高效反硝化聚磷菌株的筛选及其生物学特性马放

286　　　　　　　　　　　　　　　　　　　　　　　　　　　　　Vol.28№.620076　　　　　　　　　JournalofHarbinEngineeringUniversity　　　　　　　　　　　Jun.2007马　放1,王春丽1,2,王立立1(1.哈尔滨工业大学市政与环境工程学院,黑龙江哈尔滨150090;哈尔滨工程大学建筑工程学院,黑龙江哈尔滨150001)　:,、PHB,/SBR4H16、H19、H24Xg.,3(Pseudomonas),(Enterobacter).4,pH4,,35℃,25℃;,H16、H19H24pH6～8,XgpH7～9;4pH.:;;;pH;:V526　:A　:1006-7043(2007)06-0631-05Screeningofdenitrifyingpolyphosphate-accumulatingorganismsandtheirbiologicalcharacteristicsMAFang1,WANGChun-li1,2,WANGLi-li1(1.SchoolofMunicipalandEnvironmentalEngineering,HarbinInstituteofTechnology,Harbin150090,China;2.CollegeofCivilEngineering,HarbinEngineeringUniversity,Harbin150001,China)Abstract:Tostudythebiologicalcharacteristicsofdenitrifyingphosphate-accumulatingorganisms(DN-PAOs),foureffectiveDNPAOswereidentifiedinananaerobic/anoxicSBRreactorwithsteadyoperation.Candidateorganismswerescreenedbyaphosphateuptakeexperiment,wherenitratesweredenitrifiedtogasandthroughinspectionofmetachromaticandPolyphydroxybutyrate(PHB)granules.TheDNPAOswerePseudomonassp.H16,H19,H24andEnterobactersp.Xg.Twoimportantfactors,temperatureandpHvalue,werestudiedtoevaluatetheireffectonthegrowthandphosphorusremovalratiosofH16,H19,H24andXg.ResultsshowthattheoptimaltemperatureforthefourDNPAO'sgrowthandphos-phorusremovalisabout35℃and25℃respectively.Inidealtemperatureconditions,thebestpHvaluesforH16,H19andH24growthandphosphorusremovalrangefrom6.0～8.0;forXg,thebestpHrangeisbetween7.0～9.0.Phosphorusremovalforallfourstrainsareoptimalinamildlyalkalicondition.Growthcurvesforthesestrainswerealsodetermined.Keywords:denitrifyingphosphate-accumulatingorganisms(DNPAOs);screening;temperature;pH;growthcurve:2006-04-12.:(50521140075).:　(1963-),,,,E-mail:[e-mail].　　,,———(denitrifyingphosphate-accumulatingorganisms,DNPAOs),,()()[1-4].,,,.,,,,,.1　材料与方法1.1　　　/SBR,.1.2　(L-1):10g;3g;NaCl5g;20g;pH7.2,、.(L-1)[5]:3.32gCH3COONa·3H2O;23mgNa2HPO4·2H2O;11mgCaCL2·2H2O;152.8mgNH4Cl;81.12mgMgSO4·7H2O;17.83mgK2SO4;7gHEPES2mL;pH7.2.(L-1)[5]:CH3COONa·3H2O3.32g;K2PO425mg;NH4CL305.52mg;MgSO4·7H2O91.26mg;CaCL2·2H2O25.68mg;PIPES8.5g2mL;pH7.2.(L-1):3.0g;5.0g;KNO31.0g;pH7.4.3.1.3　、1.3.1　菌株的分离、纯化10mL90mL,,,.,.2～3d,1.(30～300),3,、.,,3,,,.,.1.3.2　菌株筛选、,,1mg/LpH7.0.30℃、140r/min.1×104r/min,,,,;30℃,24h,10mL,,PO3-4-P.50%、-β-(PHB).,PHBDNPAOs[6].1.4　DNPAOs/SBR,,,KNO3,NO-3-N60mg/L,,24hDN-PAOs,,,,4h,NO-3-N.1.5　[7-8].1.6　1.6.1　生长曲线的测定,722OD600.1.6.2　温度、pH值与菌株生长关系的确定20℃、25℃、30℃、35℃40℃,DNPAOs18h,10%5(),、48h,OD600.pH4、5、6、7、8、9、10,,30℃、24h,OD600.2　结果与分析2.1　、　　348,、,50%8,、PHB,1.