1、Disinfection and Sterilization,For the course of Medical Microbiology for MBBS foreign students, Class 2006/2011, SYSUSeptember 18, 2007Mengfeng Li （黎孟枫）, M.D. Department of Microbiology, Zhongshan School of Medicine, SYSU, Guangzhou, C,Medical Microbiology,Early needs and success for disinfection (
2、Early 1800s) a historic story,By mid-1800s, almost half of post-operative patients died of sepsis (then called “hospital disease”). A common report by surgeons was: operation successful but patient died.A hypothesis: exposing moist body tissue to oxygen sepsis; best prevention: keeping air away from
3、 wounds by means of plasters, collodion or resins.Having tried methods to encourage clean healing with no success, surgeon Joseph Lister discarded the concept of direct infection by bad air but postulated that sepsis might be caused by a pollen-like dust, although he did not know yet the “dust” was
4、living microbes. When Louis Pasteur suggested the presence of living organisms in the air, Lister made the connection with wound sepsis: the microbes in the air were likely causing the sepsis and should be destroyed before they entered the wound.Lister had previously heard that carbolic acid was use
5、d to treat sewage, and that fields treated with the affluent were freed of a parasite causing disease in cattle. He then began to clean wounds and dress them with carbolic acid.,Joseph Lister operated with carbolic acid (石炭酸）spray,The milestone:case of James Greenlees,-Aug. 12, 1865, James Greenlees
6、, 11-year-old-Struck by a horse-drown cart-Compound fractures-Taken to Royal Infirmary-Under the care of Joseph Lister,Why disinfection and sterilization?,Contagious diseasesHospital infection (e.g., OR, ID ward) or other opportunistic infection Lab contaminationEtc. Microbes:usually easy to grow in
7、 environment; but also can be inhibited or killed by certain environmental (physical or chemical) factors/conditions.,Terminology,Disinfection消毒Sterilization灭菌Bacteriostasis抑菌 Antisepsis防腐Asepsis 无菌,Disinfection（消毒）,Process of reducing or eliminating living pathogenic microorganisms in or on materia
8、ls, so they are no longer a health hazard.For example: use of alcohol before drug injection.,Sterilization（灭菌）,Process of destroying all microbial forms. A sterile object is one free of all microbial forms, including bacterial spores.More thorough than disinfection,Bacteriostasis（抑菌）,Process of inhi
9、biting the growth of microorganisms, in vivo (mostly) or in vitro. For example: bacteriostatic antibiotics,Antisepsis （防腐）,Process of inhibiting or preventing growth of microbes, mostly in vitro and not bactericidal or sporicidal (杀芽胞的 )For example: use of chemical agents on skin, other living tissu
10、es or food/beverage.,Asepsis（无菌）,A state where no living microorganism exists. For example: OR (Operating Room),Controlling Microorganisms with Physical Conditions,High Temperature (heat)Radiation Ultrasound FiltrationLow Temperature Desiccation（干燥）,High Temperature Dry heat and Moist heat protein d
11、enaturation and clotting; DNA strand breakdown,.,Dry heat：protein oxidationIncineration（焚烧）most thorough (500)disposals and corpesFlaming (burner)（烧灼） test tube opening, transferring loopHot air sterilization/Baking（干烤）160-170, 2hGlassware, china, syringes, needles, etcInfrared heat: similar to baki
12、ng,Moist heat：denaturing proteins and melt lipidsAutoclaving（高压蒸汽灭菌）Most commonly used and effective121 (103.4kPa), 15-20minkilling both vegetative organisms and endospores Boiling100 (105 with 2% Na2CO2) , 15-20mincidal for vegetative cells but not necessarily sporesRegular Steam (Arnold Sterilizer
13、)100 , 15-20mincidal for vegetative cells but not necessarily spores,Pasteurization（巴氏消毒法）to kill pathogens in readily perishable objects (milk, wine)flash method (瞬间法): 71.6, 15s holding method (持续法): 62.9, 30 minFractional sterilization（间歇蒸气灭菌法）alternating exposure and cooling time for a consecuti
14、ve period:Steam heating (100, 30 min) 30 for endospores to germinate 100, 30 min to kill germinated endospores 30-37 overnight for remaining endospores to germinate 100, 60 min to kill last remaining germinated endosporesfor sugar- or milk-containing culture media,Moist Heat vs Dry Heat,Moist heat D
15、ry heatPenetrating potency higherlowerTemp for protein clotting lowerhigher Extra heat released yesnofrom condensation,Sterilizing potency: Moist heat Dry heat,Radiation,Ultraviolet (UV) radiationmechanism: blockage of DNA replication by forming thymidine dimmersmicrobicidal activity of UV depends o
16、n:length of exposure wavelength: 200-300 nm, with the best effect of 265-266nmbulb life (4000hr)very poor penetrating powerfor air or surface disinfection(OR, ID ward, labs) causing eye damage, burnsand mutation in skin cells,Ionizing Radiation（电离辐射）X-rays, gamma rays and high-speed electrons genera
