食物种类对萼花臂尾轮虫种群增长率的影响
食物种类对萼花臂尾轮虫种群增长率的影
响
动物52(1):70—78,2006
ActaZoologicaSinica
Influenceoffood.typeonthepopulationgrowthrateoftherotifer
Brachionuscalyciflorusinshort-chronicassays
M.L0RLING",W.BEEKMAN
DepartmentofEnvironmentalSciences,AquaticEcologyandWaterQualityManagementGroup,WageningenUniversi
ty,P.0.Box8080,6700DDWageningen,TheNetherlands
AbstractThetypeoffoodgivenduringshortchronicassayswiththerotiferBrachionuscalyct,ZOrUSmightbeoneofthe
sourcesofvariationinthereproductiverateoftherotifers.Tengreenalgalspeciesweresuppliedasmonospecificdietsto
examinevariabilityinrotifergrowthrate.Inaddition,rotiferswerefedcyanobacteriaasmonospecificfoodandinmix—
tureswiththegreenalgaSeenedesmusobliquustotestthehypothesisthatcyanobacteriaareavaluablesupplementincorn—
binationwithfooditemssuchasScenedesmus.ThepopulationgrowthrateofB.calycifloruswasinfluencedsignificantly
bythedifferentgreenalgalfoodspecies.S.obliquuspromotedthehighestgrowthinB.catyci~orus(1.6perday),
whilegrowthonDesmodesmusabundanswasthelowe.st(O.3perday).Aspercentageofmaximalgrowthon
S.obliquus,thesequenceofgreenalgalfoodspeciesis:Desmodesmussubspicatus(88%),lorellavulgaris
(83%),Monoraphidiumminutum(77%),D.quadricauda(74%),S.知
lcatus(71%),S.acuminatus
(69%),S.pectinatus(64%),Chlamydomonasreinhardtii(57%)andD.abundans(19%).Thegrowthdiffer—
enceswerenotexplainedfromsizedifferencesofthealgae.ThecyanobacteriaMicrocystisaeruginosaandSynechococcus
elongatusdepressedgrowthofB.calyct,Zorusnotonlyasmonospecificfoodbutalsoinmixeddietswiththegoodfood
sourceS.obliquus.Thedetrimentaleffectappearednotrelatedtomicrocystins.Theresultsdonotsupportthehypothe—
sisthatnontoxiccyanobacteriaareavaluablesupplementincombinationwithothergreenalgalfoodspecies.Thevariation
ingrowthratesobservedincurrentstudyunderlinestheinfluencefoodspeciesmighthaveonthegrowthofB.calyclflorus
andpotentiallyontheoutcomeoftoxicitytestslActaZoologicaSinica52(1):70—78,2006J.
KeywordsRotifer,Brachionuscalyciflorus,Food,Populationgrowth
食物种类对萼花臂尾轮虫种群增长率的影响
M.LURLINGW.BEEKMAN
*
DepartmentofEnvironmentalSciences,AquaticEcologyandWaterQualityManagementGroup,WageningenUniversi
ty,P.O.Box8080,6700DDWageningen,TheNetherlands
摘要在短期慢性观测过程中,食物类型可能是造成萼花臂尾轮虫
(Brachionuscalyciflor"s)种群繁殖率变化
的一种原因.共观测了分别单独投喂1O种不同绿藻对轮虫种群增长率的影响.为
验证藻青菌是藻类饵料(如绿
藻Scenedesmus)有价值的佐剂这一假说,还用蓝细菌单独投喂或与斜生栅藻
(Scenedesmusobliquus)混合投喂
轮虫进行实验观测.结果发现食物种类对萼花臂尾轮虫种群增长率影响显着.斜生
栅藻组获得最大种群增长
(1.6/d),而Desmodesmus组增长率最低(0.3/d).以占斜生栅藻组最大增长的百分率来
示,其它几种绿藻组
种群增长由高到低依次为:Desmodesmussubspicatus88%,小球藻
(Chlorellnvulgaris)83%,单壳缝藻(Mono—
raphidiumminutum)77%,D.quadricauda74%,S.falcatus71%,S.acuminatus69%,S.pecti
natus64%,
莱茵衣藻(Chlamydomonasreinhardtii)57%,D.abundans19%.轮虫增长率的差异不能用藻类饵料的大小差异
来解释.蓝细菌(Microcystisaeruginosa)和(Synechococcuselongates)不论是单独投喂还是与优良藻类饵料(斜
生栅藻)混合投喂都对萼花臂尾轮虫种群增长有抑制作用.这种副作用似乎与微囊藻素无关.该结果不支持无
毒蓝细菌可作为与其他绿藻饵料配合使用的优良佐剂这一假说.本研究所观察到的生长变化显示了饵料种类对
ReceivedJuly11,2005;acceptedNov.16,2005 *ThisresearchwasfundedbyafellowshipfromtheRoyalNetherlandsAcademyofArtsandS
ciences
**Correspondingauthor.E—mail:mique1.1urling@wur.nl ◎2006动物ActaZoologicaSinica
M.LORL1NGeta1.:AlgaeandBrachionusgrowth71 萼花臂尾轮虫种群增长的影响,也预示了对毒性实验结果的影响[动物52(1):70—78,2006].
