Disinfection

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Table Of Contents
  1. Questions (MI 1.4, 1.5)
  2. Introduction
  3. Classification
  4. Physical Methods
  5. Chemical Methods
  6. Testing of Disinfectants

Questions (MI 1.4, 1.5)

  • Define Disinfection
  • Classify Disinfectants
  • Classify different methods of disinfection
  • Most appropriate method of disinfection in specific situations in laboratory, clinical & surgical practice.
  • Classify chemical disinfectants.
  • Various Disinfectants Used
  • Mechanism of Action of Disinfectants
  • Application of different methods in the laboratory, clinical & surgical practice
  • Tests for Disinfectant Efficiency/ Controls

Introduction

READ IN LANDSCAPE MODE ON MOBILE DEVICES

  • Sterilisation– Absolute process(all or none) of removal of all microbes and all spores from inanimate objects
  • Disinfection– Removal of pathogens from inanimate objects leaving behind non-pathogens and spores
  • Sterilisation > Disinfection

Classification

Classification of Disinfectnts

Physical Methods

  • Sunlight– Bactericidal action of UV rays (most absorbed already by stratospheric ozone)
  • Drying– Kills microbes as 80% bacterial weight is water. Spores not affected, hence only disinfects.
  • Heat– S&D. Most common method unless contraindicated(Eg.Heat labile materials). Two types-
    • Moist Heat– Better than dry heat, kills by denaturation and coagulation of proteins. Three types-
      • Below 100 C-
        • Pasteurisation
          • Used for beverages, beer, dairy products
          • Kills all non-sporing pathogens such as Mycobacterium, Brucella, Salmonella, etc. except Coxiella burnetti(heat resistant)
          • Tests: Phosphatase Test, Coliform Test, Standard Plate Test
        • Waterbath– Disinfection of Bactericidal vaccines (60 C for 1 hour), Serum and other heat labile body fluids (56 C for 1 hour)
        • Inspissation– Heating at 80-85 C for 30 minutes for 3 consecutive days. Sterilisation of medium containing Egg (Lowenstein Gensen medium for Mycobacterium tuberculosis) or serum (Loeffler’s serum slope for Corynebacterium diphtheriae)
      • At 100 C-
        • Boiling–
          • Water boiled for 15 minutes
          • Only vegetative forms are killed- Disinfection only
        • Steaming–
          • Koch’s Steamer or Arnold’s Steamer used
          • Used for culture media which decomposes at very high temperatures.
          • Steam condenses and gives off latent heat.
          • 100 C , 1 atm, 90 minutes.
        • Tyndallisation–
          • Intermittent sterilisation of sugar, serum, egg or gelatin containing culture media.
          • 100 C, 20 mins, 3 consecutive days.
      • Above 100 C-
        • Autoclave–
          • Boiling at temperatures above 100 C by raising atmospheric pressure.
          • Holding Period: 121 C, 15 minutes, 15lbs psi. Or 126 C, 10 mins, 20lbs psi, Or, 133 C , 3 min, 30lbs psi
          • Sterilizes all heat stable culture media, gloves, gowns, non-sharp surgical instruments, biohazardous wastes, Plastic and rubber Foley’s catheter, etc.
          • Contraindicated for waterproof and dry materials, PET, sharp surgical instruments,  thermocouples, Browne’s tube and autoclave tapes.
          • Physical Control- Digital displays with temperature and time
          • Chemical Control – Autoclave tapes
          • Biological- Geobacillus stearothermophiles.
  • Dry Heat- Kills by charring, oxidative damage and toxic effects of elevated electrolytes besides denaturation. Dry Heat methods are-(H-FIR)-
    • Hot Air Oven-
      • Best and most commonly used dry heat method.
      • Holding Period- 160 C for 2 hours, or 170 C for 30 Min, or 180 C for 15 min.
      • Used to sterilise glassware like test tube and petri dish, sharp surgical instruments(scalpel, scissors and forceps), and chemicals such as Fats, oils, glycerol, glove powder, grease, liquid paraffin and jellies.
      • Physical Controls- Digital displays, thermocouples, Brownie’s tube (heat sensitive red dye changes color to green)
      • Biological Controls- Spores of non-toxic strain of Clostridium tetani or Bacillus subtilis or Bacillus atropheus.
  • Flaming–
    • Done for fragile items like mouths of test tube, Inoculation loops & wires.
    • Used to decrease microbial load in surrounding air.
  • Incineration- Burning (sterilisation) anatomical & microbiological waste(Biomedical Waste-Yellow bag materials) by providing a very high temperature (870 – 1200° C) & thereby converting all waste into either ashes, flue gas (reduced waste volume) or heat.
  • Red Heat- Bacterial loops/wires heated till red hot, then cooled for 20-30 seconds
  • Radiation- Used for Heat labile solids. Control is Bacillus pumilis.
    • Ionising-
      • High penetrating radiations
      • Method of Cold Sterilisation, also disinfection.
      • Cosmic or X-rays or Gamma Rays used
      • Used to sterilise disposable plastics, antibiotics, adhesive dressing, catgut suture, bone and tissue grafts, etc.
    • Non-Ionising-
      • Low penetrating radiations
      • Method of Disinfection
      • IR and UV rays such as from sun, mercury vapor bulbs used.
      • Antimicrobial activity best at 250-260 nm
      • Used for wounds, biosafety cabinets, hospital wards, drinking water, contact lenses.
      • Disadvantage: Can cause keratoconjunctivitis and skin erythema
  • Filtration-
    • Used to separate toxins and bacteriophage from bacteria in heat labile  fluids.
    • To get bacteria free filtrates
    • Purification of water
      • Depth Filter- filtration of food and beverages. Some particles still pass through. Can be candle filters like Diatomaceous earth filters(berkefeld) or ungazed porcelain filters(Chamberland)  or asbestos and sintered glass filters(food and chemical industry).
      • Membrane Filter-
        • Most commonly used with pore size of 0.22 microns which removes most of bacteria.
        • 0.45 microns retains the coliform bacteria.
        • 0.8 microns removes air borne microbes.
        • Used in OTs, ICUs, and Transplantation Units. Serums, Vaccines, antibodies, antibiotics, biosafety cabinets,
        • HEPA- high Efficiency Particulate Air Filters- 99.97%, 0.3 microns
        • ULPA – 99.99% , 0.12 microns
        • 99% of particles bigger than 0.12 microns removed.
        • Sterilisation Control- Bevundimonas diminuta and Serratia marcescens.

