Chapter 9

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Chapter 9
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2013-03-29 04:56:11
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Controlling Microbial Growth Enviornment
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Controlling Microbial Growth in the Enviornment
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  1. Sterilization
    Removal or destruction of all microbes, includuing viruses and bacterial endospores.

    Does not apply to prions (infectious proteins).
  2. Aseptic
    Describes and environment or procedure that is free of contamination by pathogens.
  3. Disinfection
    • The use of physical or chemical agents known as disinfectants.
    • Include: UV light, heat, alcohol, and bleach, to inhibit or destroy MO, especially pathogens.
  4. Commerical Sterilization
    Does not kill all hyperthermophilic microbes in canned foods, however, do not cause disease, cannot grow and spoil food at ambient temperatures, so are of no practical concern.
  5. Are all pathogens eliminated by disinfection?
     Does not guarantee complete elimination and cannot alone inhibit endospores or some viruses. Used only when discussing tx of inanimate objects.
  6. Antisepsis
    The process when a chemical is used on skin or other tissue, chemical called antiseptic.
  7. Disinfectants vs. Antiseptics
    Often have same components, but disinfectants are more concentrated or can be left on a surface for longer periods of time.

    Some disinfectants, such as steam or concentrated bleach, are not suitable for use as antiseptics.
  8. Degerming
    Removal of all microbes from a surface by scrubbing with soap or alcohol.
  9. Sanitization
    Process of disinfecting places and utensils used by public to reduce number of pathogenic microbes to meet accepted public health standards.
  10. Disinfecting vs. Sanitizing
    The difference is the place - private vs. public - in which the activity takes place.
  11. Pasteurization
    Use of heat to kill pathogens and reduce number of spoilage MO in food and beverages.
  12. -stasis/-static
    Indicates that a chemical or physical agent inhibits microbial metabolism and growth, but doesn't necessarily kill microbes.

    Refrigeration is bacteriostatic for most bacterial species; it inhibits their growth, but they can resume metabolism when the optimal temp is restored.
  13. -cide/-cidal
    Refers to agents that destroy or permanently inactivate a particular type of microbe.

    • Virucides: inactivate viruses
    • Bactericides: kill bacteria
    • Fungicides: kill fungal hyphae, spores, yeast
    • Germicides: chemical agents that destroy pathogenic MO in general.
  14. Microbial Death
    The permanent loss of reproductive ability under ideal environment conditions.
  15. What is one technique for evaluating the efficacy of an antimicrobial agent?
    By calculating the microbial death rate, which is usually found to be constant over time for any particular MO under a particular set of conditions.
  16. Microbial Death Rate
    The time it takes for microbicidal agents to kill cells over a period of time.
  17. How is data collected to determine the microbial death rate of a MO?
    On a semilogarithmic graph. The line graph determines the most susceptible cells that die first down to the more resistant cells that require longer exposure to the agent before they die.
  18. What are the two basic categories of the actions of antimicrobial agents?
    Those that disrupt the integrity of cells by adversly altering their cell walls or cytoplasmic membranes, and those that interrupt cellular metabolism and reproduction by interfering with the structures of proteins and nucleic acids.
  19. How do cell walls maintain their cellular integrity?
    By counteracting the effects of osmosis when the cell is in a hypotonic solution.
  20. Function of Proteins
    They regulate cellular metabolism, function as enzymes in most metabolic reactions, and form structural components in membranes and cytoplasm.
  21. What does a protein's function depend on?
    The exact three-dimensional shape, which is maintained by hydrogen and disulfide bonds between amino acids.

