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石油污染胁迫下土壤潜在降污固碳微生物互作关系研究

宋佳宇 李昀照 李兴春 李丹丹 王庆宏 史权 陈春茂

宋佳宇, 李昀照, 李兴春, 李丹丹, 王庆宏, 史权, 陈春茂. 石油污染胁迫下土壤潜在降污固碳微生物互作关系研究[J]. 环境科学研究, 2023, 36(7): 1392-1403. doi: 10.13198/j.issn.1001-6929.2023.05.04
引用本文: 宋佳宇, 李昀照, 李兴春, 李丹丹, 王庆宏, 史权, 陈春茂. 石油污染胁迫下土壤潜在降污固碳微生物互作关系研究[J]. 环境科学研究, 2023, 36(7): 1392-1403. doi: 10.13198/j.issn.1001-6929.2023.05.04
SONG Jiayu, LI Yunzhao, LI Xingchun, LI Dandan, WANG Qinghong, SHI Quan, CHEN Chunmao. Potential Pollution-Reducing and Carbon-Fixing Microorganisms in Soils under Oil Pollution Stress Interaction Relationship Study[J]. Research of Environmental Sciences, 2023, 36(7): 1392-1403. doi: 10.13198/j.issn.1001-6929.2023.05.04
Citation: SONG Jiayu, LI Yunzhao, LI Xingchun, LI Dandan, WANG Qinghong, SHI Quan, CHEN Chunmao. Potential Pollution-Reducing and Carbon-Fixing Microorganisms in Soils under Oil Pollution Stress Interaction Relationship Study[J]. Research of Environmental Sciences, 2023, 36(7): 1392-1403. doi: 10.13198/j.issn.1001-6929.2023.05.04

石油污染胁迫下土壤潜在降污固碳微生物互作关系研究

doi: 10.13198/j.issn.1001-6929.2023.05.04
基金项目: 中国石油天然气集团有限公司科学研究与技术开发项目(No.2021DQ03-A4);中国石油科技创新基金项目(No.2023DQ02-0506)
详细信息
    作者简介:

    宋佳宇(1985-),女,天津人,高级工程师,硕士,主要从事石油污染生物修复技术研究,songjiayu@cnpc.com.cn

  • 中图分类号: X172

Potential Pollution-Reducing and Carbon-Fixing Microorganisms in Soils under Oil Pollution Stress Interaction Relationship Study

Funds: Scientific Research and Technology Development Program of China National Petroleum Corporation (No.2021DQ03-A4);China National Petroleum Corporation Innovation Foundation (No.2023DQ02-0506)
  • 摘要: 为探究石油污染土壤潜在降污固碳微生物关键类群及其互作响应关系,采集华北某油田开发井场表层(0~20 cm)土壤,利用荧光定量PCR及高通量测序技术开展石油污染土壤潜在降污固碳微生物群落结构及代谢功能研究. 结果表明:①石油污染胁迫下土壤微生物优势菌门为变形菌门(Proteobacteria)、放线菌门(Actinobacteriota)、绿弯菌门(Chloroflexi)、厚壁菌门(Firmicutes)、酸杆菌门(Acidobacteriota)和拟杆菌门(Bacteroidota). ②相关性网络分析表明,类诺卡氏菌属(Nocardioides)、链霉菌属(Streptomyces)、假单胞菌属(Pseudomonas)及鞘脂单胞菌属(Sphingomonas)是石油污染土壤降污固碳关键属. 同时,假单胞菌属与链霉菌属(r=−0.818,p=0.001)、类诺卡氏菌属(r=−0.811,p=0.001)以竞争关系共存;鞘脂单胞菌属与链霉菌属(r=0.895,p<0.001)、类诺卡氏菌属(r=0.916,p<0.001)以互利共生关系共存,链霉菌属与类诺卡氏菌属(r=0.895,p<0.001)以互利共生关系共存. ③Spearman相关性分析表明,烷烃降解功能基因alkB丰度与固碳功能基因cbbL(r=0.846,p=0.001)、aclB(r=0.825,p=0.001)、fhs(r=0.853,p<0.001)丰度均呈极显著正相关;芳烃降解功能基因PAH-RHDα GP丰度与固碳功能基因cbbL丰度(r=0.825,p=0.001)呈极显著正相关,与fhs(r=0.706,p=0.010)、aclB(r=0.650,p=0.022)的丰度均呈显著正相关. ④KEGG数据库功能注释结果表明,石油污染土壤中同时存在石油烃降解和固碳代谢通路,且固碳代谢通路相对丰度显著高于石油烃降解代谢通路. 研究显示,石油污染土壤中存在潜在降污固碳微生物且多为互利共生关系,石油污染胁迫下土壤微生物群落降污固碳作用可能存在协同关系.

