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Microbiology 201 - Chapter

Physical and Chemical Control of Microorganisms

Principles of microbial control

Chemical antimicrobial agents

Physical antimicrobial agents

Principles

Why do we need to control microorganisms?

• Prevention of infections and spread of disease caused by pathogens (viruses, bacteria, fungi, spores, eggs, cysts)

-Bacterial endospores are most resistant to destruction. If you kill them you can assume all others have been destroyed

• Prevention of food spoilage
• Prevention of deterioration of manufactured goods, building materials, odors
• Microbial control includes:

1. Killing microorganisms

2. Inhibiting microbial growth or metabolism

3. Physically removing microorganisms

Terminology

Disinfection Destruction of pathogenic nonsporulating microorganisms or their toxins

Disinfectant Agent applied to disinfect objects

-Dishes
-Counter tops
-Lavatories

Antiseptic Agent applied to disinfect living tissue

-Wounds
-Skin prior to injections or incisions

Sterilization Killing or removal of all microorganisms (including spores and viruses)

-Surgical instruments, syringes, dental equipment
-Injected drugs
-Catheters and surgical implants

Bacteriostatic Effect of an agent that inhibits bacterial growth (organism is not killed)

Bactericidal Effect of an agent that kills bacteria

Bactericide Agent that kills bacteria

Sporocide Agent that kills spores (bacterial endospores and fungal spores)

Fungicide Agent that kills fungi

Viricide Agent that destroys viruses

Viable Capable of living, germinating or reproducing under favorable conditions

Nonviable Dead, or unable to infect a cell if a virus

Factors that affect rate of microbial death

1. Degree of contamination (microbial load)

• Time needed to sterilize something depends on the number of organisms present (Fig. 11.2b)

Ex. Sterilization of surgical instruments with heat

Y axis =Logarithim of the number of viable microorganisms

Exposure Time

-Red line shows that more organisms = longer time needed to achieve sterility

2. Susceptibility of microorganism to antimicrobial agents (Fig. 11.2c)

• Phase of growth

-Rapidly dividing cells are easier to kill (blue line)

-Spores and inactive cells are more resistant (red line)

Exposure Time

• Species

-Some organisms are resistant to particular killing agents

Ex. Deinococcus radiodurans is resistant to radiation

3. Temperature and pH

Ex. Killing effect of ethanol may be greater at high temperature or at high or low pH

4. Concentration or dosage of antimicrobial agent

Higher concentration usually more effective, but one exception is that 70% ethanol is more effective than 100%

5. Presence of material that interferes with the action of the agent

Ex. blood, saliva, fat and oil, dirt and debris

Targets of Antimicrobial Agents

1. Proteins Disruption of structure and chemical modification of enzymes and structural proteins (Fig. 11.4)

2. Lipids Disruption of membranes (Fig. 11.3)

3. Nucleic acids Denaturation and chemical modification of bases

1. Soaps and detergents (Fig. 11.14)

• Physically removes organisms from:

Skin, food utensils, clothing, bedding

• Kills some microorganisms by disrupting membranes

Ex. Streptococcus and flu virus

2. Acids and bases

• Low or high pH kills or inhibits microorganisms

Ex. Lactic, acetic, and benzoic acid are food preservatives

3. Halogens (Chlorine and Iodine)

Ex. Chlorine bleach: food utensils, dairy equipment, clothing, bedding

Iodine: skin antiseptic for wounds and prior to surgery; also used by campers to purify drinking water

4. Alcohols

Ex. Ethanol and isopropanol (rubbing alcohol)

Skin prior to injection or blood withdrawal

5. Phenols (Fig. 11.12)

Disrupt cell membranes, denature proteins

Ex. Orthophenylphenol (in Lysol) Used in hospitals and bathrooms

Triclosan is in toothpaste, mouthwash, kitty litter, antibacterial soap

6. Oxidizing agents

Ex. Hydrogen peroxide H₂O₂ Good penetation of puncture wounds

7. Alkylating agents

React chemically with proteins and nucleic acids

Ex. Formaldehyde CH₂O Inactivates viruses and toxins

Ethylene oxide A gas and has good penetration of bedding

Highly toxic to humans, special equipment is needed (Fig. 11.18)

