Microbial Control

• Sterilization

  Destruction of all forms of life including bacterial spores
• Disinfection

  Destruction of some forms of life except bacterial spores
• Disinfectant


  • Applied to inanimate objects
• Antiseptic

  • Applied to skin; cannot be used as disinfectant
• Physical methods of sterilization

  • Moist heat

  1. Autoclave (Sterilization, Decontamination)
  2. Tyndallization
  3. Inspissation

  • Dry heat

  1. Direct Flame
  2. Dry/Hot air oven
  3. Incineration

  • Ionizing radiation
  • Filtration
• Chemical methods of sterilization

  • Peracetic acid
  • Formaldehyde vapor/Vapor phase H2O2
  • Glutaraldehyde
  • Ethylene oxide (ETO) gas
• Physical methods of disinfection

  • Boiling
  • Pasteurization
  • Non-ionizing radiation
• Chemical methods of disinfection

  • Alcohol
  • Halogens
  • Heavy metals (Hg, Cu, Ag)
  • Quaternary ammonium compounds (QUATS) (Zephiram, Cepacol)
  • Phenol
• Autoclave


  1. Operates based on the principle of steam under pressure

  2. Effective indication: 121 °C for 15lbs/in2 for 15 minutes for sterilization, 135 °C for 30lbs/in2 for 30 minutes for decontamination

  3. Biological indicator: Bacillus stearothermophilus

  
• Tyndallization

  1. Fractional discontinuous sterilization, effective indication: 100 °C for 30-60 minutes (commonly used to destroy heat-resistant endospores)

  2. Instrument: Arnold's Sterilizer
• Inspissation

  1. Thickening through evaporation, effective indication: 75-80 °C for 2 hours

  2. Instrument: Inspissator
• Dry heat

  • Oxidation of bacterial components
• Dry heat sterilization methods

  • Direct flame
  • Dry/hot air oven
  • Incineration

  
• Ionizing radiation

  works by alkylation of nucleic acid of bacteria using high energy short wavelength deep penetrating gamma rays; used for heat sensitive materials

  Biological indicator: Bacillus pumilis
• Filtration

  Based on membrane gradient by differences in particle size; used for the sterilization of heat sensitive materials
• Pore sizes for filtrationfor Water/Liquid solutions/Antibiotics/Vaccines


  • usually uses a thin membrane filter of cellulose acetate with different pore size depending on the intended purpose:
  • -0.45-0.80 μm - retains most bacteria, yeasts, and molds
  • -0.22 μm - used to filter Pseudomonas-like organisms, used for critical sterilization of parenteral solutions
  • -0.01 μm - able to retain small viruses
• Air: (HEPA filter)High Efficiency Particulate Air Filter 


  Has a pore size of 0.3 μm, used in Biological Safety Cabinet (BSC) and rooms of immunocompromised patients
• Boiling

  Destroys vegetative cells of bacteria but not their spores

  Effective indication: 100 °C for 15 - 30 minutes
• Pasteurization


  used for the preservation of alcoholic beverages such as beers, wines, and also dairy products such as milks and yogurt
• Pasteurization methods

  • Batch: 62.5 °C for 30 minutes
  • Flash: 72 °C for 15 seconds
  • Ultra High Temperature (UHT): 72 °C - 110 °C - 72 °C for 5 seconds
• Non-ionizing radiation

  Uses low energy long wavelength ultraviolet rays to disinfect heat sensitive materials as well as large spaces
• Alcohol

  Mechanism of action: dehydration, lipid dissolution, and protein denaturation 70% Alcohol not 90%:

  • Minimum Contact Time; 1-2 minutes or until completely evaporated
• Halogens


  Mechanism of action: inhibits protein function and acts as strong oxidizing agents

  • Chloride (Cl) in NaOCl: used as disinfecting agents in many laboratory and hospitals spaces, surfaces, and also in treating water for portability

  • Iodine (I2) in Betadine used as a household antiseptics and surgical antiseptics
• Heavy metals


  Mechanism of action: denaturation of enzymes and other essential bacterial proteins
• Heavy metals used as disinfectants

  • Mercury (Hg) - active ingredient or merthiolate but this is already banned in the market due to its known toxicity
  • Copper (Cu) - CuSO4 crystals are used as algaecide in swimming pools and aquarium
  • Silver (Ag) - 1% AgNO3 – used as prophylactic agent in Crede’s Prophylaxis in suspected cases of Ophthalmia neonatorum
• Quaternary ammonium compounds (QUATS)

  Mechanism of action: enzyme inhibition, protein denaturation, and disruption of plasma membrane
• QUATS used as disinfectants

  • Zephiran: Benzalkonium chloride
  • Cepacol: Cetylpyridium chloride
• Phenol/Phenolic compounds/Bisphenols

  Mechanism of action: plasma membrane destruction and enzyme denaturation
• Chemical agents commonly used as disinfectants and antiseptics

  • Alcohols
  • Aldehydes
  • Halogens
  • Heavy metals
  • Detergents
  • Phenolics
  • Gases
• Autoclave


  Principle: heat under steam pressure

  • It is at this temperature in which all microorganisms (except for prions) and their endospores are destroyed within approximately 15 minutes

  • 121 °C, 15 psi for 15 minutes – recommended for media, liquids, utensils, glass pipettes

  • 132 °C, 15 psi for 30-60 minutes – recommended for decontaminating medical wastes
• Steps in running an autoclave Step 1 to 5


  • Steps depend on the autoclave type and the manufacturer. The ff. are the general procedures

  1. Check the inside of the autoclave if there are items left from the previous cycle

  2. Place sufficient amount of water inside the chamber

  3. The materials to be sterilized are then placed inside

  4. The lid is closed using screws to ensure airtight condition, and the electric heater is switched on 5. The water then boils and the pressure is allowed to reach the designated level
• Biologic indicators


  Used to monitor the effectiveness of sterilization processes
• Ionizing radiation

  causes mutation in the DNA; destroys vegetative cells and endospores

  • Used to “pasteurize” meat products

  • Utilizes gamma rays and x-rays
• Non-ionizing radiation


  Damages cellular DNA

  • Used on exposed surfaces and rooms

  • Utilizes ultraviolet rays
• Phenols


  First widely used antiseptic and disinfectant, destroys plasma membranes and denatures cell proteins

  5% phenol with 10-30 minutes contact time is effective against mycobacteria
• Alcohol

  Denatures proteins and causes dissolution of lipid membranes, effective at 60-90% concentration
• Chlorine (Hypochlorite)


  Used at 1:10 dilution, 3 minutes contact time
• Preparation of disinfectants


  Use the dilution formula C1V1=C2V2
• Moist Heat

  coagulation of bacterial proteins including bacterial enzymes
• Control of Microorganism Using Heat Methods: Boiling water (steam)

  Temperature (in Celsius): 100

  Time required: 15 minutes

  Applications: Kills microbial vegetative form; endospores survive