关键词萼花臂尾轮虫食物种群增长
Planktonicrotifersareknowntoplayanlmpor—
tantroleintheenergyflowinlakesandponds,and numerousstudieshaveindicatedtheimportanceofro—
tifersasprimaryconsumers(e.g.,Dumont,1977; Pilarska,1977;Pourriot,1977;Walz,1995).
Becauseoftheirimportanceinzooplanktoncommuni—
tiesandaquaticfoodchainsrotifershavebeenusedas modelorganismsinfeedingandfoodchainstudies,in aquacultureresearchandecotoxicologicalresearch (e.g.,Bogdaneta1.,1980;Schlfiteretal 1987;Walz,1995;ChialandPersoone,2003). Inecotoxicologicalresearch,two—daylifecycle
testswiththerotiferBrachionuscalyciflorusin whichmultiplebroodsareproducedandtheFxgener—
ationalsoreproducesareusedextensively(e.g., Snelleta1.,1991;SnellandMoffal,1992;Radix eta1.,1999;ChialandPersoone,2003;Margdlek andB16ha,2004).Incontrasttoacuteassayswith mortalityasendpoint,thetwo—daylifecycletesthas
growthorreproductionasendpoint,whichrequires feedingofthetestorganisms.However,consider—
ablevariabilityintestresultsbetweenstudiesmight occurduetouseofdifferentfoodspeciesandgrowth conditions(ChialandPersoone,2003).Ingeneral, greenalgalspeciessuchasmembersofthecommon generaChlamydomonas,Clorella,Desmodesmus andScenedesmusarefedtoB.calyciflorus,but thereisstillnoconsensusonthenutritionaladequacy ofthefoodspecies.Theresultsofsomestudies,for example,suggestChlorellasp.tobeabetterfood thanScenedesmusspecies(e.g.,Xieta1.,2001; 2002;Flores—Burgoseta1.,2003),whereasinoth—
ersB.calyciflorusgrewbetteronScenedesmusthan onChlorella(e.g.,Rothhaupt,1990a).Compar—
isonoftheresultsfromthesestudies,however,is
difficultasdifferentmethodsforlong—termassays
wereused.
Inseveralstudies,B.calycii一.
orusnthe2d
growthassayswereofferedamonospecificdietofonly livegreenalgae(e.g.,CecchineandSnell,1999; Radixeta1.,2000,2002).However,inother studies.therotiferswerefedacombinedfoodoflive greenalgaeanddriedcyanobacteria,Spirulina (ChialandPersoone,2003,2005).Cyanobacterial speciesmightbeavaluablesupplementincombination withothergreenalgalfoodspeciesas,forexample, astimulatoryeffecthasbeenreportedfor B.calyciflorusfeda3:1mixtureofthegreenalga MonoraphidiumandthecyanobacteriumPlank. tothrix(WeithofandWalz,1995).Incontrast, mixedfeedsofthecyanobacteriumAnabaenaandthe cryptophyteCryptomonas
B.calyciflorus(Gilbert,
hamperedgrowthof
1994,1996).These
studieswereperformedwithfilamentouscyanobacte—
ria.buttoourknowledgeeffectsofmixeddietsof greenalgaewithnon—filamentouscyanobacteriaare virtuallyabsentinliterature.
