EnvironmentalMicrobiologyWhatcanyouknowfromthetitleofthiscourseshownabove?Youmusthavealreadybeentoldorleantsomethingaboutmicroorganisms.Tellmewhatarethey(e.gmicroorganisms)allwhatyouhaveknown.Hints:SARS,cold,hospital,skin/muscle(肌肉)cut,diarrhea(痢疾)

size,shape…….Question1Question2*Chapter1Introduction

*WhatisMicrobiology?Microbiologyisthestudyoforganismstoosmalltobeclearlyseenbytheunaidedeye(i.e.,microorganisms)andhowtheyareliving/working/surviving.MicroorganismsincludeBacteria(細(xì)菌)0.5~2μmArchaea(古菌)0.5~2μmFungi(真菌)2~200μmProtozoa(原生動物)mm~cmAlgae(藻類)Viruses(病毒)20~300nm*Besidetheirsmallinsize,theyare:Basicallyunicellular,theirlifedosenotdependonothercells.ItmeansthatONEcellisacompletelife,whichdistinguishesfromothermulticulturalorganisms.Diverseintheirlivinghabits(adaptation,livealmosteverywherewherelifeispossibleHighmetabolicactivities,fastreproduction.

morenumerousthananyotherkindoforganisms(geneticdiversity)globalecosystemdependsontheiractivitiesinfluencehumansocietyinmanywaysMicroorganismsarethoughttohavebeenlivingontheearthfor2.3~4.7billionyears.Howmanyyearshavehumanbeingsbeenlivingontheearth?*生物演化時間略表OriginVertebrate脊椎動物EukaryotesInvertebrate無脊椎動物Dinosaurs恐龍HumanProkaryotesonly40億年*Microbiologycoverse.g.,microbialmorphologye.g.,microbialphysiologye.g.,microbialgeneticsMicrobialtaxonomyMicrobialcytology*FieldsofMicrobiologyVirology=StudyofvirusesBacteriology=StudyofbacteriaPhycology=StudyofalgaeMycology=StudyoffungiProtozoology=StudyofprotozoaFoodanddairymicrobiologyAgriculturalmicrobiologyMicrobialecologyMedicalmicrobiologyMicrobialgeneticsandmolecularbiology*Wherearevirus?Currentsystemofclassification*WhatisEnvironmentalMicrobiology?EnvironmentalMicrobiologycoversthesamesubjectsstatedinpreviousslide,butstressesontheorganismsandtheirrolesinproducingand/oreliminatingenvironmentalproblems.SubjectincludesBiologicaltreatmentofwastes(liquid,solid,gaseous)BiogeochemicalrolesinelementcyclingMicrobialtransformationoftoxicmaterialsinenvironmentsInteractionsbetweenthem/otherorganismsMicroorganismsrelevanttopublichealthToolsandtechnologiestostudy/knowthem*BriefhistoryofMicrobiologySeehowMicrobiologyasasciencewasdevelopedandthinkwhatwecanknowfromthehistory.1Observationtools2Fightagainstfooddecayanddiseases3Fromlaboratorytonaturalenvironment*AntonyVanLeeuwenhock(1632-1723)Dutch

*DiscoveryofMicroorganismsAntonyvanLeeuwenhoek(1632-1723)Builtmorethan500microscopesfirstpersontoobserveanddescribemicroorganismsaccuratelyKnowntodayasthefatherofprotozoologyandbacteriology**Firstmicroorganismseen*GoldenAgeofMicrobiology

