细菌分类_ 欢迎光临中国免疫学信息网课件

《细菌分类_ 欢迎光临中国免疫学信息网课件》由会员上传分享，免费在线阅读，更多相关内容在教育资源-天天文库。

Faculty:Dr.AlvinFoxSuggestedreading:Murray,ThirdeditionChapter2,16-18and21.TOPIC:CLASSIFICATION&IDENTIFICATIONOFINFECTIOUSAGENTS KeyTermsIsolation(culture)Agarplate/coloniesLiquidmediatesttube-bulkIdentification&taxonomyFamilyGenusSpeciesTypeStrainBiochemical(physiological)testsMoleculartestsDNA-DNAhomology16SrRNAsequencingChemicalprofilingNonculturebaseddetectionPolymerasechainreaction-(PCR)Agglutination(antigendetection)StainSerology(antibodydetection) Bacterialidentificationinthediagnosticlaboratory1.Isolation&identificationaidstreatmentsinceinfectiousdiseasescausedbydifferentbacteriahaveavarietyofclinicalcoursesandconsequences.Susceptibilitytestingofisolateshelpsinselectionofantibioticsfortherapy.Theidentificationisbasedontaxonomicprinciplesappliedtotheclinicalmicrobiologicalsituation.4.Testsmustbeeasilylearned,lowincostandrapidlyperformed. ClassificationFamily:agroupofrelatedgenera.Genus:agroupofrelatedspecies.Species:agroupofrelatedstrains.Type:setsofstrainwithinaspecies(e.g.biotypes,serotypes).Strain:onelineorasingleisolateofaparticularspecies. StepsindiagnosticisolationandidentificationofbacteriaStep1.Samplesofbodyfluidsarestreakedoncultureplatesandisolatedcoloniesofbacteriaappearafterincubation.Observationofthesecoloniesforsize,texture,color,and(ifgrownonbloodagar)hemolysisreactions,ishighlyimportantasafirststepinbacterialidentification.Whethertheorganismrequiresoxygenforgrowthisanotherimportantdifferentiatingcharacteristic. BloodAgarPlate Step2.ColoniesareGramstainedandindividualbacterialcellsobservedunderthemicroscope. Asinglecolonyisplacedonaslideanddried.Stage1.Stainingwithcrystalviolet.Stage2.Fixationwithiodinestabilizescrystalvioletstaining.Allbacteriaremainpurpleorblue.Stage3.Extractionwithalcohol.Decolorizessomebacteria(Gramnegative)andnotothers(Grampositive).Stage4.Counterstainingwithsafranin.Grampositivebacteriaarestainedwithcrystalvioletandremainpurple.Gramnegativebacteriaarestainedpink.GramStain GramstainmorphologyShapecocci(round)bacilli(rods)spiralorcurved(e.g.spirochetes)Singleormultiplecellsclusters(e.g.streptococci)chains(e.g.streptococci)Grampositiveornegative Step3.Thebacteriaarespeciatedusingtheseisolatedcolonies. TypicalCultureLaboratoryBench Step4.Antibioticsusceptibilitytestingisperformed. TaxonomiccharacterizationofbacteriaComparisonsofspeciesinvolvecomparisonsofmultiplestrainsforeachspecies.Comparisonsareprimarilybasedonchemicalormolecularanalysis. MolecularanalysisItwouldbeidealtocomparesequencesofentirebacterialchromosomalDNA.Alternatively,genomicsimilarityhasbeenassessedbytheguanine(G)+cytosine(C)content(%GC).Thishasbeenreplacedbytwoalternatives:HybridizationSequencingspecificgenes DNA-DNAhomology1.HowwelltwostrandsofDNAfromdifferentbacteriabind(hybridize)together.Thistechniqueisemployedtocomparethegeneticrelatednessofbacterialstrains/species.IftheDNAfromtwobacterialstrainsdisplayahighdegreeofhomology(i.e.theybindwell)thestrainsareconsideredtobemembersofthesamespecies. “Goldstandard"inbacterialtaxonomySequencingof16SribosomalRNAmolecules(16SrRNA)The16SrRNAisapproximately1600nucleotidesinlength.The16SrRNAprovidesgenomicsimilarityabovethespecieslevelallowingcomparisonsofrelatednessacrosstheentirebacterialkingdom.CloselyrelatedbacterialspeciesoftenhaveidenticalrRNAsequences.5.ThetechniqueprovidescomplementaryinformationtoDNA-DNAhybridization.Thedevelopmentofprobesbasedon16Sdatahaveimprovedidentificationofsomebacterialpathogens. ChemicalanalysisSophisticatedtoolsareavailableforstudyingthestructuralcompositionofbacteria(mostcommonlyfattyacid,carbohydrateorubiquinoneprofiling)ofbacteria.Characterizationofsecretedmetabolicproducts(e.g.volatilealcoholsandshortchainfattyacids)isalsohelpful. ApproachestorapiddiagnosiswithoutpriorcultureCertainhumanpathogenseithercannotbeisolatedinthelaboratorygrowextremelypoorly.Successfulisolationcanbeslowandinsomeinstancesimpossible.3.Directdetectionofbacteriawithoutcultureispossibleforsomeorganisms. AsimpleapproachtorapiddiagnosisAntigendetectionUsedinmanydoctor'sofficesforthegroupAstreptococcus.Thepatient'sthroatisswabbedandstreptococcalantigenextracteddirectlyfromtheswab(withoutpriorbacteriologicalculture).Thebacterialantigenisdetectedbyaggregation(agglutination)ofantibodycoatedlatexbeads. BacterialDNAsequencesamplifieddirectlyfromhumanbodyfluidsThepolymerasechainreaction(PCR)canbeusedtoamplifylargeamountsofspecificgenesorportionsofgenes.Greatsuccesshasbeenachievedinrapiddiagnosisoftuberculosis. DirectmicroscopicobservationCertainclinicalsamplessuchassputumcanbedirectlyassessedforthepresenceofbacteriacanbehelpful(e.g.detectionofM.tuberculosisinsputum). SerologicidentificationSerologicidentificationofanantibodyresponse(inpatient'sserum)totheinfectingagentcanonlybesuccessfulseveralweeksafteraninfectionhasoccurred.