复合人工湿地池塘养殖生态系统细菌多样性研究姚延丹

34、、[1-3]。[4]。。DenaturedGradientGelElectrophoresisDGGE《》2010-07-022010[e-mail]*[e-mail]。EnvironmentalScience&Technology34720117Vol.34No.7Jul.2011,.-[J].,2011,34(7)50-55.YaoYan-danLiGuTaoLingetal.Bacterialdiversityinvestigationonanintegratedsystemofconstructedwetlandandpondaquaculture[J].EnvironmentalScience&Technology,2011,34(7)50-55.-122*2322221.2013062.4340003.2140812009810-DGGE。NH4+-NTPP0.05P2P4。DONH4+-N、TP。。DGGEX524Adoi10.3969/j.issn.1003-6504.2011.07.0141003-6504(2011)07-0050-06BacterialDiversityInvestigationonanIntegratedSystemofConstructedWetlandandPondAquacultureYAOYan-dan12LIGu2*TAOLing23LIXiao-li2ZHANGShi-yang2ZHAOQiao-ling2LINYu-liang21.CollegeofFisheriesandLifeScienceShanghaiOceanUniversityShanghai201306China2.YangtzeRiverFisheriesResearchInstituteChineseAcademyofFisherySciencesJingzhou434000China3.ResearchCenterforFreshwaterFisheryChineseAcademyofFisherySciencesWuxi214081ChinaAbstractThebacterialdiversityinvestigationintheintegratedsystemofconstructedwetlandandpondaquaculturewascarriedoutduringtheperiodofAug.～Oct.2009usinganalyticmethodofDGGEdenaturedgradientgelelectrophoresis.ThefindingspresentedthattemperatureandDOoftheculturepondschangedovertheinvestigationperiodandbothbacterialdensityanddiversityintheculturepondvariedwithchangeoftheseasoninadditionthelevelsofNH4+andTPinthecontrolpondweresignificantlyhigherp0.05thanthoseintherecirculationponds.CorrelationanalysispertainingtotheinvestigationrevealedthatbacterialdiversitywasremarkablyandpositivelycorrelatedwithDObutnegativelywithNH4+andTP.Itwasconcludedthatthepond-recirculationaquaculturecouldenhancebacterialdiversityinthepondsthusthepondsystembecamemorestableecologically.KeywordspondrecirculationaquaculturesystembacterialdiversityDGGEecosystem7DNA。DGGE、。DNA[5]、、、、、、、[6]。Santegoeds[7]10Teske[8]-Desulfovibriosp.Arcobactersp.。PCR-DGGE。-。11.1。、31。。450m21.5~2.0m、L×W×H30m×8.5m×0.6m0.6mCannaindica、Iristectorum、AcoruscalamusCyperuspapyrus、Thaliadealbata。L×W×H＝200m×0.54m×0.62m、、、。5P1~P51P0。。P5。P1。。P1、P3、P5(6.9±0.6)cm(7.1±1.7)gP2、P4(6.9±1.0)cm(6.4±2.2)g2000/667m25336/667m2。15%~20%。1.21.2.1P2、P4P0。20098101630~700AM50cm。2.5L35LT、pHDOORION5-StarNH4+-N、TNTP《》[9]。1.2.2250mL。3。37℃48ha×10n。1.2.3DNA0.22μm200mLTE10mmol/LTris-HCl+1mmol/LEDTApH8.010000r/min10min-20℃。DNA。DNAZhou[10]。DNA3S-20℃。1.2.4PCR341F-GC-clamp5’-CGCCCGCCGCGCGCGGCGGGCGGGGCGGGGGCACGGGGGGGGGCCTACGGGAGGCAGCAG-3’534R5’-CCGTCAATTCMTTTGAGTTT-3’。DNA16SrRNAV3。