(1857-1914)Isspontaneousgenerationofmicrobiallifeexists?Whatcausesfermentation?Whatcausesdiseases?Howcanwepreventinfectionanddisease?*LouisPasteur(1822-1895)*LouisPasteurLouisPasteurRejectedtheoryofspontaneousgenerationdemonstratedthatalcoholfermentationsandotherfermentationsweretheresultofmicrobialactivitydevelopedtheprocessofpasteurizationtopreservewineduringstoragebyheatingwinejustenoughtokillbacteria*Pasteur’sexperimentwiththeswan-neckedflaskNonsterileliquidpouredintoflaskNeckofflaskdrawnoutinflameLiquidsterilizedbyheatingLiquidcooledslowlyDustandmicroorganismstrapepedinbendLiquidremainssterileformanyyearsFlasktippedsomicroorganisms-ladendustcontactssterileliquidMicroorganismsgrowinliquid*Swan-neckedflask*Tyndall’sDust-FreeBox*JosephLister(1827-1912)*JosephLister(1827-1912)Developedasystemofantiseptic(無菌)surgery(外科手術(shù))designedtopreventmicroorganismsfromenteringwoundsSurgicalinstrumentswereheatsterilizedPhenolpreventedwoundinfectionbecauseitkilledbacteria*RobertKoch(1843-1910)*RecognitionoftheRelationshipbetweenMicroorganismsandDiseaseRobertKoch(1843-1910)establishedtherelationshipbetweenBacillusanthracisandanthraxAnnouncedthatcauseoftuberculosis(肺結(jié)核)isarod-shapedbacterium;Mycobacteriumtuberculosis*OneofKoch’sfirstmicrographsshowingBacillusanthracis*Koch’spostulatesThemicroorganismmustbepresentineverycaseofthediseasebutabsentfromhealthyindividuals.Thesuspectedmicroorganismmustbeisolatedandgrowninapureculture.Thesamediseasemustresultwhentheisolatedmicroorganismisinoculatedintoahealthyhost.Thesamemicroorganismmustbeisolatedagainfromthediseasedhost.*DevelopmentofTechniquesforStudyingMicrobialPathogensKoch’sworkledtodiscoveryordevelopmentof:agarpetridishnutrientbrothandnutrientagarmethodsforisolatingmicroorganisms*FannieHessesuggestedtheuseofagarassolidifyingagentAgarwasnotattackedbybacteriaDidnotmeltuntilreachingatemperatureof1000CRichardPetridevelopedthepetridish,acontainerforsolidculturemediathatiscurrentlyusedineverymicrolaballovertheworld.*CharlesChamberland(1851-1908)*CharlesChamberland(1851-1908)discoveredvirusesandtheirroleindiseasedevelopedporcelainbacterialfilterdiscoveredthefirstviralpathogen,TobaccoMosaicVirusTMV*Otherdevelopments…

ImmunologicalstudiesPasteurandRouxdiscoveredthatincubationofculturesforlongintervalsbetweentransferscausedpathogenstolosetheirabilitytocausediseaseAttenuatedcultureisavaccinePasteurandhiscoworkersdevelopedvaccinesforchickencholera(霍亂),anthrax(炭疽),andrabies(狂犬病)*Moredevelopments…EmilvonBehring(1854-1917)andShibasaburo

Kitasato(1852-1931)developedantitoxinsfordiphtheriaandtetanusevidenceforhumoralimmunityElie

Metchnikoff(1845-1916)discoveredbacteria-engulfing,phagocyticcellsinthebloodevidenceforcellularimmunity*–extremelyimportantconceptualdevelopmentinmicrobiology(andinmicrobialecology,too) removeorganismsfromcomplexcommunities isolatekeyprocesses obtainreproducibleresults ThismethodisstillusedtodayAttitudeofKoch’stime:“WorkwithimpureculturesyieldsnothingbutnonsenseandPenicillium

glaucum“(OscarBrefield1881)PureCultureParadigm(范例)*SirAlexanderFleming(1929),examiningexactlysuchanimpureculture(StaphylococcusculturecontaminatedbyPenicillium),ledtothediscoveryofpenicillin.AgarpetridishStaphylococcuscoloniesPenicillium

contaminantzoneofnobacterialgrowth,duetopenicillinproducedbyfungusInterferencecompetition!classicecologicalprocess*Sergei

Winogradsky

(ARussianmicrobiologist1856-1953)

