留言板

尊敬的读者、作者、审稿人, 关于本刊的投稿、审稿、编辑和出版的任何问题, 您可以本页添加留言。我们将尽快给您答复。谢谢您的支持!

姓名
邮箱
手机号码
标题
留言内容
验证码

畜禽粪便堆放地土壤中抗生素抗性基因和细菌群落的垂直分布特征

韩婉雪 王凤花 柏兆海 李文彦 王新珍 马林

韩婉雪, 王凤花, 柏兆海, 李文彦, 王新珍, 马林. 畜禽粪便堆放地土壤中抗生素抗性基因和细菌群落的垂直分布特征[J]. 中国生态农业学报 (中英文), 2022, 30(2): 268−275 doi: 10.12357/cjea.20210475
引用本文: 韩婉雪, 王凤花, 柏兆海, 李文彦, 王新珍, 马林. 畜禽粪便堆放地土壤中抗生素抗性基因和细菌群落的垂直分布特征[J]. 中国生态农业学报 (中英文), 2022, 30(2): 268−275 doi: 10.12357/cjea.20210475
HAN W X, WANG F H, BAI Z H, LI W Y, WANG X Z, MA L. Vertical distribution of antibiotic resistance genes and bacterial communities in soil of livestock manure stacking site[J]. Chinese Journal of Eco-Agriculture, 2022, 30(2): 268−275 doi: 10.12357/cjea.20210475
Citation: HAN W X, WANG F H, BAI Z H, LI W Y, WANG X Z, MA L. Vertical distribution of antibiotic resistance genes and bacterial communities in soil of livestock manure stacking site[J]. Chinese Journal of Eco-Agriculture, 2022, 30(2): 268−275 doi: 10.12357/cjea.20210475

畜禽粪便堆放地土壤中抗生素抗性基因和细菌群落的垂直分布特征

doi: 10.12357/cjea.20210475
基金项目: 国家自然科学基金面上基金项目(42077358)和中国博士后科学基金项目(2020M670484)资助
详细信息
    作者简介:

    韩婉雪, 主要研究方向为微生物分子生态学。E-mail: hanwanxue17@mails.ucas.ac.cn

    通讯作者:

    王凤花, 主要研究方向为微生物分子生态学。E-mail: fhwang@sjziam.ac.cn

  • 中图分类号: S154

Vertical distribution of antibiotic resistance genes and bacterial communities in soil of livestock manure stacking site

Funds: This study was supported by the National Natural Science Foundation of China (42077358) and the China Postdoctoral Science Foundation (2020M670484).
More Information
  • 摘要: 本文通过对养殖场猪粪和鸡粪堆放地0~100 cm土壤样品的采集和分析, 研究了长期堆放畜禽粪便对土壤中抗生素抗性基因(简称“抗性基因”)和细菌群落结构垂直分布的影响。定量PCR结果表明, 与对照土壤相比, 猪粪和鸡粪堆放增加了0~100 cm土壤中四环素类抗性基因(tetCtetGtetLtetW)、磺胺类抗性基因(sulIsulII)以及整合酶基因(intI1)的检出率和检出丰度, 说明粪肥堆放造成堆放地土壤中抗性基因污染。聚类分析结果表明, 抗性基因和intI1基因的丰度随土壤深度呈递减趋势, 且主要集中在0~30 cm土层, 表明堆放地土壤中抗性基因存在向下层土壤迁移的风险。相关性分析表明, intI1基因分别与抗性基因呈显著正相关, 说明intI1基因可能在抗性基因传播中起着重要作用。同时高通量测序结果表明, 与对照土壤相比, 猪粪和鸡粪堆放显著降低了0~10 cm和10~30 cm土层细菌群落结构的多样性, 0~30 cm土层中, 猪粪和鸡粪堆放地土壤中细菌群落结构与对照土壤的差异要高于深层土壤。此外, 方差分解分析结果表明, 土壤化学性质和细菌群落结构均影响了土壤中抗性基因的垂直分布, 且细菌群落结构的变化是其主要的影响因素。本研究可为控制畜禽粪便堆放地土壤中抗性基因污染提供科学依据。
  • 图  1  不同处理不同深度土壤tetWtetCtetGtetLsulIsulIIintI1基因的相对丰度(基因拷贝数/16S rRNA基因拷贝数)

    每一行代表一个处理, 每一列代表一个基因。基因相对丰度的数据经过lg转换。CK: 对照; PM: 猪粪堆放地; CM: 鸡粪堆放地。A row is one treatment, and a column is one target gene. The data is a lg scale of gene relative abundance. CK: control; PM: pig manure stacking site; CM: chicken manure stacking site.

