Experiment Three：Isolation and identification of pyogenic cocci（《医学微生物学》课件）PPT课件下载推荐.ppt

1、Isolation and identification of pyogenic cocciExperiment Three Objective of ExperimentExperiment n nTo master basic principle and method,which use to isolate and identify pyogenic cocci from clinical specimens.n nTo diagnose clinical disease and guide doctors to select proper medicines.Specimens（Gra

2、m-stain）Morphological characteristics（Microscopic examination）Isolation culture Smears（blood agar plate）Colonial characteristics Hemolytic reaction Pigmentation PROCEDURE PROCEDURE morphology observation colonial typical colonies stained and observe Pure culture Antibiotic susceptibility test Direct

3、 identificationIdentified method for Staphylococcin nGram-stainGram-stainn n IsolationIsolationandcultureandculturen n PureculturePureculture n n DirectDirect identifidentificationicationStaphylococci are Gram-positive cocci,typically arranged in clumps or Grape-like clustersDirect identificationn n

4、Themannitolfermentationtest.n n TheCoagulaseTestn nTheDnaseTestn nThephagetypingtestn nAnimalExperimentThe mannitol fermentation test.n nInoculatethebacteriaintoamannitolmicro-tube,incubateat370Cfor18h.S.aureuswillfermentmannitoltoproduceacid,whichcausesthemediumtoturnyellow.Test of Microbiological

5、experiment n nWhile a patient is infected by pathogenic enterobacteria（Salmonella）,how to diagnose with microbiological methods in lab?#（simple procedure）n nWhile a patient is infected by pyogenic cocci,how to diagnose with microbiological methods in lab?#（simple procedure）n nWhile a patient is infe

6、cted by influenza virus,how to diagnose with microbiological methods in lab?#（simple procedure）The Coagulase Testn nCoagulaseCoagulase is an enzyme converting fibrinogen is an enzyme converting fibrinogen into fibrin promoting blood clotting.into fibrin promoting blood clotting.n nIt might be a viru

7、lence factor with the It might be a virulence factor with the coagulated blood around the bacteria protecting coagulated blood around the bacteria protecting them from the immune system.them from the immune system.n nCoagulaseCoagulase-negative strains are often as-negative strains are often as path

8、ogenic as pathogenic as coagulasecoagulase-positive strains.-positive strains.The Slide Coagulase TestThe Tube coagulase TestLeft Left tube tube coagulasecoagulase positive positive Right tube Right tube coagulasecoagulase negative negative The Dnase Testn nInoculateInoculateDnaseDnaseagarplateswith

9、aloopsoagarplateswithaloopsothatthegrowthisinplaquesabout1cminthatthegrowthisinplaquesabout1cmindiameter.Incubateat37diameter.Incubateat370 0Covernight.FloodCovernight.Floodtheplatewith1Nhydrochlorictheplatewith1Nhydrochloricacid.Clearingaroundthecoloniesacid.Clearingaroundthecoloniesindicatesindica

10、tesDnaseDnaseactivity.Thehydrochloricactivity.Thehydrochloricacidreactswithunchangedacidreactswithunchangeddeoxyribonucleicacidtogiveacloudydeoxyribonucleicacidtogiveacloudyprecipitate.Afewotherbacteria,e.g.precipitate.Afewotherbacteria,e.g.SerratiaSerratia,maygiveapositivereaction.,maygiveapositive

11、reaction.The phage typing testn nThistestisusedtotracetheinfectiveagentinepidemiologyifnecessary.n nItisusuallynotdoneforroutineclinicalpurpose.Animal Experimentvomitexcrement remaindered foodIsolation and identification of bacteriaMeat soup mediafilter injection68w catobservationFood poisoningThe a

12、ntibiotic susceptibility testn nThistestishelpfulforthetreatmentofS.aureusinfection.n nmaterialsuuStaphaureusStaphaureus（isolatedfromthepusofisolatedfromthepusofapatientapatient）.uuSeveralkindsoffilterpaperSeveralkindsoffilterpaper（eacheachcontainsdifferentkindsofantibioticscontainsdifferentkindsofa

13、ntibiotics）uuNutrientagarplateNutrientagarplateThe antibiotic susceptibility testStreaking the staphaureus on agar plate （thoroughly covered the plate）1.Put 4 kinds of paper contained different antibiotics on the plate （each paper are far away about 2 cm）Incubate at 370C for 18-24 hours.Observe the

14、results the plates The sensitivity of the organism is indicated by the diameter of the zone of growth inhibition.Laboratory Diagnosis of Pathogenic Enterobacterial InfectionDimidiation of the enterobacteria according to the fermentation of lactose n nLactaseLactasefermentersfermenters:#saprophytican

15、dsaprophyticandcommensalcommensal EscherichiaKlebsiellaCitrobaterEscherichiaKlebsiellaCitrobaterEnterobacterEnterobactern nNonlactaseNonlactasefermentersfermenters:#pathogenspathogensSalmonellaSalmonellaShigellaShigellasomesomeCitrobacterCitrobacterProteusProteusSerratiaSerratia Colonial characteris

16、tic observationSpecimens isolation Gram Staining （SS/EMB plate）Serological identification TSI Biochemical reaction ProcedureSpecimensn nDifferentspecimensshouldbetakenDifferentspecimensshouldbetakendependingonthekindandtheprocessofdependingonthekindandtheprocessofthedisease.thedisease.bloodbloodbonemarrowbonemarrowUrineUrinestoolstoolIsolationn nCulture medium:#S.S agar n nMethod:#streak platen nResult:#Pathogenetic colonies:#middle size,red Suspect colonies:#colorless,small,opaque Biochemical reactions o