    When bonds are broken by extreme heat or certain chemicals, the protein's shape changes. These denatured proteins cease to function, bringing about cellular death.
  22. What can destroy proteins and nucleic acids?
    Chemicals, radiation, and heat. Physical or chemical agents that interfere with nucleic acids also stop protein synthesis because that portion of a ribosome that actually catalyzes the synthesis of proteins is a ribozyme (an enzymatic RNA molecule) and given that the genes of a cell or virus are composed of nucleic acids, disruption of these molecules can produce fatal mutations.
  23. What factors must you consider with the efficacy of antimicrobial methods?
    • The nature of the sites to be treated.
    • The degree of susceptiblity of the microbes involved.
    • The enviornment conditions that pertain.
  24. What factors should be considered when using agents to control microbes?
    • They should be inexpensive, fast-acting, and stable during storage.
    • A perfect agent would control the growth and reproduction of every type of microbe while being harmless to humans, animals, and objects, but every agent has limitations and disadvantages.
  25. Why do the sites to be treated by antimicrobial agents depend of the nature of the site to be treated?
    • Medical personnel must choose a method and level of microbial control based on the site of the procedure, because the site greatly affects the potential for subsequent infection.
    • You wouldnt use harsh chemicals or heat on humans, animals, or fragile objects, such as a heart valve or plastic utensils.
    • Medical instruments that penetrate the outer defenses of the body (needles and scalpels) carries a greater potential for infection, so they must be sterilized, whereas items that contact only the surface of mucous membranes or the skin may be disinfected (only immunocompromised patients require sterilization.)
  26. Why do microbial death rates vary?
    • Microbes fall along a continuum for most susceptible to most resistant to antimicrobial agents.
    • Ex. enveloped viruses (HIV) are more susceptible to antimicrobial agents and heat than are nonenveloped viruses (poliovirus), because viral envelopes are more easily disrupted than the protein coats of nonenveloped viruses.
  27. Relative Susceptibilities of Microbes to Antimicrobial Agents (Diagram)
    • Most Resistant
    •      Prions
    •      Bacterial endospores
    •      Mycobacteria
    •      Cysts of protozoa
    •      Active-stage protozoa (trophozoites)
    •      Most Gram-negative bacteria
    •      Fungi
    •      Nonenveloped viruses
    •      Most Gram-positive bacteria
    •      Enveloped viruses
    • Most Susceptible
  28. What are the most resistant microbial forms of life?
    • Bacillus and Clostridium. They can survive enviornmental extremes of temperature and acidity, and many chemical disinfectants.
    • Ex. Endospores can survive more than 20 years in 70% alcohol.
  29. Species of Mycobacterium
    • The cell walls of members of this genus, such as Mycobacterium tuberculosis, contain large amounts of waxy lipids, which allows these bacteria to survive drying and protects them from the most water-based chemicals.
    • Medical personnel must use strong disinfectants or heat to treat whatever comes into contract with tuberculosis patients, including utensils, equipoment, and patient rooms.
  30. Cysts of Protozoa
    • A protozoan cyst's wall:
    •      Prevents entry of most disinfectancts
    •      Protects against drying
    •      Shields against radiation and heat.
  31. What microbes are the most resistant than any living thing?
    Prions, infectious proteins that cause degenerative diseases of the brain.
  32. How are the effectiveness of germicides classified?
    As high, intermediate, and low, depending on their proficiency in inactivating or destroying MO on medical instruments that cannot be sterilized with heat.
  33. High-level Germicides
    • Kill all pathogens, including bacterial endospores. 
    • Use them to sterilize invasive instruments, such as catheters, implants, and parts of heart-lung machines.
  34. Intermediate-level Germicides
    • Kill fungal spores, protozoan cysts, viruses, and pathogenic bacteria, but not bacterial endospores.
    • Used to disinfect instruments that come in contact with mucous membranes but are noninvasive, such as respiratory equipment and endoscopes.
  35. Low-level Germicides
    • Eliminate vegetative bacteria, fungi, protozoa, and some viruses.
    • Used to disinfect items that only contact the skin of patietns, such as furniture and electrodes.
  36. What affects microbial death rates and the efficacy of antimicrobial methods?
    Enviornmental conditions such as temperature and pH.
  37. Do warm disinfectants or cool disinfectants work better?
    Warm disinfectants, because chemicals react faster at higher temperatures.
  38. How are antimicrobial effects of heat enhanced?
    By acidic conditions.
  39. What organic materials interfere with the penetration of heat, chemicals, some forms of radiation, and in some cases inactivate chemical disinfectants?
    • Fats, feces, vomit, blood, and intercellular secretions in biofilms (a slimy community of microbes growing on a surface.)
    • For this reason, it is important to clean objects before sterilization or disinfection so that antimicrobial agents can thoroughly contact all the object's surfaces.
  40. What methods were developed by scientists to measure the efficacy of antimicrobial agents?
    • The phenol coefficient
    • The use-dilution test
    • The Kelsey-Sykes capacity test
    • The in-use test

