微生物腸桿菌科

Chapter10

Entericbacilli第1頁Outline

GeneralpropertiesofentericbacilliEscherichia（埃希菌屬）Shigella（志賀菌屬）Salmonella（沙門菌屬）第2頁MorphologyGram-negativeRodshaped(bacilli)Mostformpiliandperitrichousflagella(周鞭毛)DonotformsporesCulturecharacteristicsFacultativeanaerobes(兼性厭氧)oraerobes(需氧)Growreadilyonsimplemedia.Colonies:circular,convex,smoothBloodagarplate:somecanformhemolyticzones.Liquidmedia:turbid.第3頁BiochemicalreactionsCatalase（觸酶）test:+Reducenitrate(硝酸鹽)tonitrite(亞硝酸鹽)Oxidase(氧化酶)test:-Lactosefermentationtest(乳糖發(fā)酵實(shí)驗(yàn))乳糖發(fā)酵實(shí)驗(yàn)可用于初步鑒別腸道致病菌和非致病菌腸道致病菌多數(shù)不發(fā)酵乳糖（沙門菌、志賀菌）腸道非致病菌多數(shù)發(fā)酵乳糖（埃希菌）第4頁AntigensOantigens:LPS,IgMHantigens:flagellin,IgGCapsularantigens:typespecific,ViantigenandKantigen.Resistance:

relativelyweakVariation第5頁Gram-negativerod-shapedbacilli第6頁MacConkeyagar

AculturemediumdesignedtogrowGram-negativebacteriaandstainthemforlactosefermentation.Itcontainsbilesalts(toinhibitmostGram-positivebacteria,exceptEnterococcusandsomespeciesofStaphylococcus),crystalvioletdye(whichalsoinhibitscertainGram-positivebacteria),neutralreddye(whichstainsmicrobesfermentinglactose),lactose(乳糖)andpeptone(蛋白胨).第7頁Macconkeyagarwithlactose(left)andnon-lactose(right)fermenters.第8頁

第9頁

SmoothRough第10頁HO第11頁Section1

Escherichia(埃希菌屬)

E.coli第12頁E.coliBiologicalcharacterizationPathogenesisandsyptomatologyPathogenicsubstancesClinicalfindingsLaboratorydiagnosisTreatment第13頁MorphologyGram-negativeRodshaped(bacilli)Mostformpiliandperitrichousflagella(周鞭毛)DonotformendosporesCulturecharacteristicsFacultativeanaerobes(兼性厭氧)Liquidmedia:turbid.Growreadilyonsimplemedia.Colonies:circular,convex,smooth.第14頁BiochemicalreactionsLactosefermentationtest(乳糖發(fā)酵實(shí)驗(yàn)):⊕(SSagar,KIagar)H2Stest:-Motilitytest:+IMViCtests:++--Antigens:Oantigens:>170typesHantigens:>50typesKantigens:>100types第15頁Gram-negativerod-shapedbacilli第16頁SSagar

(SalmonellaShigellaagar)Ingredients:Nutrientagar100ml

Lactose(乳糖)1.00gBilesalts0.85ginhibitorSodiumcitrate0.85ginhibitorSodiumthiosulfate0.85g10%ferriccitrate1ml

1%Neutralred0.25mlindicator1%brilliantgreen0.033mlinhibitor第17頁EscherichiacolionSSagar(pinkcolony)第18頁KliglerIronAgar(KIagar)Ingredients:

Peptones2.00gNaCl0.50g

Glucose

0.10g

Lactose1.00g

Ferroussulfate0.05gSodiumthiosulfate0.075gAgar0.50g0.2﹪phenolred1.25mlDistilledwater100mlSlantButtInacidcondition:yellowInbasecondition:red第19頁ReadingresultsofKIagar(sulfuretedhydrogen)第20頁2-Shigella

3-Salmonella4-E.Coli第21頁MotilitytestThetestisbasedontheflagellumofthebacteria.

Ifthebacteriahaveflagellum,thesemisolidmediumwillbecloudingandthetestwillbepositive.

