lecture8probioticbacteria：講座8益生菌.ppt

LectureOutline Introduction DefinitionofProbioticsFundamentalQuestionsRecentFindingsPossibleModesofActionRationaleforSelectingProbioticsConclusionsandFurtherDirections Sourceofnotes Verschuere L Rombaut G Sorgeloos P andVerstraete W 2000 Probioticbacteriaasbiologicalcontrolagentsinaquaculture Microb Mol Biol Rev 64 4 655 671 Introduction Asweknow aquacultureisgrowingfasterthanbeefcattleproduction 10 vs 3 Epizootics disease majorlimitingfactorinfish shrimpdevelopments Disinfectants antimicrobialdrugshavehadlittleeffectincontrollingdisease MassiveuseofantimicrobialsincreasesselectivepressureonmicrobesandencouragesnaturalemergenceofbacterialresistanceResistantbacteriathriveafternon resistantstrainshavebeenkilledandcanevenpassonresistancegenestootherbacteriathathavenotbeenexposedtoantibiotics Introduction EmphasisshouldbeplaceonpreventionMorecost effectivethancure Antimicrobials disinfectantsandpesticideslargelytreatsymptomsoftheproblemandnotthecauseAlternativestrategiesarejustcatchingonExample viavaccination Norwayhasreducedchemicaltherapeutantusefrom50MTin1987tolessthan747kgin1997 with7xconcomitantproductionincrease Other useofimmunostimulantsw orw outvaccines Introduction UseofbacteriaasafoodsourceandasabiologicalcontrolagentoffishdiseasewasfirstproposedbyYasudaandTaga 1980 VibrioalginolyticushasbeenusedasaprobioticinshrimphatcheriesinEcuadorsince1992Reducedhatcherydowntimefrom7dayspermonthtolessthan21daysperyear FAOhasnowdesignateduseofprobioticsasamajormeansforimprovementofaquaticenvironmentalqualityUltimategoal makeaquacultureproductsmoreacceptabletoconsumers DefinitionofProbiotics Manyproposed however alivemicrobialfeedsupplementwhichbeneficiallyaffectsthehostanimalbyimprovingitsintestinalbalanceHistorically terrestrialanimals genusLactobacillusDefinition above requiressomeadditionalconsiderations 1 bacteriainaquaticmediuminfluencecompositionofgutmicrobiotaandviceversa2 immediateambientenvironmenthasmuchgreaterinfluenceonmicrobiotathanwithterrestrialsInaquaticenvironments hostsandmicroorganismssharetheecosystemTerrestrials thegutrepresentsamoisthabitatinawater limitedworld DefinitionofProbiotics BigIssue1 aquaticsaresurroundedbyanenvironmentsupportingtheirpathogensindependentlyofthehostanimalResult opportunisticpathogenscanreachhighdensitiesaroundthefish shrimpSurroundingbacteriaarecommonlyingestedwiththefeedorviadrinking maximumcase filterfeeders ResearchinprobioticsbeganwithfishjuvenilesbutmoreattentionrecentlygiventoshrimpandfinfishlarvaeBigIssue2 terrestrialshaveinherentcolonizingbacteriafromthemother aquaticslargelyspawnedasaxeniceggs nofurthercontactwithparent Ambientbacteriacolonizeeggssurface younglarvaeoftenhavenodevelopedgut e g shrimp nomicrobialcommunityingut gillsorskinPoint propertiesofbacteriainambientwaterareveryimportant DefinitionofProbiotics Betterdefinition alivemicrobialadjunctwhichhasabeneficialeffectonthehostbymodifyingthehost associatedorambientmicrobialcommunity byinsuringimproveduseoffeedorbyenhancingitsnutrition byenhancingthehostresponsetowardsdisease orbyimprovingqualityoftheambientenvironmentOurfocus responsetowardsdiseaseandimprovementoftheambientenvironmentJobsofMicrobialAdjuncts 1 microbialadjunctspreventingproliferationofpathogensingutorelsewhere 2 improveddigestibility 3 deliverimprovednutritiontoaquatics 4 enhancinghostresponsetodisease acquired 5 improvedenvironmentalquality CanyouManipulateMicrobialCommunities Difficult 1 discontinuousculturecycles 2 disinfectionduringpondprep 3 suddenincreasesinnutrientsduetofeedingUnlikelyunderintensiverearingMustconsiderdeterministicfactors knownresponse salinity temp quality quantityoffeedPoint theenvironmentselectstherangeofmicrobes axiomofenvironmentalselection Stochasticfactors chance rightplace righttimeEvidence identicalculturesstartedsimultaneouslyyielddifferentassemblages CanyouManipulateMicrobialCommunities Concept insteadofaccidentalcolonization