    ***Anyone using disinfectants must consider whether higher concentrations and longer exposures may damage an object or injure a patient.***
  41. Phenol (carbolic acid)
    Founded by Lister, an antiseptic used during surgery in the late 1800s.
  42. Phenol Coefficient
    • Researchers have evaluated the efficacy of various disinfectants and antiseptics by calculating a ratio that compares a given agent's ability to control microbes to that of phenol under standardized conditions.
    • A phenol coefficient > 1.0 indicates agent more effective then phenol, and the larger the ratio, the greater the effectiveness.
    • Ex. chloramine, a mixture of chlorine and ammonia, has a phenol coefficient of 133.0 when used against the bacterium Staphylococcus aureus, and a phenol coefficient of 100.0 when used against Salmonella enterica. This indicates that chloramine is at least 133 times more effective then phenol against Stephylococcus, but only 100 times more effective against Salmonella.
  43. Use-Dilution Test
    Method of evaluating the effectiveness of a disinfectant or antiseptic against specific microbes in which the most effective agent is the one that entirely prevents microbial growth at the highest dilution.

    • Researchers:
    • Dip several metal cylinders into broth cultures of bacteria,
    • Briefly dries them at 37C,
    • Immerse into different dilutions of disinfectants.
    • After 10 minutes, each cylinder is removed, rinsed with water to remove excess chemicals, and placed into a fresh tube of sterile medium for 48 hours of incubation.

    The most effect agent is the one that entirely prevents microbial growth at the highest dilution.
  44. Kelsey-Sykes Capacity Test
    The standard alternative assessment approved by the European Union to determine the capacity of a given chemical to inhibit bacterial growth.

    • Researchers:
    • Add a suspension of bacterium to a suitable concentration of the chemical being tested.
    • At predetermined times, they move samples of mixture into growth medium containing a disinfectant deactivator.
    • After incubation for 48 hours, turbidity in the medium indicates that bacteria survived treatment. Lack of turbidity, indicating lack of bacterial reproducation, reveals the minimum time required for the disinfectant to be effective.
  45. Why is the In-Use Test method more realistic (though time-consuming) for determining the efficacy of a chemical?
    Though phenol coefficient, use-dilution, and Kelsey-Sykes capacity tests can be beneficial, they can also be misleading, giving they are measured under controlled conditions against one, or at most a few, species of microbes. But disinfectants are generally used in various enviornments against a diverse population of organisms that are often associated with one another in complex biofilms affording mutual protection.
  46. What method for evaluating disinfectants and antiseptics is the current standard in the United States?
    The use-dilution test.
  47. In-Use Test
    • Swabs are taken from actual objects, such as operating room equipment, both before and after the application of a disinfectant or an antiseptic.
    • Swabs are then inoculated into appropriate growth media, which after incubation are examined for microbial growth.
    • The in-use test allows a more accurate determination of the proper strength and application procedure of a given disinfection agent for each specific situation.
  48. What are the physical methods of microbial control?
    Exposure to extremes of heat and cold, desiccation, filtration, osmotic pressure, and radiation.
  49. What is one of the older and more common means of microbial control?
    Heat.

    High temperatures denature proteins, interfere with the integrity of cytoplasmic membranes and cell walls, and disrupt the function and structure of nucleic acids. 
  50. What can heat be used for?
    Sterilization, in which case all cells and viruses are deactivated, or for commercial preparation of canned goods.

