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1、Lecture 10Staphylococci and StreptococciBIOL 533Lecture 10Medical MicrobiologyBIOL 5331Lecture 10StaphylococciImportant human pathogenCauses both relatively minor and serious diseasesOne of the hardiest of the non-sporeforming bacteriaCan exist on dry surfaces for a long periodRelatively heat-resist
2、ant; temperature range of 18 - 40 CBIOL 5332Lecture 10StaphylococciMorphologyGram+ grape-like cluster, but in clinical specimens, can be a single cocci or diplococciGeneral physiological characteristicsNonmotileFacultatively anaerobicCatalase +Grows in media containing 10% NaClBIOL 5333Lecture 10Sta
3、phylococciRelationship to disease (only 3 important)S. aureuscauses a number of diseasesS. epidermidispresent in normal flora (normally benign, except when introduced via catheters, etc.)S. saprophyticuscauses uninary tract infectionsBIOL 5334Lecture 10StaphylococciMicrobial physiology and structure
4、Capsule may not be found growing on media, but it is usually present in vivoTeichoic acids are phosphate containing polysaccharides bound to both peptidoglycan and cytoplasmic membraneSpecies specificPoor immunogens, but when bound to peptidoglycan, get an antibody responseBIOL 5335Lecture 10Pathoge
5、nesis of S. aureusFeatures typical of staphylococci infections:Initial lesion is normally mild and localizedResults in a boilnormally, it is self-limitingCan result in systemic infectionBIOL 5336Lecture 10Pathogenesis of S. aureusStage I: encounterhumans are major reservoir for S. aureusColonize nos
6、e and are found in about 30% of individualsTransiently found on skin, oropharynx, and fecesTransmitted via:Hand contactAerosols from pneumonia patientsBIOL 5337Lecture 10Pathogenesis of S. aureusStage I, continuedCertain occupations are more prone to colonizationPhysicians, nurses, hospital workersC
7、ertain classes of patients are more prone to colonizationDiabetics, hemodialysis patients, and drug abusersBIOL 5338Lecture 10Pathogenesis of S. aureusStage II: entrynot normally through unbroken skinCan enter if large numbers have accumulated through poor hygieneBIOL 5339Lecture 10Pathogenesis of S
8、. aureusStage III: spread and multiplicationSurvival depends onNumber of organismsSite involvedSpeed with which inflammatory response is mountedImmunological competence of hostIf inoculum is small and host immunologically competent: infection normally defeatedBIOL 53310Lecture 10Pathogenesis of S. a
9、ureusStage IV: damageLocal infection leads to formation of abscess (collection of pus)In skin, boils or furunclesInterconnected abscesses are called carbunclesMay also spread in subcutaneous or submucosal tissuescellulitisBIOL 53311Lecture 10Pathogenesis of S. aureusStage IV, continuedDevelopmentinv
10、olves both host and bacterial factorsAcute inflammatory reactionProportion of bacteria survive and are capable of lysing neutrophils that engulfed themOutpouring of lysosomal enzymes that damage surrounding tissuesInflammatory area surrounded by fibrin clotBIOL 53312Lecture 10Virulence Factors of S.
11、 aureusStage IV, continuedVirulence factorsmost designed to avoid phagocytosis or survive once ingestedWall componentsSurrounded by capsule: not as effective as pneumococcus or meningococcusCell wall murein activates complement by alternative pathwayTeichoic acid also activates and involved in adher
12、enceProtein A interferes with opsonization by binding with Fc region of Abcomplement activated primary pathwayBIOL 53313Lecture 10Virulence Factors of S. aureusStage IV, continuedSecretion of enzymesCatalasehydrogen peroxide to water and oxygen (all staphylococci produce)Coagulasemakes fibrin clot (
13、wbc penetrate badly; only S. aureus)Hylauronidasedegrades connective tissues (facilitates spread; 90% of S. aureus strains)Fibrinolysin (staphylokinase)dissolve fibrin clots (virtually all S. aureus)BIOL 53314Lecture 10Virulence Factors of S. aureusStage IV, continuedSecretion of enzymesLipasesrequi
14、red for invasion into cutaneous and subcutaneous tissues (found in all S. aureus and 30% of others)Nucleaseheat stable (role is uncertain; S. aureus)PenicillinaseBIOL 53315Lecture 10Virulence Factors of S. aureusStage IV, continuedSecretion of toxinsCytolytic (membrane-damaging by pores)Alpha, beta,
