Download Microbiology: A Systems Approach

LECTURES IN
MICROBIOLOGY
Control of Microbial Growth
LESSON 7
Sofronio Agustin
Professor
Lesson 7 Topics
Controlling Microorganisms:
 Physical Means
 Chemical Means
2
Overview
Various Microbial Control Methods
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Antimicrobial Action
-static agents- inhibit growth of microbes
Ex: bacteriostatic, fungistatic
-cidal agents - destroy or kill microbes
Ex: bactericidal, fungicidal
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Antimicrobial Resistance
 Highest resistance - bacterial spores and prions.
 Moderate resistance - some bacteria, protozoan
cysts, fungal (sexual) spores, naked viruses.
 Least resistance - most bacteria, fungal
(asexual) spores and hyphae, enveloped viruses,
yeast, protozoan trophozoites.
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Comparative Resistance
Relative resistance of bacterial endospores and vegetative cells
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Terms
Sterilization
Disinfection
Antisepsis
Sanitation
Degerming
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Sterilization
 Process of complete removal or
destruction of all viable microbes including
endospores.
 Example: Use of physical or chemical
agents on surgical instruments, and
commercially packaged foods.
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Disinfection
 Use of physical or chemical agent
(disinfectant) to destroy vegetative bacteria
and other microbes except endospores.
 Example: Use of bleach solution on bench
tops.
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Antisepsis
 Use of physical or chemical agent
(disinfectant) to destroy vegetative pathogens
on living organisms.
 Example: Scrubbing surgical patients with
chemical agents.
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Sanitization
Reduction of the number of microbes to
acceptable levels established by public
health standards.
Sanitization of utensils in restaurants.
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Degerming
 Physical removal or reduction of
microbes from human skin.
Ex: Washing of hands with germicidal
soap.
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Efficacy
 Number of microorganisms
 Target microbe - bacteria, fungi,
spores, viruses.
 Temperature and pH
 Concentration of agent
 Its mode of action
 Interfering substances - solvents,
debris, saliva, blood, feces.
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Factors
Factors that influence the effectiveness of antimicrobial agents
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Mode of Action
Effects on:
Cell wall
Cell membrane
Nucleic acid synthesis
Protein synthesis
Protein function
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Cell Wall
Effects on bacteria and fungi:
Block cell wall synthesis
Degrade cellular components
Destroy or reduce its stability
Ex: Penicillin, detergents, alcohols
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Cell Membrane
Effects on all microbes including
enveloped viruses:
Bind and penetrate membrane lipids
Loss of selective permeability resulting
in leakage of cytoplasmic contents.
Ex: Surfactants - surface active agents.
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Surfactant Action
The effect of surfactants on the cell membrane.
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Nucleic Acid Synthesis
Irreversible binding to microbial DNA results in:
Ceasation of transcription and translation
Mutations
Ex: Formaldehyde and ionizing radiation
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Protein Synthesis
Ribosome binding stops translation and
prevents peptide bond formation.
Ex: The antibiotic chloramphenicol
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Protein Function
Blockage of protein (enzyme) active sites
prevents binding of substrate.
Protein denaturation.
Ex:
Heat, acids, alcohols, phenolics,
metallic ions.
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Protein Function
Effects of heat, pH, and blocking agents on protein function.
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Physical Methods
Heat
Radiation
Filtration
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Heat
Moist heat:
Coagulation of proteins
Denaturation of proteins
Dry heat:
Dehydration
Denaturation
Oxidation (burning to ashes)
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Moist Heat
Boiling water
Tyndallization
Pasteurization
Steam and pressure
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Boiling Water
Boiling at 100 ˚C for 10-30 minutes kills most
non-spore forming pathogens.
Ex:
Boiling of baby bottles and unsafe
drinking water
Note: Hepatitis B virus needs 1 hour
boiling to be killed.
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Tyndallization
Intermittent sterilization using free-flowing
steam for 30 to 60 minutes.
Ex:
Used on heat-sensitive media, canned
foods.
Note: This will not destroy spores.
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Pasteurization
 Use of moist heat at :
- 63 ˚C for 30 minutes (Classical)
- 72 ˚C for 15 seconds (HTST)
- 140 ˚C for < second (UHT)
 Used in milk industry, wineries, breweries.
 Prevents transmission of milk-borne diseases caused by:
Salmonella, Campylobacter, Listeria, Mycobacteria.
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Steam and Pressure
 Used in media preparation and glassware

