Microbial growth control170713022721.ppt

Control of Microbial Growth:
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

Definitions
Sterilization: Killing or removing all forms of
microbial life (including endospores) in a material
or an object.
Heating is the most commonly used method of
sterilization.
Commercial Sterilization: Heat treatment that
kills endospores of Clostridium botulinum the
causative agent of botulism, in canned food.
Does not kill endospores of thermophiles, which are
not pathogens and may grow at temperatures above
45o
C.

Definitions
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.
May use physical or chemical methods.
 Disinfectant: Applied to inanimate objects.
 Antiseptic: Applied to living tissue (antisepsis).
 Degerming: Mechanical removal of most microbes in a
limited area. Example: Alcohol swab on skin.
 Sanitization: Use of chemical agent on food-handling
equipment to meet public health standards and minimize
chances of disease transmission. E.g: Hot soap & water.

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Definitions
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.

Definitions
Bacteriostatic Agent: An agent that inhibits the
growth of bacteria, but does not necessarily kill
them. Suffix stasis: To stop or steady.
Germicide: An agent that kills certain
microorganisms.
 Bactericide: An agent that kills bacteria. Most do not
kill endospores.
 Viricide: An agent that inactivates viruses.
 Fungicide: An agent that kills fungi.
 Sporicide: An agent that kills bacterial endospores of
fungal spores.

Rate of Microbial Death
When bacterial populations are heated or treated
antimicrobial chemicals, they usually die at a
constant rate.

Rate of Microbial Death
Several factors influence the effectiveness of
antimicrobial treatment.
1. Number of Microbes: The more microbes present, the more
time it takes to eliminate population.
2. Type of Microbes: Endospores are very difficult to destroy.
Vegetative pathogens vary widely in susceptibility to different
methods of microbial control.
3. Environmental influences: Presence of organic material
(blood, feces, saliva) tends to inhibit antimicrobials, pH etc.
4. Time of Exposure: Chemical antimicrobials and radiation
treatments are more effective at longer times. In heat treatments,
longer exposure compensates for lower temperatures.

Physical Methods of Microbial Control:
Heat: Kills microorganisms by denaturing their
enzymes and other proteins. Heat resistance varies
widely among microbes.
 Thermal Death Point (TDP): Lowest temperature at
which all of the microbes in a liquid suspension will be
killed in ten minutes.
 Thermal Death Time (TDT): Minimal length of time in
which all bacteria will be killed at a given temperature.
 Decimal Reduction Time (DRT): Time in minutes at
which 90% of bacteria at a given temperature will be
killed. Used in canning industry.

Physical Methods of Microbial
Control:
Moist Heat: Kills microorganisms by coagulating
their proteins.
In general, moist heat is much more effective than
dry heat.
 Boiling: Heat to 100o
C or more at sea level. Kills vegetative
forms of bacterial pathogens, almost all viruses, and fungi
and their spores within 10 minutes or less. Endospores and
some viruses are not destroyed this quickly. However brief
boiling will kill most pathogens.
 Hepatitis virus: Can survive up to 30 minutes of boiling.
 Endospores: Can survive up to 20 hours or more of boiling.

Control:
Moist Heat (Continued):
Reliable sterilization with moist heat requires
temperatures above that of boiling water.
 Autoclave: Chamber which is filled with hot steam under
pressure. Preferred method of sterilization, unless material is
damaged by heat, moisture, or high pressure.
 Temperature of steam reaches 121o
C at twice atmospheric pressure.
 Most effective when organisms contact steam directly or are contained
in a small volume of liquid.
 All organisms and endospores are killed within 15 minutes.
 Require more time to reach center of solid or large volumes of liquid.

Autoclave: Closed Chamber with High
Temperature and Pressure

Control:
Moist Heat (Continued):
 Pasteurization: Developed by Louis Pasteur to prevent the
spoilage of beverages. Used to reduce microbes responsible for
spoilage of beer, milk, wine, juices, etc.
 Classic Method of Pasteurization: Milk was exposed to 65o
C for 30
minutes.
 High Temperature Short Time Pasteurization (HTST): Used today.
Milk is exposed to 72o
C for 15 seconds.
 Ultra High Temperature Pasteurization (UHT): Milk is treated at
140o
C for 3 seconds and then cooled very quickly in a vacuum chamber.
 Advantage: Milk can be stored at room temperature for several
months.

Control:
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 170o
C for sterilization. Dry heat is
transfers heat less effectively to a cool body, than moist heat.

Control:
Filtration: Removal of microbes by passage of a liquid
or gas through a screen like material with small pores.
Used to sterilize heat sensitive materials like vaccines,
enzymes, antibiotics, and some culture media.
 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. Don’t retain
spirochetes, mycoplasmas and viruses.
 0.01 um Pores: Retain all viruses and some large proteins.