Semisolidmedium第22頁E.coliSSagar:reddishcolonyKIagar:yellowslantsandbutts,

bubbleinthebottom,thelineofinoculationisblurrynoH2SSemisolid:turbidgrowth,withflagella第23頁Indoletests色氨酸吲哚對二甲基氨基苯甲醛玫瑰吲哚+E.coli第24頁Methylredtests某些細(xì)菌在糖代謝過程中，分解葡萄糖產(chǎn)生丙酮酸，丙酮酸可進(jìn)一步分解，產(chǎn)生甲酸、乙酸、乳酸等，使培養(yǎng)基旳pH降至4.5下列，當(dāng)加入甲基紅試劑則呈紅色，為甲基紅實(shí)驗(yàn)陽性。若細(xì)菌分解葡萄糖產(chǎn)酸量少，或產(chǎn)生旳酸進(jìn)一步轉(zhuǎn)化為其他物質(zhì)（如醇、酮、醚、氣體和水等），則培養(yǎng)基旳酸度仍在pH6.2以上，故加入甲基紅批示劑呈黃色，是為陰性。第25頁Voges-Proskauertest某些細(xì)菌在葡萄糖蛋白胨水培養(yǎng)基中能分解葡萄糖產(chǎn)生丙酮酸，丙酮酸縮合，脫羧成乙酰甲基甲醇，后者在強(qiáng)堿環(huán)境下，被空氣中氧氧化為二乙酰，二乙酰與蛋白胨中旳胍基生成紅色化合物，稱V-P（+）反映。大腸埃希菌V-P實(shí)驗(yàn)陰性，產(chǎn)氣腸桿菌V-P實(shí)驗(yàn)陽性。第26頁Citrateutilizationtest以枸櫞酸鈉為唯一碳源、PH值7.0旳培養(yǎng)基上，產(chǎn)氣桿菌分解枸櫞酸鈉產(chǎn)生碳酸鹽，使培養(yǎng)基由中性變?yōu)閴A性，培養(yǎng)基中批示劑溴麝香草酚藍(lán)（BTB）由淺綠色變?yōu)樯钏{(lán)色，此為枸櫞酸鹽運(yùn)用實(shí)驗(yàn)陽性。大腸桿菌因不能運(yùn)用枸櫞酸鹽，此實(shí)驗(yàn)為陰性反映。第27頁

第28頁第29頁AdhesinsExotoxinsOthervirulencefactorsEndotoxinCapsulesTypeⅢsecretionsystemsPathogenicsubstances第30頁Adhesins（黏附素）Function:facilitatethespecificadherence.Severaltypes:Colonizationfactorantigen,CFAⅠ/Ⅱ/Ⅲ（定植因子抗原Ⅰ,Ⅱ,Ⅲ）Aggregativeadherencefimbriae,AAFⅠ/Ⅲ（集聚黏附菌毛Ⅰ和Ⅲ）Bundleformingpili,Bfp（束形成菌毛）Intimin（緊密黏附素）Ppili（P菌毛）第31頁Exotoxins（外毒素）Shigatoxins(Stx-1,Stx-2)（致賀毒素Ⅰ和Ⅱ）Heatstableenterotoxins(STa,STb)（耐熱腸毒素a和b）Heatlabileenterotoxins(LT-Ⅰ,LT-Ⅱ)（不耐熱腸毒素Ⅰ和Ⅱ）Hemolysin（溶血素A）:uropathogenicE.coli(UPEC)第32頁ClinicalfindingsExtraintestinalinfections（腸道外感染）Septicemia（敗血癥）Meningitisofnewborn（新生兒腦膜炎）Urinarytractinfection(UTI)（泌尿道感染）-UPEC,urophathogenicE.coli（尿路致病性大腸埃希菌）Intestinalinfections（胃腸炎）-Diarrhea-causingE.coli第33頁第34頁Diarrhea-causingE.coliETEC:enterotoxigenicE.coli腸產(chǎn)毒型大腸埃希菌“Traveler’sdiarrhea”&Children’sdiarrheaWaterydiarrheawithoutbloodSelf-limiting(3~4days)EIEC:enteroinvasiveE.coliEPEC:enteropathogenicE.coliEHEC:enterohemorrhagicE.coliEAEC:enteroaggregativeE.coli第35頁Enterotoxigenic(ETEC)Pathogenicsubstances★EnterotoxinsHeatlabileenterotoxin,LT-ⅠHeatstableenterotoxin,ST-aColonizationfactorantigens(CFAⅠ/Ⅱ/Ⅲ)第36頁Heatlabileenterotoxin（不耐熱腸毒素）EncodedbyplasmidLT-Ⅰ:amajorvirulencefactor

