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1、消毒与灭菌消毒与灭菌本课论题：本课论题：理化因素多理化因素多 可保菌可保菌 抑菌抑菌 灭菌菌生物因素生物因素杂 能抗菌能抗菌 杀菌菌 吃菌吃菌物理消毒物理消毒灭菌法菌法生物因素生物因素对微生物微生物的影响的影响化学消毒化学消毒灭菌法菌法微生物的微生物的亚致死性致死性损伤及其修复及其修复基本概念基本概念消毒消毒 Disinfection灭菌灭菌 Sterilization杀菌作用杀菌作用 Bacteriocidic action抑菌作用抑菌作用 Bacteriostatic action抗菌作用抗菌作用 Antibiotic action防腐防腐 Antisepsis无菌无菌 Asepsis灭菌
2、法灭菌法 Antiseptic technique影影响响消消毒毒灭灭菌菌效效果果的的因因素素处理剂量：处理剂量：消毒剂的性质、浓度和作用时间消毒剂的性质、浓度和作用时间微生物的种类与数量微生物的种类与数量温度：温度：NaOH 15 过氧乙酸、环氧过氧乙酸、环氧 乙烷、戊二醛、甲醛、洗必泰、来沙尔乙烷、戊二醛、甲醛、洗必泰、来沙尔 药物颉抗：药物颉抗：卵磷脂、肥皂、阴离子去污剂能中和季铵盐卵磷脂、肥皂、阴离子去污剂能中和季铵盐类类；硫代硫酸钠可中和氯和氯化物硫代硫酸钠可中和氯和氯化物其他因素：其他因素：湿度、穿透力、表面张力湿度、穿透力、表面张力等等消毒与灭菌方法消毒与灭菌方法消消毒毒与与灭灭
3、菌菌方方法法物理法物理法化学法化学法生物学法生物学法机械除菌类机械除菌类弱杀伤类弱杀伤类擦洗擦洗、沉淀沉淀、过滤过滤强杀伤类强杀伤类低温低温（寒冷、冰冻）、（寒冷、冰冻）、干燥、渗透压干燥、渗透压、可见光可见光热力热力、紫外线紫外线、电离辐射电离辐射、超声波超声波凝固菌体蛋白凝固菌体蛋白损伤胞浆膜损伤胞浆膜干扰酶系统干扰酶系统变更核酸功能变更核酸功能酚类、醇类、重金属盐类、酸碱类、醛类酚类、醇类、重金属盐类、酸碱类、醛类酚类、醇类、表面活性剂酚类、醇类、表面活性剂氧化剂、重金属盐类氧化剂、重金属盐类染料、烷化剂染料、烷化剂抗（杀）菌物质抗（杀）菌物质活体影响活体影响抗生素、植物杀菌素、细菌素、
4、抗菌肽抗生素、植物杀菌素、细菌素、抗菌肽噬菌体、蛭弧菌噬菌体、蛭弧菌微生物的亚致死性损伤及其修复微生物的亚致死性损伤及其修复细菌损伤和修复的特征细菌损伤和修复的特征细菌的损伤和修复细菌的损伤和修复细菌芽孢的损伤和修复细菌芽孢的损伤和修复消毒：杀死或清除停留在体外传播因素上的活的病原消毒：杀死或清除停留在体外传播因素上的活的病原体，目的是切断传播途径，借以限制或歼灭传染病。体，目的是切断传播途径，借以限制或歼灭传染病。病原体病原体传播途径猪猪随时消毒随时消毒接触接触工具工具空气空气载体载体体质体质非特异抗拒力非特异抗拒力特异性免疫特异性免疫预防性消毒预防性消毒疫疫源源地地消消毒毒终末消毒终末消毒
5、消毒消毒灭菌：杀灭体外环境中某些物体上，包括细菌芽孢在灭菌：杀灭体外环境中某些物体上，包括细菌芽孢在内的一切病原体和非病原体。内的一切病原体和非病原体。应用：微生物学试验、医疗手术器械和敷料、宇宙飞应用：微生物学试验、医疗手术器械和敷料、宇宙飞船的处理、制药和食品工业等船的处理、制药和食品工业等擦洗擦洗Scrubbing沉淀沉淀SedimentationFiltration of a mixture of bacteria and viruses.If a mixture of viruses and bacteria are filtered through a bacterial-proo
6、f filter(red),the viruses will pass through into the filtrate in the flask.Filtered beer is produced by a similar process.过滤过滤Filtration超净工作台、无菌隔离器、无超净工作台、无菌隔离器、无菌操作室、试验动物室以及疫菌操作室、试验动物室以及疫苗、药品、食品等生产中干净苗、药品、食品等生产中干净厂房的空气过滤除菌厂房的空气过滤除菌液体培育基和药物除菌：液体培育基和药物除菌：糖糖培育液、特殊培育基、血清、培育液、特殊培育基、血清、毒素、抗毒素、抗生素、维生毒素、抗毒
7、素、抗生素、维生素、氨基酸等不能加热灭菌的素、氨基酸等不能加热灭菌的液体常用滤器过滤除菌液体常用滤器过滤除菌Chamberland滤菌器滤菌器未上釉的陶瓷制成的空未上釉的陶瓷制成的空心圆柱体，心圆柱体，L1、L1bis、L2、L3、L5、L7、L9、L11、L13九级。BerkefeldBerkefeld滤菌器，硅藻土加压制成的空心滤菌器，硅藻土加压制成的空心圆柱体圆柱体British Berkefeld Emergency Water Filter Independently tested to remove the following parasites and pathogenic ba
8、cteria:E.Coli 99.99%Klebsiella 99.999%Cholera 99.99%Shigella 99.99%Salmonella 99.999%Giardia Lambia 100%Live Cryptosporidium*100%Seitz 滤菌器滤菌器 金属制成金属制成玻璃滤菌器玻璃滤菌器薄膜滤菌器薄膜滤菌器Sterlitech Membrane Filters低温低温美国强生过氧化氢等离子低美国强生过氧化氢等离子低温灭菌系统温灭菌系统 STERRAD NX STERRAD NX低温低温冷冻保存冷冻保存冷冻真空干燥（冻干）冷冻真空干燥（冻干）保保菌菌STERRAD