15、 (sphingomyelinase C), delta, gamma, leukocidin (cannot lyse red blood cells)Others lyse rbc and leukocytes (referred to previously as hemolysins)Cause lysis of neutrophils leading to massive lysosomal enzyme secretionBIOL 53316Lecture 10Virulence Factors of S. aureusStage IV, continuedSecretion of
16、toxinsExfoliative toxin (scalded skin syndrome) extrachromosomalToxic Shock Syndrome toxin-1 (enterotoxin F)exotoxin secreted during growthSome produce enterotoxin B instead (role not clear)BIOL 53317Lecture 10Virulence Factors of S. aureusStage IV, continuedSecretion of toxinsEnterotoxins (A-E)foun
17、d in both S. aureus and S. epidermidisResistant to hydrolysis by gastric and jejunal enzymesStable to heating at 100C for 30 minutesMechanism of toxin activity not understood; no satisfactory animal modelStimulate intestinal peristalsis and have CNS effect; intense vomitingBIOL 53318Lecture 10Pathog
18、enesis of S. aureusTreatmentAntibioticsTypes:Methicillin, oxacillin, nafcillin, and dicloxacillin (semisynthetic penicillins resistant to -lactam hydrolysis)Majority of patients can be treated, but 10-15% S. aureus and 40% coagulase-negative staphylococci are resistant; treat with vancomycinBIOL 533
19、19Lecture 10Pathogenesis of S. aureusTreatmentAntibioticsResistance:Plasmid-borne (hydrolysis of -lactam ring)Chromosomalchange in structure of penicillin-binding proteinsBIOL 53320Lecture 10StreptococciFermentative (oxygen tolerant) Gram+ cocci in chainsSensitive to penicillinsHuman reservoirpassed
20、 from person to personBIOL 53321Lecture 10StreptococciProperties of Lancefield Groups (CHO antigens on wallsee handout)Group A: cross-reaction can lead to:Rheumatic feverGlomerulonephritisBIOL 53322Lecture 10StreptococciRecent Group A Streptococcus virulence factorsM-like proteinsbind IgM IgG (prote
21、ase inhibitor) and alpha2 macroglobulinF proteinadherence to epithelial cellsC5a peptidasedegrades C5A pyrogenic exotoxins; previously called erythrogenic toxinsBIOL 53323Lecture 10Staph and Strep ToxinsS. aureus toxic shock TSST-1S. pyogenes toxic shock TSSL-1S. pyogenes scarlet feverSPE-1 (childre
22、n, not adults; immunity)BIOL 53324Lecture 10Staph and Strep ToxinsS. aureus : Toxic Shock SyndromeFever, diffuse rashExfoliation of skin on palms and soles of feetNormally doesnt compete well in relatively anaerobic vaginal areaBIOL 53325Lecture 10Staph and Strep ToxinsS. aureus : Toxic Shock Syndro
23、meSuper-absorbent tampon:Created aerobic pocketsRemoved Mgproducing toxinAfter removed tampon, cases declined; did not disappearStill associated with wounds, rare nasal surgeryBIOL 53326Lecture 10Staph and Strep ToxinsS. pyogenes: Toxic Shock-Like SyndromeSkin or wound infection - bloodstreamDeath r
24、ate 30%; over 10-fold higher than TSSTSeen in immunocompromised peopleAlso other infections occurred: soft tissue infection with influenza symptomsHigh fatality rate because rapid development of shock and multiple organ failureBIOL 53327Lecture 10Staph and Strep ToxinsS. pyogenes: Toxic Shock-Like S
25、yndromeFeatures in common with scarlet feverOccur in healthy peopleBoth associated with high fatality rateProduce same exotoxin: streptococcal pyrogenic exotoxin (Spe)Similar in mechanism to TSST-1BIOL 53328Lecture 10Staph and Strep ToxinsComparing TSLS-1 and TSST-1:Rash, fever, shock, multiple orga
26、n failure; resemble endotoxin septic shockBoth toxins superantigensSame mechanisms of actionLimited similarity at amino acid sequence levelBIOL 53329Lecture 10Staph and Strep ToxinsTSLS-1 related to erythrogenic toxin (scarlet fever; SpeA)Serotypes:Spe ATSLS or invasive S. progenesSpe BSpe CSome str
27、ains dont produce Spe A, so Spe B or Spe C also has roleBIOL 53330Lecture 10Staph and Strep ToxinsHow do TSST-1 and SPE cause shock and multiple organ failure?Hypotheses not mutually exclusiveBIOL 53331Lecture 10Shock and Organ FailureFirst Hypothesis: same as LPS triggering release cytokines IL1,TN
28、FConsistent with role as superantigenPromote association between macrophage and helper T cellsproliferation of T cells producing high IL2 levelSecondarily produce IL1 TNFInject TSST-1 into rabbits; elevated levels IL1 TNFBIOL 53332Lecture 10Shock and Organ FailureSecond hypothesis: increase bodys se
29、nsitivity to LPS; consistent with:Acts synergistically with LPS to amplify toxic effects in vitro and in animalsConceivablelow levels leaching into blood due to lysis of resident microfloraNormally no effectPresence of Spe or TSST-1 causes an effectBIOL 53333Lecture 10Shock and Organ FailureEvidence
