第12章-细菌感染性疾病的分子机制-分子生物学课程课件.ppt

1、返回目录返回目录返回主页返回主页上一页上一页返回目录返回目录返回主页返回主页返回目录返回目录返回主页返回主页Bacterial Pathogenesis References:1.Murray,P.et al.,Medical Microbiology(5th edition)2.Samuel Baron,Medical Microbiology(4th edition)3.Wilson,B.A.et al.,Bacterial Pathogenesis:A Molecular Approach(3rd edition)4.Falkow,S.and Miller,V.L.,Molecular

2、Genetics of Bacterial Pathogenesis:A Tribute to Stanley Falkow细菌的感染与致病机制细菌的感染与致病机制InfectionInfection细菌的细菌的感染（传染）感染（传染）细菌细菌 宿主防御功能宿主防御功能 病理过程病理过程病原菌（或致病菌）病原菌（或致病菌）(pathogen)(pathogen)：能使宿主致病的细菌。能使宿主致病的细菌。非病原菌非病原菌(nonpathogen(nonpathogen)：不能造成宿主感染的细菌。不能造成宿主感染的细菌。一、感染的来源一、感染的来源 外源性感染外源性感染:传染源、转播方式与途径传染

3、源、转播方式与途径 exogenous infectionexogenous infection 内源性感染内源性感染 endogenous infectionendogenous infection二、感染的类型 （一）（一）隐性感染隐性感染inapparent inapparent infection/suclinicalinfection/suclinical infection infection （二）（二）显性感染显性感染 apparent infectionapparent infection ：(传染传染病病)按感染部位不同，分为：按感染部位不同，分为：局部感染局部感染:全身感

4、染：全身感染：毒血症毒血症 toxemiatoxemia 外毒素外毒素进入血液，引起特殊的毒性症状，进入血液，引起特殊的毒性症状，而细菌不进入血液。而细菌不进入血液。菌血症菌血症(bacteremia(bacteremia)病原病原菌侵入血流，菌侵入血流，但但未未在血流中在血流中生生长繁殖长繁殖。细菌仅短暂通过血流。细菌仅短暂通过血流。败血症败血症(septicemia(septicemia)病原病原菌侵入血流后在其中大量生长繁菌侵入血流后在其中大量生长繁殖，产生殖，产生毒性代谢产物毒性代谢产物，引起严，引起严重的全身中毒症状。重的全身中毒症状。脓毒血症脓毒血症 pyemiapyemia化脓性

5、细化脓性细菌侵入血流后在其中大量菌侵入血流后在其中大量繁殖繁殖，并可通过血流扩散至机体的其他并可通过血流扩散至机体的其他组织器官，产生组织器官，产生新新的的化脓病灶化脓病灶。内毒素血症内毒素血症 endotoxemiaendotoxemia 毒素入血毒素入血 菌入血菌入血 中毒症状中毒症状毒血症毒血症 +-+-菌血症菌血症 -+-+-败血症败血症 +脓毒血症脓毒血症 +（化脓性细菌）（化脓性细菌）新的化脓灶新的化脓灶pathogen致病菌致病菌条件致病菌条件致病菌/机会致病菌机会致病菌非致病菌非致病菌The significance of normal flora（正常菌群的重要性）const

6、itute a protective host defense mechanism（构建宿主保护机制）（构建宿主保护机制）:Competition of nutrients and receptors Metabolic substances by normal flora:e.g.,bacteriocins,antibiotics,etc.serve a nutritional function（提供营（提供营养）养）:several B vitamins and vitamin Kkeep our immune systems in tune（维持免（维持免疫系统）疫系统）normal f

7、lora share many antigenic determinants with pathogenic organisms Opportunistic pathogens（机会致病原/条件致病原）Definition:normally nonpathogenic microorganisms capable of causing infection disease in an immunosuppressed host.（非病原微生物在免疫低下宿主中能够造成感染疾病）Conditions of causing diseases by opportunistic pathogens:o A