Bacteria:centralinelementtransformationsFounderofsoilmicrobiology-isolatednitrifyingbacteria

winogradskycolumn:microbialcommunitiesdevelopalongagradientofoxygentension;methodstillusedtoday-describedoxidationofhydrogensulfide,sulfur,ferrousiron-…allleadingtotheconceptofchemoautotrophy–derivingenergyfromchemicaloxidationofinorganiccompoundsandcarbonfromCO2*Martinus

Beijerinck(1851-1931)“ThewayIapproachmicrobiology...canbeconciselystatedasthestudyofmicrobialecology,i.e.,oftherelationbetweenenvironmentalconditionsandthespecialformsoflifecorrespondingtothem”FounderoftheDutchDelftSchoolOfMicrobiologyADutchmicrobiologist*Martinus

Beijerinck(1851-1931)isolatedNfixersandSreducers‘microbialubiquity’:allmicroorganismsareeverywhere;conditionsandresourcesdeterminewhoflourishesenrichmentculture:growthmediumtailoredtosuitparticularmetabolicfunctionwithWinogradsky,recognizedthatmicrobesarethemajorplayersinelementtransformationsledtofieldofglobalbiogeochemistry*AlbertJanKluyver(1888-1956)studentofBeijerinckmicrobialphysiologycomparativeapproachunifyingmetabolicfeaturesamongmicrobesleaderoftheDutchschool

afterBeijerinckmicrobialphysiology

comparativeapproach*CorneliusBernardusvanNiel(1897-1985)*IsolatedpurplesulfurbacteriaMajorcontribution,chemistryofphotosynthesis:

2H2A+CO2CH2O+2A+H2OwhereAcanbeSulfurorOxygen…extendedmodeltophotosynthesisingreenplantsoxygenfromwater,notfromCO2Also,chemistryofdenitrification,definitionofprokaryotein1961(withR.Stanier)CorneliusBernardusvanNiel(1897-1985)*RobertE.Hungate

(1908-2004)AKA“GrampaBob”Anaerobicmethods

CowsandtermitesstudentofvanNielmethodsforisolatinganaerobesculturemethods–selectusingnaturalsubstrates,ratherthanguessesaboutwhatorganismseatmicrobiologyofgutsofrumen,termitesASMpresidentwhen“EnvironmentalMicrobiology”and“MicrobialEcology”formallyrecognized*In1953JamesWatsonandFrancisCrickpublishadescriptionofthedouble-helixstructureofDNA.Thepaperacknowledgesthattheauthorswere"stimulatedbyknowledgeoftheunpublishedexperimentalresultsof"MauriceWilkinsandRosalindFranklin,whosex-raycrystallographyimagesofDNAsuggestedthestructure.Franklindiedin1958;Watson,CrickandWilkinsareawardedtheNobelPrizeinPhysiologyorMedicinein1962.double-helixstructureofDNA*FrederickSangerdevelopedamethodtosequenceDNAin1977.Heandhisco-workersPaulBergandWalterGilbertreceivedtheNobelPrizeinChemistryin1980FrederickSanger

WalterGilbert

PaulBerg

*CarlWoeseusesribosomalRNAanalysistoidentifyathirdformoflife,theArchea,whosegeneticmakeupisdistinctfrombutrelatedtobothBacteriaandEucaryea.BacteriaArchaeaEukarya*KaryB.Mullis

MichaelSmith

KaryB.Mullisinventedthepolymerasechainreaction(PCR)methodin1983.HeandKaryB.MullisreceivedTheNobelPrizeinChemistry1993*TheInstituteforGenomicResearch(TIGR)publishedthefirstfullDNAsequenceofafree-livingorganismHaemophilus

influenzae

in1995,whichfollowedupthatachievementwitharapid-fireseriesofscientificaccomplishments,including:? Decipheringthegenomeofthesmallestbacterialgenome,Mycoplasma