17、ting more energy and penetrating power than UVto sterilize pharmaceuticals, disposable medical supplies (e.g., syringes, gloves, catheters, sutures) and foodsMicrowave penetrating non-metal materials (glass, plastics, china),Ultrasoundmore effective for gram-negative bacteriaLack of thoroughness sur
18、vivors remainFiltration sterilize heat- or chemical-sensitive solutionsnot effective for virus, ricketia, mycoplasma,Seitz filter,Desiccationstatic effect by inhibiting microbial enzymesnot effective against endosporesmainly for food reservation Low Temperature (-20 -70)inhibits microbial growth by
19、slowing down microbial metabolisma special form: lyophalization (freeze-drying), used for long-term (years) reservation of bacteria stocks fast freezing + dryingprotecting agents (glycerol, serum),Control Microorganisms with Chemical Agents (Disinfectants and Antiseptics),Antimicrobial modes of acti
20、on of disinfectants and antiseptics,Denaturation of bacterial proteins by disrupting hydrogen and disulfide bonds phenol (high conc.), alcohol, heavy-metal (high conc.), acids, alkalies, aldehydes)Damage to bacterial membrane (lipids and/or proteins), causing leakage of intracellular molecules pheno
21、l (low conc.), surfactants, dyesInterference of bacterial enzyme and metabolism oxidants, heavy-metals (low conc.), alkylating agents,Phenol and phenol derivatives altering membrane permeability and denaturing proteins0.01% - 0.05% Chlorhexidine（洗必泰） vaginal wash, OR hand-wash3% - 5% carbonic acid o
22、r 2% Lysol floor or surface disinfection,Alcohols denaturing bacterial proteins and membranes70% - 75% ethyl or isopropyl alcohol（乙醇或异丙醇） skin and thermometer disinfectionineffective against endospores and non-enveloped viruses,Heavy metals (Hg2+、Ag+) denaturing proteins and inactivating enzymes2% m
23、ercurochrome（红汞）or 0.1% merthiolate（硫柳汞） skin, mucosa and woundbacteriostatic, ineffective against endospores1% silver nitrate（硝酸银） eye drops for newborns to prevent gonococcal ophthalmia（淋菌性眼炎）,Oxidants oxidation, protein precipitation0.1% potassium permanganate（高锰酸钾） skin, fruits/vegetables 3% per
24、oxide（过氧化氢） small trauma wound, skin, mucosa0.2% - 1% peroxyacetic acid（过氧乙酸） plastics, glassware0.2 0.5 ppm cholorines water and swimming pool,Surfactants damaging bacterial membranes, inactivating enzymes, protein precipitation0.05-0.1% bromogeramine （新洁尔灭） OR hand-wash, skin, surgical instruments
25、,Alkylating agent（烷化剂） alkylating proteins and nucleic acidsformalin (formaldehyde) surface disinfection, air, surgical instrumentsglutaric dialdehyde（戊二醛） high-precision instruments, endoscopes50mg/L epoxy ethane（环氧乙烷） surgical instruments and dressing,Dyes inhibiting bacterial growth by interferin
26、g with oxidation2% - 4% methyl violet（龙胆紫） wound disinfection,Acids and alkalies destroying cell membrane and cell wall, denaturing proteins5-10ml/m3 acetic acid evaporation air disinfectionquicklime Ca(OH)2 floor and excretion (feces, urine, sputum, pus) disinfection,Effectiveness of antimicrobial
27、agents are affected by ,The concentration/intensity and nature of the disinfectant; Length of exposure;Species and number of the microbe(s); Temperature and humidity;Acidity (pH);Presence of organic substances;Presence of chemical antagonistsThe nature of the material bearing the microbes,Summary 1.
28、 Application of chemical disinfectants,Patient excretionChlorines, 5% carbonic acid, 2% LysolSkin (hands)2% Lysol, 0.2-0.4% peroxyacetic acid（过氧乙酸）for HBV, 70% ethyl alcohol, 2% mercurochrome（红汞）Mucosaoral- 3% peroxide; uri-reproductive- 0.01-0.05% Chlorhexidine（洗必泰）, 0.1% potassium permanganate（高锰酸
29、钾）; newborn eyes- 1% silver nitrate,Drinking waterChlorinesToilets, sewagequicklime Ca(OH)2Air (OR, ID ward)formalin steam (12.5-25ml/m3,12-24h),formalin 40ml + potassium permanganate 30g/m3;HBV ward- peroxyacetic acid 3g/m3 90minGlassware, china,0.5% iodophores, 0.2-0.4% peroxyacetic Rubber, metal
30、aciddevices,Potency DefinitionExamplesHigh Killing all microbes including glutaric dialdehyde（戊二醛）， endospores and TBformaldehyde peroxyacetic acid（过氧乙酸），epoxy ethane（环氧乙烷）Medium Killing all non-spore microbesalcohol, chlorines, iodophores including TBLow Killing vegetative bacteria chlorhexidine（洗必
31、泰）， and lipophilic (enveloped) bromogeramine（新洁尔灭） viruses, but resisted by endospores, TB and hydrophilic (non-enveloped) viruses,Summary 2. Potency levels of chemical disinfectants,Spore-killing disinfectinsglutaric dialdehyde（戊二醛）, formaldehyde（甲醛), Iodines, H2O2, epoxy ethane（环氧乙烷）Non spore-kill
32、ing disinfectinsalcohols, phenols, chlorhexidine（洗必泰）, bromogeramine（新洁尔灭）,Summary 3. Spore-killing effects of chemical disinfectants,Disinfection and Sterilization,For the course of Medical Microbiology for MBBS foreign students, Class 2006/2011, SYSUSeptember 18, 2007Mengfeng Li （黎孟枫）, M.D. Department of Microbiology, Zhongshan School of Medicine, SYSU, Guangzhou, C,Medical Microbiology,
