硫化物抑制潮土反硝化过程中氧化亚氮还原的菌群机制

隋维康1 李冀1 吴晓刚1 吴巧玉1 马怡茗1 张馨玉1 张晓君1

(1.上海交通大学生命科学技术学院微生物代谢国家重点实验室, 上海 200240)

【摘要】【背景】土壤中的反硝化作用形成气态产物N2O和N2,会导致氮素的气态损失,并造成温室效应。硫化物对土壤的N2O还原具有抑制作用,但其对菌群和功能基因的影响机制还不清楚。【目的】研究有无外加碳源情况下,硫化物对反硝化作用中间产物(NO、N2O)的积累、反硝化功能基因(narG、nirS、nirK和nosZ)表达量以及菌群结构的影响。【方法】分别设置不同量葡萄糖(0和1 000 mg-C/kg干重土壤)和硫化钠(0和150 mg-S/kg干重土壤)添加的交叉处理,进行室内微宇宙培养实验,利用自动化培养与实时气体检测系统检测培养过程中NO、N2O和N2的积累量,通过反转录定量PCR测定反硝化功能基因表达量,利用MiSeq技术平台基于16S rRNA基因序列的高通量测序分析样品的菌群结构。【结果】硫化钠的添加显著抑制N2O还原,但是其对于N2O积累量没有显著影响,却显著降低了NO的积累量。硫化钠的添加短时间内在转录水平上显著抑制N2O还原酶的活性,并且抑制固氮弧菌属(Azoarcus)、微枝形杆菌属(Microvirga)、剑菌属(Ensifer)、氮氢单胞菌属(Azohydromonas)、芽孢杆菌属(Bacillus)、斯科曼氏球菌属(Skermanella)、申氏杆菌属(Shinella)和西索恩氏菌属(Chthoniobacter)的基因转录,降低它们的转录本丰度,结合Kyoto Encyclopedia of Genes and Genomes (KEGG)数据库的查询结果,发现硫化钠的添加抑制了不产生N2O的N2O还原反硝化细菌的生长。【结论】堆肥或其他原因引起的土壤硫化物增加,导致反硝化过程N2O还原被抑制的原因是由于其对氧化亚氮基因转录的抑制和对不同反硝化菌的选择作用,研究结果有助于认识硫化物对氮代谢影响的微生物机制。

【关键词】 硫化物; 反硝化; 氧化亚氮还原; 菌群结构; 潮土;

【DOI】

【基金资助】 国家自然科学基金(31670105,31971526,31861133018) National Natural Science Foundation of China (31670105;31971526;31861133018)

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    References

    [1]Wang SY,Liu WY,Zhao SY,et al.Denitrification is the main microbial N loss pathway on the Qinghai-Tibet Plateau above an elevation of 5000m[J].Science of the Total Environment,2019,696:133852

    [2]Conthe M,Wittorf L,Kuenen JG,et al.Life on N2O:deciphering the ecophysiology of N2O respiring bacterial communities in a continuous culture[J].The ISME Journal,2018,12(4):1142-1153

    [3]Ji MM,Wu XG,Wu XX,et al.Effect of overuse nitrogen fertilizer on bacterial community and N2O emission from greenhouse soil[J].Microbiology China,2018,45(6):1323-1332 (in Chinese)

    [4]Ravishankara AR,Daniel JS,Portmann RW.Nitrous oxide(N2O):the dominant ozone-depleting substance emitted in the 21st century[J].Science,2009,326(5949):123-125

    [5]Tiedje JM,Sexstone AJ,Myrold DD,et al.Denitrification:ecological niches,competition and survival[J].Antonie van Leeuwenhoek,1982,48(6):569-583

    [6]Schmidt CS,Richardson DJ,Baggs EM.Constraining the conditions conducive to dissimilatory nitrate reduction to ammonium in temperate arable soils[J].Soil Biology and Biochemistry,2011,43(7):1607-1611

    [7]Liu Y,Wang JL.Reduction of nitrate by zero valent iron(ZVI)-based materials:a review[J].Science of the Total Environment,2019,671:388-403

    [8]Tugtas AE,Pavlostathis SG.Electron donor effect on nitrate reduction pathway and kinetics in a mixed methanogenic culture[J].Biotechnology and Bioengineering,2007,98(4):756-763

    [9]Akunna JC,Bizeau C,Moletta R.Nitrate and nitrite reductions with anaerobic sludge using various carbon sources:glucose,glycerol,acetic acid,lactic acid and methanol[J].Water Research,1993,27(8):1303-1312

    [10]Dong LF,Sobey MN,Smith CJ,et al.Dissimilatory reduction of nitrate to ammonium,not denitrification or anammox,dominates benthic nitrate reduction in tropical estuaries[J].Limnology and Oceanography,2011,56(1):279-291

    [11]Ogilvie BG,Rutter M,Nedwell DB.Selection by temperature of nitrate-reducing bacteria from estuarine sediments:species composition and competition for nitrate[J].FEMS Microbiology Ecology,1997,23(1):11-22