genitalium,andinvestigatingtheminimumcomplementofgenesrequiredtosupportlife.? Sequencingthefirstcompletegenomefromarepresentativeofthethirddomainoflife,theArchaea.? PlayingakeyroleindecipheringtheDNAsequenceofArabidopsisthaliana,thefirstplantgenomecompleted,aswellasthesequencesofriceandotherimportantcrops.GenomicSequence:ContributionfromTheInstituteforGenomicResearch(TIGR)*? Decipheringthegenomesequencesofmorethanthreedozenhumanpathogens,includingthebacteriathatcausepneumonia(肺炎),cholera(霍亂),syphilis(梅毒),meningitis(腦膜炎),Lymediseaseanthrax(炭疽熱)

andgingivitis(齒齦炎)

aswellastheparasitesthatcausemalaria(瘧疾),amoebicdysentery(阿米巴痢疾),andAfricanSleepingSickness(非洲昏睡病).? Findingnewwaystousegenomicsasatoolformicrobialforensics(毒物)anddemonstratingthepotentialtousemicroarraytechnologyfortumordiagnosis.? Developingahostofsoftwaretoolsthatarewidelydisseminatedinthescientificcommunitytoassemble,annotateandcomparegenomes.GenomicSequence:ContributionfromTheInstituteforGenomicResearch(TIGR)*1960s:RonaldAtlas,RichardBartha -studiesofpetroleumdegradation -ledtonewfieldofbioremediation, -extendedtomanyotherpollutants:DDT,PCBs,mercury, selenium,industrialsolvents

1970sfuel-shortage: -shortageinNfertilizer -sparkedinterestinthebiologyofnitrogenfixersOtherContributions*CurrenttrendsinEnvironmentalmicrobiology:A.spaceexploration–microbesinextremeenvironments

(hotsprings,thermalvents,lithosphere)B.moleculartechniques–diversityofmicroorganisms

(CarlWoese),newmethodstoassesspresence/abundance

ofindividualspeciesinsituC.realizationthatwithpureculture/enrichmenttechniques,

weknowsomewherebetween1-10%ofexistingmicrobial

species–lotstolearn!D.biologyofclimatechange,globalbiogeochemistry*InthefuturemicrobiologistswillbeTryingtobetterunderstandandcontrolexisting,emerging,andreemerginginfectiousdiseasesStudyingtheassociationbetweeninfectiousagentsandchronicdiseasesLearningmoreabouthostdefensesandhost-pathogeninteractionsDevelopingnewusesformicrobesinindustry,agriculture,andenvironmentalcontrol*StilldiscoveringthemanymicrobesthathavenotyetbeenidentifiedandculturedTryingtobetterunderstandhowmicrobesinteractandcommunicateAnalyzingandinterpretingtheever-increasingamountofdatafromgenomestudies*FieldsofMicrobiologyMicrobialecology-EnvironmentalmicrobiologyMedicalmicrobiologyMicrobialgeneticsandmolecularbiology*SummaryofthischapterWhatismicroorganisms,microbiology,environmentalmicrobiology?CharacteristicsofmicroorganismsThreedomainsoflifeScopeofmicrobiologyBriefhistoryofmicrobiology(trytorememberafewimportanteventsandheroesinvolved,e.g.LouisPasteur,RobertKoch……)Futuretrendsofmicrobiology(molecular)Canyouimagewhatwillbetaughtinthissubject???*AfterIntroduction,canyouimagewhatwillbetaughtinthissubject???Thisquestionspeaksitselfwhywesaysomuchinthisfirstclass.*TheEnd*Inthefirstchapter,weintroducedageneraloutlineonwhatwillbeofthissubject.Nowwestarttoexaminedetailsofmicroorganisms.Let’sseewhatmicroorganismslooklikeandwhataremadeof.