     

  • 图  1  石油污染胁迫下土壤微生物门水平上的物种组成

    Figure  1.  Species composition at the phylum level in oil-contaminated soils

    图  2  石油污染胁迫下土壤潜在降污固碳微生物相关性网络(属水平)

    注:不同节点颜色代表不同门水平物种,每个节点代表1个属,节点大小代表物种丰度大小;连线颜色代表物种相关性,红色代表正相关,蓝色代表负相关;连线粗细代表物种之间相关程度,连线越粗,物种相关性越高.

    Figure  2.  Correlation network map of potential pollution-reducing and carbon-fixing microorganisms in oil-contaminated soil (genus level)

    图  3  石油烃降解及固碳功能基因的Spearman相关性分析

    注:样本数为12. *表示在0.05水平下(双尾)显著相关,**表示在0.01水平下(双尾)显著相关.

    Figure  3.  Spearman correlation analysis between functional genes of petroleum hydrocarbon degradation and carbon fixation

    图  4  能量代谢和外源物质生物降解功能注释

    Figure  4.  Functional annotation of energy metabolism and xenobiotics biodegradation

    表  1  土壤样品理化指标测试结果

    Table  1.   Test results of physical and chemical index analysis of soil samples

    样品编号物理指标1)化学指标2)
    微团聚体
    (0.25~2 mm)
    含量/%
    微团聚体
    (<0.25 mm)
    含量/%
    含水率pHTP含量/
    (mg/kg)
    TN含量/
    (g/kg)
    TOC含量/
    (g/kg)
    硝态氮
    含量/(mg/kg)
    铵态氮
    含量/(mg/kg)
    S110.8289.1827.41%±0.67%8.37±0.373.37±0.341.45±0.0770.30±0.850.66±0.061.58±0.01
    S214.1885.8224.45%±0.89%8.65±0.102.62±3.422.15±0.3597.40±11.030.68±0.062.62±0.05
    S39.8890.1223.59%±0.73%8.82±0.043.46±0.851.15±0.0752.30±2.550.55±0.051.49±0.05
    S413.1386.8623.59%±1.52%8.81±0.303.05±1.801.40±0.0265.35±0.490.54±0.041.27±0.04
    S513.1486.8527.58%±0.19%8.19±0.160.42±0.132.05±0.0780.55±0.783.89±0.053.12±0.07
    S629.3070.7016.61%±1.64%8.79±0.082.81±1.941.30±0.0147.50±0.020.67±0.031.50±0.02
    S715.5984.4025.90%±0.60%9.06±0.061.82±1.531.30±0.0360.40±0.140.70±0.061.76±0.05
    S815.6784.3323.27%±1.94%8.83±0.082.80±2.191.85±0.0792.95±2.760.85±0.061.67±0.11
    S913.9886.0226.77%±0.76%8.49±0.373.67±0.541.25±0.0761.80±0.420.78±0.071.71±0.03
    S1014.0985.9126.52%±0.94%8.31±0.230.63±0.261.50±0.0167.60±1.270.82±0.062.29±0.01
    S1122.4177.599.00%±0.01%8.85±0.060.16±0.233.15±0.07120.75±5.020.63±0.041.15±0.01
    S1225.4074.6017.47%±0.07%8.46±0.223.72±0.440.85±0.0737.75±2.191.24±0.071.20±0.06
    注:1)物理指标分析测试方法参照《土壤检测 第20部分:土壤微团聚体组成的测定》(NY/T 1121.20—2008);2)化学指标分析测试方法参照《土壤农业化学分析方法》[16].
    下载: 导出CSV

    表  2  土壤样品总石油烃含量

    Table  2.   Total petroleum hydrocarbon content in soil samples

    样品编号总石油烃含量/
    (mg/kg)
    样品编号总石油烃含量/
    (mg/kg)
    S143.04±9.72S7444.10±17.11
    S244.18±3.11S8478.50±11.72
    S386.43±9.06S9616.30±7.41
    S4247.53±13.51S10784.85±14.28
    S5417.20±6.19S111 027.75±15.91
    S6432.60±11.78S121 266.00±16.26
    下载: 导出CSV

    表  3  石油烃降解及固碳功能基因的引物设计

    Table  3.   Primers design of functional genes for petroleum hydrocarbon degradation and carbon fixation

    基因名称引物名称引物序列(5´-3´)qPCR产物长度/bp
    alkB alkB-1f AAYACNGCNCAYGARCTNGGNCAYAA[17] 550
    alkB-1r GCRTGRTGRTCNGARTGNCGYTG[17]
    PAH-RHDα GN PAH-RHDα GN F GAGATGCATACCACGTKGGTTGGA[18] 305
    PAH-RHDα GN R AGCTGTTGTTCGGGAAGAYWGTGCMGTT[18]
    PAH-RHDα GP PAH-RHDα GP F CGGCGCCGACAAYTTYGTNGG[18] 290
    PAH-RHDα GP R GGGGAACACGGTGCCRTGDATRAA[18]
    cbbL cbbLG1F GGCAACGTGTTCGGSTTCAA[19] 1 100
    cbbL1106R CRTGRATVCCRCCIGAIGCIACVG[20]
    cbbM RubII331f AACAACCARGGYATGGGYGA[20] 300
    RuIIR2 TGRCCIGCICGRTGRTARTGCA[21]
    aclB 892F TGGACMATGGTDGCYGGKGGT[22] 300
    1204R ATAGTTKGGSCCACCTCTTC[22]
    fhs fhs1 GTWTGGGCWAARGGYGGMGAAGG[23] 250
    FTHFS-r GTATTGDGTYTTRGCCATACA[23]
    下载: 导出CSV