sterilization.
Ex: Autoclave and pressure cooker
 Autoclave setting at 121 ˚C, 15 p.s.i. pressure for
at least 15 minutes effectively destroys spores.
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Autoclave
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Dry Heat
Hot air
Incineration
Requires greater temperature and
exposure time than moist heat
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Hot Air
Ex: Hot air oven
Effective at 150˚C to 180˚C for 2-4 hrs
Useful for sterilization of glasswares
and oils
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Incineration
Destroys microbes to ashes or gas
Bunsen flame - up to 1870˚C
Ex: Sterilization of loops and needles.
Furnace - 800˚C to 6500˚C
Ex: Incineration of animal carcasses.
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Incinerator
An infrared incinerator using flame to burn or oxidize materials into ashes.
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Cold and Desiccation
Cold temperatures - reduce microbial
activity except psychrophiles.
Desiccation or dehydration - removal of
water stops metabolic activity of microbes.
 Lyophilization –a freeze drying method used to
preserve microbes and vaccines.
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Radiation
Ionizing:
- Gamma rays (High energy)
- X-rays (Intermediate energy)
- Cathode rays (Lowest energy)
Nonionizing:
Ultraviolet light
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Radiation Effects
Ionizing radiation:
Ejects orbital electrons from an atom
High energy - penetrates liquids and
solids effectively.
Nonionizing radiation:
Raises atoms to a higher energy level
Low energy - less penetrating
UV - formation of pyrimidine dimers
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Cellular Effects of Radiation
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Ultraviolet Radiation
UV radiation cause the formation of pyrimidine dimers on DNA.
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Applications of Radiation
Ionizing radiation:
Alternative sterilization method
Materials sensitive to heat or chemicals
Some foods (fruits, vegetables, meats)
Nonionizing radiation:
Alternative disinfectant
Germicidal lamp in hospitals, schools, food
preparation areas (inanimate objects, air, water)
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Gamma Irradiation
(a) Gamma radiation machine used to sterilize fruits,
vegetables, meats, fish, and spices (b) Radora symbol
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UV Irradiation
UV treatment system used to disinfect water.
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Filtration
Removes microbes and spores from liquids
and air
Perforated membrane of varying pore sizes v
(0.22 to 0.45 um)
Applications:
Liquids that are sensitive to heat (serum,
vaccines, media)
HEPA filtration of operating rooms etc.
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Membrane Filter
(a) Membrane filtration system.
(b) Membrane filter close-up.
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Chemical Methods
Chemical agents commonly used in healthcare
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Classes of Chemical Agents
Halogens
Phenolics
Surfactants
Hydrogen peroxide
Detergents and soaps
Heavy metals
Aldehydes
Gases
Dyes, acids, and alkali
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Halogens
Chlorine -disinfectant and antiseptic
Disrupt sulfhydryl groups in amino acids
Iodine - topical antiseptic
Disruption is similar to chlorines
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Phenol and Phenolics
 Differ by functional groups attached to
their aromatic rings.
Disrupts cell walls and membranes.

 Ingredient in soaps up to kitty litter
 Examples: Hexachlorophene, Triclosan
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Phenolics
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Alcohols
Dissolve membrane lipids, disrupt cell surface
tension, denatures proteins.
Used for skin degerming.
Examples: Ethyl alcohol, Isopropyl alcohol
70% concentration the best and most effective
concentration to use.
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Hydrogen Peroxide
Colorless and caustic liquid
Form hydroxyl free radicals
Effective against anaerobes
Applications:
Skin and wound cleaner
Quick method for sterilizing medical
equipment
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Soaps and Detergents
Quaternary ammonium (quats):
Cationic
Bind and disrupt cell membrane
Low-level disinfectant in the clinical setting
Soaps:
Fatty acids, oils, sodium or potassium salts
Cleaning and degerming agents
More effective if mixed with germicides
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Detergent Action
Positively charged region of the detergent binds with bacteria
and the uncharged region integrates into the cell membrane
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Hand Scrubbing
Comparison of efficacy between nongermicidal and
germicidal soaps used in hand scrubbing.
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Heavy Metals
Mercury and Silver:
Inactivate proteins
Oligodynamic action
Preservatives in cosmetics and ophthalmic
solutions.
Silver nitrate as prophylactic against
Ophthalmia neonatorum.
Silver sulfadiazine for burn patients and
catheter tips.
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Oligodynamic Action
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Aldehydes
Formaldehyde and Glutaraldehyde:
Crosslink with proteins on the cell surface
Disinfectant for surgical and dialysis
instruments
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Aldehyde Effect
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Gases
Reacts with functional groups of DNA and
proteins.
Sterilizes and disinfects plastic materials
Example: Ethylene oxide
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Gas Sterilization
Ethylene oxide sterilization chamber
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Dyes
Effective against Gram positive bacteria
Used as Ointments
Example: Crystal violet
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Acids and Alkali
Prevents spore germination and vegetative
growth.
Used as food preservative
Examples: Acetic acid, Ammonium
hydroxide
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