*Chapter2

StructureofBacteria*Macromolecules(大分子)

protein,polysaccharide,lipids,DNA,RNA,96%(dw%)Monomers:(單體及前體)

aminoacidsandprecursors,sugarsandprecursors3%Inorganicions:

K+,Mg+2,Ca2+,Fe2+,Mn2+,Cu2+、Co2+、Zn+2……1%Chemicalcompositionofaprokaryoticcell2.1*OCCCCBaseHHorOHHHHHCH2OPOHOHOSugar:

RNA–ribose(OH)DNA–deoxyribose(H)MacromoleculesNucleoside:base+sugarBases:adenine(A),cytosine(C),

guanine(G),

thymine(T)RNAusesuracil(U)

insteadofthymineRememberthatcarboninsugarisnumberedinsequence!!!Howtheyarenumbered?*C胞嘧啶U尿嘧啶T胸腺嘧啶G鳥嘌呤A腺嘌呤圖1c核糖圖1b核苷酸*ATCGQuestion:ifacellDNAcontainsmoreG+CthanA+T,isitharderoreasiertoseparatetwocomplementarychains?

Hydrogenbond*chainscomplementarytoeachother*氨基酸一般結(jié)構(gòu)肽鍵Macromoleculesprotein*2.2SizeandshapeofBacteriaAveragebacteria0.5-2.0umindiam.Surfacearea~12μm2Volumeis~4μmSurfaceareatovolumeis3:1TypicaleukaryotecellSA/Volis0.3:1FoodentersthroughSA,quicklyreachesallpartsofbacteriaEukaroytesneedstructures&organelles*Approximatesizeofmicro-organismscomparedtoatypicaleukaryoticcell.*Therearetwotypesofcells:prokaryoticvs.eukaryotic*ShapesofBacteriaCoccus(球菌)Chain=StreptoccusCluster=StaphylococcusBacillus(桿菌)Chain=StreptobacillusCoccobacillus(短桿菌)Vibrio=curved(弧菌)Spirillum(螺旋菌)Spirochete(螺旋體)Square(方形)Star(星形)*Streptococci(鏈球菌)showingdividingmanner*Tetradococci(四疊球菌)*Staphylococci(葡萄球菌)*

球菌

(coccus)的分裂方式于群體形態(tài)之間的關(guān)系沿著同一平面分裂；分裂后兩個細(xì)胞連在一起的稱為雙球菌(Diplococcus)

沿著兩個相互垂直的平面分裂；四個細(xì)胞連在一起的稱為四聯(lián)球菌(Tetrads)沿著三個平面分裂；子細(xì)胞聚集在一起的稱為葡萄球菌(Staphylococcus)細(xì)胞沿三個互相垂直的面分裂，分裂后每八個細(xì)胞特征性地疊在一起呈一立方體的稱為八疊球菌(Sarcinalutea)沿著同一平面分裂；分裂后兩個以上細(xì)胞連在一起的稱為鏈球菌(Streptococcus)單球菌(Coccus)*Squarecell:Cyanobacteria*2.3BacterialStructures*Flagella(鞭毛)Motility-movementSwarming(聚集)occurswithsomebacteriaSpreadacrossPetriDishProteus(多變的)speciesmostevidentArrangementbasisforclassificationMonotrichous(單身鞭毛);1flagellaLophotrichous(叢生鞭毛);tuftatoneendAmphitrichous(端生鞭毛);bothendsPeritrichous(周身鞭毛);allaroundbacteriaObservePictureinMicroLab.****Pili(纖毛)ShortproteinappendagessmallerthanflagellaAdherebacteriatosurfacesE.colihasnumeroustypesK88,K99,F41,etc.AntibodiestowillblockadheranceF-pilus;usedinconjugationExchangeofgeneticinformationFlotation;increaseboyancyPellicle(scumonwater)Moreoxygenonsurface*F-PilusforConjugation（結(jié)合）*CapsuleorSlimeLayer（莢膜/粘液）Glycocalyx(多糖)-PolysaccharideonexternalsurfaceAdherebacteriatosurfacePreventsphagocytosis(嗜菌作用)Preventwaterfromloss*Cytoplasm（細(xì)胞質(zhì)）80%Waterbyweight,20%salts-proteinsOsmotic(滲透壓)shockimportantDNAiscircular(環(huán)狀DNA),haploid(單鏈)Moreefficient;growsquickerMutations(變異)allowadaptationtoenvironmentquickerPlasmids(質(zhì)粒);circularDNAindependenttochromosome(獨(dú)立于染色體的環(huán)狀DNA);Antibioticresistance(抗抗生素)Noorganelles細(xì)胞器(mitochondria,Golgi,etc.)Thepartinsidethemembrane*CellMembrane（細(xì)胞膜）BilayerphospholipidWatercanpenetrateFlexibleNotstrong,ruptures(斷裂)easilyOsmoticpressurecreatedbycytoplasm磷脂的化學(xué)結(jié)構(gòu)(左)及其示意圖(右)親水端疏水端親水端疏水端*親水端疏水端脂肪酸甘油*嵌入蛋白磷脂*醣蛋白醣脂類固醇*真菌和古細(xì)菌質(zhì)膜中磷脂組成的區(qū)別DifferencesofmembranelipidsbetweenArchaeaandotherorganisms細(xì)菌和真菌古細(xì)菌細(xì)菌和真菌的質(zhì)膜由無分支的直鏈脂肪酸通過酯鍵與甘油連接；古細(xì)菌則由帶有支鏈脂肪酸與甘油連接。酯鍵醚鍵Branches*CellWall

