微生物实验5 肠道杆菌

EntericbacteriaDemonstrationEntericbacteria(E.coli,

Shigellae,Salmonellae,Proteus)Morphology&stainingGrowthcharacteristicsonSS

agarplateandEMB

agarplate)Biochemicalreaction:kliger’sironslant,UreasetestOperationWidaltestLaboratorydiagnosisofentericbacteriamannitol

fermentationtest（Continued）AntimicrobialSusceptibilitytest（Continued）

VideoAnaerobicbacteria22023/3/28ClassificationEnterobacteriaceaeShigellaEscherichiaSalmonellaProteusKlebsiellaETEC,EIEC,EPEC,EHEC,EAECS.dysenteriae,S.flexneri,S.boydii,S.sonneiS.typhi,S.enterica……32023/3/28BasiccharacteristicsGram-negativerodEasytoculture,formcircular,convex,smoothcolonieswithdistinctedgesonbasicmediaBiochemicalreaction:activeComplxantigenicstructureResistance:notstrongEasytomutate42023/3/28Widaltest

beusedtohelpmakea

presumptive

diagnosisofentericfeverPrinciple：S.typhiOantigenS.typhiHantigenS.paratyphiAHantigenS.schottmuelleri

HantigenKnownantigensUnknownantibodyinpatientserumagglutination52023/3/2812345678S.typhiO0.2ml0.2mlS.typhiH0.2ml0.2mlS.paratyphiAH0.2ml0.2mlS.SchottmuelleriH0.2ml0.2mlInitialdilution1:201:401:801:1601:3201:6401:1280ControlSalineFinaldilution1:401:801:1601:3201:6401:12801:2560Agtest

AbknownunknownSerum1:10

1:200.2ml×4

0.8ml1.6mlSaline0.8mlmixing

0.8mlmixing

1:400.2ml×4

Saline0.8ml1.6ml0.8mlmixingSaline0.8ml

1：80

……Addingantigen1.DilutingAntibody(serum)2.Addingantigen3.Incubatingat37℃,

60’(at45℃,

30’)4.Readingtheresults62023/3/28LaboratorydiagnosisofpathogenicEntericbacteria(stool)2suspectedcolonies(Small,colorless,transparent)InoculatedinKligler’sironslantSpecimen:stooldifferentialandselectivemediumSSagarplateBiochemicalreactionKligler’sironslantserologictestSlideagglutinationCultivated72023/3/28SS

agar

plate(Shigella&Salmonella)

——adifferentialandselectivemediumusedforinitialisolationofentericbacteriaContains：basicmedium,lactose,sodiumcitrate,sodiumthiosulfate,ironcitrate,bile

salts,brilliantgreen,neutral

red(pH6.8-8.0,red→yellow）Usage：lactose,neutral

red：Lactosefermented:fermentlactosewithsufficientacidproductiontochangetheneutralredformredcoloniesLactosenotfermented:colorlesscoloniesSodiumcitrate,brilliantgreen,bile

salts:inhibitE.coliResult:E.coliShigellae&SalmonellaeRed,large,opaquecoloniesSmall,colorless,transparentcolonies82023/3/28EMB

agar

plate

Contains：basicmedium,lactose,eosin

,methyleneblueUsage：Lactosefermented:fermentlactosetoproduceacid,thenmakeeosin

andmethylenebluetobeaBLACKcompound.Lactosenotfermented:colorlesscoloniesResult:E.coliShigellae&Salmonellaelarge,black，opaquecoloniesSmall,colorless,transparentcolonies92023/3/28双糖管接种方法观察上层含乳糖部分发酵乳糖致病菌红色，非致病菌黄色；H2S试验大肠杆菌无黑色肖氏沙门菌出现黑色然后观察下层发酵葡萄糖（产酸产气）及动力乳糖,Fe2+固体葡萄糖半固体指示剂:酚红（pH6.8-8.4,黄红）102023/3/28Biochemicalreaction

（Kligler’sironslant改良克氏双糖铁琼脂斜面）OberservationLactose（10）Glucose（1）Ferroussulfateindicator:phenolredpH6.8-8.4Yellow→redGlucoselactoseH2S++-+SlantYellowRedblackBottomYellowandbubbleYellowblack112023/3/28InocuatingKligler’sironagarslantInoculatingneedleRemovingcellsfromssagarplatewithaneedleStabneedletobottomofthetubeAstheneedleiswithdrawnfromtheagar,streakitacrossthefaceoftheslantfrombottomtothetopIncubateat37℃122023/3/28

Lactose

GlucoseH2S

UreaseE.coli

＋

+S.dysenterias－

+-S.typhi－

++/-S.paratyphiA－

+-Proteus－++132023/3/28LaboratorydiagnosisofpathogenicEntericbacteria(stool)Specimen:stooldifferentialandselectivemediumSSagarplateBiochemicalreactionKligler’sironslantserologictestSlideagglutination142023/3/28ReadingtheresultsofWidaltestExaminingthecontroltubesfirst:negativeTheagglutinationofOantigenasa“matt”or“carpet”atthebottom.TheagglutinationofHantigensloose,woolyorcottony（＋＋＋＋）liquidclear,allantigenareagglutinateatthebottom.（＋＋＋）liquidclear,mostofantigen(75%)areagglutinateatthebottom（＋＋）liquidsemitransparent,partofantigenareagglutinateatthebottomobviously.（＋）liquidturbid,alittleantigenareagglutinateatthebottom.（－）negative,sameascontrolAgglutinationtiter:thehighestdilutionof“++”152023/3/28Interpretationofwidaltestbaseline：S.typhiOantibody≥1：80，S.typhiHantibody≥1：160,S.paratyphiAHantibody≥1：80Arisein