Chapter 9: Physical Methods of Microbial Control

Filtration

0.22 μm filter will filter out microorganisms

What factors might affect the efficacy of physical and chemical control agents/mechanisms?

1) concentration and type of chemical used 2) intensity and nature of physical agent 3) length of exposure to agent 4) temperature/pH 5) number of organisms 6) types of organism 7) nature of material bearing the microorganism

Phenol (derivatives phenolics) - intermediate to low

1. Cellular poisons, disrupt cell walls and membranes. 2. Denatures proteins. 3. Effective on some bacteria, viruses and fungi. 4. Not effective against spores. 5. pretty acidic, no longer used as an antiseptic, but derivatives are used as disinfectants.

What are the chemical Methods of microbial control?

1. Chemical disinfectants: a. Halogens - Chlorine and Iodine - Chloramines, Iodaphors, Hypochlorites b. Phenolics & Derivatives: - Chlorhexidine c. Alcohols: - Ethyl - Isopropyl d. Quats: - Detergents e. Heavy Metals - Mercury tincture > Thimerosal - Silver Nitrate f. Peroxides - Hydrogen - Oxygen 2. Chemical Sterilants: a. Aldehydes: - Glutaraldehyde - Formaldehyde b. Gases - Ethylene Oxide

What are 3 considerations for sterilization?

1. Cost 2. Effectiveness 3. Method of disposal

What methods fall under Heat?

1. Dry heat: - Hot air oven - incineration - leads to sterilization 2. Moist heat: - boiling water - steam (under pressure: autoclave) 3. Pasteurization: - mild heat - aseptic packaging - ultra high temp method) - leads to elimination of pathogens or inhibition of growth and division

Chlorhexidine - intermediate to low

1. Effective against most bacteria, viruses and fungi. 2. Destroys bacterial membrane and makes cells permeable. 3. Low toxicity. 4. hand scrubs 5. mouthwash after periodontal work 6. antiseptic 7. +charged, binds to - charged cell wall

From least to most, list the resistance of microorganisms.

1. Enveloped viruses 2. Most gram-positive bacteria 3. Nonenveloped viruses 4. Fungi and fungal spores 5. most gram-negative bacteria 6.Protozoan trophozoites 7. Protozoan cysts 8. Staphylococcus and Pseudomonas 9. Mycobacterium 10. Bacterial endospores 11. Prions

What are the physical methods of microbial control?

1. Heat 2. Low Temp 3. Radiation 4. Filtration

What methods fall under Radiation?

1. Ionizing (X-rays) 2. Nonionizing (Gamma Rays) 3. Both lead to sterilization

What factors affect the activity of germicidal chemicals?

1. Length of Exposure -Most compounds require adequate contact time to allow the chemical to penetrate and act on microbes present. 2. Composition of Material Being Treated -Smooth, solid objects are more reliably disinfected than those with pores or pockets. -Large amounts of organic material can hinder penetration. -Adequate cleaning of instruments and other reusable materials must precede the use of a germicide or sterilant.

What methods fall under Low temp?

1. Lyophilization (Freezing) 2. Desiccation (Drying) 3. Both lead to elimination of pathogens or inhibition of growth and division

What methods fall under Filtration?

1. Membranes (liquid) 2. HEPA Filter (Air) 3. Both lead to sterilization.

What are the 3 types of microbial control methods?

1. Physical agents 2. Chemical agents 3. Mechanical removal methods

Why is an autoclave so great?

1. Pressure allows steam to get hotter than 100oC 2. Moist heat is very effective at denaturing proteins and destroying living things - better conductor of heat, penetrates cells better and denatures proteins.

How does cold treatment work?

1. Principal benefit of cold treatment is to slow growth of cultures and microbes in food during processing and storage: -Cold merely retards the activities of most microbes. 2. Most microbes are not adversely affected by gradual cooling, long-term refrigeration, or deep-freezing. 3. Temperatures from -70°C to -135°C can preserve cultures of bacteria, viruses, and fungi for long periods.

Ethylene Oxide - high

1. Sterilant, kills endospores. 2. highly reactive, damages biological molecules and destroys their function. 3. Used to sterilize plastics and delicate instruments that cannot withstand autoclaving 4. works well with pre- packaged sterile devices

What are the methods of microbial control?

1. Sterilization 2. Disinfection 3. Decontamination/ Sanitization 4. Antisepsis/Degerming

What are the 4 cellular targets of physical and chemical agents?

1. The cell wall 2. The cytoplasmic membrane 3. Cellular synthetic processes (DNA, RNA) 4. Proteins

What affects bacterial death rate?