Peptido-glycan(肽-糖)polymer(aminoacids+sugars)Unique(特別)tobacteriaSugars;NAG&NAMN-acetylglucosamine(N-乙酰氨基葡萄糖)N-acetymuramicacid(N-乙酰胞壁酸)DformofAminoacidsusednotLformHardtobreakdownDformAminoacidscrosslinkNAG&NAM**D-丙氨酸D-谷氨酸間二氨基庚二酸L-丙氨酸圖20肽聚糖的亞單位NAG：N-乙酰葡萄糖胺NAM：N-乙酰胞壁酸氨基糖氨基酸*CellwallstructureofGramnegativebacteriaG-細(xì)菌細(xì)胞壁結(jié)構(gòu)細(xì)胞質(zhì)胞圍細(xì)胞膜肽聚糖外膜*裂解酶敏感鍵**生長點(diǎn)胞內(nèi)合成前體，通過膜運(yùn)送到胞外合成薄壁*外膜內(nèi)膜*CellwallstructureofGrampositivebacteria革蘭氏陽性菌(G+)的細(xì)胞壁*CellWallSummaryDetermineshapeofbacteriaStrengthpreventsosmoticrupture20-40%ofbacteriaUniquetobacteriaSomeantibioticseffectdirectlyPenicillin*CellWallTeichoicAcids磷壁(酸)質(zhì)Gram+onlyGlycerol,Phosphates,&RibitolAttachmentforPhages**Lipopolysaccharide(LPS)EndotoxinorPyrogenFevercausingToxinnomenclatureEndo-partofbacteriaExo-excretedintoenvironmentStructureLipidAPolysaccharideOAntigenofE.coli,SalmonellaG-bacteriaonlyAlcohol/Acetoneremoves***LPS(cont’d)FunctionsToxic;killsmice,pigs,humansG-septicemia;deathduetoLPSPyrogen;causesfeverDPTvaccinationalwayscausesfeversAdjuvant;stimulatesimmunityHeatResistant;hardtoremoveDetection(alltopical&IVproducts)Rabbits(measurefever)Horseshoecrab(AmoebocytesLyseinpresenceofLPS)*LPS(cont’d.)AppearanceofColoniesMucoid=Smooth(lotsofLPSorcapsule)Dry=Rough(littleLPSorcapsule)OAntigenofSalmonellaandE.coli2,000differentOAgsofSalmonella100’sdifferentOAgsofE.coliE.coliO157OAgsdifferinSugars,notLipidA*Endospores(芽孢、內(nèi)生芽孢)ResistantstructureHeat,irradiation,coldBoiling>1hrstillviableTakestimeandenergytomakesporesLocationimportantinclassificationCentral,Subterminal,TerminalBacillusstearothermophilus(強(qiáng)耐高溫)-sporesUsedforqualitycontrolofheatsterilizationequipmentBacillusanthracis-sporesUsedinbiologicalwarfare*Bacillusanthracis(炭疽熱細(xì)菌