9、 100S过氧化氢气过氧化氢气浆低温灭菌系统浆低温灭菌系统RefrigerationFreezingDesiccationFreeze drying(lyophilization)Freezing-Freezing is employed to more completely stave off organismal deterioration-For organisms that survive the freezing process,freezing constitutes a reasonably good long-term preservation method with prev
10、ention of deterioration increasing as temperatures are lowered-Prevention of damage during freezing is accomplished by mixing cultures with glycerol-Lower temperatures result in greater long-term storage:-20C is the temperature of a standard freezer-80C is a good temperature for long-term storage of
11、 microorganisms-180C is the temperature of liquid nitrogen,a common medium for long-term storage of microorganisms 干燥和渗透压干燥和渗透压干燥和渗透压干燥和渗透压吸取菌体水分，造成生理干燥，使吸取菌体水分，造成生理干燥，使细菌的生命活动停止。细菌的生命活动停止。干燥干燥不能作为有效的灭菌方法，只能用于保存食不能作为有效的灭菌方法，只能用于保存食品、饲料、谷类、皮张、药材等。品、饲料、谷类、皮张、药材等。高温高温高温高温间歇灭菌法间歇灭菌法湿热灭菌法湿热灭菌法干热灭菌法干热灭菌法火
12、焰灭菌法火焰灭菌法热空气灭菌法（干烤）热空气灭菌法（干烤）灼烧灼烧和和焚烧焚烧煮沸灭菌煮沸灭菌巴氏消毒法巴氏消毒法流通蒸汽灭菌法流通蒸汽灭菌法高压蒸汽灭菌法高压蒸汽灭菌法灭灭菌菌低温维持：低温维持：63-65，30min.30min.高温瞬时：高温瞬时：71-7271-72，15s15s超高温：超高温：132132，1-2s1-2s Dry heat-To effect sterilization dry heat typically requires higher temperatures than moist heat-It also is less penetrating and req
13、uires longer exposure-Nevertheless,application of dry heat is cheap and easy-Typically one bakes materials in an oven at 171C for at least one hour160C for at least two hours121C for at least 16 hours-Note that the length of exposure necessary is inversely proportional to the temperature(lower temps
14、 require longer exposures)-Note additionally that the times given are for once the indicated temperature has been achieved;this is true for all methods of disinfection and sterilization:the clock does not begin until the material has been exposed to the antimicrobial-Incineration is another common m
15、ethod of dry heat sterilization,e.g.,such as the flame incineration of an inoculating loop Moist heat-Moist heat is more effective than dry heat at a given temperature or length of exposure-Moist heat is also more penetrating than dry heat-However,to achieve sterilization employing moist heat requir
16、es rather elaborate equipment,i.e.,the employment of an autoclave-Just as with dry heat,temperature and length of exposure are inversely related and penetration is absolutely critical-With moist heat,penetration is even more of a problem since penetration of heat without associated penetration of mo