30、 to support role for LPS in TSST and TSLSInjecting TSST-1 or Spe is lethal to rabbitsInjecting exfolatin and concanavalin A not lethal to rabbits Both elicit T cell proliferation, but dont enhance sensitivity to LPSBIOL 53334Lecture 10Shock and Organ FailureEvidence to support role for LPS in TSST a
31、nd TSLSTSST-1 not lethal to gnotobiotic animalsWouldnt expect leakage, but still T cell responseTherefore, both suggest T cell proliferation not as important as synergy of LPSNot conclusive; difficult to prove same level T cell stimulation, proliferation occurred in all casesBIOL 53335Lecture 10Shoc
32、k and Organ FailureThird hypothesis:TSST-1 can act directly on endothelial cellsDamage causes malfunction in circulatory system, which creates hypotensionData: swelling associated with massive leakage of fluid from capillaries is marked symptom of both TSST and TSLSCould also be result of action of
33、blood vessels by cytokines, coagulation, or complement cascadeBIOL 53336Lecture 10Staph and Strep ToxinsMortality of S. pyogenes vs. S. aureusTSLS higher than TSSTSLS strains enter bloodstreamTSS, only the toxin circulatesS. pyogenes known to be invasive; killed by PMNs and macrophage if ingestedBIO
34、L 53337Lecture 10S. pyogenes InvasivenessStrategies for evading phagocytosis; (1):M protein binds H factor better than factor BLeads to degradation of C3bTherefore, prevents opsonization by C3b and formation of C3 convertaseBIOL 53338Lecture 10S. pyogenes InvasivenessStrategies for evading phagocyto
35、sisData supporting:M mutants yield more susceptible to phagocytosis; less virulent than wild typeAb against M protective80 serotypes of M; possibly evades host antibodies by changing serotype; however, no data to support this hypothesisBIOL 53339Lecture 10S. pyogenes InvasivenessStrategies for evadi
36、ng phagocytosis; (2):Protease cleaves C5aChemoattractant stimulates oxidative burstSome activation of complement could occur in spite of M protein becauseLysis releases wall components that activate complementStreptococci could protect themselves- C5a peptidaseData supporting:C5a mutants less virule
37、nt that wild type in animalsBIOL 53340Lecture 10S. pyogenes InvasivenessStrategies for evading phagocytosis; (3):M like proteinsSequence and structural similarity to MCOOH embedded; NH2 exposedMost similar to M and each other at carboxy endThese proteins bind Fc portion of IgG and IgABIOL 53341Lectu
38、re 10S. pyogenes InvasivenessStrategies for evading phagocytosisM like proteins; possible rolesCoat with host proteinless likely detected as invader by complement and immune systemAdherence for body cells that contain Ab on surfaceAlso can bind host protease inhibitor such as 2 macroglogulinHost use
39、s protease inhibitor to protect against proteases released by phagocytesBIOL 53342Lecture 10S. pyogenes InvasivenessStrategies for evading phagocytosis; (4):F proteinbinds fibronectinAdherence of bacteria to tissuesEvasion of immune systemSummary of invasivenessNo direct evidence M-like proteins inv
40、olved in virulenceFound in impetigo strains, not always in severe invasive strainsNeed mutant studies to answer questionsBIOL 53343Lecture 10S. pyogenes VirulenceRegulation of S. pyogenes virulence genesExpression M, C5a peptidase, and some M-like proteins; regulated at transcriptional levelResponds
41、 to CO2 levelsIncreased CO2 causes increased productionBIOL 53344Lecture 10S. pyogenes VirulenceRegulation of S. pyogenes virulence genesRegulatory genemry transcriptional activator; sequence analysis shows it is part of two-component systemSensornot foundActivatorAlso known that speA gene on temper
42、ate phageBIOL 53345Lecture 10S. pyogenes PathogenesisTreatment and preventionTSST, TSLS are medical emergenciesSurgical debridement of wounds prevents further production of toxinAntibiotics; penicillinToxic effects TSST-1 countered by intravenous rehydration; counter hypotensionBIOL 53346Lecture 10S
43、treptococcal TreatmentPreventionVaccine possibleTarget against MPossible problems# serotypes, but severe invasive disease caused by fewAB against M cross-reacts with heartBIOL 53347Lecture 10Streptococcal Sequellae HypothesesFirst: Autoimmune theoryEpitopes that cross react with epitopes on cardiac