).Gramstain.Thecellshavecharacteristicsquaredends(端齊).Theendosporesareellipsoidal(橢圓)shapedandlocatedcentrallyinthesporangium.Thesporesarehighlyrefractile(折射)tolightandresistanttostaining.*Bacillusthuringiensis.PhasePhotomicrograph

ofvegetativecells,intracellularspores(light)and

parasporalcrystals(dark)(類芽孢晶體).*G+vs.G-G+ThickercellwallTeichoicAcidsG-Endotoxin-LPSWhicharemoresensitivetoPenicllin?Alcohol/Acetoneaffectswhichmore?*Prokaryotesvs.EukaryotesCellWallTeichoicAcidsLPSEndosporesCircularDNAPlasmids*EukaryoteCellStructure*Chapter3MicrobialGrowth*MicrobialGrowthmeansincreaseinnumberofcells,notcellsize.Itisveryconfusingwithreproduction(繁殖)inotherformsoflife(e.g.animals).*GrowthrequirementsPhysicalChemical1Temperature2pH3Osmoticpressure(water)4Light1Carbon2Nitrogen3Sulfur4Phosphorus5Inorganicelement6Oxygen7growthfactors*TemperatureMinimumgrowthtemperatureOptimumgrowthtemperatureMaximumgrowthtemperaturePhysicalRequirementsforGrowth:1temperature*專性嗜冷兼性嗜冷中溫型嗜熱極端嗜熱Arctic/antarctic

Sea/refrige

mammal/soils

compost

Spring/seavolcano

*Question:Isitalwayssafeiffoodsarestoredinrefrigeratorforlongtime?

*Figure6.2*pHMostbacteriagrowbetweenpH6.5and7.5MoldsandyeastsgrowbetweenpH5and6AcidophilesgrowinacidicenvironmentsPhysicalRequirementsforGrowth:2pH*PhysicalRequirementsforGrowth:3osmoticpressureOsmoticPressure(=wateravailability)Hypertonic(高滲)environments,increasesaltorsugar,causeplasmolysis(質(zhì)壁分離)Extremeorobligate(專性)halophilesrequirehighosmoticpressureFacultative(兼性)halophilestoleratehighosmoticpressure*PlasmolysisFigure6.4CellinnormalosmoticpressureenvironmentCellinhypertonicenvironment*PhysicalRequirementsforGrowth:4LightLight(=radiation)Necessaryforphototrophicbacteria(usinglightasenergysource)Radiationindifferentwavelengthhasdifferenteffects(ultraviolet,x-ray,andgamma-raykillsbacteriawhilegreenlightinducesdevelopmentoflifecycle,e.g.mushrooms)*CarbonStructuralorganicmolecules,energysource(recallthechemicalconstituentsofcells)Chemoheterotrophs(化能異養(yǎng)型)useorganiccarbonsourcesAutotrophs(自養(yǎng)型)useCO2ChemicalRequirementsforGrowth:1carbon*NitrogenInaminoacids,proteinsMostbacteriadecomposeproteinsSomebacteriauseNH4+orNO3

AfewbacteriauseN2innitrogenfixationSulfurInaminoacids,thiamine(硫胺素=Vb1),biotin(生物素VH)MostbacteriadecomposeproteinsSomebacteriauseSO42

orH2SChemicalRequirementsforGrowth:*PhosphorusA.InDNA,RNA,ATP,andmembranesB.PO43

isasourceofphosphorusTraceelementsA.InorganicelementsrequiredinsmallamountsB.UsuallyasenzymecofactorsChemicalRequirementsforGrowth*Oxygen(O2)ChemicalRequirementsforGrowth:Obligateaerobes專性需氧菌O2isnecessaryFacultativeanaerobes兼性厭氧菌GrowbetterifO2ispresentObligateanaerobes專性厭氧菌O2istoxicAerotolerantanaerobes微耐氧菌O2isnotnecessarybuttolerableMicro-aerophiles微需氧菌O2isneededatlowpartialpressure*Singletoxygen:O2boostedtoahigher-energystateSuperoxidefreeradicals(超氧化物自由基):O2

