Lecture5-2MicrobialGrowthCHAPTER6BROCKBIOLOGYOFSchoolofLifeScienceandBiotechnologyJiaoTongIII.MeasuringMicrobialShanghaiJiaoTongGrowthHowtocountcellnumbersinaDirectcountofmicrobialTotalcountViablecounttecountingIndirectcountofmicrobialgrowthTurbidimetricShanghaiJiaoTongShanghaiShanghaiJiaoTongDirectmeasurementsofmicrobialgrowth:totalandviablecountsTotalandviablecountingmethodseachhavetheirstrengthsandweaknesses.總菌計(jì)數(shù)與活菌 ShanghaiJiaoTongShanghaiJiaoTongDirectMeasurements1:TotalcellcountsDirectmicroscopiccount:Thenumberofcellsinapopulationcanbemeasuredbycountingasampleunderthemicroscope.直接顯微,Withliquidsamples,specialcountingShanghaiJiaoTongDirectmicroscopiccountingprocedureusingthePetroff-Haussercountingchamber ShanghaiJiaoTongShanghaiJiaoTongShanghaiJiaoTongDeadcellsandlivingcellsareallcountedSmallcellsaredifficulttocountNeedphasecontrastmicroscopeNeedhighcellconcentration,106cells/ml(why?)PrecisionisdifficulttoachieveMotilecellsmustbeimmobilizedbeforecountingShanghaiJiaoTongShanghaiShanghaiJiaoTong DirectMeasurements2:ViableCount活菌記數(shù)ShanghaiJiaoTongtetecount/colonycount):Twomethodsofatetecount/colonycount):ShanghaiJiaoTongShanghaiJiaoTongDilutingcellsuspensionsbeforeWiththe temethods,itisimportantthatthenumberofcoloniesdevelo onthe notbetoolarge.Toobtaintheappropriatecolonynumber,thesampletobecountedmustalmostalwaysbeShanghaiJiaoTongDilutingcellsuspensions tewillbemembranefiltrationprocedure.Membranewithdifferentporesizesareusedtotrapdifferentmicroorganisms.IncubationtimesformembranesalsovarywiththemediumandShanghaiJiaoTongLimitationsof tingcounting平 Singlecolonymayarisefrommorethanonecell(cellcfu/mlcolony-formingunit/ml菌落形成單位ToomanyInoculumSuitabilityofIncubationLengthofSizeofShanghaiJiaoTongShanghaiShanghaiJiaoTongMembrane-filteredsamplesgrownonavarietyofmedia.StandardnutrientmediaforatotalbacterialFecalcoliformmediumfordetectingfecalcoliformsthatformbluem-EndoagarfordetectingE.coliandothercoliformsthatproducecolonieswithagreensheen.mol.nghaiShanghaiJiaoTongIndirectMeasurements:Turbidity濁度Principle:acellsuspensionlookscloudy(turbid)totheeyebecausecellsscatterlightpassingthroughthesuspension.Themorecellspresent,themorelightscatteredandhencethemoreturbidthesuspension.ShanghaiJiaoTongShanghaiJiaoTongTurbidityTurbiditymeasurementsofmicrobial(a)MeasurementsofturbidityaremadeinShanghaiJiaoTongLimitationsofturbiditycountingDeadcellsandlivingcellsarenotParticlesinculturedisturbmeasurementNotgoodathighShanghaiJiaoTongContinuousculturethechemostat連續(xù)培養(yǎng):恒化器CanwemaintainapopulationatexponentialgrowthFixedvolumeofcultureGrowthrateandgrowthyieldarecontrolledsimultaneously,notyoucando,FlowsystemofconstantContinuousadditionofmediumandcontinuousremovalofSteadystateGrowthrateandgrowthyieldcanbemanipulatedindependentlyContinuousculture-thechemostatInsuchadevice,thepopulationdensityiscontrolledbytheconcentrationoflimitingnutrientinthereservoir,andthegrowthrateiscontrolledbytheflowrate.BothparameterscanbesetbytheRelationshipbetweennutrientconcentration,growthrate,andgrowthyieldinabatchculture.Atlownutrientconcentrationsbothgrowthrateandgrowthyieldareaffected.ShanghaiJiaoTongGrowthrateandgrowthyieldcanbecontrolledindependentlyinachemostatGrowthratecontrolFlowrateofthefreshmediumintothesystem-dilutionGrowthyieldcontrol(populationdensity)MaintainingapopulationatitsexponentialgrowthphaseforalongtimeShanghaiJiaoTongSteady-staterelationshipsintheThedilutionrateisdeterminedfromtheflowrateandthevolumeoftheculturevessel.稀釋率由流速及培養(yǎng)器體積決定.Athighdilutionrates,growthcannotbalancedilutionandthepopulationwashesSteady-staterelationshipsintheAlthoughthepopulationdensityremainsconstantduringsteadystate,thegro