    表  4  土壤样品的Alpha多样性指数

    Table  4.   Alpha diversity index of soil samples

    样品编号Sobs指数Shannon-Wiener指数Ace指数Coverage
    S12 8826.163 693.990.98
    S22 6886.173 412.540.98
    S32 4025.283 388.580.98
    S42 3915.743 150.500.98
    S51 3024.541 845.580.99
    S62 3006.192 869.560.98
    S72 3114.073 244.610.98
    S82 4804.183 517.450.98
    S92 7945.983 709.610.98
    S103 0156.533 833.540.98
    S112 0916.282 512.450.99
    S121 7705.472 283.400.99
    下载: 导出CSV

    表  5  石油污染胁迫下土壤潜在降污固碳微生物网络节点属性

    Table  5.   Attribute table of potential pollution-reducing and carbon-fixing microbial network nodes in oil-contaminated soil

    节点物种名称节点物种
    所属门
    连通性度中
    心性
    Acinetobacter Proteobacteria 2 0.08
    Arthrobacter Actinobacteriota 7 0.28
    Bacillus Firmicutes 2 0.08
    Blastococcus Actinobacteriota 5 0.20
    C1-B045 Proteobacteria 1 0.04
    Cavicella Proteobacteria 1 0.04
    Dietzia Actinobacteriota 2 0.08
    Georgenia Actinobacteriota 2 0.08
    Gordonia Actinobacteriota 2 0.08
    Hydrogenophaga Proteobacteria 2 0.08
    KCM-B-112 Proteobacteria 2 0.08
    Marmoricola Actinobacteriota 7 0.28
    MND1 Proteobacteria 6 0.24
    Nocardioides Actinobacteriota 6 0.24
    norank_f_67-14 Actinobacteriota 5 0.20
    norank_f_JG30-KF-CM45 Chloroflexi 3 0.12
    norank_f_norank_o_norank_c_KD4-96 Chloroflexi 4 0.16
    norank_f_norank_o_Vicinamibacterales Acidobacteriota 5 0.20
    norank_f_Rhodocyclaceae Proteobacteria 1 0.04
    norank_f_Vicinamibacteraceae Acidobacteriota 3 0.12
    Pseudomonas Proteobacteria 4 0.16
    RB41 Acidobacteriota 3 0.12
    Skermanella Proteobacteria 3 0.12
    Sphingomonas Proteobacteria 4 0.16
    Streptomyces Actinobacteriota 8 0.32
    Thiobacillus Proteobacteria 2 0.08
    下载: 导出CSV

    表  6  石油烃降解及固碳功能基因丰度

    Table  6.   Abundance of functional genes for petroleum hydrocarbon degradation and carbon fixation

    样品编号功能基因丰度
    cbbLcbbMaclBfhsalkBPAH-RHDα GNPAH-RHDα GP
    S18.87×1042.79×1074.38×1063.31×1067.02×1041.28×1061.59×105
    S22.59×1042.94×1061.08×1061.22×1061.24×1042.83×1051.02×104
    S31.03×1054.75×1072.42×1061.52×1061.97×1054.31×1067.98×104
    S42.15×1051.15×1077.28×1061.04×1075.00×1051.92×1063.41×106
    S58.90×1041.33×1077.67×1061.09×1072.67×1052.46×1062.49×104
    S63.15×1054.25×1067.12×1061.11×1072.54×1055.08×1043.29×105
    S74.68×1047.40×1062.63×1061.43×1068.66×1041.13×1063.16×104
    S81.43×1051.95×1075.66×1064.15×1063.72×1053.11×1061.16×105
    S98.87×1042.76×1073.26×1062.99×1068.52×1041.02×1067.48×104
    S103.10×1052.37×1071.36×1079.83×1062.09×1057.75×1052.76×105
    S118.68×1051.46×1082.35×1072.39×1075.19×1057.31×1047.32×105
    S123.17×1055.15×1072.71×1071.69×1085.33×1051.06×1052.83×105
    下载: 导出CSV

    表  7  6个土壤样品中一级代谢通路的相对丰度

    Table  7.   Relative abundance of primary metabolic pathways of six samples

    一级代谢通路相对丰度/%
    S1S3S4S10S11S12
    细胞过程5.105.014.644.624.754.48
    环境信息处理5.445.465.235.205.255.12
    遗传信息处理5.565.535.346.095.495.64
    人类疾病4.044.674.163.694.474.03
    新陈代谢77.3376.7277.9278.0377.4978.10
    机体系统2.532.622.722.372.552.63
    下载: 导出CSV
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