Infection, Sterilization & Disinfection


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Chain of Infection, Various physical & chemical methods used for sterilization & disinfection

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Infection, Sterilization & Disinfection

  1. 1. A S S T I N F E C T I O N E P R D I S I N F E C T I O N I L S I Z A A N T I S E P T I C I O - K A L P E S H N
  2. 2. INFECTION A pathological condition due to the growth of microorganisms in host
  3. 3. Chain of Infection Causative agent Susceptible Host Reservoir Portal of Entry Portal of Exit Mode of Transmission
  4. 4. Causative agents / Infectious agent: A microbial organism with the ability to cause disease. Examples: Bacteria Viruses Fungi Protozoa Reservoir / Sources: A habitat within which microorganisms lives, grows and multiplies. Examples: Human beings Animals Soil Water
  5. 5. Portal of Exit: A place of exit providing a way for a microorganism to leave the reservoir. Examples: Upper respiratory tract Saliva from the oral cavity Sneezing Coughing Urogenital tract Semen Vaginal secretions Infected urine Gastrointestinal tract Feces / diarrhoea from the bowel Vomitus Skin and mucous membranes Discharges from infected skin lesions and infected wounds Blood Infected blood
  6. 6. Mode Of Transmission: Method of transfer by which the organism moves or is carried from one place to another. Transmission of Infection Direct Indirect Direct contact Droplet spread Air-borne Vehicle-borne Vector-borne (mechanical or biologic)
  7. 7. Direct Transmission: Direct contact • Person-to-person contact • Contact with soil or vegetation harboring infectious organisms. Droplet spread • Spray with relatively large, short-range aerosols produced by sneezing, coughing, or even talking. • Droplet spread is classified as direct because transmission is by direct spray over a few feet, before the droplets fall to the ground. • Eg. Pertussis and meningococcal
  8. 8. Indirect Transmission: Transfer of an infectious agent from a reservoir to a host by suspended air particles, inanimate objects (vehicles), or animate intermediaries (vectors). Airborne transmission: • Occurs when infectious agents are carried by dust or droplet nuclei suspended in air. • Airborne dust → settles on surfaces → become resuspended by air currents • Droplet nuclei: less than 5 microns in size. • Droplets fall to the ground within a few feet Droplet nuclei may remain suspended in the air for long periods of time and may be blown over great distances.
  9. 9. Vehicles: • Food • Water • Biologic products (blood) • Fomites: inanimate objects such as handkerchiefs, bedding, or surgical scalpels Vectors: • Mosquitoes • Fleas and ticks • Carry an infectious agent through mechanical means or may support growth or changes in the agent.
  10. 10. Portal Of Entry: An opening allowing the microorganism to enter the host. Examples: Upper respiratory tract inhalation (eg. influenza) Urogenital tract sexual contact (eg. Chlamydia) Gastrointestinal tract ingestion (eg. gastroenteritis) Skin and mucous membranes open wound or punctures ulcers Blood Needlestick injury (eg. hepatitis B) Tubes placed in body cavities such as urinary catheters
  11. 11. Susceptible Host: A person who cannot resist a microorganism invading the body, multiplying, and resulting in infection. Susceptibility of a host depends on: • Genetic or constitutional factors • Specific immunity
  12. 12. Important Terminologies • ASEPSIS: The state of being free of living pathogenic microorganisms • STERILIZATION: Process by which an article, surface or medium is made free of all living microbes either in vegetative or spore state. • DISINFECTION: Process of reducing or eliminating living pathogenic microorganisms in or on materials, so they are no longer able to cause disease. The agent used to achieve this state is called Disinfectant.
  13. 13. • ANTISEPTICS: Chemical disinfectants which can be easily applied on skin or mucus membrane to prevent infection by inhibiting the growth of pathogens • BACTERICIDAL AGENTS: Agents which are able to kill bacteria. • BACTERIOSTATIC AGENTS: Agents which inhibit the multiplication of bacteria but does not necessarily kill them. • DECONTAMINATION: Process of rendering an article or area free of danger from contaminants.
  14. 14. Various agents used in Sterilization & Disinfection Physical agents Chemical agents
  15. 15. PHYSICAL AGENTS CHEMICAL AGENTS Sunlight Alcohols: Ethanol, isopropyl alcohol Drying Aldehydes: Formaldehyde, Glutaraldehyde Dry heat : Hot air oven, Incineration, Flaming Dyes Moist heat: Boiling, Pasteurization, tyndallisation, Autoclaving Halogens Filtration Phenolic compounds Radiation Ionizing, Non Ionizing Gases Ethylene oxide, Formaldehyde, beta propiolactone(BPL). Ultrasonic vibration Metallic salts and surface active agents
  16. 16. Physical Methods of Sterilisation Sunlight • Direct sunlight is a natural method of sterilization of water in tanks, rivers and lakes. • Direct sunlight has an active germicidal effect due to the combined effect of ultraviolet and heat rays. Drying • 4/5th of the bacterial weight is due to water. • Therefore drying in air has deleterious effect on many bacteria. • This is unreliable method, spores are unaffected by drying.