Theaimofthecurrentstudywastoexaminethe effectoftypeoffoodgivenduringshortchronicas—
sayswiththerotiferBrachionuscalyci. orusonthe
reproductiverateoftherotifers.AccordingtoSnell
andMoffat(1992)growthratesshouldnotbeless thanthe0.65acceptabilitythresholdforshortchron—
ictestsinordernottoinfluencetheoutcomeofthe toxicityassays.Consequently,feedorganisms shouldbeofgoodnutritionalvaluetoavoidunsatisfac—
toryreproduction.Inasmuchasthetypeofgreenal—
galfoodmightseriouslyaffectthegrowthof B.calyciflorus,inthecurrentstudy,10greenalgal speciesasavariableinthegrowthrateoftherotifers werecomparedsimultaneouslyina2-dgrowthassay. Inadditiontotheeffectofthetypeofgreenalgalfood alsotheeffectofthenon—filamentouscyanobacteria
MicrocystisaeruginosaandSynechococcuselongatus asmonospecificfoodandinmixtureswiththegreen algaScenedesmusobliquusisstudied.Thiswasdone totestthehypothesisthatcyanobacteriaareavalu—
ablesupplementincombinationwithfooditemssuch asScenedesmus.
1Materialsandmethods
1.1Organisms
Thegreenalgaeandcyanobacteriausedinthe currentstudyarelistedinTable1.Semi—continuous
stockculturesweremaintainedin300mlcellulose plugclosedErlenmeyerflaskscontaining150mlof modifiedWC(WoodsHoleChu)一medium(Larling
andBeckman,1999)withvitaminsadded(Biotin B1andB12at50ng/L,ThiamnineHCLat
100ng/L).Flaskswithgreenalgaewerekeptincli—
matecontrolledroomat20?incontinuouslightof
60一ttmolquanta/m/sandeverythreeweeksforat leastthreemonthspriortotheexperiments,aliquots weretransferredseparatelyintofreshsterilemedium. Flaskswithcyanobacteriawerekeptat20?andin
45ttmolquantam/m2/slightina16:8hlight:dark rhythm.
TherotiferBrachionuscalyci疗.
orusPallaswas
obtainedfromMicrobiotestsInc.(Nazareth,Bel—
glum)andwassuppliedascysts.Cystswerekept refrigerateduntiluse.
72动物52卷
FoodtypesusedinExp11:
100%cyanobacterium一0%S.obliquus;75%一25%;50%一50%;25%一75%;
100%;75%;50%;25%;0%S.obliquus
1:NorwegianInstituteforWaterResearch 1.2Exp.I:effectofgreenalgae
Thefirstexperimentevaluatedtheeffectof10 differentgreenalgalspecies(Table1)onrotifer growthrate.Toobtainexponentiallygrowingalgae withgoodnutritionalvalue,aliquotsofgreenalgae fromthesemi..continuousstockculturesweretrans.. ferredpriortotheexperimentinto100一mlErlenmey—
erflasks:containing50mlWCmediumwithvita—
minsaddedandthatwereclosedwithacellulose plug.Algaldensitiesandparticlesizedistributions weredeterminedinthesizerange2.5—35btmequiv—
alentsphericaldiameterusingaCoulteMultisizer
II(capillary100p.morificewidth,CoulterElec—
tronicsLimited,Luton,England).Eachflaskre—
ceivedidenticalinitialconcentrationofalgae.i.e.5 ×100um/m1.Theseflaskswereplacedonarotat—
ingshakingtable(60r/m)at24?incontinuous
lightof100一t~molquanta/m/s.Afterthreedaysthe algaeweretransferredagainatidenticalconcentra—
tionsof5×10.um/mlinto100mlErlenmeyer'sthat contained50一mlWC—medium.Afterthreedaysthe algaewereusedintheexperiment.MeanwhileB. calycifloruscystswerehatchedin100一mlsuspensions
ofeachofthecorrespondinggreenalgaeintheabove mentionedWC-medium.Thehatchingflaskswere placedonarotatingshakingdeviceat24?incontin—
UOUSlightof100一ttmolquanta/m2/s.Thecystswere hatchedinthedesiredfoodinasmuchasashort—
prefeedingperiod(2hafterhatching)hasbeen foundtohaveapronouncedpositiveeffectongrowth rates(ChialandPersoone,2005).
