DONH4N实现短程硝化过程中生物膜特性赵青

39320183ENVIＲONMENTALSCIENCEVol．39No．3Mar．2018DO/NH+4-N111*11121．1001242．100011．DO/NH+4-N10.27%、52.12%93.54%FISHCLSM、AOBNOBEEM．AOBNOB．6～25μm．AOB/NOB15.56．EPS．EPSNOB．EPS．X703.5A0250-3301201803-1278-08DOI10．13227/j．hjkx．2017080552017-08-072017-09-012015ZX07202-0131991～、[e-mail]*[e-mail]lmDuringtheTransitionProcessofCompleteNitrificationandPartialNitrificationZHAOQing1BIANWei1LIJun1*WANGWen-xiao1SUNYi-qi1LIANGDong-bo1ZHANGShu-yan21．CollegeofArchitectureandCivilEngineeringBeijingUniversityofTechnologyBeijing100124China2．ChinaEnergySavingandEmissionＲeductionLimitedCompanyofShenhuaGroupBeijing100011ChinaAbstractTheobjectiveofthestudywastoinvestigatethechangeofbiofilmcharacteristicswhenimplementingtheprocedureofpartialnitrification．AratiocontrolstrategyDO/NH+4-Nwastakentoachievepartialnitrificationandbiofilmsampleswereobtainedat10.27%52.12%and93.54%ofthenitriteaccumulationrate．TheamountandspatialdistributionoftotalbacteriaammoniaoxidizingbacteriaAOBandnitriteoxidativebacteriaNOBwereobservedbyfluorescenceinsituhybridizationFISHandconfocallaserscanningmicroscopeCLSMthroughathree-dimensionalexcitationemissionmatrixEEMtoobservethesecretionandcompositionchangesofextracellularpolymersubstances．ＲatiocontrolsuccessfullyenrichedAOBandachievedpartialnitrificationunderconditionswhenNOBwasnotcompletelywashed．Heterotrophicbacteriaandnitrifyingbacteriacoexistinthebiofilm．Theheterotrophicbacteriawereintheouterlayerbutnitrifyingbacteriaweredistributedinthebiofilmsurfaceat6-25μm．Duringtheprocessofshort-rangenitrificationtheAOB/NOBvaluegraduallyincreasedandthestableoperationperiodwasashighas15.56．DuringtheoperationofthereactorEPSandmicrobialflorachangesarecloselyrelated．WhenmicrobialactivitydecreasedEPSsecretiondecreased．DuringthestableoperationperiodofpartialnitrificationNOBandotherbacteriathatarenon-resistanttohighnitritenitrousaciddeclinedandthefluorescenceintensityofaromaticprotein-likebacteriadecreased．Howeverthethree-dimensionalfluorescencespectrashowedthatthechemicalcompositionofEPSwasnotobviousduringtheprocessofpartialnitrification．KeywordspartialnitrificationbiofilmnitrifyingbacteriaextracellularpolymericsubstancesAOBNOB．25%1．．AOBNOBAOBNOB．、pH、FA、FNA、HＲT、DO．2movingbedbiofilmreactorMBBＲ．AOBDONH+4-NAOBDONH+4-N．Bartrolí3Jemaat4DO3DO/NH+4-NNH+4-N100%．．．EPS．EPS．EPSPNPS70%～80%20%～30%、．EPS、．5SBBＲEPSEPS．EPSEPS．．FISHDNA、6．FISHCLSM．CLSM、7．FISHCLSMMBBＲ．EEMEPS．EPS．11.11．××18cm×12cm×18cm4.3L．．DOpHDO、pH．＞0.96g·cm－3Ф25mm×10mm＞400m2·cm－3．1L．．1Fig．