  17. 17. Heat • Most reliable method of sterilization • Dry heat: Oxidative damage & toxic effect of elevated levels of electrolytes • Moist heat: Denaturation & coagulation of proteins. • Time required for sterilization: inversely proportional to temp. of exposure Dry Heat: 1. Flaming 2. Incineration 3. Hot air oven
  18. 18. FLAMING  Inoculation loop or Wire, the tip of Forceps and spatulas are held in a bunsen flame till they are red hot.
  19. 19. INCINERATION     Combustion at high temp This is an excellent method of destroying materials such as contaminated cloths, dressings, animal carcasses and pathological materials. Plastics such as PVC & polythene sterilized by this method. Polystyrene emits dense black smoke on incineration.
  20. 20. HOT AIR OVEN • The oven utilizes dry heat to sterilize articles • Low penetrating power • A holding period of 160 C for 1 hr is desirable. • At 180 C cotton plugs get charred • Cutting instruments used in surgery sterilized at 150 C for 2 hrs • Oven heated by electricity. • Fan fitted for even distribution of air. • There is a thermostat controlling the temperature. • Double walled insulation keeps the heat in and conserves energy.
  21. 21. Recommended temp & duration:     160ºC for at least 45 mins. 170ºC for at least 18 mins. 180ºC for at least 7.5 mins. 190ºC for at least 1.5 mins. Uses:   To sterilize Glassware, Forceps, Scissors, Scalpels, Swabs Pharmaceuticals products like Liquid paraffin, dusting powder, fats and grease.
  22. 22. Sterilization Control: Clostridium tetani used as microbiological test Paper strips impregnated with spores: inserted in suitable packs Sterilized Removed & inoculated into Thioglycolate broth Incubated at 37ºC for 5 days
  23. 23. Moist Heat: 1. Temp below 100°C: Pasteurisation Water bath Inspissator 2. Temperature at 100°C: Boiling Koch/Arnold’s steamer (Steam sterilizer) Tyndallisation 3. Steam under pressure (above 100°C): Autoclave
  24. 24. TEMPERATURE BELOW 100 C Pasteurisation:   Process of killing of pathogens in the milk but does not sterilize it . Milk is heated at 63 C for 30 mins. (Holder method) or at 72 C for 15-20 Sec. (Flash process)  Followed by rapid cooling to 13 C  Non-sporing bacteria destroyed  Coxiella burtenii heat resistant & survives Pasteurisation
  25. 25. Water Bath:   To inactivate non sporing bacteria for the preparation of vaccines - Special vaccine bath at 60 C for one hour is used Serum or body fluids containing coagulable proteins can be sterilized by heating for 1 hr at 56 C in a water bath for several successive days. Hot water bath
  26. 26. Inspissator:      Sterilizes by heating at 80-85 C for half an hour for 3 successive days Media such as Lowenstein-Jensen & Loefller’s serum sterilized in Inspissator. Non-sporing bacteria killed by moist heat at 60 C for 30 mins Temp of 80 C for 5-10 mins destroy vegetative forms of bacteria, yeasts & moulds Viruses inactivated rapidly at 60 C
  27. 27. TEMPERATURE AT 100 C Boiling:  Kills vegetative forms of bacterial pathogens at 90-100 C.  Process hastened by adding 2% Sodium bicarbonate.  Not recommended for sterilization of surgical instruments  Sporing bacteria exposed for prolonged period.  Hepatitis virus: Can survive up to 30 minutes of boiling.  Endospores: Can survive up to 20 hours or more of boiling
  28. 28. Steam sterilizer:      Steam is generated using a steamer (Koch/ Arnold) Consists of a Tin cabinet Has a conical lid to enable the drainage of condensed steam Perforated tray above water: ensures materials are surrounded by steam. For routine sterilization exposure of 90 mins is used
  29. 29. Tyndallisation:    For media containing sugar and gelatin exposure of 100 C for 20 min for 3 successive days is used Process is also termed as Intermittent Sterilization Principle: - 1st exposure kills vegetative bacteria - In intervals spores germinate into vegetative cells which are killed on subsequent heating
  30. 30. STEAM UNDER PRESSURE Autoclave Principle:    Steam under pressure (above 100 C) has better killing power as compared to dry heat: better penetration power Bacteria more susceptible to moist heat: bacterial protein coagulates rapidly Steam coming in contact with cooler surfaces condenses to water & liberates latent heat to that surface