    [12]Brunet RC,Garcia-Gil LJ.Sulfide-induced dissimilatory nitrate reduction to ammonia in anaerobic freshwater sediments[J].FEMS Microbiology Ecology,1996,21(2):131-138

    [13]Mazéas L,Vigneron V,Le-Ménach K,et al.Elucidation of nitrate reduction pathways in anaerobic bioreactors using a stable isotope approach[J].Rapid Communications in Mass Spectrometry,2008,22(11):1746-1750

    [14]Behrendt A,de Beer D,Stief P.Vertical activity distribution of dissimilatory nitrate reduction in coastal marine sediments[J].Biogeosciences,2013,10(11):7509-7523

    [15]He TT.Effects of garbage compost and its compound fertilizer on farmland CO2 and CH4 emissions flux[D].Beijing:Master’s Thesis of Capital Normal University,2007(in Chinese)

    [16]Li LJ.Effects of long-term compost application on soil health in Quzhou station[D].Beijing:Doctoral Dissertation of China Agricultural University,2017 (in Chinese)

    [17]He TT,Hua L,Xu ZJ,et al.Influence of urban waste compost and compound fertilizer based on waste compost on flux of CO2 and CH4 from agricultural soils[J].Journal of Agro-Environment Science,2007,26(3):1153-1158 (in Chinese)

    [18]Yang LC,Cosolini SI.A case study on converting organic farm waste vegetables to biogas using a cartridge design anaerobic digester[J].Applied Biochemistry and Biotechnology,2019,189(2):638-646

    [19]Janssen AJH,Lens PNL,Stams AJM,et al.Application of bacteria involved in the biological sulfur cycle for paper mill effluent purification[J].Science of the Total Environment,2008,407(4):1333-1343

    [20]Liu CS,Zhao DF,Yan LH,et al.Elemental sulfur formation and nitrogen removal from wastewaters by autotrophic denitrifiers and anammox bacteria[J].Bioresource Technology,2015,191:332-336

    [21]Matsuto T,Zhang X,Matsuo T,et al.Onsite survey on the mechanism of passive aeration and air flow path in a semi-aerobic landfill[J].Waste Management,2015,36:204-212

    [22]Montalvo S,Huiliñir C,Borja R,et al.Anaerobic digestion of wastewater rich in sulfate and sulfide:effects of metallic waste addition and micro-aeration on process performance and methane production[J].Journal of Environmental Science and Health,Part A:Toxic/Hazardous Substances and Environmental Engineering,2019,54(10):1035-1043

    [23]Liang J.Disintegration dynamics of different green manures and their effects on soil nutrients[D].Changsha:Master’s Thesis of Hunan Agricultural University,2018 (in Chinese)

    [24]Zhang XQ,Jing DD,Yang ZP,et al.Improvement of green manure alfalfa appropriate returned on dryland wheat yield and soil nutrient[J].Acta Agriculturae Boreali-Sinica,2019,34(S1):221-227 (in Chinese)

    [25]Qu HK,Gao YH,Han JR.Study on the reducing effect of nitrogen fertilizer in rice under returning alfalfa[J].Agricultural Equipment&Technology,2018,44(1):22-24(in Chinese)

    [26]Decatanzaro JB,Beauchamp EG,Drury CF.Denitrification vs dissimilatory nitrate reduction in soil with alfalfa,straw,glucose and sulfide treatments[J].Soil Biology and Biochemistry,1987,19(5):583-587

    [27]Rütting T,Boeckx P,Müller C,et al.Assessment of the importance of dissimilatory nitrate reduction to ammonium for the terrestrial nitrogen cycle[J].Biogeosciences,2011,8(7):1779-1791

    [28]Wisniewski K,di Biase A,Munz G,et al.Kinetic characterization of hydrogen sulfide inhibition of suspended anammox biomass from a membrane bioreactor[J].Biochemical Engineering Journal,2019,143:48-57

    [29]Qiu SJ,Ju XT,Lu X,et al.Improved nitrogen management for an intensive winter wheat/summer maize double-cropping system[J].Soil Science Society of America Journal,2012,76(1):286-297

    [30]Tam TY,Knowles R.Effects of sulfide and acetylene on nitrous oxide reduction by soil and by Pseudomonas aeruginosa[J].Canadian Journal of Microbiology,1979,25(10):1133-1138

    [31]Lu WW,Zhang HL,Shi WM.Dissimilatory nitrate reduction to ammonium in an anaerobic agricultural soil as affected by glucose and free sulfide[J].European Journal of Soil Biology,2013,58:98-104

    [32]Caffrey JM,Bonaglia S,Conley DJ.Short exposure to oxygen and sulfide alter nitrification,denitrification,and DNRA activity in seasonally hypoxic estuarine sediments[J].FEMS Microbiology Letters,2019,366(1):fny288

    [33]Lu H,Huang HQ,Yang WM,et al.Elucidating the stimulatory and inhibitory effects of dissolved sulfide on sulfur-oxidizing bacteria (SOB) driven autotrophic denitrification[J].Water Research,2018,133:165-172