    Figure  1.  Relative abundances (gene copy number/16S rRNA gene copy number) of tetW, tetC, tetG, tetL, sulI, sulII and intI1 genes in different soil depths of different treatments

    图  2  不同处理不同土层细菌优势门分类水平的相对丰度

    CK: 对照; PM: 猪粪堆放地; CM: 鸡粪堆放地。CK: control; PM: pig manure stacking site; CM: chicken manure stacking site.

    Figure  2.  Relative abundances of the dominant bacterial phyla in different soil layers of different treatments

    图  3  不同处理不同土层细菌α多样性(A)和基于Bray-Curtis距离的主坐标分析(PCoA)的细菌群落结构组成(B)

    CK: 对照; PM: 猪粪堆放地; CM: 鸡粪堆放地。CK: control; PM: pig manure stacking site; CM: chicken manure stacking site.

    Figure  3.  Soil bacterial α diversity (A) and principal coordinate analysis (PCoA) based on the Bray-Curtis distance (B) showing the profile of bacterial community structure in different soil layers of different treatments

    图  4  细菌群落结构和土壤化学性质对抗性基因分布的方差分解分析(VPA) (A)以及intI1基因丰度、抗性基因丰度和土壤化学性质间的Pearson相关性分析(B)

    右图中, SOM、TN和TC分别为有机质含量、总氮含量和总碳含量。圆圈大小代表相关系数大小。蓝色表示正相关, 红色表示负相关。星号代表存在显著相关性。*: P<0.05; **: P<0.01; ***: P<0.001。In the right figure, SOM, TN and TC are contents of organic matter, total nitrogen and total carbon. The circle size represents the correlation coefficient. Blue indicates a positive correlation, and red indicates a negative correlation. The asterisk represents a significant correlation. *: P<0.05; **: P<0.01; ***: P<0.001.

    Figure  4.  Effects of bacterial community and soil chemical properties on the variations of resistance genes (A), and Pearson’s correlation among the intI1 gene abundance, resistance gene abundances, and soil chemical properties (B)

    表  1  不同畜禽粪便堆放地的土壤化学性质

    Table  1.   Soil chemical properties in different treatments

    土层深度
    Soil depth (cm)
    处理
    Treatment
    pH有机质含量
    Organic matter content (g∙kg−1)
    总碳含量
    Total carbon content (g∙kg−1)
    总氮含量
    Total nitrogen content (g∙kg−1)
    碳氮比
    C/N
    0~10CK7.87±0.07b17.81±1.89b15.00±0.20b1.07±0.06a14.08±0.63b
    PM8.76±0.28a29.48±5.18a20.27±2.83a1.43±0.42a14.53±2.02b
    CM8.00±0.17b13.72±2.36b19.60±1.56a1.53±0.45a17.08±4.16a
    10~30CK7.90±0.05b13.89±1.03a13.30±0.44b0.87±0.06a15.38±0.85b
    PM8.54±0.32a17.21±2.90a15.80±1.33ab0.97±0.21a16.64±2.03b
    CM8.40±0.13a7.94±2.02b18.62±3.97a0.65±0.25a30.45±7.71a
    30~50CK7.90±0.10a6.92±1.12c9.73±0.85b0.53±0.06b18.28±0.49a
    PM8.00±0.19a9.71±1.49b13.13±2.20a0.67±0.12b19.75±1.72a
    CM7.93±0.10a14.17±0.88a12.30±0.87ab1.23±0.06a10.00±1.01b
    50~70CK7.88±0.08a8.46±0.44a10.77±0.74a0.53±0.06b20.40±3.14a
    PM7.92±0.04a8.73±0.92a13.13±2.21a0.60±0.10ab21.90±1.39a
    CM7.73±0.04b7.93±1.84a5.23±0.12b0.77±0.15a7.03±1.58b
    70~100CK7.95±0.10b7.96±0.30a16.37±0.96a0.40±0.00b40.92±2.40a
    PM8.15±0.12a8.27±0.81a18.67±3.76a0.47±0.12ab36.29±0.06a
    CM7.68±0.08c6.42±0.30b4.63±0.06b0.60±0.17a8.12±2.05b
      CK: 对照; PM: 猪粪堆放地; CM: 鸡粪堆放地。不同字母代表同一土层各处理间差异显著(P<0.05)。CK: control; PM: pig manure stacking site; CM: chicken manure stacking site. Different letters represent significant differences among treatments in the same soil layer (P<0.05).
    下载: 导出CSV
  • [1] ZHAO Y, SU J Q, AN X L, et al. Feed additives shift gut microbiota and enrich antibiotic resistance in swine gut[J]. Science of the Total Environment, 2018, 621: 1224−1232 doi: 10.1016/j.scitotenv.2017.10.106
    [2] ZHANG Q Q, YING G G, PAN C G, et al. Comprehensive evaluation of antibiotics emission and fate in the river basins of China: source analysis, multimedia modeling, and linkage to bacterial resistance[J]. Environmental Science & Technology, 2015, 49(11): 6772−6782 doi: 10.1016/j.ecoenv.2019.02.023
    [3] WANG J, BEN W W, YANG M, et al. Dissemination of veterinary antibiotics and corresponding resistance genes from a concentrated swine feedlot along the waste treatment paths[J]. Environmental Science & Technolog, 2016, 92/93: 317−323 doi: 10.1016/j.envint.2016.04.020
    [4] HEUER H, SCHMITT H, SMALLA K. Antibiotic resistance gene spread due to manure application on agricultural fields[J]. Current Opinion in Microbiology, 2011, 14(3): 236−243 doi: 10.1016/j.mib.2011.04.009
    [5] QIAO M, YING G G, SINGER A C, et al. Review of antibiotic resistance in China and its environment[J]. Environment International, 2018, 110: 160−172 doi: 10.1016/j.envint.2017.10.016
    [6] 石晓晓, 郑国砥, 高定, 等. 中国畜禽粪便养分资源总量及替代化肥潜力[J]. 资源科学, 2021, 43(2): 403−411 doi: 10.18402/resci.2021.02.17