  31. 31. Components:  Autoclave consists of a vertical or a horizontal cylinder. (Gunmetal or stainless steel in supporting iron case)  Lid: fastened by screw clamps  The lid is provided with: Discharge tap: for air & steam Pressure gauge: to measure the pressure  A safety valve is present to permit the escape of steam from the chamber
  32. 32. Procedure:  Sufficient water added  Articles to be sterilised are placed on tray & autoclave is heated  Lid screwed tight with discharge tap open  When no more air comes out, discharge tap is closed  Sterilisation can be carried out under following conditions: 121 C for 15 mins. 126 C for 10 mins. 134 C for 3 mins.  Heater turned off & autoclave allowed to cool  Discharge tap opened to allow the entry of air in autoclave
  33. 33. Sterilization control: 1. Spores:  Spores of Bacillus stearothermophilus used as test organism  Spores get killed on exposure of 12 mins at 121 C  Spores impregnated on paper strips: packed in envelopes → inserted in different parts of autoclave → Sterilization → inoculated into suitable medium → incubated 2. Thermocouples: Records temp directly by potentiometer 3. Chemical indicators
  34. 34. Filtration Filtration helps to remove bacteria from heat labile liquids such as serum and solutions of sugar, Antibiotics. Uses: • To sterilize serum and solutions of sugar, Antibiotics for preparation of culture media • To obtain bacteria-free filtrates of clinical samples for virus isolation • Separation of Toxins & bacteriophages from bacteria • Filter discs to retain bacterial cells from liquid Types: • Candle filters • Asbestos filters • Sintered glass filters • Membrane filters
  35. 35. CANDLE FILTERS   Widely used for purification of water for Industrial & drinking purpose Two types 1. Unglazed ceramic filter: Eg. Chamberland filter & Doulton filter 2. Diatomaceous earth filters: Eg. Berkefeld filter & Mandler filter
  36. 36. ASBESTOS FILTER  Disposable, single-use discs  High adsorbing capacity  Carcinogenic  Eg: Seitz filter, Sterimat filter
  37. 37. SINTERED GLASS FILTER    Prepared by heat fusing powdered glass particles of graded size Low absorptive property & can be Cleaned easily brittle, expensive.
  38. 38. MEMBRANE FILTERS  Made of cellulose esters or other polymers Uses:  Water purification & analysis  Sterilization & sterility testing  Preparation of solutions for parenteral use
  39. 39. Radiation Two types: 1. Non-ionising: IR, UV 2. Ionising: Gamma, high energy electrons Non- Ionising radiation: • Electromagnetic rays with longer wavelength • Absorbed as heat • Infrared rays: Can be considered as hot air sterilisation • IR: rapid mass sterilisation of prepacked Syringes and catheters • UV: disinfecting enclosed areas like operation theatres, laboratories
  40. 40. IONISING RADIATIONS:  X- rays, gamma rays & cosmic rays.  Lethal to DNA & other vital constituents  High penetrative power  No appreciable increase in the temperature – Cold sterilisation  Sterilise plastics Syringes, swabs, catheters, animal feed, grease, fabrics, metal foils Ultrasonic & Sonic Vibrations   Bactericidal Microorganisms vary in their sensitivity, hence no practical value in sterilisation and disinfection
  41. 41. Chemical Methods Chemicals can be used as Antiseptics & Disinfectants Potency of disinfectants depends on:  Concentration of substance  Time of action  pH of medium  Temperature  Nature of organism  Presence of extraneous material
  42. 42. Characteristics of Ideal Antiseptic or Disinfectant:            Wide spectrum of activity (effective against all microbes) Active in presence of organic matter Effective in acid or alkaline media Rapid action High penetrating power Stable Should not lead to corrosion of metals Should not cause local irritation No interference with healing Should not be toxic if absorbed into circulation Inexpensive & easily available & easy to use
  43. 43. Chemical agents act in one of the following ways:  Protein coagulation  Disruption of the cell membrane   Removal of free Sulphydryl groups (required for enzyme action) Substrate competition: compound resembling essential substrate.