Control:
Low Temperature: Effect depends on microbe and
treatment applied.
 Refrigeration: Temperatures from 0 to 7o
C. Bacteriostatic
effect. Reduces metabolic rate of most microbes so they
cannot reproduce or produce toxins.
 Freezing: Temperatures below 0o
C.
 Flash Freezing: Does not kill most microbes.
 Slow Freezing: More harmful because ice crystals disrupt cell
structure.
 Over a third of vegetative bacteria may survive 1 year.
 Most parasites are killed by a few days of freezing.

Control:
Radiation: Three types of radiation kill microbes:
1. 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.
Disadvantages: Penetrates human tissues. May cause genetic
mutations in humans.

Control:
Radiation: Three types of radiation kill microbes:
2. 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, eyes. Doesn’t
penetrate paper, glass, and cloth.

Chemical Methods of Microbial Control
Types of Disinfectants
1. 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).
 Biphenols (pHisoHex): Effective against gram-positive 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 times after applied, and
remain active in the presence of organic compounds.

2. Halogens: Effective alone or in compounds.
A. Iodine:
 Tincture of iodine (alcohol solution) was one of first antiseptics
used.
 Combines with 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

2. Halogens: Effective alone or in compounds.
B. Chlorine:
 When mixed in water forms hypochlorous acid:
Cl2 + H2O ------> H+ + Cl- + HOCl
Hypochlorous acid
 Used to disinfect drinking water, pools, and sewage.
 Chlorine is easily inactivated by organic materials.
 Sodium hypochlorite (NaOCl): Is active ingredient of bleach.
 Chloramines: Consist of chlorine and ammonia. Less
effective as germicides.

Chemical Methods of Control
3. 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 cause proteins to
coagulate.
 Ethanol: Drinking alcohol. Optimum concentration is 70%.
 Isopropanol: Rubbing alcohol. Better disinfectant than ethanol.
Also cheaper and less volatile.

4. 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 until recently.
B. Mercury
 Organic mercury compounds like merthiolate and
mercurochrome are used to disinfect skin wounds.
C. Copper
 Copper sulfate is used to kill algae in pools and fish tanks.

4. Heavy Metals:
D. Selenium
 Kills fungi and their spores. Used for fungal infections.
 Also used in dandruff shampoos.
E. Zinc
 Zinc chloride is used in mouthwashes.

5. Quaternary Ammonium Compounds (Quats):
 Widely used surface active agents.
 Cationic (positively charge) detergents.
 Effective against gram positive bacteria, less effective against
gram-negative bacteria.
 Also destroy fungi, amoebas, and enveloped viruses.
 Zephiran, Cepacol, also found in our lab spray bottles.
 Pseudomonas strains that are resistant and can grow in presence of
Quats are a big concern in hospitals.
 Advantages: Strong antimicrobial action, colorless, odorless,
tasteless, stable, and nontoxic.
 Diasadvantages: Form foam. Organic matter interferes with
effectiveness. Neutralized by soaps and anionic detergents.

6. Aldehydes:
 Include some of the most effective antimicrobials.
 Inactivate proteins by forming covalent crosslinks with
several functional groups.
A. 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.
 Also used in mortuaries for embalming.

6. Aldehydes:
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.
 Sporicidal in 3 to 10 hours.
 Commonly used to disinfect hospital instruments.
 Also used in mortuaries for embalming.

7. Gaseous Sterilizers:
 Chemicals that sterilize in a chamber similar to an autoclave.
 Denature proteins, by replacing functional groups with alkyl
groups.
A. Ethylene Oxide:
 Kills all microbes and endospores, but requires exposure of 4 to
18 hours.
 Toxic and explosive in pure form.
 Highly penetrating.
 Most hospitals have ethylene oxide chambers to sterilize
mattresses and large equipment.

8. Peroxygens (Oxidizing Agents):
 Oxidize cellular components of treated microbes.
 Disrupt membranes and proteins.
A. Ozone:
 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.
 Highly reactive form of oxygen.
 Made by exposing oxygen to electricity or UV light.

B. Hydrogen Peroxide:
 Used as an antiseptic.
 Not good for open wounds because quickly broken down by
catalase present in human cells.
 Effective in disinfection of inanimate objects.
 Sporicidal at higher temperatures.
 Used by food industry and to disinfect contact lenses.
C. Benzoyl Peroxide:
 Used in acne medications.

D. Peracetic Acid:
 One of the most effective liquid sporicides available.
 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.

Efficiency of Different Chemical
Antimicrobial Agents

Microbial growth control170713022721.ppt

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