8、lteration of colonization sites（定殖位点的改变）o Declination of host immune system function（宿主免疫系统功能的下降）Bacterial Pathogenesis（细菌致病机制）（细菌致病机制）Introduction（简介）（简介）Host Susceptibility（宿主易感性）（宿主易感性）Pathogenic Mechanisms（致病机制）（致病机制）Virulence Factors（毒力因子）（毒力因子）Steps in Successful Infection（决（决定细菌能够成功感染宿主的几步）定细

9、菌能够成功感染宿主的几步）Introduction of Bacterial Pathogenesis（细菌致病机理）（细菌致病机理）1.Infection（感染）:growth（生长）and multiplication(复制/繁殖)of a microbe in or on the body with or without the production of disease.2.The capacity of a bacterium to cause disease reflects its relative“Pathogenicity（致病性）.”3.Virulence（毒力）is th

10、e measure of the pathogenicity of a microorganism.4.Pathogenesis refers both to the mechanism of infection and to the mechanism by which disease develops.Host Susceptibility（宿主易感性）（宿主易感性）1.Susceptibility to bacterial infections=Host Defenses（宿主防御）vs Bacterial Virulence（细菌毒力）2.Host Defenses:-Barriers

11、 屏障(skin皮肤&mucus黏液)first line-Innate Immune Responses 先天性免疫应答(complement,macrophages&cytokines)the early stage-Adaptive Immune Responses 适应性免疫应答(Ag-specific B&T cells)the later stage3.Host defenses can be comprised by destructing barriers or defective immune response.e.x.Cystic Fibrosis（囊肿性纤维化）=poor

12、 ciliary（纤毛）function=NOT clear mucus efficiently from the respiratory tract（呼吸道）=Pseudomonas aeruginosa=serious respiratory distress（呼吸窘迫）.Obligate pathogens are more virulent and can cause diseases in a normal person.Opportunistic pathogens are typically members of normal flora and cause diseases w

13、hen they are introduced into unprotected sites,usually occur in people with underlying conditions.Entry into the human body:infection:sheddingThe most frequent portals of entry-Mucus -SkinRoutes:Ingestion,inhalation,trauma,needles,catheters,arthropod bite,sexual transmissionPathogen Disease 柯赫法则（Koc

14、h postulates）又称证病律，通常是用来确定侵染性病害病原物的操作程序。科赫法则(Kochs postulates)包括：1 在每一病例中都出现相同的微生物，且在健康者体内不存在；2 要从寄主分离出这样的微生物并在培养基中得到纯培养（pure culture）；3 用这种微生物的纯培养接种健康而敏感的寄主，同样的疾病会重复发生；4 从试验发病的寄主中能再度分离培养出这种微生物来。如果进行了上述4个步骤，并得到确实的证明，就可以确认该生物即为该病害的病原物。罗伯特 科赫（Robert Koch，18431910），德国医生和细菌学家，世界病原细菌学的奠基人和开拓者。1905 年，科赫以举

15、世瞩目的开拓性成绩，问心无愧地摘走了诺贝尔医学及生理学奖。Stanley Falkow,Ph.DMicrobiologist and a professor of microbiology and immunology at Stanford University School of MedicineThe father of molecular microbial pathogenesis,which is the study of how infectious microbes and host cells interact to cause disease at the molecula

16、r level.He formulated molecular Kochs postulates,which have guided the study of the microbial determinants of infectious diseases since the late 1980s.President of the ASM(1997-1998);elected member of the Institute of Medicine,the National Academy of Sciences,and the National Academy of Arts and Sci

17、ences.Fellow of AAAS,UK-FRS(Foreign member)The prestigious Lasker Award for medical research(2008)医学科学特殊贡献奖：Stanley Falkow建立了一套分辩致病微生物引起宿主疾病的基因的准则，即毒力因子（virulence factor）。Molecular Kochs postulates applied to microbial pathogenicity（Falkow S.Rev Infect Dis.1988）被研究的疾病表象或性状应该与某类或某种病原微生物相关。另外，相关的毒力基因应