startwithaprobioticthatiswelladaptedtoprevailingenvironmentalconditionsThisisprobablybetterthancompetingwithadominant well established non probioticLong termexposureisoftenrequiredtoachieveaprobioticeffectDoestheprobiotichavetobecontinuouslyintroducedtotheculture Evidence inmostcases yes atleastwithLactobacillussp MostfishcontainaspecificintestinalmicrobiotaestablishedatthejuvenilestageUnlessthehosthasbeenexposedtoalimitedrangeofmicroorganismsinitsdevelopment asingleadditionwon tresultinlong termcolonization RecentFindings Asmentioned itallstartedwithYasudaandTaga 1980 UsuallyaddedinfeedortoculturewateraspreventativeagentsagainstinfectionbypathogenicbacteriaNutritionalbenefitsareusuallysecondaryTypicalgenera Lactobacillus Vibrio Bacillus PseudomonasThefollowingisasummaryoffindingsbasedonvariousaquaticspecies RecentFindings fisheggs larvae Forincubators useofantibioticsmustbeminimalAntibioticsdon trepresentcontrol instead unfavorablealterationofmicrobiotaGoal establishcolonizationontheeggpriortopathogencolonization i e nosubstrate This inturn affectssubsequentgutcolonizationOnceinitialfeedinghasstarted probioticstypicallyaddedtoculturewaterorculturemediumoflivefeeditems e g algae rotifers etc Result improvedsurvival fastergrowthMechanism Productionofantibioticsorsiderophores metalsequesterers RecentFindings finfish Digestivetractoffinfishcontains108cells g Ringoetal 1995 Forcod Gadusgadus gutiscolonizedbysimilarbacteriaasfoundineggs HansenandOlafsen 1999 Putativeprobioticsaddedassoonaspossibleafterhatchinginordertocolonizegutpriortofeeding RingoandVadstein 1998 TurbotanddabharborbacteriacapableofsuppressinggrowthofV anguillarum Ollsonetal 1992 V alginolyticuswaseffectiveinreducingdiseasecausedbyAeromonassalmonicidainAtlanticsalmon Austinetal 1995 RecentFindings finfish Kennedyetal 1998 showedadditionofaGram positiveprobioticincreasedsurvival sizeuniformity andgrowthrateofsnook reddrum spottedseatroutandstripedmullet Grametal 1999 reportedastrainofPseudomonasfluorescensreducedmortalityof40grainbowtroutinfectedwithpathogenicV anguillarumGarcia de la Bandaetal 1992 addedStreptococcuslactisandLactobacillusbulgaricustorotifersandArtemiasp naupliiandrecorded6xhighersurvivalatweaningthanuntreatedcontrolsNikoskelainenetal 2003 showedimmuneenhancementinrainbowtroutviaLactobacillusrhamnosussupplementedinfeeds RecentFindings shrimp Broadapplicationinhatcheries butfewin depthstudiespublishedoftenusedasfoodsource e g soilbacteriaforP monodonnauplii improvedsurvival 57 vs 0 after13daysagainstV anguillarumimprovedsurvivalofL vannameiPL sinoculatedwithV alginolyticus non pathogenic vs oxytetandcontrol GarriquesandArivalo 1995 Griffith 1995 reportedthatfollowingtheintroductionofprobioticsinEcuadorin1992 hatcherydown timebetweenbatcheswasreducedfrom7dayspermonthto21daysperyear productionvolumesincreasedby35 andantimicrobialusedecreasedby94 InshrimphatcheriesinNewCaledonia astrainofPseudoalteromonaspiscicidawasfoundtoinhibitthegrowthofVibriosp Saulnieretal 2000 RecentFindings bivalavemolluscs MostresearchhasfocusedonnutritionalcontributionstomollusclarvaemostworkinvitrowhereinautochthonousstrainshavebeenisolatedfromscallopsandhaveshownsomeinhibitiontoVibriosp andAeromonashydrophilaBacillussp andLactobacillussp showntodepurateoysters Crassostreavirginica againstV vulnificus Williamsetal 2001 PartII ModesofActionofProbioticBacteria ModesofAction Manypublicationsaboutprobioticshaveemergedinaquacultureinthelastdecademostbasedonempiricalarguments simple modesofactionwerelargelycircumstantialseveralmodesofactionhavebeenproposedasaresultofhumanandagriculturalapplications ModesofAction Infoonterrestrialinvestigationshasbeenusedforaquatics esp Lactobacillussp oneproblem distinctionbetween1 theintrinsicabilityoftheprobingtopositivelyinfluencethehostand2 itsabilitytoreachandmaintainitselfinthelocationwheretheeffectistobeexertedPoint doesnotmatterifitproducessiderophoresorinhibitorycompoundsinthegutifit sneveringestedcanbeveryimportant