Control%20of%20Microbial%20Growth - PowerPoint PPT Presentation

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Title: Control%20of%20Microbial%20Growth


1
Control of Microbial Growth
2
  • Introduction
  • Early civilizations practiced salting, smoking,
    pickling, drying, and exposure of food and
    clothing to sunlight to control microbial growth.
  • Use of spices in cooking was to mask taste of
    spoiled food. Some spices prevented spoilage.
  • In mid 1800s Semmelweiss and Lister helped
    developed aseptic techniques to prevent
    contamination of surgical wounds.
  • Before then
  • Nosocomial infections caused death in 10 of
    surgeries.
  • Up to 25 mothers delivering in hospitals died
    due to infection

3
  • Definitions
  • Sterilization Killing or removing all forms of
    microbial life (including endospores) in a
    material or an object.
  • Disinfection Reducing the number of pathogenic
    microorganisms to the point where they no longer
    cause diseases. Usually involves the removal of
    vegetative or non -endospore forming pathogens.

4
  • Sepsis Comes from Greek for decay or putrid.
    Indicates bacterial contamination.
  • Asepsis Absence of significant contamination.
  • Aseptic techniques are used to prevent
    contamination of surgical instruments, medical
    personnel, and the patient during surgery.
  • Aseptic techniques are also used to prevent
    bacterial contamination in food industry.

5
  • Disinfectant Applied to inanimate objects.
  • Antiseptic Applied to living tissue
    (antisepsis).
  • Degerming Mechanical removal of most microbes in
    a limited area.
  • Sanitization Use of chemical agent on
    food-handling equipment to meet public health
    standards and minimize chances of disease
    transmission.

6
General Considerations in Sterilization
Disinfection
  • 1) Population unknown.
  • 2) Determination of level of kill.
  • 3) Heterogeneity within population.
  • 4) Troublesome forms.
  • 5) Environmental factors.
  • 6) No Universal Method.
  • 7) Careless handling.
  • 8) Effect on Bacteria as the principal criteria.
  • 9) Critical, semi-critical, and non critical
    items.

7
  • Physical Methods of S/D
  • Heat
  • Kills microorganisms by denaturing their enzymes
    and other proteins.
  • Heat resistance varies widely among microbes.
  • Thermal Death Point (TDP) the temperature at
    which all of the test microorganisms in a 24 hour
    broth culture are killed in ten minutes.
  • Thermal Death Time (TDT) the time required to
    kill all microorganisms of a known population at
    a given temperature under specific conditions.
  • Decimal Reduction Time (DRT) Time in minutes at
    which 90 of bacteria at a given temperature will
    be killed.

8
  • Moist Heat
  • Kills microorganisms by coagulating their
    proteins.
  • In general, moist heat is much more effective
    than dry heat.
  • Boiling Heating to 100oC at sea level. Kills
    vegetative forms of bacterial pathogens, almost
    all viruses, and fungi and their spores within 10
    minutes or less.
  • - Hepatitis B virus Can survive up to 30
    minutes of
  • boiling.
  • - Endospores Can survive up to 20 hours or
    more of
  • boiling.

9
  • Reliable sterilization with moist heat requires
    temperatures above that of boiling water.
  • Steam at atmospheric pressure
  • Steam at 100C has 540 calories of latent heat as
    compared to the 80 calories for boiling water at
    that temperature.
  • - Used to disinfect pipes, vats, in diaries and
    breweries.

10
  • Steam under pressure( Autoclaving)
  • At a temperature of 121C of moist heat for 15
    minutes spores are destroyed.
  • In order to obtain this T saturated steam is
    placed under a pressure of 15 Psi (15 libra/inch2
    or 1.05 kg/cm2).
  • Pressure (Psi) TC Time (minutes)
  • 15 121 15
  • 20 126 10
  • 30 134 3

11
Autoclave Closed Chamber with High Temperature
and Pressure
12
  • Pasteurization Developed by Louis Pasteur to
    prevent the spoilage of beverages. Used to
    reduce microbes responsible for spoilage of milk,
    juices, beer, wine, etc.
  • Classic Method of Pasteurization exposure to a
    T of 65oC for 30 minutes.
  • High Temperature Short Time Pasteurization
    (HTST) Used today. Exposure to a T of 72oC for
    15 seconds.
  • Ultra High Temperature Pasteurization (UHT)
    Exposure to a T of 140oC for 3 seconds and then
    cooling very quickly in a vacuum chamber.
    Advantage can be stored at room temperature for
    several months.
  • Fractional sterilization (Tyndalization )

13
  • Dry Heat Kills by oxidation effects.
  • Direct Flaming Used to sterilize inoculating
    loops and needles. Heat metal until it has a red
    glow.
  • Incineration Effective way to sterilize
    disposable items (paper cups, dressings) and
    biological waste.
  • Hot Air Sterilization Place objects in an oven.
    Require 2 hours at 170oC for sterilization. Dry
    heat transfers heat less effectively to a cool
    body, than moist heat.