Theexperimentwasrunin6一welledcoveredcul—
tureplatespositionedfortwodaysonarotatingshak—
ingtable(60r/m)indarknessinaclimate-con—
trolledroomat24?.Eachwellwasfilledwith10一
mlgreenalgalfoodsuspensionsatidenticalconcentra—
tionsof10m/ml(?5mgC/L).F0reachfood
type(Table1)sixwellswereusedofwhichthree receivedsixfreshlyhatched(2—3hold)BTa—
chionus,whilethreeotherreplicatesreceivednoro—
tifers.Thenewbornrotiferswerecollectedinadish
with15mlfreshWC—mediumandwerecountedand
transferredfromthedishtothecultureplatesundera dissectingmicroscopeat20×magnificationusinga
preparedglassfinetippipette.Theexperimentwas runfor48hoursafterwhichchlorophyl1.aconcentra. tionsweredeterminedin1.5一mlsamplesfromeach
wellusingaPHYTo—PAMphytoplanktonanalyzer
(HeinzWalzGmbH,Effeltrich,Germany).Thisis anon—destructivemethodinwhichchlorophyllcon—
centrationsareusedtoestimatealgaldisappearance ratesandtoestimaterotiferclearancerates(Lfirling andVerschoor,2003).C1earancerates(,ml/ ind./h)werecalculatedusingtheequation:
l期M.LORLINGeta1.:AlgaeandBrachionusgrowth73 CR:f\ln(At1)一In(Atife)At)×
whereAc0ntrolisthealgalconcentration(aschloro—
phyll—a(ChlFn)int~g/LI1)after2daysincontrols, Ar0tiferisthealgalconcentrationintherotifertreat—
ments,?,isthetime(h),Vistheculturevolume (m1)andNisthenumberofanimals.ForNmean valueswereestimatedusingthepopulationgrowth rateandtheexponentialgrowthmode1.Algaldisap—
pearancerates(Dr,perday)wereestimatedfrom: [In(Chla?盯fe)一In(Chlacontro1)].
r=——————————————?————————————一,where Dristhealgaldisappearancerate(perday),Chla. cantrolisthechlorophyll—aconcentration(int~g/L)af—
ter2daysincontrolsandChlarotiferisthechlorophyll—
aconcentration(int~g/L)after2daysinpresence
ofrotifers.
Thenadding0.5mlofLugol'sfixativetoeach wellterminatedtheexperiment.Thenumbersofro—
tiferswerecountedunderadissectingmicroscopeand populationgrowthrateswerecalculatedfrom: :
,where.theristher——————————————一,
populationgrowthrate(perday),N2isthenumber ofanimalsafter2daysandN1theinitialnumberof rotifers(=6).
1.3Exp.1I:effectofcyanobacteria
Asecondexperimentwasruntostudytheeffect onrotifergrowthrateofmixturesofcyanobacteria withS.obliquusthatincluded0%,25%,50%, 75%and100%cyanobacteria(Table1).The cyanobacteriawereharvestedfromone..weekoldcu1.. turesmaintainedatidenticalconditionsasthe cyanobacterialstockcultures(20?andin45/tmol
quanta/m/slightina16:8hlight:darkrhythm), S.obliquusfromacultureasoutlinedabove.Each food—typewasrunintriplicateat10/tm'/m1.The experimentwasruninsix—welledcultureplatesin
darknessinaclimate—controlledroomat20?1?.
After2days,thenumbersofrotiferswerecounted underadissectingmicroscope.Growthrateswere calculatedasoutlinedabove(Exp.I).
1.4Statistics
Populationgrowthratesandalgaldisappearance ratesinExp.1wereanalyzedbyone—wayANoVA
inthestatisticaltoolpackSPSS10.1.0(SPSS.