    SHI X X, ZHENG G D, GAO D, et al. Quantity of available nutrient in livestock manure and its potential of replacing chemical fertilizers in China[J]. Resources Science, 2021, 43(2): 403−411 doi: 10.18402/resci.2021.02.17
    [7] 刘春, 刘晨阳, 王济民, 等. 我国畜禽粪便资源化利用现状与对策建议[J]. 中国农业资源与区划, 2021, 42(2): 35−43

    LIU C, LIU C Y, WANG J M, et al. The current situation of resource utilization of livestock and popultry manure in China and the countermeasures and suggestions[J]. Chinese Journal of Agricultural Resources and Regional Planning, 2021, 42(2): 35−43
    [8] JI X L, SHEN Q H, LIU F, et al. Antibiotic resistance gene abundances associated with antibiotics and heavy metals in animal manures and agricultural soils adjacent to feedlots in Shanghai; China[J]. Journal of Hazardous Materials, 2012, 235/236: 178−185 doi: 10.1016/j.jhazmat.2012.07.040
    [9] WICHMANN F, UDIKOVIC-KOLIC N, ANDREW S, et al. Diverse antibiotic resistance genes in dairy cow manure[J]. mBio, 2014, 5(2): e01017
    [10] TANG X J, LOU C L, WANG S X, et al. Effects of long-term manure applications on the occurrence of antibiotics and antibiotic resistance genes (ARGs) in paddy soils: Evidence from four field experiments in south of China[J]. Soil Biology and Biochemistry, 2015, 90: 179−187 doi: 10.1016/j.soilbio.2015.07.027
    [11] 綦峥, 齐越, 李芳, 等. 畜牧场土壤中重金属与抗生素抗性基因的分布规律研究[J]. 生态毒理学报, 2021, 16(1): 204−214

    QI Z, QI Y, LI F, et al. Distribution of heavy metals and antibiotic resistance genes in the soil of livestock farms[J]. Asian Journal of Ecotoxicology, 2021, 16(1): 204−214
    [12] CHEN Q, AN X, LI H, et al. Long-term field application of sewage sludge increases the abundance of antibiotic resistance genes in soil[J]. Environment International, 2016, 92/93: 1−10 doi: 10.1016/j.envint.2016.03.026
    [13] FORSBERG K J, PATEL S, GIBSON M K, et al. Bacterial phylogeny structures soil resistomes across habitats[J]. Nature, 2014, 509(7502): 612−616 doi: 10.1038/nature13377
    [14] SU J Q, WEI B, OUYANG W Y, et al. Antibiotic resistome and its association with bacterial communities during sewage sludge composting[J]. Environmental Science & Technology, 2015, 49(12): 7356−7363
    [15] WANG F H, CHEN S M, QIN S P, et al. Long-term nitrogen fertilization alters microbial community structure and denitrifier abundance in the deep vadose zone[J]. Journal of Soils and Sediments, 2021, 21(6): 2394−2403 doi: 10.1007/s11368-021-02931-0
    [16] HAN X M, HU H W, SHI X Z, et al. Impacts of reclaimed water irrigation on soil antibiotic resistome in urban parks of Victoria, Australia[J]. Environmental Pollution, 2016, 211: 48−57 doi: 10.1016/j.envpol.2015.12.033
    [17] HAN X M, HU H W, CHEN Q L, et al. Antibiotic resistance genes and associated bacterial communities in agricultural soils amended with different sources of animal manures[J]. Soil Biology and Biochemistry, 2018, 126: 91−102 doi: 10.1016/j.soilbio.2018.08.018
    [18] ZHANG R R, GU J, WANG X J, et al. Contributions of the microbial community and environmental variables to antibiotic resistance genes during co-composting with swine manure and cotton stalks[J]. Journal of Hazardous Materials, 2018, 358: 82−91 doi: 10.1016/j.jhazmat.2018.06.052
    [19] GOU C L, WANG Y Q, ZHANG X Q, et al. Effects of chlorotetracycline on antibiotic resistance genes and the bacterial community during cattle manure composting[J]. Bioresource Technology, 2021, 323: 124517 doi: 10.1016/j.biortech.2020.124517
    [20] 鲁如坤. 土壤农业化学分析方法[M]. 北京: 中国农业科技出版社, 2000