14
  • Low Temperature
  • Effect depends on microbe and treatment applied.
  • Refrigeration Temperatures from 2o to 8oC.
    Bacteriostatic effect. Reduces metabolic rate of
    most microbes so they cannot reproduce or produce
    toxins.
  • Freezing Temperatures below 0oC.
  • Flash Freezing Does not kill most microbes.
  • Slow Freezing More harmful because ice crystals
    disrupt cell structure.
  • Freeze- drying (lyophilization)
  • Over a third of vegetative bacteria may survive
    1 year.
  • Most parasites are killed by a few days of
    freezing.

15
  • Filtration
  • Used to sterilize heat sensitive materials like
    vaccines, enzymes, antibiotics, and some culture
    media.
  • Removal of microbes by passage of a liquid or gas
    through a screen like material with small pores.
  • High Efficiency Particulate Air Filters (HEPA)
    Used in operating rooms and burn units to remove
    bacteria from air.
  • Membrane Filters Uniform pore size. Used in
    industry and research. Different sizes
  • 0.22 and 0.45um Pores Used to filter most
    bacteria. Dont retain spirochetes, mycoplasmas
    and viruses.
  • 0.01 um Pores Retain all viruses and some large
    proteins.

16
  • Ultrasonication
  • High frequency electrical energy ? transducers ?
    high frequency sound waves (mechanical)
  • High frequency sound waves are above the human
    auditory level.
  • - Cell disintegration
  • - Sterilization
  • - Cleansing purposes (ultrasonic cleaners) to
    remove soil and organic matter esp. from
    glassware.

17
  • Desiccation
  • In the absence of water, microbes cannot grow or
    reproduce, but some may remain viable for years.
  • Susceptibility to desiccation varies widely
  • Neisseria gonnorrhea Only survives about one
    hour.
  • Mycobacterium tuberculosis May survive several
    months.
  • Viruses are fairly resistant to desiccation.
  • Clostridium spp. and Bacillus spp. May survive
    decades.

18
Radiation Forms of Radiation
19
  • Radiation
  • Three types of radiation kill microbes
  • Ionizing Radiation Gamma rays, X rays, electron
    beams, or higher energy rays.
  • - Have short wavelengths (less than 1 nanometer).
  • - Dislodge electrons from atoms and form ions.
  • - Cause mutations in DNA and produce peroxides.
  • - Used to sterilize pharmaceuticals and
    disposable medical supplies.
  • - Food industry is interested in using ionizing
    radiation.

20
  • Their microbicidal action is due to the
    ionization of water.
  • Ionized water interacts with nonionized water to
    produce hydroxyl radicals
  • Radiation
  • H2O ---------? H2O e
  • H2O H2O ---? OH- H3O
  • - Disadvantages Penetrates human tissues. May
    cause genetic mutations in humans.

21
  • Ultraviolet light (Nonionizing Radiation)
  • Wavelength is longer than 1 nanometer.
  • Damages DNA by producing thymine dimers, which
    cause mutations.
  • Used to disinfect operating rooms, nurseries,
    cafeterias.
  • Disadvantages Damages skin and eyes. Does not
    penetrate paper, glass, and cloth.

22
  • Microwave Radiation
  • - Wavelength ranges from 1 millimeter to 1
    meter.
  • Heat is absorbed by water molecules.
  • May kill vegetative cells in moist foods.
  • Bacterial endospores, which do not contain water,
    are not damaged by microwave radiation.
  • Solid food is unevenly penetrated by microwaves.
  • Trichinosis outbreaks have been associated with
    pork cooked in microwaves.