1. The number of microorganisms (more takes longer to destroy) 2. The types of microorganisms in the population (usually you are targeting an ecosystem or mixture of many things) 3. The temperature and pH of the environment 4. The concentration of the "killing" agent 5. The mode of action of the "killing" agent 6. The other materials or solvents present (blood, saliva, feces can inhibit the actions of disinfectants.

Aldehydes - high

1. Very reactive, disrupts biological molecules such as proteins and nucleic acids and destroys their function. Effective against endospores. glutaraldehyde: broad spectrum, kills rapidly, used to sterilize a wide variety of medical/dental instruments formaldehyde (gas) - dissolved in water (formalin) - used to disinfect surgical instruments and preserve specimens

Halogens - Intermediate

1. chlorine (bleach, chloramines) - combine with water to produce hypochlorous acid, denature proteins. Water disinfection, wound treatment 2. iodine (free I2 in solution, Iodophors) - interferes with metabolic functions, can kill endospores. Used as antiseptics and disinfectants.

Alcohols - intermediate

1. effective against bacteria, fungi (not endospores, many viruses) 2. denatures proteins 3. damages cell membranes 4. not good for wounds (so don't suffer!) 5. ethanol, isopropanol, 60- 95% 6. alcohol based hand sanitizers

Quats (quaternary ammonium compounds)

1. low 2. detergents are synthetic chemicals 3. usually cationic 4. often derivatives of ammonium chloride 5. alter cell membranes - lysis

How does Pasteurization work?

1. mild heat, reduce the number of microbes in food - 63 oC for 30 minutes -reduces number of pathogens - increases shelf life! 2. ultra high-temperature pasteurization - milk is brought into contact with steam at 140 oC for one or two seconds 3. with aseptic packaging, milk has a long shelf life with no refrigeration required!

Soap - low

1. not antiseptic, but good for mechanical removal of microbes 2. soap emulsifies oils, rinses debris, oil off of skin. De- germing agent 3. soap - fatty acids combined with NaOH or KOH 4. surfactants - emulsify and solubilize particles clinging to the surface. 5 some microbial destruction due to alkalinity

Define Fungicide

A chemical that can kill fungal spores, hyphae, and yeasts.

Define Bactericide

A chemical that destroys bacteria except those in the endospore stage.

Define Decontamination/ Sanitization

A cleansing technique that mechanically removes microorganisms as well as other debris to reduce contamination to safe levels. Important to restaurants, dairies, breweries, and other commercial entities that handle large numbers of soiled utensils/containers. Common Uses: cooking utensils, dishes, bottles, and cans must be sanitized for reuse. Examples: Soaps, detergents, commercial dishwashers

Define Sterilization

A process that destroys or removes all viable microorganisms (including viruses). Generally reserved for inanimate objects as it would be impractical or dangerous to sterilize parts of the human body. The term should be used only in the strictest sense to refer to materials that have been subjected to the process of sterilization (there is no such thing as slightly sterile). Common Uses: surgical instruments, syringes, commercially packaged food. Examples: Heat (autoclave), sterilants (chemical agents capable of destroying endospores)

What type of control agent would be used to achieve sterility?

A sporicide

How does osmotic pressure work?

Adding large amounts of salt or sugar to foods creates a hypertonic environment for bacteria, causing plasmolysis. Pickling, smoking, and drying foods have been used for centuries to preserve foods. Osmotic pressure is never a sterilizing technique.

How is the cytoplasmic membrane affected by antimicrobial agents?

Agents physically bind to lipid layer of the cytoplasmic membrane, opening up the cytoplasmic membrane and allowing injurious chemicals to enter the cell and important ions to exit the cell. Dissolves/opens up plasma membrane and ruptures cell. Agents used: detergents

Define Sporicide

An agent capable of destroying bacterial endospores. It can be a sterilant because it can destroy the most resistant of all microbes.

What is the cellular target of Antimicrobials?

Antimicrobials have a range of cellular targets: Least selective agents tend to be effective against the widest range of microbes (heat and radiation). Selective agents target only a single cellular component (drugs).

Define Virucide

Any chemical known to inactivate viruses, especially on living tissue.

Define Asepsis

Any practice that prevents the entry of infectious agents into sterile tissues and thus prevents infection.

Why do antiseptic and drugs often have microbistatic effects?

Because microbicidal compounds can be toxic to human cells.

Define Aseptics

Chemical agents applied directly to exposed body surfaces, wounds, and surgical incisions to destroy or inhibit vegetative pathogens

Define Germicide/Microbicide:

Chemical agents that kill microorganisms.

