不同环境中脱氮硫杆菌的分离培养及其去除硝酸盐的研究牛建敏

　　　200927、＊牛建敏　李睿华(,　210093)　:从湖水、底泥、土壤、厌氧污泥等不同环境中采集菌种,培养、分离得到10种脱氮硫杆菌菌株。在28℃下,采用脱氮硫杆菌标准培养基通过摇瓶实验比较了所得到的10株脱氮硫杆菌在生长繁殖、对硝酸盐氮去除速率、培养液pH值变化等方面的差异。结果显示:从厌氧污泥筛选而来的脱氮硫杆菌菌株相比其它菌株表现出良好的生长繁殖和反硝化能力,该菌株在接种后第48小时即达到对硝酸盐氮100%的去除率,去除速率为143.30mg/(L·d),pH从7.09下降到6.68;其次是从厌氧发酵污泥上清液富集培养的菌株,在接种后第4天达到对硝酸盐氮100%的去除率,去除速率为71.65mg/(L·d);从湖泊底泥和莲塘底泥富集的菌株相比湖水、土壤也有较好的反硝化效果。为如何利用环境中的脱氮硫杆菌、获得活性更高、脱氮稳定性更好的工程菌株提供了指导。:脱氮硫杆菌;分离;培养;硝酸盐;反硝化;环境ISOLATIONANDCULTUREOFTHIOBACILLUSDENITRIFICANSDERIVEDFROMDIFFERENTENVIRONMENTSANDTHEIRCAPACITIESOFREMOVINGNITRATENiuJianmin　LiRuihua(StateKeyLaboratoryofPollutionControl&ResourceReuse,SchooloftheEnvironment,NanjingUniversity,Nanjing210093,China)Abstract:TendifferentstrainsofThiobacillusDenitrificanswereisolatedfromlake,soilandanaerobicsludgeetc.Experimentswereconductedat28℃usingconicalflaskwithenrichmentmediaofThiobacillusDenitrifi-cans.Thesestudiesincludethegrowthsofthese10strainsofThiobacillusDenitrificans,NO3--Nremovalve-locitiesandthechangesofpHduringthedenitrificationprocesses.Theresultsshowthatthiobacillusdenitrifi-cansscreenedfromanaerobicsludgeexhibitthebestdenitrificationcapacities.TheNO3--Nremovalefficiencybythisstrainreaches100%in48hoursandtheNO3--Nremovalvelocityis143.30mg/(L·d).ThepHdur-ingthisprocessdecreasesfrom7.09to6.68.Thestrainscreenedfromsupernatantofanaerobicfermentationsludgerequires4dfor100%NO3--NremovalafterinoculationandtheNO3--Nremovalvelocityis71.65mg/(L·d);Thestrainsscreenedfrombottomsludgeoflakeandpondalsoshowabetterdenitrificationcapa-citesthanstrainsfromlakeandsoil.ItishelpfulforhowtouseThiobacillusDenitrificansinnaturalenviron-mentandfindengineeringmicrobeswhicharehigh-activeandhavestabledenitrificationcapacities.Keywords:thiobacillusdenitrificans;isolation;culture;nitrate;denitrification;environment＊(WTWER0717)　　,、,2090,103.2%0.53%,5%55.01%[1],,。、,,。。,、,226DOI:10.13205/j.hjgc.2009.s1.057　　　200927,NO3-N2。,,,,[2]。,,,、、[3-5]。,。,,[6]。,NO2-NO3-,,,,NO2-NO3-N2。/(SLAD),Koening[7]SLADNO3--N400mg/L,5.17h100%;1991Montferland2384～3103m3/dSLAD,,ρ(COD)/ρ(N)1.2,NO3--N30mg/L90%[8]。,、,、、。1　1.1　菌种的采集、、、,10,T.d.1～T.d.10。1　T.d.1T.d.2T.d.3T.d.4T.d.5T.d.6(15cm)T.d.7T.d.8(15cm)T.d.9T.d.101.2　菌种的培养、分离与筛选、、,,[9],100mL5mL。