采油废水厌氧处理系统的微生物群落特征

83Vol．8No．320143ChineseJournalofEnvironmentalEngineeringMar．201412341．1000072．1634533．1000854．063020。DGGE。MethanomethylovoransthermophilaMethanolineatarda。M．tarda。。EpsilonproteobacteriaNitrospiraDeltaproteobacteria。SyntrophusDeltaproteobacteriagroupTA。。X703A1673-9108201403-0807-08CharacteristicsofmicrobialcommunityinanaerobicproducedwatertreatmentsystemMuJian1KuangLi2GaoYingxin3ZhangDandan41．PetroChinaCompanyLimitedBeijing100007China2．EnvironmentalMonitoringandEvaluationCenterDaqingOilfieldCo．Ltd．Daqing163453China3．StateKeyLaboratoryofEnvironmentalAquaticChemistryＲesearchCenterforEco-EnvironmentalSciencesChineseAcademyofSciencesBeijing100085China4．TangshanGraitTechnologyCo．Ltd．Tangshan063020ChinaAbstractMicrobialcharacteristicsintheanaerobictankofafull-scaleproducedwatertreatmentplantcapableofanaerobichydrocarbonremovalwereanalyzedandcomparedtothoseintheinfluentproducedwaterusingcultiva-tion-independentmolecularmethods．BoththeDGGEandcloninganalysisindicatedthatthemethanogensintheanae-robicbiofilmwasderivedfromtheinfluentproducedwater．ItisfoundthatthearchaealcommunityintheanaerobictankandtheproducedwastewaterwasmainlycomposedofmethanogensincludingtheMethanomethylovoransther-mophilaandMethanolineatarda．Ithadabundanthydrogen-utilizingM．tardaintheanaerobicbiofilm．Butthebacte-rialcommunitystructureintheproducedwaterwasobviouslydifferentfromthatintheanaerobicbiofilm．ThemainbacterialpopulationintheinfluentwasEpsilonproteobacteriawhileitwasNitrospiraandDeltaproteobacteriainthean-aerobicbiofilm．Notablymanysyntrophicbacteria-relatedcloneswerefoundamongtheDeltaproteobacteria．OneDeltaproteobacteriaclonewasrelatedtoSyntrophussp．whichiscommonlyfoundinmethanogenichydrocarbon-de-gradingconsortia．AnumberofDeltaproteobacteriacloneswereassignedtothecloneclustergroupTAmembersofwhichpredominateinvariousmethanogenicconsortiathatdegradearomaticcompounds．TheseresultssuggestedthatitexistsuchkindofmicrobialcommunityrelatedtotheanaerobicdegradationofpetroleumhydrocarbonmediatedbythemethanogensintheanaerobicwastewatertreatmentsysteminDaqingOilfield．Keywordsproducedwastewateranaerobictreatmentmethanogenanaerobichydrocarbondegradation8632012AA0634015117844712L03ESPC2013－02－192013－04－031960～、。[e-mail]、。、、、、。8。。、1。。2-4。。。。。2080、。。-TPH、CODPAH。。。11．12001430m×18m×5．5mHＲT12h5000tCOD0．72～0．75g/L·d。。———2009。3。DNA。1．2DNADNAFastDNASPIN。DNA1．5%。1．3PCＲ-DGGE16SrＲNAV3341F5'-CCTACGGGAGGCAGCAG-3'534Ｒ5'-ATTACCGCGGCTGCTGG-3'5DNA341F5'40bpGC5'-CGCCCGCCGCGCGCGGCGGGCGGGGCGGGGGCACGGGGGG-3'。