    MO vary in their suseptibility to heat, as a result scientists have developed concepts and terminology to convey these differences in susceptibility.
  51. Commercial Sterilization
    Hyperthermophilic prokaryotes remain viable but are harmless because they cannot grow at the normal (room) temperatures in which canned foods are stored.
  52. Thermal death point
    The lowest temp that kills all cells in a broth in 10 minutes.
  53. Thermal death time
    The time it takes to completely sterilize a particular volume of liquid at a set temperature.
  54. Decimal reduction time (D)
    • When measuring the effectiveness of heat sterilization, researchers calculate the time required to destroy 90% of the microbes in a sample.
    • This concept is especially useful to food processors to eliminate all the endospores of anaerobic Clostridium botulinum.
  55. Moist heat
    Is more effective than dry heat because water is a better conductor of heat then air. Used to disinfect, sterilize, sanitize, and pasteurize. Kills cells by denaturing proteins and destroying cytoplasmic membranes.
  56. What moist heat methods are used for controlling microbes?
    Boiling, Autoclaving (pressure cooking), Pasteurization, and Ultrahigh-temperature sterilization.
  57. Boiling
    • Boiling time is a critical factor.
    • Kills vegetative cells of bacteria and fungi, the trophozoites of protozoa, and most viruses within 10 minutes at sea level.
    • Effective for sanitizing restaurant tableware or disinfecting baby bottles.
  58. How long can bacterial endospores withstand boiling?
    For more than 20 hours, not recommended when true sterilization is required.
  59. Autoclaving
    True sterilization using heat requires higher temps than that of boiling water. To achieve required temp, pressure is applied to boiling water to prevent the escape of heat in steam. 
  60. Autoclave
    A laboratory equipment to sterilize chemicals and objects that can tolerate moist heat. Consists of a pressure chamber, pipes to introduce and evacuate steam, valves to remove air and control pressure and temp gauges to monitor the procedure.

    As steam enters an autoclave chamber, it forces air out, raises the temp of the contents, and increases the pressure, until a set temp and pressure are reached.
  61. How long does it take for an autoclave with the temp at 121C and 15psi above normal pressure take to destroy all microbes in a small volume?
    About 10 minutes.
  62. What is the margin of safety used for an autoclave during sterilization?
    15 minutes.
  63. What substances require extra time to sterilize in an autoclave?
    Solid substances, such as meat because it takes longer for heat to penetrate to their centers.
  64. Sterilization in autoclaves require that steam be able to contact all liquids and surfaces that might be contaminated with microbes.
    • Solid objects must be wrapped in porous cloth or paper, not sealed in plastic or aluminium foil, which are impermeable to steam.
    • Containers of liquids must be sealed loosely enough to allow steam to circulate freely, and all air must be forced out by steam.
  65. What means do scientists use to ensure that an autoclave has sterilized its contents?
    • A common one is a chemical that changes color when the proper combination of temp and time have been reached.
    • A color indicator can be impressed in a pattern on tape or paper so that the word sterile or a pattern or design appears.
    • Use of plastic beads that melt when proper conditions are met.
    • A biological indicator of sterility uses endospores of the bacterium Bacillus stearothermophilus impregnated into tape. After autoclaving, the tape is asceptically inoculated into sterile broth. If no bacterial growth appears, the original material is considered sterile.
    • Another variation is that the endospores are on a strip in one compartment of a vial that also includes a growth medium containing a pH color indicator. After autoclaving, a barrier between the two compartments is broken, putting the endospores into contact with the medium. The absence of a color change after incubation indicates sterility.
  66. Moist Heat Treatments of Milk
    • Historical (batch) pasteurization: 63C for 30 mins
    • Flash pasteurization: 72C for 15 seconds
    • Ultrahigh-temp pasteurization: 134C for 1 sec
    • Ultrahigh-temp sterilization: 140C for 1-3 sec
  67. Is pasteurization sterilization?
    No. Thermoduric (heat-tolerant) and thermophilic (heat-loving) prokaryotes survive pasteurization, but they do not cause spoilage over the relatively short times during which properly refrigerated and pasteurized foods are stored before consumption. Such prokaryotes are generally not pathogenic.
  68. Pasteurization
    • Method developed by Louis Pasteur of heating beer and wine just enough to destroy the MO that cause spoilage without raising the temp so much that the taste was ruined.
    • The combination of time and temp required for effective pasteurization varies with the product.
  69. What type of moist heat treatments do we use for pasteurizing milk today?
    Flash Pasteurization, in which milk flows through heated tubes that raise its temp to 72C for only 15 seconds.
  70. Ultrahigh-Temperature Sterilization
    • Involves flash heating milk or other liquids to rid them of all living microbes. This process involves passing the liquid through superheated steam at 140C for 1-3 seconds, and then cooling rapidly.
    • Treated liquids can be stored indefinitely at room temp without microbial spoilage, thoguh after months of storage chemical degradation results in flavor changes.
    • Ex. small packages of dairy creamer served in restaurants.
  71. Dry Heat
    For substances such as powders and oils that cannot be sterilized by boiling or with steam, or for materials that can be damaged by repeated exposure to steam (such as some metal objects), sterilization can be achieved by the use of dry heat, as occurs in an oven.
  72. What is the ultimate means of sterilization?
    Complete incineration.
  73. Why is hot air an effective sterilizing agent?
    Because it denatures proteins and fosters the oxidation of metabolic and structural chemicals, howver, in order to sterilize, dry heat requires higher temps for longer times than moist heat because dry heat penetrates more slowly.
  74. In food preperation and storage, what is the most convenient method of microbial control?
    Refrigeration (0C-7C) or freezing (<0C).

