全 文 ：第27卷第5期
2008年10月
生态科学
EcologicalS ience
27(5)：341-345
Oct．2008
Organicaggregate—-attachedbacterialcommunity
compositionandy amicsoftheshallowhypertrophic
freshwaterLakeTaihu，revealedbyT—RFLPanalysis
TANGXiang．min91一，GAOGuan91andQINBo．qian91’
J．StateK yLaboratoryo／LakeScienceandEnvironment,NanjingInstituteofGeographyandLimnology,ChineseAcademyofScience8，
EastBeijingRoad73,Nanfing210008,China．
2．GraduateSchoolfChineseAcademyofSciences,Beijing100039China
Keywords：bacterialcommunitycomposition；organicagg egates；shallowlake；T-RFLP；microbialecol gy
太湖有机聚集体上附着细菌群落结构与动态的T-RFLP分析
汤祥明“2，高光1，秦伯强1’
1中国科学院南京地理与湖泊研究所，南京210008
2中国科学院研究生院，北京100039
【摘要】 有机聚集体(organicggregates)，是指由浮游动(植)物的残体、粪便颗粒及各种有机碎屑、活的自养及异养微生物以
及无机颗粒等由于物理的、化学的或生物的作用聚集而成的颗粒物。人们对水生态系统中有机聚集体的认识始于20世纪50年
代的海洋学研究。细菌是有机聚集体最重要的组成部分之一。有机聚集体在水体中物质与能量循环中的作用很大程度上是靠附
着其上的异养细菌而起作用的。目前，有机聚集体的概念在水生态系统中已被广泛接受，由于其独特的物理、化学及生物组成，
以及复杂的形成、转化过程，使其在水生态系统中具有重要的生态学作用。然而，有关浅水湖泊中有机聚集体上细菌群落的研
究目前尚未见报道。近年来，基于DNA多聚酶链式反应(PCR)的末端限制性片段长度多态性(T．RFLP)技术是一种新兴的
研究微生物多态性的分子生物学方法。该技术由于具有快捷、高分辨率、高通量和不依赖于培养等优点而被广泛应用于微生物
群落结构的时空演替研究。本研究采用T-RFLP技术，研究了太湖梁溪河入湖河口(SiteA)和贡湖湾(SiteB)2006年6月至
2007年5月一年间有机聚集体上附着细菌群落组成的时空变化规律。T-RFLP分析检测到这两个采样点共有187个独特的末端
限制性片段(T-RFs)，月平均T-RFs分别42．7和44．9。t检验显示它们没有显著性差异(P>o．05)。虽然河口的营养盐浓度要显著
高于贡湖湾(P显示，除了春季外，河口和贡湖湾有机聚集体上细菌群落结构有明显的不同。在T．I球LP分析附着细菌群落组成及季节变化的
基础上，采用多元统计方法研究环境因子与附着细菌群落组成变化的相关性。典型对应分析(cCA)结果表明诸多环境因子中，
DIP、DIN和水温与有机聚集体上细菌群落结构的变化具有显著的相关性(P<0．05)。
关键词：细菌群落结构；有机聚集体；浅水湖泊；T-RFLP；微生物生态学
中图分类号：Q938．1 文献标识码：A 文章编号：1008-8873(2008)05—341-05
收稿日期：2008—09—10收稿，2008．10-10接受
基金项目：国家自然科学基金(40573062)；中国科学院知识创新工程重要方向性项目(kzcx2一yw一419)
作者简介：汤祥明(1976一)，男，博士研究生，研究方向为环境微生物分子生态学。
·通讯作者，E-mail：aiabaCalniglas．∞．∞
万方数据
i丝 竺查型堂里!旦!竺星!里垒!墨!!塑堡 兰!鲞
1 Introduction
Organicaggregates(oA)havebecome‘hotspots”in
termsofenergyflux，仃ansformationofnu rients，and
biogeochemicalcyclingofelementsin aquatic
environments【1’21．Whileorganicaggr gatesinmarine，
deeplakeandloticsystemshavebeen
well-characterized．1ittleisknownabouttheorganic
aggregate—attachedbacterialcommunities(o≯mc)and
theirecologicalimportanceinlargeshallowakes【2，31．
Terminalrestrictionfragmentlengthpolymorphism
(T—RFLP)analysis[4Jhasbeenproventobeaconsistent,
high—resolution．and high·throughput
cultivation—independenttechniquetomonitormicrobial
communitystructures．Terminalr strictionfragment
(T—RF)patternsar generatedfromisolatedDNA，
targetingheconserved16SrRNAgene．Thisparticular
techniqueallowsquickanalysisof bacterial
communitieswithoutcul ivation．
Inthistudy，thestructureandynamicsofthe
OA—attachedbacterialcommunitiesin thelarge
subtropicalsh loweutrophicLakeTaihuwere
investigatedusingT—RFLPanalysis．Forcomparisonof
intra—habitatdifferencesofOA．attachedbacterial
communitystructure，twosi esereselectedfor
investigationoveraperiodoftwelvemonths．Thetwo
sitesdifferstronglyin theireutrophicationand
microbialfoodwebdynamicsl引．Themajoraimofthe
presentstudywasto(1)explorethest uctureand
dynamicsoftheOABCinLakeTaihu，andto(2)to
revealpossiblesite—specificdifferencesin the
interactionbetweenOABCandenvironmentalfactors．