Chemical Methods

  • FDA Classification of Chemical Disinfectants-
    • High-
      • Kills Virus, Fungi, Bacteria, Mycobacteria, Spores
      • Eg. ETO, Plasma, Aldehyde
    • Intermediate-
      • Kills Virus, Fungi, bacteria, Mycobacteria,  +/- spores
      • Eg. Halogens, Phenolic compounds
    • Low level-
      • Kills Virus, Fungi, bacteria, +/- Mycobacteria, +/- spores
      • Eg. Alcohols, Dyes, Surface Active Agents, Heavy metal salts.
  • Phenol-
    • Good action on GP, GN, and Mycobacterium tuberculosis.
    • Poor action on spores and viruses, hence main disinfectant action.
    • Damages cell membrane, causes cellular lysis, denaturation of proteins.
    • Disinfectant phenol derivatives (Phenolics)- Lysol(surface disinfectant), cresol(sharps), xylenol, ortho phenyl phenol, chloroxylenol(Dettol), chlorhexidine(savlon)
  • Halogens(Chlorinated compounds)-
    • Free chlorine or hypochlorites which release hypochlorous acid (HOCl)
    • Good action on GP and GN and viruses.
    • Poor action on spores and Mycobacterium tuberculosis, hence mainly surface disinfectant
    • Large blood spills- 0.5% NaClO
    • Small blood spills- 0.05% NaClO
    • Liquid pretreatment/lab disinfection- 1% NaClO
    • Surface disinfectant- 0.5% NaClO
    • Clostridium difficile- 0.5% NaClO(Sporicidal)
  • Iodinated Compounds-
    • Good antiseptics but may cause skin disorders and allergies.
    • E.g. Tincture of Iodine= (2%) water-ethanol solution, Betadine= Povidone-Iodine solution. 
  • Iodophors-
    • Water soluble, preanesthetic antiseptic prepared by coupling iodine with organic carriers.
    • E.g. Povidone, Chlorhexidine.
    • Chlorhexidine is bactericidal and sporostatic
    • 4% CHG is effective against Clostridium difficile.
    • Present is handwash, handrub and bodywash.
  • Alcohols-
    • Disinfectant & Antiseptic
    • Good action on enveloped virus & Microbes through denaturation of proteins and cell membrane damage
    • 70% Ethyl alcohol is used as surgical spirit, hand rub.
    • 70 % Isopropyl alcohol is better as it spreads and penetrates better and can’t be misused.. Used to disinfect surgical thermometer and small surgical instruments
  • Aldehydes-
    • Used to disinfect and sterilise
    • Acts by alkylation of proteins and nucleic acids
    • Formaldehyde is toxic, irritant, corrosive and carcinogenic
    • Formalin(40% formaldehyde) is used to preserve anatomical specimens, preparation of toxoids from toxins, and earlier used for fumigation of Ots
    • Glutaraldehyde is less toxic and used for disinfection(15 mins) and sterilisation(12 hrs) of fibre optic endoscopes(2%). Activated by alkalinisation and lasts only 14 days.
    • 0.55 % Orthopthalaldehyde used for disinfection & sterilisation of fibre optic scopes. Unlike glutaraldehyde it does not need activation and is more stable.
  • Ethylene Oxide(ETO)-
    • Highly penetrating, irritant, explosive, corrosive, gaseous chemical sterilant (10-20%)
    • Acts by alkylation of nucleic acids and combines with cellular proteins