17、isture is simply dry heat and dry heat is less effective as an antimicrobial than moist heat;consequently,things that block moisture penetration can inhibit sterilizing efficacy-Boiling water is the most common form of application of moist heat but is not capable of killing endospores or killing all
18、 viruses Autoclave-An autoclave is a high pressure device used to allow the application of moist heat above the normal-atmosphere boiling point of water-Exposure to 121C for 15+minutes is typically sufficient to sterilize-Again,the material must be 121C before the clock starts-Large items,large volu
19、mes,and items that are poorly penetrated by steam may take much longer than 15 minutes to sterilize Strong visible light-Strong visible light can negatively affect bacterial viability so excessive exposure to strong visible light should be avoided when ones goal is culture preservation-Practical app
20、lication of strong visible light to disinfection involves the use of sun light to dry laundry(i.e.,that“old-fashioned”alternative to a cloths dryer)Ultraviolet(UV)radiation-UV light is not terribly penetrating but is good for disinfecting surfaces and air-This is accomplished by a long-term exposure
21、 to UV light-On the down side,UV light is damaging to human tissue as well as to many plastics 辐射辐射赫兹电波赫兹电波红外线红外线可见光线可见光线紫外线紫外线X X射线射线宇宙线宇宙线射线射线射线射线射线射线高能质子高能质子阳光阳光电离电离辐射辐射中子中子粒粒子子辐辐射射电电磁磁波波辐辐射射辐辐射射Ionizing radiation-Ionizing radiation is radiation that ionizes water;this temporarily turns water int
22、o an oxidizing agent-Different types of ionizing radiation display different degrees of penetration-One potential application of ionizing radiation is as an antimicrobial is in food preservation 超声波和微波超声波和微波 超声波：频率在超声波：频率在2 2万万-20-20万万hzhz的声波称为超声波。细的声波称为超声波。细菌和酵母菌在超声波作用下于几特殊钟内死亡。小菌和酵母菌在超声波作用下于几特殊钟内死
23、亡。小型病毒、芽抱对超声波有抗拒力。主要用于裂解细型病毒、芽抱对超声波有抗拒力。主要用于裂解细胞，提取细胞组分等。胞，提取细胞组分等。微波：从几百兆赫至几十万兆赫频率的无线电波称微波：从几百兆赫至几十万兆赫频率的无线电波称为微波。微波灭菌，主要是利用微波的加热作用完为微波。微波灭菌，主要是利用微波的加热作用完成（交变电场，分子旋转振动、分子相互作用产生成（交变电场，分子旋转振动、分子相互作用产生摩擦力，分子克服摩擦力产生热量，使物品升温，摩擦力，分子克服摩擦力产生热量，使物品升温，起加热灭菌作用）。起加热灭菌作用）。微波灭菌法，需时短、加热匀整，可用于非金属微波灭菌法，需时短、加热匀整，可用于
24、非金属医药用品等的灭菌。医药用品等的灭菌。化学消毒灭菌法化学消毒灭菌法化化学学消消毒毒灭灭菌菌法法防腐消毒剂防腐消毒剂化学治疗剂化学治疗剂用于用于杀灭杀灭体外体外病原微生物病原微生物的化学药物称为的化学药物称为消毒剂消毒剂；用于用于抑制抑制微生物生长繁殖微生物生长繁殖的化学药物称为的化学药物称为防腐剂防腐剂或或抑菌剂抑菌剂，两者并无严格的，两者并无严格的界限，统称为界限，统称为防腐消毒剂防腐消毒剂用于消除宿主用于消除宿主体内体内病原微病原微生物或其他寄生虫的化学生物或其他寄生虫的化学药物药物消消毒毒剂剂的的种种类类使菌体蛋白使菌体蛋白质变性质变性如酚类、醇类、重金属如酚类、醇类、重金属盐类、酸
25、碱类、醛类；盐类、酸碱类、醛类；损伤胞浆膜损伤胞浆膜如酚类如酚类(低浓度低浓度)、表面、表面活性剂、醇类等脂溶剂活性剂、醇类等脂溶剂干扰细菌的酶干扰细菌的酶系统和代谢系统和代谢如氧化剂、重金属盐如氧化剂、重金属盐类类(低度低度)变更核酸功能变更核酸功能如染料、烷化剂等如染料、烷化剂等The ideal disinfectantAn ideal disinfectant should Be fast acting even in the presence of organic substances,such as those in body fluid;resistant to inactiva