2000).InExp.1I,meanpopulationgrowthrates werecomparedrunningtwo—wayANOVAwith
cyanobacterialspeciesandfoodcompositionasthe fixedfactors.Forcomparisonsamongtreatmentsand speciesalsoone—wayANoVAwererun.Differences
betweenmeansweredistinguishedbyTukey'spost hoccomparisontest(P<0.05).InExp.I,rela—
tionshipsbetweengreenalgalparticlevolume,algal disappearancerates,rotiferclearanceratesandpopu—
lationgrowthwereestimatedbynon—linearregression
usingasigmoidmodel(four-parameterGompertfunc—
tion)inthetool-packSigmaPlot2000,version6.O0. 2Results
2.1Exp.I:effectofgreenalgae
The10differentgreenalgalfoodspeciesexerted distincteffectsonthepopulationgrowthrateofB. calyciflorus(Fig.1).Theone—wayANoVAindi—
catedthatgrowthratesweresignificantlydifferent betweenfoodtypes(F9.2o=155.3,P<0.001). Tukey'spost—hoccomparisonrevealed8homogenous groupsthatweresignificantlydifferent(Fig.1). WiththeexceptionofD.abundansallgreenalgal speciessupportedgoodtoexcellentgrowthin B.calyciflorus.Thehighestgrowthrateswereob—
servedwhenS.obliquuswasofferedasfood. Growthasapercentageofthemaximum(on
S.obliquus)growthrateforothergreenalgaevaried between19%(D.abundansasfood)and88%
(D.subspicatusasfood).Formostspeciesthevari—
ancecoefficientswerelow(<7%).however,for
D.abundansitwas47%(Fig.1).Greenalgaldis—
appearancerateswerealsosignificantlydifferent (F9.2o:41.6,P<0.001)betweenfood.types (Fig.2).Tukey'spost—hoccomparisonrevealed6
homogenousgroupswiththehighestdisappearance rateforS.obliquusandthelowestforD.abundans. C.reinhardtiiandS.pectinatus(Fig.2).Esti—
matedclearanceratesdifferedsignificantly(F9.2o 3.05,P=0.018),butTukey'stestrevealedthat onlytheclearanceratesofrotifersfedC.reinhardtii weresignificantlylowerthanthosefeedingon S.acuminatus(Fig.3).Meflnrotiferclearance ratesvariedbetween1.5and7"lmicroliter/rotifer/h (Fig.3).Themeanparticlevolumesofthegreenal—
gaeshowedconsiderablevariation(Table2).Be—
causethedatadidnotsatisfyassumptionsforANO—
VA.separatet.testswererun.Thesetestsshowed thatparticlevolumesofD.quadricaudaand S.pectinatusweresignificantlylargerthanofallthe othergreenalgae,whilethoseofM.minutumand Chlorellavulgariswerethesmallest(Table2). Non—linearregressionanalysesyieldednosignificant relationshipbetweentheparticlevolumeandthealgal disappearancerate(r2di=0.00,F=0.82,P= 0.528),clearancerates(r2adi=0.00,F=0.02,P =0.994)orrotifergrowthrates(r2di=0.00,F= 0.93,P=0.481).Algaldisappearanceratesand rotifergrowthrates(r2di=0.597,F:5.45,P= 0.038)aswellasalgaldisappearanceratesandclear—
ancerates(ra2di=0.578,F=5.12,P=0.043)
weresignificantlycorrelated,butclearanceratesand populationgrowthratesshowednocorrelation(ad2i :0.190,F=1.70,P=0.265).
74动物52卷
?
蒸
荟
蓍
善
Greenalgalf0odspecies
Fig.1Populationgrowthrate(perday)oftherotiferBrachionuscalyciflorusinshort?chronic
assays(2-d)fed
withtendifferentgreenalgalspecies Errorbarsindicateonestandarddeviation(n=3).Similarsymbolsabovebarsindicatehomog
eneousgroupsthatarenotsignificantly different(Tukey'sposthoccomparison,P<0.05). ?
矗
e
罢
8
l
O
一
0.2
-
0-4
-
0.6
-
0.8
c,
Greenalgalf0odspecies
Fig.2Disappearancerate(perday)oftendifferentgreenalgalspeciesusedasfoodforth