HeatlabileLikecholeragen(霍亂腸毒素)1subunitA(toxic)+5subunitB(binding)SubunitB:bindtoGM1-ganglioside（神經(jīng)節(jié)苷脂）SubunitA:Activatesadenylcyclase(腺苷環(huán)化酶)ATP→cAMPSecretionwater/ions第37頁霍亂腸毒素旳作用機(jī)制第38頁Heatstableenterotoxin（耐熱腸毒素）EncodedbyplasmidSTa:amajorvirulencefactor

HeatstableFunctions:Activatesguanylylcyclase(鳥苷環(huán)化酶)cGMP

Secretionwater/ions第39頁LT-Ⅰ&STa

TheresultisthestimulationofchloridesecretionoutofthecellandtheblockageofNaClabsorption.Theneteffectistheaccumulationofwaterandelectrolytesintothebowellumen.第40頁環(huán)境和食品衛(wèi)生學(xué)上，“大腸菌群數(shù)”常被用作糞便污染旳檢測指標(biāo)≤3個大腸菌群/1L飲水≤5個大腸菌群/100ml瓶裝汽水、果汁衛(wèi)生學(xué)指標(biāo)我國飲用水衛(wèi)生原則：第41頁第42頁第43頁Section2Shigella

第44頁第45頁第46頁Biological

characterizationNoflagellaGlucosefermentationtest:+Lactosefermentationtest:-H2Stest:-Motilitytest:-Antigens:O

andKantigensS.dysenteriae(GroupA)：痢疾志賀菌S.flexneri(GroupB)：福氏志賀菌S.boydii(GroupC)：鮑氏志賀菌S.sonnei(GroupD)：宋內(nèi)志賀菌第47頁第48頁Shigella(small,colorless,transparentcolony)E.coli(large,reddishcolony)第49頁Shigellasonneionbloodagar.Mediumsized,greycoloniesthatcausenoalterationoftheblood.第50頁2-Shigella

3-Salmonella4-E.Coli第51頁MotilitytestThetestisbasedontheflagellumofthebacteria.

Ifthebacteriahaveflagellum,thesemisolidmediumwillbecloudingandthetestwillbepositive.

Semisolidmedium第52頁ShigellaSSagar:flaxencolonyKIagar:redslantsandyellowbutts,nobubbleinthebottom,thelineofinoculationisclearnoH2SSemisolid:lineargrowth,withoutflagella第53頁P(yáng)athogenesis★PathogenicsubstancesInvasivenessEndotoxinExotoxinClinicalfindingsBacillarydysentery(菌痢)HumansareonlyknownreservoirsOral-fecaltransmissionAcuteChronic:>2M第54頁InternalizedshigellaeinduceapoptosisofmacrophageandreleaseofthebacteriaAttractedbythecytokinesreleasedbymacrophageActivatestheinvasiongenesonthevirulenceplasmidMcell3.Exotoxin2.Endotoxin1.Invasiveness第55頁靶細(xì)胞：回腸末端和結(jié)腸部位旳黏膜上皮細(xì)胞一方面侵犯派伊爾淋巴結(jié)旳M細(xì)胞致病過程：黏附→Ⅲ型分泌系統(tǒng)→細(xì)胞膜內(nèi)陷、細(xì)菌內(nèi)吞→胞內(nèi)生長繁殖→擴(kuò)散膿血黏液便：壞死旳黏膜、死亡旳WBC、細(xì)胞碎片、纖維蛋白和血液1、侵襲力：第56頁2、毒素：內(nèi)毒素作用于腸壁通透性增長（腹瀉）粘膜炎癥、潰瘍（膿血黏液便）局部全身內(nèi)毒素血癥（發(fā)熱、神志障礙、中毒性休克）外毒素