23
Chemical Methods of S/D
  • Qualities of a universal disinfectant
  • Destroys all forms within a practical period of
    time
  • Not irritant, allergenic or toxic
  • Makes effective contact and not bypassed by
    dilution
  • Readily soluble
  • Chemically stable, no disagreeable odor, non
    discoloring and leaves no stains
  • Not expensive

24
  • Important Factors
  • Characteristics of microorganisms (chemical make
    up)
  • Concentration of disinfectant
  • PH of disinfectant (neutral to slightly acidic)
    affects degree of ionization
  • Interfering matter
  • 5) Time

25
Types of Disinfectants
  • Phenols and Phenolics
  • Phenol (carbolic acid) was first used by Lister
    as a disinfectant.
  • Rarely used today because it is a skin irritant
    and has strong odor.
  • Used in some throat sprays and lozenges.
  • Acts as local anesthetic.
  • Phenolics are chemical derivatives of phenol
  • Cresols Derived from coal tar (Lysol).
  • Bisphenols (Hexachlorophene) Effective against
    gram-positive bacteria like staphylococci and
    streptococci. Used in nurseries. Excessive use
    in infants may cause neurological damage.
  • Destroy plasma membranes and denature proteins.
  • Advantages Stable, persist for long time after
    application, and remain active in the presence of
    organic compounds.

26
  • Alcohols
  • Kill bacteria, fungi, but not endospores or naked
    viruses.
  • Act by denaturing proteins and disrupting cell
    membranes.
  • Evaporate, leaving no residue.
  • Used to mechanically wipe microbes off skin
    before injections or blood drawing.
  • Not good for open wounds, because it causes
    proteins to coagulate.
  • Ethanol Optimum concentration is 70.
  • Isopropanol Rubbing alcohol. Better
    disinfectant than ethanol. Also cheaper and less
    volatile.

27
  • Halogens
  • Effective alone or in compounds.
  • A. Iodine
  • Tincture of iodine (alcohol solution) was one of
    first antiseptics used.
  • - Combines with the amino acid tyrosine in
    proteins and denatures proteins.
  • - Stains skin and clothes, somewhat irritating.
  • Iodophors Compounds with iodine that are slow
    releasing, take several minutes to act. Used as
    skin antiseptic in surgery. Not effective
    against bacterial endospores.
  • Betadine, Isodine, Wescodyne, Prepodyne.

28
  • B. Chlorine
  • When mixed with water it forms hypochlorous acid
  • Cl2 H2O ------gt H Cl- HOCl
  • Used to disinfect drinking water, pools, and
    sewage.
  • Chlorine is easily inactivated by organic
    materials.
  • Sodium hypochlorite (NaOCl) Is the active
    ingredient of bleach.
  • Chloramines Consist of chlorine and ammonia.
    Less effective as germicides.

29
  • Surfactants
  • Property of concentrating at interfaces
  • Posses hydrophobic and hydrophilic groups
  • Lower surface tension and increase miscibility of
    molecules
  • A. Soap
  • Na or K salts of fatty acids
  • degerm skin mechanically

30
  • B- Detergents
  • Superior to soap because they do not form
    precipitates or deposits with water minerals
  • Anionic (COO-, SO4-)
  • Cationic (NH4, phosphonium, sulfonium)
  • Quaternary ammonium ion compounds four organic
    groups covalently linked to nitrogen
  • Interact with cell membrane of microorganisms.

31
  • Alkylating Agents
  • Substitute alkyl groups for the hydrogen of
    reactive groups of enzymes, nucleic acids, and
    proteins
  • Ethylene oxide
  • - Chemosterilizer
  • - Colorless, soluble in water and common
    organic solvents.
  • - Explosive (mixed with CO2 or N2)
  • - Toxic
  • - Slow action influenced by concentration of
    gas, relative
  • humidity, time of exposure, and
    temperature (4 hours at 50-
  • 56C and 6-12 hours at RT).
  • - Optimum humidity is 30.

32
  • Used to sterilize heat sensitive items.
  • Gas must be dissipated before the item can be
    used (24 hours).
  • Exposure time is reduced by 50 for each doubling
    of ETO concentration.
  • likewise the activity of ETO doubles with each T
    increase of 10C.
  • BetaPropiolactone Fumigant. (carcinogen)

33
  • Aldehydes
  • Include some of the most effective
    antimicrobials.
  • Inactivate proteins by forming covalent
    crosslinks with several functional groups.
  • Formaldehyde gas
  • Excellent disinfectant.
  • Commonly used as formalin, a 37 aqueous
  • solution.
  • Formalin was used extensively to preserve
    biological
  • specimens and inactivate viruses and
    bacteria in
  • vaccines.
  • Irritates mucous membranes, strong odor.