18、该存在于所有致病微生物中，但不存在于非病原微生物中；毒力基因的特异性失活会导致致病微生物在动物模型中毒力/致病性的下降；将毒力基因重新导入毒力基因失活的微生物中能够恢复其在动物模型中的致病力；毒力基因必须在感染过程中进行表达；免疫力必须是保护性的 具有以上特征的基因为毒力基因 存在问题：适合的动物模型；微生物的分子操作（基因修改）Spectrum of virulencePoliomyelitis（脊髓灰质炎）（脊髓灰质炎）in a child0.1-1%of infections are clinically apparentRubella（风疹）（风疹）50%of infections a

19、re clinically apparentRabies（狂犬病）（狂犬病）100%of infections are clinically apparentThe iceberg concept of infectious diseaseasymptomatic infectionclassical clinical diseaseless severe disease1.Transmissibility（传播能力）（传播能力）2.Adherence to host cells（粘附到宿主）（粘附到宿主）3.Invasion of host cells and tissue（入侵宿主细胞和组

20、织）（入侵宿主细胞和组织）4.Evasion of the host immune system（逃避宿主免疫系（逃避宿主免疫系统）统）5.Toxigenicity（毒性）（毒性）A bacterium may cause diseases by 1.Destroying tissue(invasiveness)（侵染力）（侵染力）2.Producing toxins(toxigenicity)（毒性）（毒性）3.Stimulating overwhelming host immune responses（刺（刺激产生压倒性的宿主免疫反应）激产生压倒性的宿主免疫反应）Characteristi

21、cs of Pathogenic Bacteria（病原菌的特点）（病原菌的特点）Pathological Mechanisms of Bacterial Infections1.Bacteria-mediated Pathogenesis 细菌介导的细菌介导的致病机制（直接）致病机制（直接）2.Host-mediated Pathogenesis 宿主介导的宿主介导的致病机制（间接）致病机制（间接）3.Bacterial virulence factors（细菌毒力因子）（细菌毒力因子）=bacterial factors causing diseases（细菌因子导致（细菌因子导致疾病）疾

22、病）Adopted from Samuel Baron“Medical Microbiology”Bacterial Virulence MechanismsBacterial virulence factorsAdhesins Pili(fimbriae)菌毛 Nonfimbrial adhesins 非菌毛粘附素 Invasion of host cells Tissue damage Growth byproducts Tissue-degrading enzymes ImmunopathogenesisToxins Exotoxins(cytolytic enzymes and A-B

23、 toxins);enterotoxins;superantigens;endotoxin and other cell wall componentsAntiphagocytic factorsIntracellular survivalAntigenic heterogeneity Antigenic variation Phase variationIron acquisition Siderophores Receptors for iron-containing moleculesResistance to antibioticsEncapsulation（形成荚膜）（形成荚膜）(I

24、nhibition of phagocytosis and serum bactericidal effect)Antigenic mimicry（抗原模拟）（抗原模拟）Antigenic masking（抗原屏蔽）（抗原屏蔽）Antigenic or phase variation（抗原变异）（抗原变异）Intracellular multiplication（胞内复制）（胞内复制）Escape phagosome（吞噬体逃脱）（吞噬体逃脱）Inhibition of phagolysosome fusion（抑制吞噬溶酶体融合）（抑制吞噬溶酶体融合）Resistance to lysoso

25、mal enzymes（溶酶抗性）（溶酶抗性）Production of anti-immunoglobulin protease（产生抗免疫球蛋白（产生抗免疫球蛋白酶）酶）Inhibition of chemotaxis（抑制趋化作用）（抑制趋化作用）Destruction of phagocytes（破坏吞噬细胞）（破坏吞噬细胞）Microbial defenses against host immunologic clearance（微生物抵御宿主免疫清除）（微生物抵御宿主免疫清除）Mechanisms for escaping phagocytic clearance and intr