Mytilusedulis BlueMuscle canselectivelyingest digestmicrobes ModesofAction Also ifthecandidateprobiontcannotproliferateinthegutitprobablywon texertastrongeffectwillneedcontinuousapplicationthroughthedietorviathewater summary theymustreachthelocationwhereeffectisdesired PossibleModesofAction productionofinhibitorycompoundscompetitionforchemicals availableenergycompetitionforadhesionsites exclusion enhancementoftheimmuneresponseimprovementofwaterqualityinteractionwithphytoplanktonasourceofmacro andmicro nutrientsenzymaticcontributiontodigestion 1 productionofinhibitorycompounds Releaseofchemicalshavingabactericidalorbacteriostaticeffectultimateresult competitiveedgefornutrients energyproductionsites inhostintestine onitssurface orinculturemediumproducts antibiotics bacteriocins siderophores lysozymes proteases hydrogenperoxide organicacids pHchange exactcompoundisseldomidentified hence theterm inhibitory 1 productionofinhibitorycompounds Lactobacillussp producesbacteriocins toxins marinebacteriaproducebacteriolyticenzymesagainstV parahaemolyticusAlteromonassp producesmonastatin showntobeinhibitoryagainstAeromonashydrophilainhibitoryeffectshavebeenshownbyprobioticsagainstaquaculturepathogensnodemonstrationunderinvivoconditions oops moreresearchrequired Didn tyoumentionthislasttime 2 CompetitionforChemicalsorAvailableEnergy Explainshowdifferentmicrobialpopulationsexistinsameecosystemitislikelythatitoccursinthemammaliangut butproofislackingapplicationoftheprinciplesofcompetitiontonaturalsituationsisnoteasymicrobialsituationinecosystemsisusuallycontrolledbyheterotrophscompetingfororganicsubstratesasbothcarbonandenergysourcesifyouknowthefactorsaffectingmicrobialcompositionofthemicrobiota youcanmanipulateit 2 CompetitionforChemicalsorAvailableEnergy Allmicroorganismsrequireironforgrowthsiderophore lowmwferricion specificchelatingagentsdissolveprecipitatedFeandmakeitavailableformicrobialgrowthsiderophoresscavengeFeandmakeitunavailabletootherspeciesthisoccursattissuelevelprobioticsproducingsiderophorescanoutcompetepathogensforFe thuslimitingpathogengrowthworksbestwithpathogensthatalsoproducesiderophores e g V anguillarum 3 CompetitionforAdhesionSites Competitionforgutadhesionsiteswouldlimitcolonizationadhesiontoentericmucusisnecessaryforbacteriatobecomeestablishedinfishintestinesthisisprobablythefirstprobioticeffectadhesioncanbespecific basedonadhesinandreceptormolecules ornon specific basedonphysiochemicalfactors totalprobioticeffectisprobablyamixtureofsitecompetition productionofinhibitorycompoundsandnutrient energycompetition 4 EnhancementofImmuneResponse Remdefinitionofanimmunostimulant Chemicalcompoundsthatactivatetheimmunesystemsofanimalsandrenderthemmoreresistanttoinfectionsbyviruses bacteria fungiandparasites Immuneresponsevariesinanimalslacticacidbacteriaadministeredorallymayinduceincreasedresistancetoentericinfectionsproblem onlyshownwithspecificcellcompoundsordeadcellsgoodindications butnoproof 5 ImprovementofWaterQuality ProposedasamodeofactionasaresultofmonitoringwaterqualityafteradditionofprobioticsusuallyassociatedwithBacillussp Hook gram bacteriaarebetterconvertersoforganicmatterbacktoCO2thangram thus phytoplanktonbloomsaremoreeasilymaintained interestingresearcharea monitor DOC POCsofar hasn tbeendemonstrated RationaleforSelectingProbiotics Verschuereetal 2000 BackgroundInfo ProfoundknowledgeofcultureCriticalreviewofavailableliteratureCharacterizationofabioticandbioticenvironmentimpactingcultureMustknowrelationshipbetweenmicrobiotaandhostwell understoodRelationshipbetweenmicrobiotaandcarryingcapacityofenvironment AcquisitionofPutativeProbiotics Areputativeprobiotics isolatedfromhostorhostenvironment betterthanthosefromothersources Putativesmightbebetterduetofacttheyarealreadyadheredtogutwall i e couldblockpathogenadhesion Ifanon pathogenicbacteriumexistsathighdensityintypicalculturewater itwillbewell adaptedtoprevailingconditionsandcanprobablycompeteefficientlywithpathogensfornutrien