4细菌的感染,诊断,防治,消毒,灭菌

《4细菌的感染,诊断,防治,消毒,灭菌》由会员分享，可在线阅读，更多相关《4细菌的感染,诊断,防治,消毒,灭菌（92页珍藏版）》请在装配图网上搜索。

1、Bacterial InfectionNormal flora Opportunistic pathogenPathogenic bacteria(virulent bacteria)DysbacteriosisExotoxins EndotoxinsSeparation&Culture of bacteriaBacteriemiaSepticemiaPyemiaToxemiaEndotoxemiaCarrier Colony Toxoids AntitoxinNot all bacteria cause disease Normal flora The normal flora are ba

2、cteria which are found in or on our bodies on a semi-permanent basis without causing disease.There are more bacteria living in or on our bodies,than we have cells of our own.A human body contains around 1013 cells.The human body is home to around 1014 bacteria.One fourth of fecal weight is made of b

3、acteria!The normal flora are particularly important in the large intestine.Normal flora are also found in the nose,mouth,throat,skin.Opportunistic bacteria can be part of the normal flora but not pathogenic for normal persons,cause disease only in immunocompromised persons,or under specific conditio

4、ns.Pathogenic bacteria(virulent bacteria)Normally not members of the normal flora,have mechanisms to promote their own growth in the expense of hosts tissue or organs functionThe normal flora protect us from disease by:1.Competing with invaders for space and nutrients.2.Producing compounds(bacterioc

5、ins)which kill other bacteria.3.Lowering the pH so that other bacteria cant grow.In addition to the above ways of protecting us from disease our normal flora help us in other ways.Of course,there is a down side as well.The Good Side Produce vitamins we are unable to produce such as vitamin B12.Boost

6、 our immune system Germ free animals(born and raised in a germ free plastic tent)are very susceptible to disease when removed from the germ free environment because their immune system is underdeveloped.Help digest food.The Bad Side If the normal flora escape from their normal location,they can caus

7、e disease.For example,Escherichia coli,commonly found in the intestine,can cause urinary tract infections if introduced into the bladder.Immunosuppression can allow otherwise harmless bacteria to cause disease.AIDS,some cancer treatments and transplant rejection drugs all suppress the immune system

8、and allow the normal flora to cause occasionally serious disease.Three conditions correlate with opportunistic infection:1.Immune System Compromise2.Dysbacteriosis3.Change of Inhabiting Places:Pathogenesis is a multi-factorial process which depends on the nature of the species or strain(virulence fa

9、ctors),the Pathway of Bacterial Entrance(gateway)and the immune status of the host,as well as the number of organisms in the initial exposure and the Environmental conditions.Bacterial Pathogenesis1.Adherence Factors Surface hydrophobicity,pili,Capsule&glycocalyx Bacterial Biofilms 2.Invasion of Hos

10、t Cells&Tissues:Enzymes3.immune evasion:Capsule&glycocalyx,IgA1 Proteases,Antigenic variation,Interference of complement activity Intracellular pathogenicity.Bacterial Virulence Factors4.Toxins Exotoxins EndotoxinsExotoxins l Cholera toxin and E.coli labile toxinADP-ribosylation of regulator adenyla

11、te cyclase activation cyclic AMP active ion and water secretion diarrhea Normal condition(Acetylcholine)15 neutralizevaccinationEndotoxinsn Lipopolysaccharide(LPS):a toxic lipid A a core polysaccharide O antigen polysaccharide side chains n Cell wall lysis required n Formaldehyde and heat resistant

12、n Poor antigen as free molecule n Endotoxin effects Endotoxin effects l Fever-pyrogen 1 microgram/kg l Leukopenia and leukocytosis necrosis l Endotoxemia and shock l Disseminated intravascular coagulation(DIC).Exotoxins Toxins produced by bacteria and secreted to the outside of the bacteria cell(dif