1Schematicdiagramofexperimentaldevice、200dNAＲ5%．FISHAOBNOB1%±1%．NH+4-NNH4Cl60mg·L－1CODCH3COONa90mg·L－1KH2PO42mg·L－1NaHCO3pH7.8～8.2．．MBBＲHＲTDONH+4-NDO/NH+4-N．1.2NH+4-NNO－2-NN-1--NO－3-NDO、pHTWTW．NAＲ=NO－3-N/NO－2-N+NO－3-NNO－3-N、NO－2-NNH+4-N．MLSS、MLVSS45kHz120W2～3min10000g15min．1.31.3.1FISHFISHCLSM、AOBNOB．1×PBS34%4℃1972139～3h－20℃6．46℃2～3h20min89．FV1200OlympusJapan．101115'．1FISHTable1MaincharacteristicsoftheFISHprobeusedintheexperiment/nm/nmEUB338Mostbacteria525.0488.1FAMEUB338ⅡBacteriallineagesnotcoveredbyprobeEUB338andEUB338Ⅲ525.0488.1FAMEUB338ⅢBacteriallineagesnotcoveredbyprobeEUB338andEUB338Ⅱ525.0488.1FAMNSO190β-subgroupammoniaoxidizingbacteria595.0560.8Cy3NIT3Nitrobacter700.0646.8Cy51.3.2CLSM、Hoechst33342propidiumiodidePI12．fluoresceinisothiocyanateFITC．Hoechst33342PIFITC．、、2．1×PBS3Hoechst3334210μL0.01mg·mL－130min200r·min－1PI1μL0.02mg·mL－130minFITC10μL10mg·mL－11h1×PBS3．FV1200．2CLSMTable2MaincharacteristicsoffluorescentdyesusedintheCLSM/nm/nm/mg·mL－1Hoechst33342450.0402.80.01PI595.0560.80.02FITC525.0488.1101.3.313．1.3.4EPS1.3.3PBS80℃0.5hEPS．PN、PSPNPSF-7000Ex200～400nm5nmEm240～600nm2nm5nm1200nm·min－1．22.1MBBＲDO7～8mg·L－1．2MBBＲ2NO－3-N、NO－2-N、NH+4-NNAＲFig．2VariationofNO－3-NNO－2-NNH+4-NandNAＲNO－3-N、NO－2-N、NH+4-NNAＲ．DO2.0mg·L－1NH+4-N8.0mg·L－1DO/NH+4-N=0.25±0.05．20dNAＲ20d08213DO/NH+4-N90.54%DO/NH+4-NMBBＲ．1～20d21～60d．20dNO－3-N46.83mg·L－14.58mg·L－1NO－2-N2.69mg·L－146.03mg·L－1．NO－3-N、NO－2-N3.62±0.1mg·L－146.64±0.2mg·L－1NAＲ90%AOBNOB．NAＲ=10.27%2d、NAＲ=52.12%10dNAＲ=93.54%45d．1．NAＲ=10.27%2．NAＲ=52.12%3．NAＲ=94.53%T．A．AOBN．NOB3FISH3DFig．3FISH3Dimagesofsimples2.2EUB338、EUB338Ⅱ、EUB338ⅢFAMNSO190β-Cy3NIT35'Cy5630nm670nm．FV10-ASW4.20Viewer．33DT1、T2、T3NAＲ10.27%、52.12%94.53%3A1～A3、N1～N33AOBNOB．FV12003D．3．．CLSM4aEPS．、AOB、NOB．DO．．DO．CLSMMBBＲ30μm6～25μmAOBNOB．3A1～A3N1～N3AOBNOB14．DODO．182139．．、EPS、EPS4a．．3．4b．、．DO、．4b、DO．aFITCbHoechst33342PI4CLSMFig．4CLSMfluorescentstainingimages2.33、AOBNOB．AOBNOB．2dNAＲ10.27%AOBNOB3N1N2NOB3N3NOBDO/NH+4-NNOBNOBBian2＞200dNOB．AOB．AOBNOBNOBAOBAOB．NAＲ10.27%、52.12%93.54%、AOBNOB．15Wagner16ProteobacteriaBacteroidetesAOBNOB．β-α-β-AOB、NOB、17NitrosomonasNitrosospiraAOBβ-α-23%18NOBNitrobacter．NitrospiraNOBphylumNitrospira．NitrosomonasNitrosospiraAOBNOBNitrobacterNitrospira“AOB/NOB”．ShannonSimpsonShannonSimpson．3AOB/NOB、Shannon、Simpson．3NAＲβ-68.17%α-23.11%3.99%β-AOBα-NOB．β-α-70%．NitrosomonasNitrosospiraNitrosomonas0.088h－128213DO/NH+4-N3Table3CharacteristicsofmicrobialpopulationduringdifferentperiodsofpartialnitrificationNAＲ/%β-/%α-/%AOB/%NOB/%NitrosomonasNi