志賀毒素植物NS功能紊亂（腹痛、里急后重）干擾蛋白合成，導(dǎo)致上皮細(xì)胞損傷第57頁所致疾病——細(xì)菌性痢疾急性細(xì)菌性痢疾急性典型：發(fā)熱、腹痛、里急后重、膿血粘液便急性中毒型：小兒常見，全身性中毒癥狀慢性細(xì)菌性痢疾：病程遷移二月以上傳染源：病人和帶菌者傳播途徑：糞—口傳播第58頁

中毒性痢疾病人第59頁

Specimens:Freshstool,mucusflecks,andrectalswabs.LargenumbersoffecalleukocytesandsomeRBCsmayoftenbeseenmicroscopically.

Culture:Differentialandselectivemediaasusedforsalmonellae.Serology:Multipledrugresistance.LaboratorydiagnosisTreatment:第60頁Humansaretheonlyreservoirforshigellae.Transmissionofshigellae:water,food,fingers,feces,andflies.Mostcasesoccurinchildrenunder10yearsofage.Preventionandcontrolofdysentery:Sanitarycontrolofwater,foodandmilk;sewage

disposal;andflycontrol.Isolationofpatientsanddisinfectionofexcreta.Detectionofsubclinicalcasesandcarriers.Preventionandcontrol

第61頁Salmonella第62頁第63頁MorphologyGram-negativeRodshaped(bacilli)Mostformpiliandperitrichousflagella(周鞭毛)DonotformendosporesCulturecharacteristicsFacultativeanaerobes(兼性厭氧)Growreadilyonsimplemedia.Bacterialcharacterization第64頁BiochemicalreactionsLactosefermentationtest(乳糖發(fā)酵實(shí)驗(yàn)):-Glucosefermentationtest:+/⊕H2Stest:+/-Motilitytest:+Antigens:Oantigens,Hantigens,Viantigens第65頁第66頁第67頁SalmonellaonSSagar（colourlesscolonywithablackcenter）第68頁SalmonellaonSSagar第69頁2-志賀菌

3-沙門菌

4-大腸埃希菌第70頁MotilitytestThetestisbasedontheflagellumofthebacteria.

Ifthebacteriahaveflagellum,thesemisolidmediumwillbecloudingandthetestwillbepositive.

Semisolidmedium第71頁SalmonellaSSagar:flaxencolonywithblackcenterKIagar:yellowslantsandbutts,nobubbleinthebottom,thelineofinoculationisblurryproduceH2SSemisolid:turbidgrowth,withflagella第72頁P(yáng)athogenicsubstancesInvasiveness(侵襲力)——小腸黏膜（M細(xì)胞）Pili（特異性菌毛）SPI-ⅠandSPI-Ⅱ（Ⅲ型分泌系統(tǒng)）Acidtoleranceresponse,atr（耐酸應(yīng)答基因）Oxidase（氧化酶）,SOD（超氧化物歧化酶）Viantigen（微莢膜）Endotoxin(內(nèi)毒素)：fever,leukopenia,shockEnterotoxin(腸毒素):likeETECenterotoxin.第73頁沙門菌(紅色)入侵培養(yǎng)旳人體細(xì)胞第74頁ClinicalfindingsEntericfever(腸熱癥)Foodpoisoning(食物中毒)：最常見Septicemia(敗血癥)Carrierstate(無癥狀帶菌者)第75頁膽囊-----腸道-------糞排菌皮膚----血栓出血--玫瑰疹腎-----尿肝脾-----腫大骨髓------受克制傷寒和副傷寒旳致病過程傷寒和副傷寒桿菌小腸上部粘膜腸系膜