26、acellular survivalMechanisms for escaping phagocytic clearance and intracellular survivalMechanisms for escaping phagocytic clearance and intracellular survivalSteps in successful infection Sex comes before disease acquire virulence genes（基因交换）Sense environment（感知环境）and Switch virulence genes on and

27、 off（基因开关）Swim（游动）to site of infection Stick（粘附）to site of infection Scavenge nutrients（攫取营养）especially iron Survive stress（生存压力）Stealth（隐形）avoid immune system Strike-back（反击）damage host tissues Subvert（颠覆）host cell cytoskeletal and signalling pathways Spread（扩散）through cells and organs Scatter（散播）B

28、acterial Sex acquiring virulence genes Bacteria have three ways of exchanging DNA Transformation cells take up naked DNA Transduction phages carry DNA Conjugation cells mate through specialised appendagesBacterial Sex Mobile genetic elements Transposons ST enterotoxin genes Virulence Plasmids e.g.TT

29、SSs in Shigella,Yersinia;toxins in Salmonella,E.coli,anthrax Phage-encoded virulence e.g.botulinum toxins,diphtheria toxin,shiga-like toxin(linked to lysis),staphylococcal toxins,TTSS substrates in Salmonella.Bacterial Sex Pathogenicity Islands Concept originated from study of uropathogenic E.coli s

30、trains Defining Features Carriage of(many)virulence genes Presence in pathogenic versus non-pathogenic strains Different G+C content from host chromosome Occupy large chromosomal regions(10-100 Kb)Compact distinct genetic units,often flanked by DRs,tRNAs,ISs Presence of(cryptic)mobility genes Unstab

31、le,prone to deletionBacterial Sex Pathogenicity Islands often encode secretion systems LEE region in EPEC Spi1,Spi2 in Salmonella Cag in H.pylori can also encode adhesins,siderophores,toxins Uropathogenic E.coli(Pai I,II,IV,V)Yersinia spp.(HPI)V.cholerae(VPI or TCP-ACF element)Sense environment Bact

32、eria can sense changes in environment e.g.in temperature,nutrient availability,osmolarity,cell density(“quorum sensing”).In simplest cases,change in intracellular concentration of ion linked directly to gene expression e.g.fall in intra-cellular iron levels triggers de-repression of diphtheria toxin

33、 gene In more complex cases,sophisticated signal transduction cascades allow bacteria to regulate gene expression in response to environmental cues the pathogen as an information processorSwitch virulence factors on and offA multi-layered hierarchy Changes in DNA sequence Gene amplification Genetic

34、rearrangements e.g.Hin flip-flop control of flagellar phase variation Transcriptional Regulation Activators and Repressors(helix-turn-helix motif)mRNA folding and stability Translational Regulation Post-translational Regulation Stability of protein,controlled cleavage Covalent modifications e.g.phos

35、phorylation in two-component sensor-regulator systemsSwim Many bacterial pathogens are motile Enterics,Campylobacter,Helicobacter,spirochaetes Motility crucial for virulence in some cases Usual organelle of motility=flagellum Variants Twitching motility SwarmingStick To avoid physical and immunologi

36、cal removal,bacteria must adhere to cell surfaces and extracellular matrixe.g.in respiratory,gastrointestinal and genitourinary tracts solid surfacese.g.teeth,heart valves,prosthetic material other bacteria Direct interaction Molecular bridging via e.g.fibronectin Adherence often combined with manip

37、ulation of host cell signalling and cytoskeleton Invasion Intimate adherenceStick Common adherence mechanisms（共同机制）Capsules and slime Biofilm formation Gram-positive adhesins（革兰氏阳性粘附素）MSCRAMMs(microbial surface components recognizing adhesive matrix molecules),e.g.protein A Fimbriae Gram-negative ad

38、hesins(CHO and protein receptors)（革兰氏阴性粘附素）Fimbriae,Afimbrial adhesins(FHA,Pertactin etc.)Outer Membrane Proteins Types III-IV secretionStickScavenge nutrientse.g.ironFree iron levels very low in body fluids（游离铁在体液中含量非常低）Acute phase response causes further drop Iron overload increases susceptibility