5min,28℃4d,120r/min。4d,,。5,,2%,;,,;。,4℃。1.3　菌种的生长繁殖2,1.00mL1.00mL,[10]。328℃48h。,。1.4　脱氮硫杆菌对硝酸盐氮的去除实验T.d.1～T.d.10,T.d.1～T.d.10,NaOHpH7,100mL5mL,10,28℃,120r/min,24hpH。1.5　分析方法[11];[11];pHPHB-4pH;HITACHIH-7650。2　2.1　菌株培养与鉴定2,T.d.10、T.d.9、T.d.2、T.d.4,T.d.10,,,。3,T.d.1、T.d.3、T.d.6、T.d.8;T.d.5、T.d.7。,,,、,,。5,。227　　　200927T.d.10,。T.d.49,[12],,,,,0.4μm×0.8μm,1,[13],、、、。1　(T.d.4)2.2　菌株的生长繁殖2T.d.1、T.d.4、T.d.6、T.d.7、T.d.9、T.d.102。,3,T.d.105.80×108/mL,T.d.74.67×105/mL。,T.d.10,T.d.9,T.d.4T.d.1()、T.d.6(15cm)。2　/mLT.d.15.60×106T.d.43.20×107T.d.68.70×106T.d.74.67×105T.d.91.30×108T.d.105.80×1082.3　不同脱氮硫杆菌菌株对NO3--N的去除210,T.d.1～T.d.10。310。T.d.1048h100%,143.30mg/(L·d);T.d.9()、T.d.4()、T.d.2()、T.d.6()4、6、7、9100%。10,T.d.1()、T.d.3()、T.d.5()、T.d.7()、T.d.8(15cm):87.68%,92.11%,60.33%,42.67%,87.51%。,T.d.10,T.d.9()。,;,。2　T.d.1～T.d.10NO3--N3　T.d.1～T.d.10NO3--N2.4　pH的变化4T.d.1、T.d.4、T.d.9、T.d.10pH。T.d.1pH7.21,1087.68%,pH6.55,T.d.3、T.d.5、T.d.7、T.d.8;T.d.4pH7.23,6100%,pH6.63,T.d.2,pHpH7.20、6.61。T.d.9pH7.05,4100%,pH6.65;T.d.1048h100%,pH7.096.68。228　　　200927T.d.1～T.d.10pH,,BisogniDriscoll(1)[14]。0.844S2O2-3+NO-3+0.434H2O+0.347+0.0865CO2-3+0.0865NH+40.0865C5H7O2+0.5N2+1.689SO2-4+0.697H+(1),,pH7.0,,pH,CaCO3,。,。4　pH3　1)、、,;10,T.d.10()5.80×108/mL,,T.d.7()4.67×105/mL,3。T.d.9(),1.30×108/mL;10,T.d.10,143.30mg/(L·d),pH7.096.68;、、T.d.9、T.d.4、T.d.2,71.65,47.7740.94mg/(L·d)。;2),,,。[1],.[M].:,1990:118-120.[2],.[J].,2005,25(2):88-92.[3],.[J].,2005(2):52-54.[4],,,.[J].,2005,9:56-57,71.[5],,.[J].,2004,24(2):30-34.[6],,,.、[J].,2004,36(4):423-425,429.[7]KoeningA.LiuLH.Autotrophicdenitrificationoflandfillleachateusingelementalsulphur[J].WatSciTech,1996,34(5-6):469-476.[8]ZhangTC.Nitrateremovalinsulfur:Limestonepondreactors[J].JournalofEnvironmentalEngineering,2002,128(1):74—84.[9]BaalsruudK,BaalsruudKS.StudiesonThiobacillusdenitrificans[J].ArchivfurMikrobiologie,1954,Bd.20,S.34-62.[10],,.[M].:,2004(1):34-36.[11][M].:,2002(4).[12].[M].:,2003.[13]R.E.,N.E...[M].:,1984.[14]BisogniJJ,DriscollCT.Denitrificationusingthiosulfaeandsul-fide[J].J.Enviro.Engineer.Divi,AmerSociofCivEngin,1977,103:593-604.　　210093　E-mail　[e-mail]2009-01-16229