2 MaterialsandMethods
2．1Samplecollection
Samplesw recollectedfromtwositesofLakeTaihu
(Fig．1)：ashallow(maximumdepth，3．3)，turbulent，
eutrophicIakeIocatedin asternChina[61．Thetwosites
representdifferenteco ypes，includingaeutrophicarea
(rivermouthofMeiliangBay,SiteA)，anda
macrophytedominatedrea(GonghuBay,SiteB1．
Surfacewater(top50cm)inthetwositeswascollected
monthlyformJune2006toMay2007．AndtheOA
samplesw reobtainedbycentrifugationaspreviously
de cribed[31．Thetwositesdifferedinanumberof
chemical，physicalandbiologicalvariables．Thevariables
werem asuredaccordingtostandardmethods用．
喏
～节⋯。虹ity
N
≮秽争
k一、／P
@8、——!一
Fig．1Mapofthenorthernpa tofLakeTaihushowingthe
locationsoftheamplingsitesintherivermouthofLiangxi
River(SiteA)andtheGonghuBay(SiteB)
2·2 DNAextraction．PCRamplification
TotalDNAforOA．associatedmicroorganismsWas
extractedusingproteinaseKsodiumdodecylsulfate
(SDS)andcetyltrimethylammoniumbro ide(CTAB)
concomitantwithchloroformextractionand
isopropanolprecipitationacc rdingtotheprevious
protocol【6J．CrudeDNAextractswerepurified
immediatelywiththeE．Z．N．A．9Cycle．PureKit
(Omega)．PCRWasperformedwithCy5labeled
eubacterialforward primer 8F
(5’-AGAGTTTGATCMTGGCTCAG．3’)anduniversal
reverse primer 1492R
(5’．GGTTACCTTGTTACGACTT．3’)【91，whichare
targetedto16SrDNAuniversally—conservedregionsto
permittheamplificationofnapproximately1,500bp
fragment．
2．3EnzymedigestionandT—RFLP
ThePCRproductswerepurifiedanddigested
simultaneouslywithworestrictionenzymesHaelIIand
MvnI(MmFer entas，Germany)．Thedigestionswere
performedfor3 hat37。Cin20plreactionv lumes
containing10plPCRproducts，2pl10×buffers，6山
sterileMilli-Qwater,and1肛l(1OU／I．tleach)ofthe
万方数据
5期 汤祥明，等：太湖有机聚集体上附着细菌群落结构与动态的T-RFLP分析 343
Table1 Physical，chemical,andbiologicalparametersdeterminedforthetwosamplingsitesofLakeTaihuintheperiodfrom
June2006toMay2007．ThedataofthetwosampHngsiteswerecomparedbyt-test．
restrictionenzymes．Thedigestedproductswere
purifiedbythanolprecipitationandthepelletWaSthen
resuspendedin20灶ofsterileMilli-Qwater．The
T-RFLPpatternswereanalysedwithaBeckmanCoulter
’CEQ8000GeneticAnalysisSy tem,usingtheCEQ
DNASize。StandardKit．600．T．RFsizesandT—RF
quantities(peakheights弱rfulweredeterminedwith
theCEQ8000geneticanalysissoftwareV9．0usinga
signaldetectionhresholdof100rfu．Binarym trices
werecreatedwherebythepresence(1)orabsence(0)of
peaks．TherelativeabundanceofT·RFsWaSdetermined
bycalculatingtheratiobetweenthepeakareaofeach
peakandthetotalpeakareaofa11peakswithino e
sample．Ratioswerec nvenedi topercentages，andthe
resultsareusedforcomparingtheOABCwiththe
AFLPprogramoftheCEQ8000software．
2．4Statisticalanalysis
Thestatisticalan ysesw reperformedseparatelyfor
dataofSiteAandSiteBbyusingStatisticalPackage
fortheSocialSciences(sPss)forWindows15．0．T-test
WaSusedtotestthedifferencesofparametersbetween
thetwosims．CanonicalCorrespondenceAnalysis
(CCA)WaSappliedforevealingrelationshipbetween
themembersoforganicaggregate-attachedbacterial
communitycompositionsandthemajorenvironmental
variablescontrollingthecompositionsofdifferent