13、ferent from the endotoxin)acting on cell surface By binding to certain receptors Possess some degree of host cell specificity denatured by formaldehyde treatment to generate toxoids which lack toxic activity but still induce protective immunity when used as vaccines.Many exotoxins have an A-B subuni

14、t structure A subunit-provides the toxic activity B subunit-mediates adherence of the toxin complex to a host cell SummaryEndotoxins(LPS)n derived from Gcell walls and are often liberated when the bacteria lyse.n heat-stable,n three main regions l a toxic lipid A l a core polysaccharide l O antigen

15、polysaccharide side chains n The pathophysiologic effects of LPS are similar.Pathway of Bacterial EntrancenContact:direct or indirect(Sexual contact)nInhalation:transmitted by the respiratory route(Airborne droplets)nIngestion:ingested and transmitted through intestinal tract to the outside and get

16、new infections by contaminating of food and water,which is called fecal-oral spread()nInoculation&blood transfusionnAnimal vectors:transmitted by animal vectors()Response to InfectioninfectionxdiseaseInnate immunityno diseaserecoveryadaptive immunityre-infectionno diseasex Immunity of extracellular

17、bacterial infection:antibodies(IgG,IgM,SIgA);phagocytes(neutrophils);complement;humoral immunity mainly.Immunity of intracellular bacterial infection:cell-mediated immunity(delayed-type hypersensitivity,DTH response(DTH)involving TH1and macrophages)mainly.No diseaseHost defenseBacterial infectionBal

18、ance between Infection and Immunity(immunity)Disease develops only in the right host and under the right conditionsDiseaseHostPathogenEnvironmentDiseaseHost defenseBacterial infectionEnvironmental conditions can help tilt the balance Environmental signals often control the expression of the virulenc

19、e genes.Common signals,include:Temperrature/Iron availability:C diphtheriae/low ion/Osmolality/Growth phase/pH/Specific ionsSources of InfectionB.Endogenous Infection:opportunistic pathogen infections A.Exogenous Infection PatientsCarriersAnimalsTypes of Bacterial InfectionAccording to infectious st

20、ate:nInapparent infection:without clear clinic symptoms.nApparent Infection:have evident clinic symptoms.lAccording to infectious sites(Local infection and Generalized or systemic infectionlBacteriemia:bacteria circulate but not multiply in the blood.lSepticemia:Bacteria circulate and multiply in th

21、e blood.lPyemia:produce septicemia with multiple abscesses in internal organs.lToxemia:Bacteia multiply at invading location and do not enter blood stream,but the exotoxins enter blood and cause corresponding toxic symptoms lEndotoxemia:multiply at location or in blood stream,release a lot of amount

22、 endotoxin released from bacterial cell rupture n Carrier:Principles of Diagnosis and Prevention of Bacterial InfectionnManifestations of Infection:Signs and symptoms vary according to the site and severity of infection.Diagnosis requires a composite of information,including history,physical examina

23、tion,radiographic findings,and laboratory data.nMicrobial Causes of Infection:Infections may be caused by bacteria,viruses,fungi,and parasites.The pathogen may be exogenous(acquired from environmental or animal sources or from other persons)or endogenous(from the normal flora).1.The quantity materia

24、l must be adequate.2.Collect from appropriate site.3.Prepare site to minimize contamination.4.Whenever possible,collect specimens prior to antibiotics.5.Transport system maximized for pathogen survival should be used.General guidelines for specimen collectionBody fluid for specimen collectionnBlood

25、septicemia nCerebrospinal fluid bacterial meningitisnPeritoneal(abdominal)nPleural(chest)nSynovial(joint)nPericardial(heart)nUrine Sample collection devicesMicroscopy and Stains:cellular morphology and stain may permit preliminary identification.Separation&Culture of bacteria:Isolation of infectious

26、 agents frequently requires specialized media.Nonselective(noninhibitory)media permit the growth of many microorganisms.Selective media contain inhibitory substances that permit the isolation of specific types of microorganisms.Colony morphology can sometimes be useful in bacterial identificationIde