PCR25μL10×Buffer2.5μL10mmol/LdNTP0.3μL10mol/L0.5μL5U/μLTaq-5134DNA0.3μL50ng/μLDNA1.0μL19.9μL。PTC-200PCRPCRtouchdown-PCRtd-PCR。94℃5min94℃30s65℃30s72℃90s0.5℃201172℃5min。1%。1.2.5DGGEPCR。10%=37.5：120%~50%100%7mol/L40%20μL。1×TAE20mmol/LTris+10mmol/L+0.5mmol/LEDTApH7.460℃80V12h。。1.3Quantityone4.1.0。Shannon-WienerH’H’H′=-Si=1ΣniNlnniN1SniiN。One-WayANOVAPearsonCorrelationSPSS13.0。22.1831.4℃1020.0℃1.23~2.19mg/LpH7.4~8.0NH4+-N、TNTP2。NH4+-NTPP0.05。2.21.1×105cfu/mL1。3。。P25.7×103~2.1×104cfu/mLP48.7×103~1.7×104cfu/mL。2.3DGGE2.3.1DGGE4527。DGGEOperationTaxonomyUnitOUT[11]。。4DGGE、。。41234579。5。6。68P2。DGGE。2.3.21Shannon-WienerH′5。P2P4。。DOR=0.67P0.05NH4+-NR=-0.84P0.05、TPR=-0.80P0.05。33.1。[12]。。NH4+-NTPTN。TN。、/。TN。TNTN5.0~10.5mg/L。NH4+-NTP。。2006。[13]。20062007TP40.4%-533446.2%。NH4+-NNH4+-N。NH4+-NTP。3.2。。。、[14-16]。。[17]。[18]。[19]。。。。。810。。。[20-21]。3.3DGGE、。[22-23]。、DNA、PCR、DGGE[2224]。DGGE。。。[25-26]。DONH4+-N、TP。NH4+-N、TP。。。P4P2P2。DGGE-。。41NH4+-NTP2。3。。DONH4+-N、TP。[][1].ZL270051740.8[Z].2008917.[2]LinYFJingSRLeeDYetal.Nutrientremovalfromaquaculturewastewaterusingaconstructedwetlandssystem[J].Aquaculture2002209169-184.[3]SindilariuPDCarstenSReinhardR.Treatmentofflow-throughtroutaquacultureeffluentsinaconstructedwetland[J].Aquaculture200727092-104.[4].[J].200120231-39.LiuWei-dongSuHaoDengLi-kang.Applicationsofmicroorganismsinaquaculture[J].FisheriesScience200120231-39.inChinese[5]MuyzerGDe-WaalECUitterlindenAG.Profilingofcomplexmicrobialpopulationsbydenaturinggradientgelelectrophoresisanalysisofpolymerasechainreaction-am－plifiedgenescodingfor16SrRNA[J].AppliedandEnvi－ronmentalMicrobiology199359695-700.547[6]GongMLRenNQXingDF.Applicationofdenaturinggradientgelelectrophoresisandtemperaturegradientgelelectrophoresisinmicrobialmolecularecology[J].ActaMi－crobio1Sinica2004446843-848.[7]SantegoedsCMNoldSWaldDM.Denaturinggradientgelelectrophoresisusedtomonitertheenrichmentcultureofaerobicchemoorganotrophicbacterialfromahotspringcyanobacterialmat[J].AppliedandEnvironmentalMicrobi－ology199662113922-3928.[8]TeskeASigalevichPCohenYetal.Molecularidentifi－cationofbacteriafromacoculturebydenaturinggradientgelelectrophoresisof16SribosomalDNAfragmentsasatoolforisolationinpurecultures[J].AppliedandEnviron－mentalMicrobiology1996624210-4215.[9].[M]..:2002.[10]ZhouJBrunsmATiedjeJM.DNArecoveryfromsoilsofdiversecomposition[J].ApplMicrobiolBiotechnol1996622316-322.[11].PCR-DGGE[J].200626111828-1832.SuJun-fengMaFangWangHong-yuetal.ApplicationofPCR-DGGEtoresolvemicrobialdiversityinbio-ceram－icreactor[J].A