members．Alldatawerelog@+1)transformed．m
CCAWaSperformedwiththesoftwareCANOCO4．5
(SCIENTIASoftware)byusingunimodalmethod
becauseDe臼endedCorrespondenceAnalysis(DCA)
tunonspeciesvariablesindicatedthatthelengthofthe
firstaxisWaS>2．Thesignificanceofthefirstordination
andcanonicalaxestogetherwasassessedin
permutationtestswith499unrestrictedMonteCarlo
permutations．
3 ResultsandDiscussion
3．1Majorecologicaldifferencesatthetwostudy
sites
Mainecologicalparametersmea uredatthetwositesof
LakeTaihud ringtheone·years mplingperiodwere
summarizedinTable1．Majorbetween-sitesdifferences
inchemo—physicalparametersw eobservedforpH，
dissolvedoxygen(DO)，conductivity(Cond)，secchi
万方数据
344 生态科学EcologicalScience 27卷
depth(SD)，andconcentrationsofmajornutrients
(Table1)．Theconcentrationsofotalphosphorus(TP)，
totalnitrogen(T№，anddissolvedcarbon(DeC)were
significantlyhigheratSiteAthanthoseatSiteB．These
witllin·lakedifferencesofTP．TNandother
chemo-physicalparametersw efromthegeographic
environmentalgradients．SiteAislocatedinahJighly
eutrophicareaattherivermouth；theeutrophicationis
dueto nitrogenandphosphorusf omdomestic
wastewaterdischargedfromtheLiangxiRiver(Fig．1)．
SiteBislocatedinGonghuBay，whichischaracterized
bysubmersedmacrophytecommunitiesandclearwater，
withrelativelylowphytoplanktonconcentrations(Fig．1，
Table11．
3．2VariationsinOABC
AnalysisofthemicrobialcommunitieswimT-RFLP
identifiedatotalof187operationaltaxonomicun ts
(OT∽．ItdemonstratesthaOAmicrohabitatsharbor
diversebacterialcommunities．Thedynamicsofthe
numberofuniqueT-RFsareshowni Fig．2．The
numberofuniqueT-RFs(orOTUs)isan ndicatorof
thediversityofoA．attachedbacterialcommunity．T．1己F
sizesbetween60．640ntwereincludedintheresult．The
mostabundantT-RFsizeswererecordedb tween
60-400nt．AmongtheT-RFs，someofth mmayhave
beenderivedfromcyanobacteriabecausethebacterial
primers8F and1492Rcouldalsoamplify
cyanobacterial16S RNAgenes．Althoughhenutrient
leveldiffersignificantly,therearenosignificantly
differencesinmicrobiald versityatthetwostudied
sites(ThemonthlymeannumberofT-RFsis42．7and
44．9．respectively；尸>0．05)．Thisresultindicatedthat
thenutrientl velat hetwositesi nothelimitedfactor
．controlledth OA．．attachedmicrobiald versity．The
abundanceofT-RFsin ummerissignificantlylower
thanthatofotherseasons(ANO、，A，P<0．05)，which
demonstratedth thhightemperaturendalgaebloom
mightavea negativenfluenceonOA．attached
bacterialdiversity．Moredetailedstudiesshouldbe
examined．
AlthoughthedifferenceoftheOA．attached
bacterialdiversitiesisminoratthetwosites，the
compositionofhebacteriaIcommunitiesdiff redalot．
Distinctspatialdifferenceswerevidenti heCCA
plotofOABC(Fig．31．Mostsamplesfthetwosites
wereseparateddistinctly．OnlyMarchandApril
samplescollectedfromtheRiverMouthandJatmary，
FebruaryndMarchsamplescollectedfromthe
GonghuBayweregroupedintoacluster．Horizontal
heterogeneityof bacteroplanktonmicrobial
communitiesexistedbetweenhabitatshasbeen
observedinLakeTaihuinpreviousstudy[10-121．111e
observedOABCheterogeneitywithinhabitatsmay