    LU R K. Soil Agricultural Chemical Analysis Method[M]. Beijing: China Agricultural Science and Technology Press, 2000
    [21] WANG F H, QIAO M, CHEN Z, et al. Antibiotic resistance genes in manure-amended soil and vegetables at harvest[J]. Journal of Hazardous Materials, 2015, 299: 215−221 doi: 10.1016/j.jhazmat.2015.05.028
    [22] SUZUKI M T, TAYLOR L T, DELONG E F. Quantitative analysis of small-subunit rRNA genes in mixed microbial populations via 5'-nuclease assays[J]. Applied and Environmental Microbiology, 2000, 66(11): 4605−4614 doi: 10.1128/AEM.66.11.4605-4614.2000
    [23] WALTERS W, HYDE E R, BERG-LYONS D, et al. Improved bacterial 16S rRNA gene (V4 and V4-5) and fungal internal transcribed spacer marker gene primers for microbial community surveys[J]. mSystems, 2016, 1(1 doi: 10.1128/msystems.00009-1
    [24] BOLYEN E, RIDEOUT J R, DILLON M R, et al. Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2[J]. Nature Biotechnology, 2019, 37(8): 852−857 doi: 10.1038/s41587-019-0209-9
    [25] CALLAHAN B J, MCMURDIE P J, ROSEN M J, et al. DADA2: High-resolution sample inference from Illumina amplicon data[J]. Nature Methods, 2016, 13(7): 581−583 doi: 10.1038/nmeth.3869
    [26] GLÖCKNER F O, YILMAZ P, QUAST C, et al. 25 years of serving the community with ribosomal RNA gene reference databases and tools[J]. Journal of Biotechnology, 2017, 261: 169−176 doi: 10.1016/j.jbiotec.2017.06.1198
    [27] OKSANEN J, BLANCHET F, FRIENDLY M, et al. Vegan: Community Ecology Package. R package version 2.4–3[EB/OL]. Vienna, AU: R Core Development Team. [2017]. https://CRAN.R-project.org/package=vegan
    [28] UDIKOVIC-KOLIC N, WICHMANN F, BRODERICK N A, et al. Bloom of resident antibiotic-resistant bacteria in soil following manure fertilization[J]. PNAS, 2014, 111(42): 15202−15207 doi: 10.1073/pnas.1409836111
    [29] CHENG W, CHEN H, SU C, et al. Abundance and persistence of antibiotic resistance genes in livestock farms: A comprehensive investigation in eastern China[J]. Environment International, 2013, 61: 1−7 doi: 10.1016/j.envint.2013.08.023
    [30] LIU M S, ZHANG W H, WANG X G, et al. Nitrogen leaching greatly impacts bacterial community and denitrifiers abundance in subsoil under long-term fertilization[J]. Agriculture, Ecosystems & Environment, 2020, 294: 106885
    [31] WANG F H, HAN W X, CHEN S M, et al. Fifteen-year application of manure and chemical fertilizers differently impacts soil ARGs and microbial community structure[J]. Frontiers in Microbiology, 2020, 11: 62 doi: 10.3389/fmicb.2020.00062
    [32] ZHANG R M, LIU X, WANG S L, et al. Distribution patterns of antibiotic resistance genes and their bacterial hosts in pig farm wastewater treatment systems and soil fertilized with pig manure[J]. Science of the Total Environment, 2021, 758: 143654 doi: 10.1016/j.scitotenv.2020.143654
    [33] MA L P, LI A D, YIN X L, et al. The prevalence of integrons as the carrier of antibiotic resistance genes in natural and man-made environments[J]. Environmental Science & Technology, 2017, 51(10): 5721−5728 doi: 10.1021/acs.est.6b05887
  • 加载中
图(4) / 表(1)
计量
  • 文章访问数:  198
  • HTML全文浏览量:  69
  • PDF下载量:  27
  • 被引次数: 0
出版历程
  • 收稿日期:  2021-07-19
  • 录用日期:  2021-09-03
  • 网络出版日期:  2021-11-09
  • 刊出日期:  2022-02-08

目录

    /

    返回文章
    返回