8. Heavy metals

Mercury (Hg), silver (Ag), gold (Au), copper (Cu), arsenic (As), zinc (Zn)

Inactivate proteins

High toxicity to humans limits use

9. Other agents

• Natural plant products

Thymol from thyme (a phenol)

Topical antiseptic

Internally as antihelmintic

Eugenol from cloves (also a phenol)

Dental antiseptic and analgesic

• Sulfites (SO₃^--) and nitrites (NO₂^--)

-Food preservatives (vegetables, wine, meat)

Agent                                                Mode of Aciton

Heat	Denatures proteins, nucleic acids and organic metabolites; disrupts membranes
Refrigeration	Slows growth and metabolism
Drying	Lack of water hinders metabolism and growth
Irridiation	High energy atomic particles or waves promote chemical reactions that damage proteins, nucleic acids, membranes and other organic compounds
Filtration	Physically reomves microorganisms from gases and liquids

Heat Sterilization

1. Dry heat

• Oven: 171^OC --1 hr, 160^OC -- 2 hr, or 121^OC --16 hr
  
  Metal, glass, powders and oils

• Flame

Inoculating loops and mouth of culture tubes,, flasks and bottles

2. Moist heat (steam, boiling water)

• Good penetration
• Pressure increases boiling point of water above 100^OC
• Autoclave Uses steam and high temperature (Fig. 11.5)

15 lb/in2, 121^OC, 15-20 min

Kills all microorganisms (cells, spores and viruses)

• Sterilization indicators included along with stuff being sterilized

Ensure that proper sterilization conditions were achieved

Indicator tape. "STERILE" appears when conditions were met

Endospore strip. Contains bacterial endospores. If sterilization conditions were not achieved, endospores will germinate and grow (indicated as turbidity in the medium after incubation--this takes time but is the ultimate test of sterility because endospores are the hardest to destroy)

Pasteurization

• Invented by Louis Pasteur to preserve wine
• Uses temperature below 100^OC
• Foods are heated to kill pathogens and reduce the number of spoilage organisms

Temperature is not high enough to "cook" the food

Preserves natural taste and texture

• Not sterilization. Nonpathogenic organisms and spores may still be alive
• Used to kill pathogens in heat sensitive food like milk, cheese and beer

Ex.

Listeria

Salmonella

Mycobacterium tuberculosis in raw milk killed by:

Flash method 71.6^OC for 15 sec

Holding method 62.9^OC for 30 min

Refrigeration

• 5^OC or freezing at -20^OC or lower
• Prevents spoilage of foods
• Slows growth and enzyme catalyzed reactions
• Doesn't kill microorganisms

Drying

• Used for fruits, vegetables, grains, meat and fish

Freeze-drying (lyophylization)

• Food is frozen, then placed under a vacuum

-Water evaporates, food stays frozen until dry

-Preserves color and flavor

• Used for instant coffee, fruit and other foods

Radiation (Fig. 11.17)

• Ultraviolet light (Fig. 11.7c, 11.9 and 11.10)

-Damages nucleic acids

-Doesn't penetrate glass, clothing, paper

-Effective against airborne pathogens in hospitals and animal holding facilities

-Reduces number of organisms on surfaces

Ex. in operating rooms

• Ionizing radiation: x-rays, gamma rays (Fig. 11.17a and 11.18)

-Sterilizes foods like milk

-Kills pathogens in meat

Ex. Salmonella and Campylobacter on chicken

• Microwave radiation

-Raises the temperature of things that contain water

-Endospores resistant as they contain very little water

Filtration

• Removes microorganisms by passage of solution or gas through a filter
• For heat-sensitive materials

Ex. Growth media with heat sensitive nutrients or antibiotics

Some beer is filtered rather than pasteurized

• Membrane filters can have various pore sizes that remove different sizes of microorganisms (Fig. 11.11)
• High-efficiency particulate air filters (HEPA) are used in ventilation systems in hospitals and some industrial manufacturing facilities

Osmotic pressure

• High concentrations of salt or sugar preserve food

Ex. Salted meat, jelly

• Causes water to move by osmosis through the membrane out of cell
• Water is the solvent for dissolved substances in cell and is necessary fro metabolism

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Last updated: March 6, 2007 /jdh