AＲCH344F5'-ACGGGGYGCAGGCGCGA-3'5'GC519Ｒ5'-GWATTACCGCGGCKGCTG-3'616SrＲNA。50μLPCＲ1．5mmol/LMgCl21×PCＲ、200μmol/L、10pmol、1．25UTaq1μLDNA。PCＲPCＲ95℃10min95℃1min、65℃45s0．5℃、72℃1min2095℃1min、55℃45s、72℃1min1072℃5min。PCＲ95℃10min95℃1min、55℃1min、72℃1min3072℃5min。。DGGEBio-ＲadD-Code30μLPCＲ10%w/v30%～50%100%7mol/L40%1×TAE60℃25V20min150V6h。20minGelDoc2000TMBio-ＲadBio-ＲadQuantityOne4．3．0。1．416SrＲNA27F5'-AGAGTTTGATCCTGGCTCAG-3'1492Ｒ5'-GACGGGCGGTGTGTAC-3'716SrDNAPCＲ。8083A571FUA1406Ｒ816SrDNAPCＲ。PCＲ95℃10min95℃1min、55℃1min、72℃1min3072℃10min。PCＲ16℃2h。－80℃100μL10μL30min42℃90s1～2minLB890μL37℃170r/min1h。100μL20μL0．1mol/LIPTG100μL20mg/mLX-gal50μg/mLLB30min37℃18h。5mLLB37℃、170r/min18h1μL。1．5DNABioEditＲDPIIGeneBankClustalXMEGA3．1neighbor-joiningJukes-CantorBootstrap1000。chimeraＲDPIICHIMEＲA_CHECKBellerophon。DOTUＲ97%operationaltaxonomicunitOTU9。22．11。HＲT12hCOD270mg/L。80mg/LCOD30%7mg/LTPH25%。20mg/L。。。2．22．2．1DGGEDGGE1a。1Table1QualityoftheproducedwaterbeforeandafteranaerobicandaerobictreatmentpH7．207．30℃50．0050．00CODmg/L270．00187．00TPHmg/L28．0021．00TDSmg/L1400．00－SO2－4mg/L25．504．70。。。1b。112DGGEFig．1DenaturinggradientgelelectrophoresisDGGEprofilesforarchaealandbacterialcommunitiesininfluentproducedwaterlane1andanaerobiclane2samplesDGGE2。3DGGEMethanosarcinales。DGGEEpsilonpro-teobacteriab5b6BetaproteobacteriaThermotogae。2．2．2DGGE180～200bp90882DGGETable2NucleotidesequencesimilarityofsequencedDGGEbandsa1WL-22a2a3b1b2DXH4-98ε-b3ε-ε-b4724B10S32ε-b5β-b6DGGE16SrＲNA。4APABBPBB。212DOTUＲ97%OTU2642OTU65OTU。1。16SrＲNA、1011。MethanosarcinalesMethanomicrobiales2Methanomethylovoransthermophila16SrＲNA≥98%。M．thermophila68．8%56．8%。Methanolineatarda34．1%10．4%。M．tardaH2。2。Epsi-lonproteobacteria74%Nitrospira40%Deltaproteobacteria20%3。。Epsilon-proteobacteriCampylobacterales。48%OTUBP-B106BP-B134BP-B105BP-B121BP-B82BP-B164BP-B71Helicobacter-aceaeSulfuricurvumsp．EU498374S．kujiense。EpsilonproteobacteriaCampylobacteraceae41213。Nitrospira40%。17．6%BB-HB76BB-B51BB-LB6BB-B47Thermodesulfovibrio。H21415。。2AP、ABFig．2PhylogenetictreeofarchaealclonesretrievedfromtheinfluentAPandanaerobicbiofilmABsamples01833BBFig．3PhylogenetictreeofbacterialclonesretrievedfromthebiofilmBBDeltaproteobacteria。11．8%BB-B38BB-B40BB-B2BB-B20BB-B93BB-HB117GroupTABB-B38Syntro-phorhabdaceaeSJA-51BB-B4011884BPFig．4PhylogenetictreeofbacterialclonesretrievedfromtheinfluentBPWCHB1-27BB-HB11792%BB-B2、BB-B20BB-B93Syntro-phorhabdusaromaticivorans。OTUBB-LB39Syntrophussp．。Deltapro-teobacteriaSyntrophobactersulfatireducensS．fumaroxidansBB-HB110SyntrophaceaeD15_21