27、ntification of bacteria:qCultural characteristics:Growth characteristics under various conditions,lBiochemical characterization:utilization of carbohydrates and other substrates,enzymatic activitylSerology Identification A high or rising titer of specific IgG antibodies or the presence of specific I

28、gM antibodies may suggest or confirm a diagnosis.lGenomic MethodsAntimicrobial Susceptibility:Microorganisms,particularly bacteria,are tested in vitro to determine whether they are susceptible to antimicrobial agents.Microbiologic ExaminationMicroscopy and Stains Direct examination of stained or uns

29、tained preparations by light(bright field)microscopy is a relatively simple:l with a 100 oil immersion objective,l a 5 to 10 eyepiece,l optimal lighting.The two most important methods,the Gram and acid-fast techniques,to classify as well as stain the organism.l employ staining,l decolorization,l cou

30、nterstaining Examples of Gram StainsGram Positive Rods and CocciGram Negative Rods and CocciExamples of Acid-Fast StainsFully Acid-Fast Rods(Mycobacterium species)Partially Acid-Fast Rods(Nocardia species)types of media for routine culture:The standard medium for specimens:blood agar,-Usually made w

31、ith 5%sheep blood.-Most aerobic and facultatively anaerobic organisms will grow on blood agar A second necessary medium:Chocolate agar,-a medium containing heated blood with or without supplements,-Some organisms that do not grow on blood agar,including pathogenic neisseria and haemophilus,will grow

32、 on chocolate agar.CultureSelective medianSelective media is one that grows only certain microorganisms while inhibiting(or preventing)others from growing,that is to say,the media has certain chemicals that allow one organism to grow but another organism cannot grow with those ingredients in the med

33、ia,thus,is selective for the organisms that can grow in that media.Therefore,most commonly grows only one type of organism.Differential medianDifferential media is one that distinguishes one microorganism from another,it can grow more than one microorganism,but depending on how each organism reacts

34、to the media(like turns red)it differentiates from another microorganism.In this type of media if you are trying to distinguish between 2 types of microorganisms,both should grow but they will have different reactions to the media,and thru their visible reactions you can tell them apart(differentiat

35、e them).qGrowth on selective-differential media,such as Salmonella-Shigella(SS)medium,eosin-methylene-blue(EMB)and MacConkey agarTThe select effect of the media in suppressing unwanted gram-positive organisms is exerted by bile salts or bacteriostatic dyes in the agar.TThe differential ability of th

36、ese media is based on lactose fermentation:normal flora positive(colored colonies)and pathogens negative(colorlesscolonies).Separation of bacteria:plate streaking.Bacteriologic plate streaking.Colony:the visible growth of bacteria on solid growth media.Ideally,the colony is the progeny of one,or at

37、most,a few bacteria.A colony will usually contain millions of bacterial cells.Colony morphology can sometimes be useful in bacterial identification.Colonies are described as to such properties as size,shape,texture,elevation,pigmentation,effect on growth medium.ColonyGrowth on blood agar to test for

38、 hemolytic properties-hemolytic:incomplete lysis of red blood cells,resulting in a greenish halo around the colony-hemolytic:complete lysis of red blood cells,resulting in a clear halo around the colony-hemolytic:non-hemolytic1.Cultural characteristics:lunique nutritional requirements,lpigment produ

39、ction,lhemolytic properties Identification of BacteriaMacConkeys AgarContains lactose and a pH indicator,E.coli ferment lactose to produce acid,which turns the pH dye red.So,E.coli colonies appear red.2.Biochemical characterizationIdentification of Bacterial the ability to attack various substrates

40、l or to produce particular metabolic products 2.Biochemical characterizationTriple Sugar Iron AgarIdentification of BacteriaA simple approach to rapid diagnosis(as an example of antigen detection)is used in many doctors offices for the group A streptococcus.The patients throat is swabbed and strepto