26、tion;Be effective against all types of infectious agents without destroying tissues or acting as a poison if ingested;broadly active;Easily penetrate material to be disinfected without damaging or discoloring the material;not poisonous or otherwise harmful;Be easy to prepare and stable even when exp
27、osed to light,heat,or other environmental factors;penetrating;not damaging to non-living materials;Be inexpensive and easy to obtain and use;stable;easily prepared;Not have an unpleasant odor.not unpleasant to work with“b.Note that different disinfectants typically have different strengths and weakn
28、esses,and often what disinfectant one uses depends on circumstances;c.Remember,the best thing one can do if unsure or ignorant of disinfectant abilities is to read the label测定消毒剂杀菌力的方法测定消毒剂杀菌力的方法石炭酸系数试验：亦称石炭酸系数试验：亦称“酚系数酚系数”，是以伤，是以伤寒杆菌作试验菌，在确定条件下待测杀毒剂寒杆菌作试验菌，在确定条件下待测杀毒剂对伤寒杆菌在规定时间内杀死所需的最大稀对伤寒杆菌在规定时间内杀
29、死所需的最大稀释度。释度。最小抑菌浓度试验：系用培育基将待测消毒最小抑菌浓度试验：系用培育基将待测消毒剂配制成一系列不同的稀释度，在每一稀释剂配制成一系列不同的稀释度，在每一稀释度中接种等量的细菌，经培育后以浊度测定度中接种等量的细菌，经培育后以浊度测定细菌的生长状况。细菌的生长状况。琼脂平皿试验：琼脂平皿试验：死亡曲线测定法：先将消毒剂与受检菌混合，死亡曲线测定法：先将消毒剂与受检菌混合，每隔确定时间取出计数残存的活菌数，再依每隔确定时间取出计数残存的活菌数，再依据菌数画成曲线。据菌数画成曲线。生物因素对微生物的影响生物因素对微生物的影响抗抗（杀杀）菌菌物物质质抗抗 生生 素素化学治疗剂化学
30、治疗剂磺胺类磺胺类抗菌增效剂抗菌增效剂异烟肼异烟肼呋喃类呋喃类细菌素细菌素植物杀菌素植物杀菌素抗菌肽抗菌肽干扰细菌细胞壁的合成干扰细菌细胞壁的合成损伤细菌胞浆膜损伤细菌胞浆膜影响细菌蛋白质的合成影响细菌蛋白质的合成改变核酸代谢改变核酸代谢活体影响活体影响噬菌体噬菌体蛭弧菌蛭弧菌Protein synthesis inhibitors 噬菌体噬菌体Sizes and Shapes of BacteriophagesT4LyticCycle Bacteria w PhagesA bacterial lawn inoculated with a range of bacteriophage Lyti
31、c and Lysogenic Cycles of a Bacteriophage噬菌体的应用噬菌体的应用细菌的鉴定和分型细菌的鉴定和分型检测未知细菌检测未知细菌分子生物学工具分子生物学工具用于生物防治用于生物防治危害抗生素和发酵工业危害抗生素和发酵工业蛭弧菌蛭弧菌食菌蛭弧菌食菌蛭弧菌Bdellovibrio bacteriovorus(bdello:leech,vibrio:curved,bacteriovorus:bacteria eater Bdellovibrio in a variety of environments-from the aquatic to the human-an
32、 indication that they are rather common,perhaps even in our guts.Theyre around 0.2-0.5 microns wide and 0.5-2.5 microns long,small enough to get through that glass filter.But culturing them is difficult.Unlike other bacteria,they cannot make their own food from the raw materials of conventional media.They depend on prey bacteria to survive.GrowthofBdellovibrio bacteriovorusDSM50701Tinagarculture.(A)BdellovibrioplaqueintheleftagarofDSMZmedium257.(B)Phase-contrastmicrographofasinglehostcellalongwithseveralBdellovibriocells.Bar=5m.