lachenmeier, D. W, Kanteres, F, & Kuballa, T. Ethyl carbamate in alcoholic beverages from Mexio (tequila, mescal, and stool) and Guatemala (cuxa): market survey and risk assessment [J]. International Journal of Environmental Research and Public Health, 2009, 6(1): 349-360.
[31] Sheik, C. S, Mitchell, T. W, & Rizvi, F. Z. Exposure of soil microbial communities to chromium and arsenic alters their diversity and structure [J]. PLoS One, 2012, 7(6): e40059.
[32] Martins, G., Lauga, B., & Miot-Sertire, C. Characterization of epiphytic bacterial communities from grapes, leaves, bark and soil of grapevine plants grown, and their relations[J]. PLoS One, 2013, 8(8): 73013.
[33] Wawrik, B, Kerkhof, L, & Kukor, J. Effect of different carbon sources on community composition of bacterial enrichments from soil [J]. Applied and Environmental Microbiology, 2005, 71(11): 6776-6783.
[34] Kamma, M., Mburu, H., & Blanchart, E. Effects of organic and inorganic fertilization on soil bacterial and fungal microbial diversity in the Kabete long-term trial, Kenya [J]. Biology and Fertility of Soils, 2011, 47(3): 315-321.
[35] 陈伟, 姜中武, 胡艳丽,等. 苹果园土壤微生物生态特征研究 [J]. 水土保持学报, 2008, 22(3):168-171.
CHEN Wei, JIANG Zhongwu, HU Yanli, et al. Study on biological character of rhizosphere microorganism in apple orchard [J]. Journal of Soil & Water Conservation, 2008,22(3): 168-171.
[36] 陈大勋, 李双霖, 李剑兴. 福建省主要果园土壤酶活性与微生物特性 [J]. 福建农林大学学报(自然版), 1991, (2): 207-211.
CHEN Daxun, LI Shuanglin, LI Jianxing. Enzyme activities and microbe properties of the major orchard soils in Fujian [J]. Journal of Fujian Agriculture & Forestry University, 1991 (2): 207-211.
[37] 张翔, 朱洪勋, 孙春河, 等. 长期施肥对土壤微生物和腐殖质组分的影响 [J]. 华北农学报, 1998, 13(2):87-92.
ZHANG Xiang, ZHU Hongxun, SUN Chunhe, et al. Effects of long-term fertilizer application on soil microorganisms and humus compositions [J]. Acta Agriculturae Boreall-Sinica, 1998, 13(2): 87-92.
[38] Baddam, R., Reddy, G. B., & Raczkowski, C. Activity of soil enzymes in constructed wetlands treated with swine wastewater [J]. Ecological Engineering, 2016,(91): 24-30.
[39] 魏玉洁, 邹弯, 马文瑞, 等. 应用高通量测序技术研究新疆产区葡萄果实、叶片及果园土壤微生物多样性 [J]. 食品科学, 2018, 39(6): 162-170.
WEI Yujie, ZHOU Wan, MA Wenrui, et al. Microbial diversity of berries, leaves and soil of grapevine plants grown in Xinjiang analyzed by high-throughput sequencing [J]. Food Science, 2018, 39(6): 162-170.
[40] Tan, Y, Cui, Y, & Li, H. Rhizospheric soil and root endogenous fungal diversity and composition in response to continuous Panax notoginseng cropping practices [J]. Applied and Environmental Microbiology, 2017,(194): 10-19.
[41] Cordero-Bueso, G, Arroyo, T, & Serrano, A. Influence of the farming system and vine variety on yeast communities associated with grape berries [J]. International Journal of Food Microbiology, 2011, 145(1): 132-139.
[42] 曹宏杰, 倪红伟. 土壤微生物多样性及其影响因素研究进展 [J]. 国土与自然资源研究, 2015,(3): 85-88.
CAO Hongjie, NI Hongwei. Research advances in soil microbial diversity and its impact factors [J]. Territory & Natural Resources Study, 2015,(3): 85-88.
[43] 牛世全, 龙洋, 李海云, 等. 应用IlluminaMiSeq高通量测序技术分析河西走廊地区盐碱土壤微生物多样性 [J]. 微生物学通报, 2017, 44(9): 2067-2078.
NIU Shiquan, LONG Yang, LI Haiyun, et al. Microbial diversity in saline alkali soil from Hexi Corridor analyzed by Illumina MiSeq high-throughput sequencing system [J]. Microbiology, 2017, 44(9): 2067-2078. (in Chinese)
[44] Zahran H H. Diversity, adaptation and activity of the bacterial flora in saline environments [J]. Biology and Fertility of Soils, 1997, 25(3):211-223.