Define Bacteristatic

Chemical agents that prevents growth of bacteria on tissues or on objects in the environment

Define Fungistatic

Chemicals that inhibit fungal growth

Define Lyophilization

Combination of freezing and drying Method of preserving microorganisms in a viable state for many years Pure cultures are frozen instantaneously and exposed to a vacuum that removes water, avoiding the formation of ice crystals.

Define Semicritical Devices

Come into contact with mucosal membranes Must receive high-level disinfection

How does dry heat work?

Dehydrates the cell, removing water necessary for metabolic reactions Denatures proteins Increases the stability of some protein conformations, requiring higher temperatures At very high temperatures, oxidizes cells, burning them to ashes not as effective as moist heat needs higher temps, longer times glassware, metal, objects that won't melt can be sterilized in a dry heat oven incineration: burns organisms to destroy them inoculating loops, biohazard waste ash, gas

Define Noncritical Devices

Do not touch the patient or only touch intact skin Require only low-level disinfection

What are some other Antimicrobial agents?

Dyes: -Active against gram-positive bacteria and fungi -Limited application because they stain and have a narrow spectrum of activity Acids and Alkalis: -Very low or very high pH can destroy or inhibit microbial cells -Limited in applications due to their corrosive, caustic, and hazardous nature -Ammonium hydroxide used in detergents, cleaners, and deodorizers -Organic acids used in food preservation

How does filtration work?

Effective method to remove microbes from air and liquids: Fluid is strained through a filter with openings large enough for the fluid to pass, but too small for microbes. Also used are thin membranes of cellulose acetate, polycarbonate, and plastics whose pore size is carefully controlled. Charcoal, diatomaceous earth, or unglazed porcelain are also used. Pore sizes can be controlled to permit true sterilization by trapping viruses or large proteins. Used to prepare liquids that cannot withstand heat such as serum, blood products, vaccines, drugs, IV fluids, enzymes, and media Alternative method for decontaminating milk and beer without altering its flavor Important step in water purification Unable to remove soluble molecules (toxins) that can cause disease High efficiency particulate air (HEPA) filters are used in hospital rooms and sterile rooms.

How does Radiation work?

Energy emitted from atomic activities and dispersed at high velocity through matter or space: Gamma rays and X rays: ionizing; rip apart DNA, passes through solids Ultraviolet radiation: non-ionizing; better for surface disinfection, thymine dimers (germicidal lamps)

Traits of Bacterial Endospores

Exhibit greatest resistance Destruction of spores usually requires temperatures above boiling Resistance varies

Define Critical Medical Devices

Expected to come into contact with sterile tissues Must be sterilized before use

What is the method for selecting a microbicidal chemical?

Germicides are evaluated in terms of their effectiveness in destroying microbes in medical and dental settings: -High-level germicides kill endospores and can be used as sterilants. -Intermediate-level germicides kill fungal, but not bacterial spores, resistant pathogens, and viruses. -Low-level germicides eliminate only vegetative bacteria, vegetative fungal cells, and some viruses. Rapid action, even in low concentrations Solubility in water or alcohol and long-term stability Broad-spectrum action without being toxic to human and animal tissues Penetration of inanimate surfaces to sustain a cumulative or persistent action Resistance to becoming inactivated by organic matter Noncorrosive or nonstaining properties Sanitizing and deodorizing properties Affordability and ready availability No chemical can completely fulfill all of these requirements: Glutaraldehyde and hydrogen peroxide approach this ideal.

Germicidal effectiveness

High level germicide: kill spores, and can be sterilants. Good for medical devices like catheters, heart-lung equipment, implants. Intermediate level germicides: kill fungal (not bacterial) spores, resistant pathogens such as mycobacterium, and viruses. Used for things that come into contact with mucous membranes (thermometers, respiratory equptment). Low-level germicides: kill vegetative cells, fungal cells, some viruses. Used to clean things that only come into contact with skin surface (electrode straps, furniture)

How do you know if a microbe is dead?

If it can no longer reproduce, we assume that it is dead.

How is cellular synthesis affected by antimicrobial agents?

Inhibits protein synthesis, damages genetic code, damages proteins needed for growth and metabolism, etc. Agents can interrupt the synthesis of proteins via the ribosomes, inhibiting proteins needed for growth and metabolism and preventing multiplication. Agents can change genetic codes (mutation) Agents used: Formaldehyde, radiation, ethylene oxide

Heavy Metals - low

Metals (Hg, Ag, and others) bind to functional groups of proteins and inactivate them. Effective against some bacteria, viruses fungi Organic mercury tinctures - antiseptics, preservatives silver nitrate solutions - topical germicides and ointments

What is the primary targets of microbial control?

