河流沉积物中反硝化细菌的分离及脱氮除磷研究副本

1,2,23,2,1,1(11,475001;21,100094):,20,,,.,,,50%10,F106312%,16SrDNA,(Alcaligenesfaecalis);F10,100mgPL,10d,,7612%9318%.:;;;;:X172:A:025023301(2009)0120091205:2008201202;:2008203220:(2006BAD17B05);(XK10019440);(XK100190553):(1980),,,,,E2mail:[e-mail]3,E2mail:[e-mail]olatedfromRiverSedimentandItsNitrogenandPhosphorusRemovalCapacityfromRiverWaterWANGLin1,2,LIJi2,KANGWen2li2,CHENYun2zeng1,GUOTing2zhong1(1.EnvironmentalandHealthyEngineeringResearchCenter,HenanUniversity,Kaifeng475001,China;21CollegeofResourcesandEnvironmentalSciences,ChinaAgriculturalUniversity,Beijing100094,China)Abstract:20strainsofbacteriawereisolatedfromriversedimentusingenrichmentculturemediumfordenitrification,andthedenitrificationintensitywasdetermined.F10,oneofbacteriastrain,wasidentifiedhavingthehighestdenitrifyingintensity,andfurtherusedtotestitsroleintheremovingofnitrogenandphosphorusfromwastewaterinlaboratory.Bycheckingtheindividualmorphology,colonyculturecharacteristics,DNAsequencingand16SrDNAgenebank,F10wasidentifiedasAlcaligenesfaecalis,anditsdenitrifyingintensitywas6312%.ThehighestremovalofTN(7612%)andTP(9318%)wereobservedina10daysperiodwithanadditionofF10atarateof100mgPL.Keywords:denitrifyingbacteria;eutrophication;riversediment;domesticwastewater;bioremediation,[14].[59][10,11][1214].,,,...,,.11.1,,TN9146mgPL(N),TP0196mgPL(P).510g,,30.45d,20%,34,.:KNO32g,5g,K2HPO41g,MgSO47H2O012g,1000mL,121,20min.1.2.2mL,,,,50,30120091ENVIRONMENTALSCIENCEVol.30,No.1Jan.,2009©1994-2011ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.[15],.1.3,.,.20%,30,3600nm,(D),,D,.1.4,16SrDNADGGE,,.1.5DNAPCR2DGGE1.5.1DNA8mL,8000rPmin,4,20min,,,015mLDNA(1molPLTris2HCl,pH810).DNABenzylChloride[16].1.5.2PCRPCRAmpliTaqGoldTM(PerkinElmer,AppliedBiosystems,NJ,USA).357F517R,52CCTACGGGAGGCAGC2AG2352ATTACCGCGGCTGCTGG23.PCR:9510min;25,931min,501min,721min30s,,931min,501min,725min.(50L):1LdNTP(10mmolPL)5L10PCRGoldBuffer4LMgCl2(25mmolPL)015L357F(45pmolPL)015L517R(45pmolPL)1LDNA(10ngPL)015LTaq(2UPmL),.PCR2%,,DNABLAST,.1.5.3DGGEDCode(Bio2Rad,Laboratories,Hercules,Calif)PCRDGGE[17].6%12%,20%55%(7molPL,40%100%),61,200V5h,SYBRGREEN(MolecularProbes,Eugene,Ore)30min,[18].1.6.25L,20L,,,.,4000rPmin10min,,018%3,20L,1005001000mgPL,3.,5,,,.1.7(TN):,(TP):[19].22.1,314d,,.2.2,20.TN,20,,,50%10(1),F10,6312%.F10,.2.3F10F10,2930©1994-2011ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.(016112)m(017216)m,,1.2.4F1002d,(2),25d,,,,F10,;57d,,8d,,.,[20].,5d.2.5DGGE1F10Fig.1IndividualmorphologycharacterofstrainF102Fig.2GrowthcurveofdenitrifyingbacteriawhenitwasculturedDNA,DNA2%,16SrDNAV3PCR,PCRDGGE,3,DGGE.3F10DGGEFig.3DGGEprofileofV3regionof16SrDNAfromF102.6DNA,DNA,DGGE.DGGE,F10.16SrDNANCBI16SrDNA,F10.1,.16SrDNA.:Alcaligenesfaecalis(AY866407),9919%,F10(Alcaligenesfaecalis),EU169605.,Nishio[21]OKK17.Potivichayanon[22]391:©1994-2011ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.[23],COD.Joo[24].2.7F102.7.1F10,.TN4,F103,TN,.10d,,100mgPL,TN7216%.,500mgPL1000mgPL100mgPL,10d,,TN,,TN.4Fig.4Totalnitrogenchangesbetweentreatments2.7.2TP5,,3TP.,TP10d,,TP,100mgPL9318%,,10,,TP.,F10.5Fig.5Totalphosphoruschangesbetweentreatments3(1),20,,,,F10,6312%.F1016SrDNA,(Alcaligenesfaecalis).,F1025d,,,57d,,.(2)F10,100mgPL,,7612%9318%.F10,,.,,TNTP5d,510d,,,TNTP.,.,.,,.(3),,,,.,4930©1994-2011ChinaAcademicJournalElectronicPublishingHouse.Allrightsreserved.:[1],.[J].,2006,36(5):28232.[2]StouthamerAH,deBoerAPN,vanderOostJ,etal.EmergingprinciplesofinorganicnitrogenmetabolisminParacoccusdenitrificansandrelatedbacteria[J].AntonievanLeeuwenhoekInternationalJournalofGeneralandMolecularMicrobiology,1997,71:33241.[3]PaiSL,ChongNM,ChenCH.Potentialapplicationofaerobicdenitrifyingbacteriaasbioagentsinwastewatertreatment[J].BioresourceTechnology,1999,68:1792185.[4]SchmidtI,SliekersO,SchmidM,etal.Newconceptsofmicrobialtreatmentprocessesforthenitrogenremovalinwastewater[J].FEMSMicrobiologyReviews,2003,27:4812492.[5]OvezB.BatchbiologicaldenitrificationusingArundodonax,Glycyrrhizaglabra,andGracilariaverrucosaascarbonsource[J].ProcessBiochemistry,2006,41:128921295.[6]AslanS,CakiciH.Biologicaldenitrificationofdrinkingwaterinaslowsandfilter[J].JournalofHazardousMaterials,2007,148:2532258.[7]RoccaCD,BelgiornoV,MericS.HeterotrophicPautotrophicdenitrification(HAD)ofdrinkingwater:prospectiveuseforpermeablereactivebarrier[J].Desalination,2007,210:1942204.[8]FernandezN,AlvarezRS,FieldJA,etal.Microbialcommunitydynamicsinachemolithotrophicdenitrificationreactorinoculatedwithmethanogenicgranularsludge[J].Chemosphere,2008,70:4622474.[9]SuC,PulsRW.Removalofaddednitrateincottonburrcompost,mulchcompost,andpeat:Mechanismsandpotentialuseforgroundwaternitrateremediation[J].Chemosphere,2007,66:91298.[10]HoshinoT,TeraharaT,TsunedaS,etal.Molecularanalysisofmic