41、coccal antigen extracted directly from the swab(without prior bacteriological culture).The bacterial antigen is detected by aggregation(agglutination)of antibody coated latex beads.Serologic identification of an antibody response(in patients serum)to the infecting agent can only be successful severa

42、l weeks after an infection has occurred.3.3.Serology Identificationuse of antibodies of known specificity to detect antigens present on whole bacteria or free in bacterial extracts Identification of BacteriaSerological methods used to detect both antigen and antibody in specimensnThe fastest and mos

43、t specific wayqImmunofluorescence microscopy,FACSqEnzyme-immunoassay ELISA,Western blotqRadioimmunoassay quantitate antigen-antibody complexFACS(fluorescence activated cell sorter)4.Genomic Methodsn16S DNA sequencingqLabeled probes specific for the 16S rRNA of a species are added,and the amount of l

44、abel on the double-stranded hybrid is measured.q This technique is widely used for the rapid identification of many organisms.nFluorescence in situ hybridization(FISH)nPolymerase Chain Reaction(PCR)nNucleic Acid Sequence Analysis nChecker board DNA hybridization(DNA microarray)Identification of Bact

45、eriaELISA(Enzyme-Linked Immunosorbent Assay)E.coli 16S RNAl The 16S rRNA of each species of bacteria has stable(conserved)portions of the sequence.l Many copies are present in each organism.FISH PCR(polymerase chain reaction)BacteriamRNAcDNADNA microarrays37oC cDNA labeledw/fluorescein tag25oC cDNA

46、labeledw/rhodamine tagDNA array 6000 genes“Transcriptome”SummarynDirect microscopyqFastqGive some hints on the type of bacteriaqLow sensitivitynCultureqHigh sensitivityqCan make definitive IDqSlowqOnly works on culturable bacterianSerological assayqFastqHigh specificityqCan detect both antigen and a

47、ntibodyqEasy(can be used at chairside or bedside)qLow sensitivitynGenomic based assayqFastqHigh sensitivity and specificityqWorks on both culturable and non-culturable bacteriaqEspecially useful for detecting slow growing bacteria such as T.b.qRequires technical expertiseqFalse positive or negativeP

48、revention of Bacterial InfectionArtificial active immunitynToxoids:a modified form of the toxin that preserves its antigenicity but has lost its toxicity.This has been spectacularly successful with tetanus and diphtheria.nInactivated vaccines:nAttenuated live vaccines:nSpecial vaccines:polysaccharid

49、e vaccine,subunit vaccine,(conjugate vaccine,bio-engineered vaccine,chemical vaccine,synthetic vaccine),nucleic acid vaccine,idiotype vaccine,autovaccine,etc.Artificial active immunityArtificial passive immunity nAntitoxin:e.g.Tetanus antitoxin and diphtheria antitoxin.It is raised in the horse.It i

50、s most important to give an intented recipient of equine serum a prior test dose to exclude hypersensitivity subjects who may have been sensitized by a previous dose of equine serum.nPooled immunoglobulin:It contains the normal repertoire of antibodies for an adult,and can protect against hepatitis

51、A,and measles.nSpecific immunoglobulin:Preparations of specific immunoglobulin are available for passive immunization against tetanus,hepatitis B,rabies,varicella-zoster.nCytokine Sterilization and DisinfectionA.SterilizationA physical or chemical process that completely destroys or removes all micr

52、obial life,including spores.B.Disinfection treatment to destroy harmful microorganisms.C.Preservation The prevention of multiplication of microorganisms in formulated products,including pharmaceuticals and foods.Sterilization and Disinfection BactericidalA specific term referring to the property by

53、which a biocide is able to kill bacteria.Bactericidal action differs from bacteriostasis only in being irreversible;ie,the“killed”organism can no longer reproduce,even after being removed from contact with the agent.BacteriostaticA specific term referring to the property by which a biocide is able t