  44. 44. ALCOHOLS      Ethanol /Isopropyl alcohol are frequently used As skin antiseptics: denature proteins No action on spores Concentration recommended 60-90% in water Isopropyl alcohol preferred: better fat solvent, more bactericidal, less volatile Uses:  Disinfection of clinical thermometer.  Methanol: effective against fungal spores → Used for treating cabinets & incubators
  45. 45. ALDEHYDES  Formaldehyde & Glutaraldehyde are frequently used Formaldehyde:  Bactericidal, sporicidal & has a lethal effect on viruses  Used to preserve anatomical specimens  Destroying Anthrax spores in hair and wool  10% Formalin+0.5% Sodium tetraborate is used to sterilise metal instruments  Formaldehyde gas: irritant & toxic when inhaled used for sterilizing instruments & heat-sensitive catheters, fumigating wards, laboratories
  46. 46. Glutaraldehyde:  Action similar to formaldehyde  Effective against Tubercle bacilli, fungi & viruses  Less irritant & toxic to eyes & skin  Used to treat corrugated rubber anesthetic tubes, Face masks, Plastic endotracheal tubes, Metal instruments and polythene tubing
  47. 47. DYES Bacteriostatic in nature Aniline dyes:  Are Brilliant green, Malachite green & Crystal violet  Active against Gram positive bacteria  No activity against tubercle bacilli. Hence, malachite green used in LJ medium  Non-toxic & non-irritant to tissue but inhibited by organic material like pus Acridine dyes:  Impair the DNA complexes in bacterial cell  Effective against Gram positive  Important dyes are Proflavine, Acriflavine, Euflavine, Aminacrine
  48. 48. HALOGENS Iodine:  Iodine in aqueous and alcoholic solution has been used widely as a skin disinfectant  Actively bactericidal with moderate action against spores  Active against Tubercle bacilli & viruses Chlorine:  Chlorine and its compounds have been used as disinfectants in water supplies, swimming pools, food & dairy industries  Used commonly as hypochlorites (ClO)  Wide spectrum of action against viruses
  49. 49. PHENOLS       Obtained by distillation of coal tar at 170-270 C Damage cell membrane releasing cell contents Phenolic derivatives have been widely used as disinfectants for various purposes in hospitals Lysol, cresol: active against wide range, not inactivated by organic matter, but toxic to humans Hexachlorophene: toxic, should be handled with care Chlorhexidine: non-toxic antiseptic, most active against Gram +ve Uses:  Various combinations are used in the control of pyogenic cocci in surgical & neonatal units in hospitals.  Aqueous solutions are used in treatment of wounds
  50. 50. GASES Ethylene Oxide:  Colourless, Highly penetrating gas with a sweet ethereal smell.  Effective against all types of microorganisms including viruses and spores  Inflammable & explosive at high concentration  Alkylate carboxyl, hydroxyl & amino group in protein and also react with RNA & DNA  Toxic, mutagenic & carcinogenic Uses:  Specially used for sterilising heart-lung machines, respirators, sutures, dental equipments, books and clothing.  Also used to sterilise Glass, metal and paper surfaces, plastics, soil, some foods and tobacco.
  51. 51. Formaldehyde gas:   Widely employed for fumigation of operation theatres and other rooms When formaldehyde vapour is generated, doors should be sealed & left unopened for 48 hrs Beta Propiolactone (BPL):  Low penetrating power but more efficient than formaldehyde for fumigation  Used in fumigation  For sterilisation 0.2% BPL is used  Has a rapid biocidal activity, but carcinogenic  Very effective against viruses
  52. 52. SURFACE ACTIVE AGENTS & METALLIC SALTS Surface active agents:  Substances which reduce the surface tension  Used as wetting agents, detergents & emulsifiers  Classified as: Anionic, Cationic, Nonionic & Amphoteric  Cationic compounds act on PO4 group in cell membrane  Cations are widely used in the form of quaternary ammonium compounds.  Markedly bactericidal, active against Gram positive organisms.  No action on spores, tubercle bacilli, viruses  Anionic compounds (soaps): moderate action  Soaps from saturated fatty acids: against Gram –ve  Soaps from unsaturated fatty acids: against Gram +ve
  53. 53. Metallic Salts:  Salts of heavy metals: greater action  The salts of silver, copper and mercury are used as disinfectants.  Act by coagulating proteins  Organic compounds: marked bacteriostatic, weak bactericidal and limited fungicidal activity  Mercuric chloride: highly toxic  Copper salts: fungicides
  54. 54. TESTING OF DISINFECTANATS Rideal-Walker test:  Activity of disinfectant to be tested is compared to the activity of phenol of varying concentration Chick-Martin Test:  Disinfectant is tested in the presence of organic matter
  55. 55. Thank You