44、myosin and sarcolemmal membrane proteinsThus, T cells or antibodies could attack tissueInflammatory response damages heart valvesBIOL 53348Lecture 10Streptococcal Sequellae HypothesesGlomerulonephritisHigh levels Ab to streptococcal Ag circulating in blood stream causes AgAb complexes to accumulate
45、in kidneyInflammatory response attacks kidney interfering with kidney functionBIOL 53349Lecture 10Streptococcal Sequellae HypothesesData supportingAg-Ab complexes visible in people with glomerulonepheritisglomeruliDecrease in C3 and other complement components also seen; supports hypothesis that inf
46、lammatory response is occurringSecond: Toxins cause sequellaeBIOL 53350Lecture 10Streptococcal Sequellae HypothesesMain argument againstTime lag between initial infection and development of rheumatic fever (RF; several weeks) or glomerulonephritis (10 days)Normally, if due to toxin, within a weekCan
47、didates for toxin most likely to cause glomerulonephritis: streptococcal O, streptokinase, or SpeBIOL 53351Lecture 10Glomerulonephritis HypothesesStreptococcal O cytotoxinMechanism and aa sequence similarity to pneumolysinPore-forming toxinInjected into lab animals; damages heartTherefore, may have
48、role in RFAlso, very immunogenic; maybe Ab damageBIOL 53352Lecture 10Glomerulonephritis HypothesesStreptokinasePlasminogenplasminTherefore causes symptoms similar to glomerulonephritis in animalsInteresting, but not provenBIOL 53353Lecture 10Rheumatic Fever HypothesesSpe RF strains produce Spe; othe
49、rs dontEnhances cardiotoxicity caused by Streptococcal O in animalsHavent explained long time lagBIOL 53354Lecture 10Rheumatic Fever HypothesesMysterious feature of RF unexplainedTreated with antibiotics for as late as 9 days after symptoms, still protected against RFAfter 9 days, toxic products sho
50、uld be circulating and immune response underwayRecurrence of diseaseNormally, infection results from different strainSome result from same strainRF symptoms take as long to develop as in originalBIOL 53355Lecture 10Rheumatic Fever HypothesesMysterious feature of RF unexplainedIf caused by autoimmune
51、 response, would expect faster responsePossible explanation: previously exposed produces primed immune systemBIOL 53356Lecture 10Streptococcal SequellaeTreatment and preventionStrep throat self-limitingTreat with antibiotics and prevent RF and glomerulonephritisOnly S. pyogenes strains cause RF or G
52、Not all people colonized actually contract RF or GBecause of the seriousness, treat any strain with antibioticsBIOL 53357Lecture 10Streptococcal SequellaeTestsBlood agarHemolysisBacitracin sensitivityRapid test and culture test both yield high number of false negativesFor patients who are allergic t
53、o penicillin, use erythromycinBIOL 53358Lecture 10Streptococcal SequellaePrevious damageEven heart murmurDentists recommend prophylactic penicillin before dental workKill bacteria escaping into blood stream from mouth (oral bacteria are susceptible to penicillin)Reduces chance of colonizationBIOL 53
54、359Lecture 10Staphylococcal EnterotoxinsNormal enterotoxins cause diarrheaWater loss from small intestine mucosaCause c-GMP or c-AMP levels in mucosal cells to riseBIOL 53360Lecture 10Staphylococcal EnterotoxinsStaph toxins operate in a different mode; hypotheses include:1) Stimulates vagus nerve en
55、dings in stomach lining that control emetic (vomiting) responseIf hypothesis correct, its a neurotoxin, not enterotoxin2) SuperantigenIL2Administering IL2 to human volunteers produces many of the same symptoms (nausea, vomiting, fever, malaise)BIOL 53361Lecture 10Staphylococcal EnterotoxinsNeither h
56、ypothesis conclusively provenCould be a combination of bothSeven serotypes: SEA, SEB, SEC1, SEC2, SEC3, SED, SEE30 kDa proteins share considerable aa similarity and single internal S-SBIOL 53362Lecture 10Staphylococcal EnterotoxinsSE closely related to SpeGenes coding for SEC1 and Spe A have 60% nuc
57、leotide identitySE produced in different amounts by different strainsentA produces much less toxin than entBDifferences in promoter strengthBIOL 53363Lecture 10Staphylococcal EnterotoxinsSEB on integrated plasmidSEA on lysogenized phageOne strain: entB, entC, plasmid-borne SED: entD on 27.6 kb plasmidBIOL 53364Lecture 10Lecture 10Questions?Comments?Assignments.BIOL 53365