54、o inhibit bacterial multiplication;multiplication resumes upon removal of the agent.DisinfectionnPhysical MethodsqHeatqRadiation qFiltrationqUltrasonic and SonicqFreezingSterilization and Disinfection.HeatnHeat energy can be applied in three ways,in the forms of qMoist heat(either boiling or autocla

55、ving)qDry heatqPasteurization Moist Heat:far more rapid and effective in sterilization than dry heat,n A temperature of 100C will kill all but spore forms of bacteria within 23 minutes in laboratory-scale cultures;121C for 15 minutes is utilized to kill spores.n Autoclaves or pressure cookers are us

56、ed for this purpose.n Autoclaves are usually operated at 121C,which is achieved with a pressure of 15 lb/sqn Its use：any materials that are not damaged by heat and moisture,l heat-stable liquids l swabs,l most instruments,l culture media,l rubber gloves.HeatDry Heat:nrequired:2 hours at 180nIts use:

57、lsterilization of glassware,lincineration of disposable objects lpassage of bacteriologic needles,coverslips,or small instruments through the name of a Bunsen burner.n The most widely used type of dry heat:is the hot-air oven.Heat.Heat Pasteurization,which is used primarily for milk,consists of heat

58、ing the milk to 62 for 30minutes followed by rapid cooling.(“Flash”pasteurization at 72 for 15 seconds is often used).This is sufficient to kill the vegetative cells of the milk-borne pathogens,eg,M bovis,Salmonella,Streptococcus,Listeria,and Brucella,but not to sterilize the milk.Used for foods(mil

59、k,fruit Drinks,wine)and fragile medical equipment,to kill all vegetative bacteria of significance in human disease，but not spores.RadiationnThe two types of radiation used to kill microorganisms areqUltraviolet(UV)light:The most effective bactericidal wavelength is in the 240 to 280 nm range,with th

60、e optimum at about 260nm,q Ionizing Radiations:have higher energy and penetrating power than UV radiation Ultraviolet Radiation(UV)Mechanism:UV light causes direct damage to DNA,formation of the pyrimidine dimers,As a result,DNA replication is inhibited and the organisms cannot grow.Use of UV light

61、in medicine is limited l its poor ability to penetratel safety,because UV radiation can damage the cornea and skin,So,It must be remembered that workers exposed to it must be appropriately protected.its main application:l air of hospital sites l laboratory facilities.RadiationIonizing radiation Mech

62、anism:carries far greater energy than UV light.It,too,causes directdamage to DNA and produces toxic free radicals and hydrogen peroxide from water within the microbial cells.widely used in industrial processes,including sterilization ofl many disposable surgical supplies such as gloves,plastic syrin

63、ges,l specimen containers,l some foodstuffs,.Radiation.FiltrationThe most commonly used filter is composed nitrocellulose and has a pore size of 0.22m.This size will retain all bacteria and spores.Filtration is the preferred method of sterilizing certain solutions,especially those containing heat-la

64、bile components such as serum,plasma,or trypsin.Ultrasonic and Sonic Vibrations Microorganisms vary markedly in their sensitivity to sonic and ultrasonic vibrations.(The most susceptible are the gram-negative rods,and among the most resistant are the staphylococci.)Although sonic vibrations are leth

65、al to many members of the exposed bacterial population,there are numerous survivors.treatment with sonic vibration is of no practical value in sterilization and disinfection.Freezing Although many bacteria are killed by exposure to cold,freezing is not a reliable method of sterilization.This method

66、is widely used for the preservation of bacterial cultures.nChemical Methods:Chemical agents act primarily by one of three mechanismsnDisruption of the lipid-containing cell membranenModification of proteinnModification of DNAnDisruption of the lipid-containing cell membraneqAlcohols:It acts mainly by disorganizing the lipid structure in membranes,but it denatures proteins as well.qCationic detergents,particularly the quaternary ammonium compounds(“quats”)such as benzalkonium chloride,their hydro