Peroxideanion(過氧化物陰離子):O22

Hydroxylradical羥基(OH

)WhyO2istoxictosomebacteriaSomebacteriadonothavetheseenzymes

*OrganicGrowthFactorsOrganiccompoundsobtainedfromtheenvironmentVitamins,aminoacids,purines(嘌呤),pyrimidines(嘧啶)ChemicalRequirementsforGrowth:*CultureMedium:NutrientspreparedformicrobialgrowthSterile:NolivingmicrobesInoculum:IntroductionofmicrobesintomediumCulture:Microbesgrowingin/onculturemediumCultureMedia*ComplexpolysaccharideUsedassolidifyingagentforculturemediainPetriplates(平板),slants(斜面),anddeeps(深層培養(yǎng))GenerallynotmetabolizedbymicrobesLiquefiesat100°CSolidifies~40°CAgar*Chemicallydefinedmedia:exactchemicalcompositionisknownComplexmedia:Extractsanddigestsofyeasts,meat,orplants,e.g.Nutrientbroth(營養(yǎng)肉湯)Nutrientagar(營養(yǎng)瓊脂)CultureMedia*ExamplesofCultureMedia**ReducedmediaContainchemicals(thioglycollate

巰基乙酸鹽orcystine(胱氨酸)orascorbate(抗壞血酸)toremoveO2HeatedtodriveoffO2AnaerobicCultureMethods*AnaerobicjarAnaerobic

Culture

Methods鈀*AnaerobicchamberAnaerobic

Culture

Methods*CandlejarO2-packetSomebacteriarequirehighCO2*Enhancethegrowthofcertainwantedorganismsbutsuppressunwantedmicrobes.SelectiveMediaFigure6.9b,cMakeiteasytodistinguishcoloniesofdifferentmicrobes.DifferentialMedia*EncouragesgrowthofdesiredmicrobeAssumeasoilsamplecontainsafewphenol-degradingbacteriaandthousandsofotherbacteriaInoculatephenol-containingculturemediumwiththesoilandincubateTransfer1mltoanotherflaskofthephenolmediumandincubateTransfer1mltoanotherflaskofthephenolmediumandincubateOnlyphenol-metabolizingbacteriawillbegrowingEnrichmentMedia*Anall-purposed(rich)mediumisrichinawidevarietyofnutrients(includingmanygrowthfactors)andwill,therefore,supportthegrowthofawiderangeofbacteria.All-purposed(rich)medium*AMinimalmediumsuppliesonlytheminimalnutritionalrequirementsofaparticularorganism.MinimalMedium*SummaryofCommonly-UsedConstituentsofMediaSubstanceFunction CompositionSource

AGAR SolidifyingagentImpurepolysaccharidemarinealgaePEPTONES nutrientAnimal/Plantproteinscow,soy EXTRACTS nutrientAnimal/Bacteriapastecow,yeastBODYFLUIDS hormones Blood animals BUFFERS pH K2PO4;NaHPO3;CaCO3 - REDUCTANTSe-source thioglycolate -SELECTIVESbacteriostatAntibiotics,sodiumazidevariesINDICATORS pHbromothymolblue,phenolred-WATER hydration H2O(DI&tap) -*ApureculturecontainsonlyonespeciesorstrainAcolonyisapopulationofcellsarisingfromasinglecellorsporeorfromagroupofattachedcellsAcolonyisoftencalledacolony-formingunit(CFU)Afewwords*StreakPlate(平板劃線)*Mediumsterilization*Sterilization:alllivingcells,viablespores,virusesarekilledorremovedfromobjectorhabitatthough:Irradiation:destroys/distortsnucleicacidsX-raysµwaves.UVcommonlyusedonobjectsurfacesFiltration:physicalremovalfromliquidorgas