    [34]Molstad L,Dörsch P,Bakken LR.Robotized incubation system for monitoring gases (O2,NO,N2O N2) in denitrifying cultures[J].Journal of Microbiological Methods,2007,71(3):202-211

    [35]Yang LQ,Zhang XJ,Ju XT.Linkage between N2O emission and functional gene abundance in an intensively managed calcareous fluvo-aquic soil[J].Scientific Reports,2017,7:43283

    [36]Liu BB,FrostegårdÅ,Bakken LR.Impaired reduction of N2O to N2 in acid soils is due to a posttranscriptional interference with the expression of nos Z[J].m Bio,2014,5(3):e01383-14

    [37]Dees PM,Ghiorse WC.Microbial diversity in hot synthetic compost as revealed by PCR-amplified r RNA sequences from cultivated isolates and extracted DNA[J].FEMSMicrobiology Ecology,2001,35(2):207-216

    [38]Yang LQ.N2O production processes and the abundance and expression of relative functional genes in calcareous fluvo aquic soil[D].Beijing:Doctoral Dissertation of China Agricultural University,2017 (in Chinese)

    [39]Nadkarni MA,Martin FE,Jacques NA,et al.Determination of bacterial load by real-time PCR using a broad-range(universal) probe and primers set[J].Microbiology,2002,148(1):257-266

    [40]Bru D,Sarr A,Philippot L.Relative abundances of proteobacterial membrane-bound and periplasmic nitrate reductases in selected environments[J].Applied and Environmental Microbiology,2007,73(18):5971-5974

    [41]Kandeler E,Deiglmayr K,Tscherko D,et al.Abundance of nar G,nir S,nir K,and nos Z genes of denitrifying bacteria during primary successions of a glacier foreland[J].Applied and Environmental Microbiology,2006,72(9):5957-5962

    [42]Henry S,Baudoin E,López-Gutiérrez JC,et al.Quantification of denitrifying bacteria in soils by nir K gene targeted real-time PCR[J].Journal of Microbiological Methods,2004,59(3):327-335

    [43]Hallin S,Lindgren PE.PCR detection of genes encoding nitrite reductase in denitrifying bacteria[J].Applied and Environmental Microbiology,1999,65(4):1652-1657

    [44]Henry S,Bru D,Stres B,et al.Quantitative detection of the nos Z gene,encoding nitrous oxide reductase,and comparison of the abundances of 16S r RNA,nar G,nir K,and nos Z genes in soils[J].Applied and Environmental Microbiology,2006,72(8):5181-5189

    [45]Zhang QP,Wu YQ,Wang J,et al.Accelerated dysbiosis of gut microbiota during aggravation of DSS-induced colitis by a butyrate-producing bacterium[J].Scientific Reports,2016,6:27572

    [46]Moeller AH,Suzuki TA,Phifer-Rixey M,et al.Transmission modes of the mammalian gut microbiota[J].Science,2018,362(6413):453-457

    [47]Segata N,Izard J,Waldron L,et al.Metagenomic biomarker discovery and explanation[J].Genome Biology,2011,12(6):R60

    [48]Zhang X,Davidson EA,Mauzerall DL,et al.Managing nitrogen for sustainable development[J].Nature,2015,528(7580):51-59

    [49]Yoon S,Song B,Phillips RL,et al.Ecological and physiological implications of nitrogen oxide reduction pathways on greenhouse gas emissions in agroecosystems[J].FEMS Microbiology Ecology,2019,95(6):fiz066

    [50]Sanford RA,Wagner DD,Wu QZ,et al.Unexpected nondenitrifier nitrous oxide reductase gene diversity and abundance in soils[J].Proceedings of the National Academy of Sciences of the United States of America,2012,109(48):19709-19714

    [51]Yoon S,Nissen S,Park D,et al.Nitrous oxide reduction kinetics distinguish bacteria harboring clade I Nos Z from those harboring clade II Nos Z[J].Applied and Environmental Microbiology,2016,82(13):3793-3800

    [52]Hein S,Witt S,Simon J.Clade II nitrous oxide respiration of Wolinella succinogenes depends on the Nos G,-C1,-C2,-H electron transport module,Nos B and a Rieske/cytochrome bc complex[J].Environmental Microbiology,2017,19(12):4913-4925

    [53]Aelion CM,Warttinger U.Low sulfide concentrations affect nitrate transformations in freshwater and saline coastal retention pond sediments[J].Soil Biology and Biochemistry,2009,41(4):735-741

    [54]Aelion CM,Warttinger U.Sulfide Inhibition of nitrate removal in coastal sediments[J].Estuaries and Coasts,2010,33(3):798-803

    [55]Sørensen J,Tiedje JM,Firestone RB,et al.Inhibition by sulfide of nitric and nitrous oxide reduction by denitrifying Pseudomonas fluorescens[J].Applied and Environmental Microbiology,1980,39(1):105-108

This Article

ISSN:0253-2654

CN: 11-1996/Q

Vol 47, No. 10, Pages 3114-3125

October 2020

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摘要

  • 1 材料与方法
  • 2 结果与分析
  • 3 讨论与结论
  • 参考文献