Microorganisms capable of causing infection or spoilage that are constantly present in the external environment and on the human body. The targeted population often contains mixtures of microbes with extreme differences in resistance and harmfulness. Bacterial endospores have traditionally been considered the most resistant microbial entities.

Which method is generally faster at killing microbes: Moist heat or Dry heat?

Moist heat

How does moist heat work?

Operates at lower temperatures and shorter exposure times to achieve the same effectiveness as dry heat Microbicidal effect is the coagulation and denaturation of proteins

Define Microbial Death

Permanent termination of an organism's vital processes: Microbes have no conspicuous vital processes, therefore death is difficult to determine Permanent loss of reproductive capability, even under optimum growth conditions has become the accepted microbiological definition of death.

Define Aseptic techniques

Practiced in healthcare; range from sterile methods to antisepsis

Which microbial form has the highest resistance?

Prions then Bacterial endospores

How are proteins affected by antimicrobial agents?

Some agents are capable of denaturing good proteins (breaking of protein bonds, which results in breakdown of the protein structure). Works by denaturation, breaking peptide bonds.Agents May attach to the active site of a protein, preventing it from interacting with its chemical substrate. Agents used: moist heat, alcohol, phenolics

How is the cell wall affected by antimicrobial agents?

The chemicals damage and disrupt the cell wall, causing the cell to rupture. Agents used: Chemicals, Detergents, Alcohol

Define Sepsis

The growth of microorganisms in the blood and other tissues.

Define Disinfection

The physical process or a chemical agent to destroy vegetative pathogens but not bacterial endospores. Removes harmful products of microorganisms (toxins) from material. Normally used on inanimate objects because the concentration of disinfectants required to be effective is harmful to human tissue. Common Uses: boiling food utensils, applying 5% bleach solution to an examining table, immersing thermometers in an iodine solution between uses. Examples: bleach, iodine, heat (boiling)

Define TDP

Thermal Death Point: The lowest temperature required to kill all microbes in a sample in 10 minutes.

Define TDT

Thermal Death Time: The shortest length of time required to kill all test microbes at a specified temperature

Define Antisepsis/ Degermation

To reduce the number of microbes on the human skin. A form of decontamination but on living tissue. Involves scrubbing the skin (mechanical friction) or immersing it in chemicals (or both). Examples: alcohol, surgical hand scrubs

Traits of Vegetative Cells

Vary in sensitivity to heat Death times vary from 50°C for 3 minutes to 60°C for 60 minutes

Define Desiccation

Vegetative cells directly exposed to normal room temperature gradually become dehydrated. Some microbes are killed by desiccation; on others desiccation has no effect

Boiling Water

boiling: 100oC for 30 min. Kills everything EXCEPT some endospores. >6 hours is required to get all the spores! However, as soon as you remove the items from the boiling water they are no longer sterile...

Methods of Chemical Control

can be gases, liquids or solids terms: disinfectants, antiseptics, sterilants, preservatives (chemicals that inhibit the deterioration of substances) if the chemical is dissolved in pure water: AQUEOUS if the chemical is dissolved in alcohol or water/alcohol mixture: TINCTURE

Define Antiseptic

chemical agents applied directly to exposed body surfaces (skin and mucous membranes), wounds, and surgical incisions to prevent vegetative pathogens: Uses: 1.Preparing the skin before surgical incisions with iodine compounds 2. Swabbing an open root canal with hydrogen peroxide 3. Ordinary hand washing with a germicidal soap

Hydrogen Peroxide

high level germicide forms oxygen free radicals, highly reactive and toxic usually used at 1.5 - 3% primarily used as an antiseptic good for infections caused by anaerobic bacteria

Radiation

ionizing (x-rays and gamma rays) - rip apart DNA, passes through solids non-ionizing (UV light) - better for surface disinfection, thymine dimers (germicidal lamps)

Pasteurization

mild heat, reduce the number of microbes in food - 63 oC for 30 minutes reduces number of pathogens increases shelf life! ultra high-temperature pasteurization - milk is brought into contact with steam at 140 oC for one or two seconds with aseptic packaging, milk has a long shelf life with no refrigeration required!

Dry Heat

not as effective as moist heat needs higher temps, longer times glassware, metal, objects that won't melt can be sterilized in a dry heat oven incineration: burns organisms to destroy them inoculating loops, biohazard waste ash, gas

Define Bactericidal

substance that kills bacteria

What is the goal of any sterilization process?

the destruction of bacterial endospores; Any process that kills endospores will invariably kill all less resistant microbial forms.