34
  • B. Glutaraldehyde
  • Less irritating and more effective than
    formaldehyde.
  • One of the few chemical disinfectants that is a
    sterilizing agent.
  • A 2 solution of glutaraldehyde (Cidex) is
    Bactericidal, tuberculocidal, and viricidal in 10
    minutes but sporicidal in 3 to 10 hours.
  • Commonly used to disinfect hospital instruments.

35
  • Heavy Metals
  • Include copper, selenium, mercury, silver, and
    zinc.
  • Oligodynamic action Very tiny amounts are
    effective.
  • A. Silver
  • 1 silver nitrate used to protect infants
    against gonorrheal eye infections (ophthalmia
    neonatorum) until recently.
  • B. Mercury
  • Organic mercury compounds like merthiolate and
    mercurochrome are used to disinfect skin wounds.

36
  • C. Copper
  • Copper sulfate is used to kill algae in pools
    and fish tanks.
  • D. Selenium
  • Kills fungi and their spores. Used for fungal
    infections.
  • Also used in dandruff shampoos.
  • E. Zinc
  • Zinc chloride is used in mouthwashes.
  • Zinc oxide is used as antifungal agent in paints.

37
  • Oxidizing Agents
  • Oxidize cellular components of treated microbes.
  • Disrupt membranes and proteins.
  • A. Ozone
  • Highly reactive form of oxygen.
  • Made by exposing oxygen to electricity or UV
    light.
  • Used along with chlorine to disinfect water.
  • Helps neutralize unpleasant tastes and odors.
  • More effective killing agent than chlorine, but
    less stable and more expensive.

38
  • B. Hydrogen Peroxide
  • Used as an antiseptic.
  • Not good for open wounds because it is quickly
    broken down by catalase present in human cells.
  • The active Oxidant is the free hydroxyl radical
  • Effective in disinfection (3-6) of inanimate
    objects.
  • Sporicidal (10-25) at higher temperatures.
  • Used by food industry and to disinfect plastic
    implants, contact lenses, and surgical
    prostheses.
  • Plasma Gas Vaporization of H2O2 with the
    generation of microwave or radiofrequency energy.
    It is a sterilant
  • C. Benzoyl Peroxide
  • Used in acne medications.

39
  • D. Peracetic Acid
  • One of the most effective sporicidal liquid
    available.
  • Generates acetic acid and Oxygen, both of which
    are nontoxic
  • Sterilant
  • Kills bacteria and fungi in less than 5 minutes.
  • Kills endospores and viruses within 30 minutes.
  • Used widely in disinfection of food and medical
    instruments because it does not leave toxic
    residues.

40
  • Chlorhexidine
  • - Antiseptic at 4
  • - Used for surgical scrub and hand washing.
  • - Hibiclens, Hibitane.
  • Parachlorometaxylenol (PCMX)
  • - Active against gram positive bacteria.
  • Triclosan
  • - Active against bacteria. It is a common
    antiseptic in deodorant soap.

41
  • High-Level disinfectants
  • Used for items involved with invasive procedure
    that cannot withstand sterilization procedures
    (certain types of endoscopes, surgical
    instruments with plastic or other components that
    cannot be autoclaved).
  • Disinfection of these and other items is most
    effective if treatment is preceded by cleaning
    the surface to remove organic matter.
  • Examples of high level disinfectants include
    treatment with moist heat, glutaraldehyde,
    hydrogen peroxide, peracetic acid, chlorine
    dioxide, and other chlorine compounds.

42
  • Intermediate Level disinfectants
  • Alcohols, iodophors, and phenolics are used to
    clean surfaces or instruments in which
    contamination with bacterial spores and other
    highly resilient organisms is unlikely
    (semicritical items).
  • Examples of items treated include flexible
    fiberoptic endoscopes, laryngoscopes, vaginal
    specula, anesthesia breathing circuits, and
    others.

43
  • Low-Level disinfectants
  • Quaternary ammonium compounds are used to treat
    noncritical instrument and devices such as blood
    pressure cuffs, ECG electrodes, and stethoscopes.
  • Although these items come into contact with
    patients, they do not penetrate through mucosal
    surfaces into sterile tissues.
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