correlatetoth ircorrespondinge v ronmentalfactorsin
thepresentstudy．
口RiverMouth
JunJuAugsepoctNovDecJanFebMatAprMay
Samplingtime
Fig．2SeasonalchangesinthenumberofT-RFLPpeaksand
theconcentrationsofDIPatthetwosamplings tes：SiteA(o)
andSiteB(A)inLakeTalhu．
3．3Majorenvironmentalvariab esregulatingoABC
ResultsofCCAillustratedthathedifferencesi
OABCbetweenthetwosamplingsitesarerelatedo
thenvironmentalfactors(Fig．3)．DIP，DINand
watertemperaturecontributedsignificantlyto
varianceinOABC伊<0．05)．Theeigenvaluesofth
firstandsecondaxiswere0．252and0．23l，
respect vely．Inthepresentstudy，thetrophicstate
wasthemostimportantenvironmentalfac orin
structuringtheOABC(Fig．3)．Thetwoaxes
explained33．3％oftheobservedvariationinOABC，
however．whichind atedthatotherfactorssuchas
protozoaandzooplanktonsmayplayanimportant
ffectinstructuringtheOABC．
一．．一里一dI凸
御
御
m
伪
伽
∞
∞
柚
∞
o
∞
加
∞
∞
柏
∞
∞
∞
0
卫母mo正JL年j_，o．oz
万方数据
5期 汤祥明，等：太湖有机聚集体上附着细菌群落结构与动态的T-RFLP分析 345
Fig．3Canonicalcorrespondenceanalysis(ccA)biplots
showingdifferentOABCin relationto themain
environmentalfactorsatthetwositesinLakeTaihu．Samples
collectedmonthlyfromJune2006toMay2007atRiver
Mouth(▲)andGonghuBay(o)．SampHngtimesare
曩djacenttothesymboh．
Nutrientconcentrationandwatertemperaturemay
directlyinfluencebacterialbiomassaswellas
communitycompositionthr ugheffectsongrowth．In
ourstudy，thelowestconcentrationsofDIPwere
observedduringspring(Fig．2)．Althoughalgaend
bacteriaproducealkalinephosphatasesinordertouse
organicformsofP，thiscouldnotimmediately
compensatetheffectsofDIPdepletion．Severalstudies
haveshownthatwatertemperaturendeficiencyof
DIPcouldplayanimportantroleinregulating
bacterioplanktoncommu itycompositionsinLake
Taihu【510121．Togetherwithpr viousstudies，ourdata
suggeststhatnutrientconcentrationndwater
temperaturelsostructuresth spatialandtemporal
variationsofOABC．
Inconclusion，thisstudyisamongthefirstreports
onaggregate—associatedbacterialdynamicsinalarge，
shallowaquaticsystem．Ourresultsdemonstrateh t
OAmicrohabitatsharbordiversebacterialcommunities．
Hori2：ontalheterogeneityofOABCexistedb tween
habitatssuggestthathetrophicstateplayakeyrolein
sustainingOABCstructures．
References
IlJAzamFandRichardAL．2001．Seasnowmicrocosms叨．
Nature，414：495-498．
【2l SimonM，C-rossartH—P’SchweitzerB，eta1．2002．
Microbialecologyoforganicaggregatesin aquatic
ecosystems[J】AquaticMicrobialEco ogy,28：175-211．
【313 BfickelmarmU，ManzW’NeuT It,eta1．2000．
Characterizationofthemicrobialcommunityofloticorganic
aggregates(rivernowt)intheElbeRiverofGermanyby
cultivationndmolecularmethods[J】．FEMSMicrobiology
Ecology，33(2)：157-170．
【4】LiuWT，MarshTL，ChengH，eta1．1997．Chamaedzation
ofmicrobiald versitybydeterminingterminalrestriction
fragmentlengthpolymorphismsofgenesncoding16S
rRNA[J】．AppliedandEnvironmentalMicrobio ogy，63(11)：
4516-4522．
15lGaoG，QinBQ，SommarugaKeta1．2007．The
bacterioplanktonofLa eTaihu,China：abundance，biomass，
andproduction【J】．Hydrobiologia，581：177·188．
【6】QinBQ，XuPZ，WuQL，eta1．2007．Environmentalissu s
ofLakeTaihu,China【J】．Hydrobiologia，581：3-14．
【71 Jin,XCandTuQY(Eds．)．1990．mStandardMethods