·632·　　　　　　　　　　　　　　　　　　　　　　281　Table1　Resultsofscreening/%PHBH1254,H1562,H1661,,H1998,,H2497,H2781,,Xa60,,Xg59H16、H19、H24、Xg,PHB,,H16、H19、H24、Xg.1H16.1　H16Fig.1　ThephotoofmetachromaticgranuleinspectionofH162.2　DNPAOs4hNO-3-NPO3-4-P,2.2　DNPAOsFig.2　NandPremovalratesofDNPAOs　　2,4,H19、H2488%86%.,NO-3,-[9].,O2,NO-3.H16、H19、H24Xg.2.3　H16、H19、H24、Xg2.,3(Pseudomonas),(Enter-obacter).2　Table2　IdentifyingfeaturesofstrainsH16H19H24Xg、、、、、、、、、、、、、、、、、、、+++--+++---++++++---+++-+++-M.R----V.P---+++-+--------:“+”,“-”.·633·6　　　　　　　　　　　　,:2.4　H16、H19、H24Xg2.4.1　菌株H16、H19、H24、Xg的生长43。3,H16、H19、H24、Xg,2h,;H16、H19、H2414h,16h;Xg,16h,18h.H16、H1916h,H24、Xg20h18h.3　H16、H19、H24、XgFig.3　GrowthcurvesofstrainH16,H19,H24andXg2.4.2　温度对菌株H16、H19、H24和Xg生长及除磷效能的影响　　5,4.4　H16、H19、H24、XgFig.4　EffectoftemperatureongrowthandPremovalrateofH16,H19,H24andXgH16H2415～35℃,;H19Xg20～35℃,;435℃,45℃;H16、H19Xg25℃;H2415～35℃,80%.2.4.3　pH对菌株H16、H19、H24和Xg生长及除磷效能的影响　　pH5,5.·634·　　　　　　　　　　　　　　　　　　　　　　285　pH4H16、H19、H24、XgFig.5　EffectofpHongrowthandPremovalrateofH16,H19,H24andXg,4DNPAOspH6～10,H16、H19H24pH6～8,XgpH7～9.,pH8,,;.,[10].,4pH.3　结　论1)、、PHB.2)4DNPAOs,35℃,,20～30℃,4,25℃.4.3)pH4DNPAOs.4pH,,pH,.pH,,pH.:[1]KUBAT,SMOLDERG,vanLOOSDRECHTM,etal.Biologicalphosphorusremovalfromwastewaterbyanae-robic-anoxicsequencingbatchreactor[J].WaterSciTechnol,1993,27(5-6):241-252.[2]JOHONWANA,TOMOTAKAD,SATOSHIT,etal.Characterizationofdenitrifyingphosphate-accumulatingorganismscultivatedunderdifferentelectronacceptorconditionsusingpolymerasechainreaction-denaturinggradientgelelectrophoresisassay[J].WaterRes,2002,36:403-412.[3]KUBAT,VANLOOSDRECHTM,HEIUNENJJ.PhosphorusandnitrogenremovalwithminimalCODre-quirementbyintegrationofdenitrifyingdephosphatationanddenitrificationinatwo-sludgesystem[J].WaterRes,1996,30(7):1702-1710.[4]MERZOUKIM,BERNETN,DELGENESJP,etal.BioloicaldenitrifyingphosphorusremovalinSBR:effectofaddednitrateconcentrationandsludgeretentiontime[J].WatSciTech,2001,43(3):191-194.[5]MERZOUKIM,DELGENESJP,BERNETN,etal.Polyphosphate-accumulatinganddenitrifyingbacteriaiso-latedfromanaerobic-anoxicandanaerobic-aerobicsequen-cingbatchreactors[J].CurrentMicrobiology,1999(38):9-17.[6]　,,,.[J].,2003,29(8):33-35.LUONing,LUOGuyuan,JIFangying,etal.Aicroor-ganismgroupsinbiologicalPandNremovalprocess[J].WaterandWastewaterEngineering,2003,29(8):33-35.[7],.[M].:,2001.[8]RE,NE.[M]:9.:,1995.[9]VLEKKEGJFM,COMEAUY,OLDHAMWK.Bi-ologicalphosphateremovalfromwastewaterwithoxygenornitrateinsequencingbatchreactors[J].EnvironmentalTechnologyLetters,1998(9):791-796.[10],.p