39、 to infectionMany different bacterial systems for scavenging iron（许多细菌系统针对游离铁离子）Siderophores chelate available iron&transport it into bacteria Iron can be scavenged direct from host iron-binding proteins,e.g by lactoferrin-binding proteins Often co-ordinately regulated e.g.by fur locus in E.coli Som

40、e pathogens avoid the problem by cutting out need for iron,e.g.Treponema pallidumIron used to regulate aggressive virulence factors（铁离子可以调控毒力基因表达）Diphtheria toxin(DtxR repressor)Shiga-like toxin Pseudomonas aeruginosa exotoxin ASurvive Stress In addition to nutrient-limitation stress,pathogens face

41、many other stresses Acid stress within stomach Heat shock during fever Oxidative stress within phagocytes Stress response proteins,such as chaperonins feature as immunodominant antigens Detoxification proteins play a role in virulence,e.g.periplasmic Cu,Zn-superoxide dismutases Infectious dose for e

42、nteric pathogens much lower in achlorhydria(no need to overcome acid stress)Stealthavoid immune system IgA proteases metalloproteases active against IgA Immunoglobulin-binding proteins e.g.protein A of S.aureus Resist complement,opsonisation Capsule(usually polysaccharide)Lipopolysaccharide Surface

43、proteins and OMPs Antigenic mimicry e.g.sialic acid capsule of group B meningococcusStealthavoid immune system Antigenic or phase variation Involves surface structures such as proteins,LPS,capsules Variety of mechanisms slip-strand mispairing flip-flop cassettes Adopt cryptic niche inside phagocytes

44、 in biofilm67700 67710 67720GAAGTGCATTTAACTT*GGGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGGTAATGAAGTGCATTTAACTTGGGGGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGTAATGAAGTGCATTTAACTT

45、*GGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGTAATGAAGTGCATTTAACTT*GGGGGGGGGGGGTAATHomopolymeric tract in Campylobacter jejuniStrike-back Damage host tissues Endotoxin Exotoxins Toxins acting on cell membranes Toxins active inside cells Supe

46、rantigensEndotoxin of Gram-negativescytopl.mem.peptidoglycanouter mem.Gram-negative cellLipid ACore polysaccharideO sidechainThe toxic partHelps solubilise Lipid ASomatic antigenLipopolysaccharideLipopolysaccharide(LPS)(LPS)Strike-back Endotoxin Actions of Endotoxin Pyrogenicity（致热原性）Leucopenia then

47、 leucocytosis（白细胞减少）Hypotension（低血压）“Gram-negative Shock”Life-threatening complication of septicaemia（败血症）e.g.in meningococcal infection,in ITU or oncology patients Endotoxic shock seen with dirty intravenous equipment Most of the effects of endotoxin are mediated by tumour necrosis factor（肿瘤坏死因子）At

48、tempts at therapy using anti-endotoxin or anti-TNF antibodiesStrike-back Membrane-Damaging Exotoxins Many bacterial toxins form pores in eukaryotic cell membranes,producing oligomeric rings,e.g.streptolysin O of Streptococcus pyogenes listeriolysin of Listeria monocytogenes alpha-toxin of S.aureusOt

49、her toxins,such as phospholipases,degrade components of the membrane e.g.Clostridium perfringens alpha toxinStrike-back Toxins active inside cells Toxins often consist of translocation and binding B subunit that delivers the active A subunit into the host cell cytoplasm Example of AB toxin:diphtheri

50、a toxinan ADP-ribosyltransferaseAB5 ToxinsSubvert inject proteins into host cells to subvert the cytoskeleton and signal-transduction pathways:manipulating e.g.Rho GTPases and the cytoskeleton to induce membrane ruffling and bacterial invasion preventing uptake by phagocytic cells,e.g.Yersinia spp.a