    • 37-43 C, 4-5 hours. Carcinogenic and Inflammable- hence aerated for 10-12 hours.
    • Sterilises heat sensitive items such as disposable plastics, Heart lung machine, respirators, dental equipment’s, catheters, sutures, etc.
    • Biological indicator- Bacillus golbigi, Bacillus atrophaeus, Bacillus subtilis.
  • Surface Active Agents-
    • Cationic Surfactants include quaternary ammonium compounds like Cetrimide, benzalkonium Chloride, etc. which kill most gram positive bacteria, and enveloped virus. Spores and Mycobacterium tuberculosis can’t be killed- Disinfectant. Strong antimicrobial, weak detergent.
    • Anionic surfactants like common soaps are active at acidic pH and have weak antimicrobial but strong detergent activity
    • Amphoteric surfactants like TEGO compounds used as antiseptics in dental practice which have strong antimicrobial and detergent properties.
  • Oxidising Agents-
    • Mainly H2O2 and Peracetic acid is used
    • H2O2 is a strong OA which can disinfect as well as sterilise ventilators, soft contact lenses, tonometer biprism, etc. 3-6% is effective against most microbes. Catalase +ve require 10%. Vaporised H2O2 is used in plasma sterilisation.
    • Peracetic Acid is sporicidal even at <1%, low temperature for objects like endoscopes. Used along with H2O2 to disinfect hemodialyzers.
  • Plasma Sterilisation-
    • Contains ions, photons, free e, and neutral uncharged particles created from H2O2 or mixture of H2O2 and peracetic acid.
    • Microbicidal and Sporicidal activity by oxidative damage at low temperature, < 50 C, 1 hour.
    • Used in sterilisation of surgical instruments, heat labile, disposables and corrosive metals.
    • Biological indicators- Geobacillus stearothermophiles and Bacillus subtilis.
  • Heavy Metals-
    • Damages cell membranes and inactivates proteins.
    • Silver sulphadiazine used for burns
    • Silver nitrate used for Ophthalmia neonatorum
  • Dyes-
    • Inhibition of cell wall, protein & nucleic acid synthesis
    • Aniline (base) -> Gentian violet & methyl violet
    • Acridine (acidic) -> acridine, acroflavin

Testing of Disinfectants

  • Phenol Coefficient Test-
    • Also known as Rideal Walker Test
    • Ratio= Highest dilution of test disinfectant, which kills S.typhi in a given time / highest dilution of phenol which kills S.typhi in same time.
    • If Ratio > 1, the test disinfectant is less effective than phenol
    • Only phenolic compound can be tested.
    • Does not assess the activity of disinfectant in presence of organic matter
  • Chick Martin Test
    • Modification of Phenol Coefficient Test
    • Test disinfectant acts in the presence of organic matter like feces.
  • Kelsey-Sykes Test
    • Capacity test
    • Tests the capability of test disinfectant to retain its activity over increasing load of microbes
  • Kelsey and Maurer Test-
    • Whether the chosen disinfectant is effective for use in the hospital or not.
    • Also called Maurer’s In-use Test
    • Most common contaminant- Pseudomonas.