forObservationndAnalysisofLakeEutrophication【MI．
2n正edn．Beijing：ChinaEnviro mentalSciencePr ss．
138-272(inChinese)．
18lZhouJ，BrimsM八andTiedjeJM．1996．DNArecovery
fromsoilsofdiversecomposition【J】．Appliedand
EnvironmentalMicrobiology,62(2)：316-3 2．
【9l NewtonRJ，KentAD，TriplettEW’eta1．2006．Microbial
communitydynamicsin a humiclake：differential
persistenceofcomlnonfreshwaterphylotyp懿叨．
EnvironmentalMicrobiology,8(6)：956．970．
【10】WuQL，ChanYW，XuKD，eta1．2007．Intra-habitat
heterogeneityofm erobialfoodwebstructureunderthe
regimeof utrophicationandsedimentresuspensioninthe
largesubtropicalsh lowLakeTaihu,China【J】．
Hydrobiologia，581(1)：241-254．
111lWu,QL，ZwartGWUJ，订a／．2007．Submersedma rophytes
playakeyroleins仇Icmmgbacterioplanktoncommu ity
compositioninthelarge，shallow,subtropicalTaihuLake，
China田．EnvironmentalMicrobiology,9(11)：2765-2774．
【121XingPandKongFX．2007．Intra—habitatheterogeneityof
environmentalfactors、regulatingbac erioplankton
communitycompositioninI．．akeTailuChina[J】．Aquatic
MicrobialEcology,48(2)：113—122．
万方数据
太湖有机聚集体上附着细菌群落结构与动态的T-RFLP分析
作者： 汤祥明， 高光， 秦伯强， TANG Xiang-ming， GAO Guang， QIN Bo-qiang
作者单位： 汤祥明,TANG Xiang-ming(中国科学院南京地理与湖泊研究所,南京,210008;中国科学院研究
生院,北京,100039)， 高光,秦伯强,GAO Guang,QIN Bo-qiang(中国科学院南京地理与湖泊
研究所,南京,210008)
刊名： 生态科学
英文刊名： ECOLOGICAL SCIENCE
年，卷(期)： 2008,27(5)
被引用次数： 1次

参考文献(12条)
1.Lin W T;Marsh T L;Cheng H Characterization of microbial diversity by determining terminal
restriction fragment length polymorphisms of genes encoding,16S rRNA[外文期刊] 1997(11)
2.Bockelmann U;Manz W;Nen T R Characterization of the microbial community of lotic organic
aggregates (river snow).in the Elbe River of Germany by cultivation and molecular methods[外文期刊
] 2000(02)
3.Simon M;Grossart H-P;Schweitzer B Microbial ecology of organic aggregates in aquatic ecosystems[外
文期刊] 2002
4.Zhou J;Bruns M A;Tiedje J M DNA recovery from soils of diverse composition[外文期刊] 1996(02)
5.Jin,X C;Tu Q Y The Standard Methods for Observation and Analysis of Lake Eutrophication 1990
6.Qin B Q;Xu P Z;Wu Q L Environmental issues of Lake Taihu,China[外文期刊] 2007
7.Gao G;Qin B Q;Sommaruga R The bacterioplankton of Lake Taihu,China:abundance,biomass,and
production[外文期刊] 2007
8.Newton R J;Kent A D;Triplett E W Microbial community dynamics in a humic lake:differential
persistence of common freshwater phylotypcs[外文期刊] 2006(06)
9.Xing P;Kong F X Intra-habitat heterogeneity of environmental factors regulating bacterioplankton
community composition in Lake Taihu,China[外文期刊] 2007(02)
10.Wu Q L;Zwart G;Wu J Submersed macrophytes play a key role in structuring bacterioplankton
community composition in the large,shallow,subtropical Taihu Lake,China[外文期刊] 2007(11)
11.Wu Q L;Chert Y W;Xu K D Intra-habitat heterogeneity of microbial food web structure under the
regime of eutrophication and sediment resuspension in the large subtropical shallow Lake Taihu,China
[外文期刊] 2007(01)
12.Azam F;Richard A L Sea snow microcosms[外文期刊] 2001

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