    These processes decrease microbial metabolism, growth, and reproduction because chemical reactions occur more slowly at low temps, and because liquid water in not available at subzero temps.

    *Psychrophilic (cold-loving) microbes can multiply in refrigerated food and spoil its taste and suitability for consumption.*
  75. Refrigeration halts growth of most pathogens, which are predominantly mesophiles. What are some notable exceptions?
    • Listeria, which can reproduce to dangerous levels in refrigerated food.
    • Yersinia, which can multiply in refrigerated blood products and be passed onto the blood recipients.
  76. Why is slow freezing more effective than quick freezing?
    • Because slow freezing allows ice crystals to have time to form and puncture cell membranes by inhibiting microbial metabolism, though MO also vary in their susceptiblity to freezing.
    • Ex. cysts of tapeworms perish after several days in frozen meat, many vegetative bacterial cells, bacterial endospores, and viruses can survive subfreezing temps for years.
  77. Dessication (drying)
    • Inhibits microbial growth because metabolism requires liquid water. Drying inhibits the spread of most pathogens, including the bacteria that causes syphilis, gonorrhea, and the more common forms of bacterial pneumonia and diarrhea. However, most molds can grow on dried raisins and apricots, which have as little as 16% water content.
    • Has been used for thousands of years to preserve such foods as fruit, peas, beans, grain, nuts, and yeast.
  78. Lyophilization
    • A technique combining freezing and drying, to preserve microbes and other cells for many years.
    • Scientists instantly freeze a culture in liquid nitrogen or frozen carbon dioxide (dry ice), then subject it to a vacuum that removes frozen water through a process called sublimation, in which the water is transformed directly from a solid to a gas.
    • Prevents formation of large, damaging ice crystals. 
    • Although not all cells survive, enough are viable to enable the culture to be reconstituted many years later.
  79. Filtration
    • The passage of fluid (either a liquid or a gas) through a sieve designed to trap particles - in this case, cells or viruses - and separate them from the fluid.
    • Researches often use a vacuum to assist the movement of a fluid through a filter.
    • Traps microbes larger than the pore size, allowing smaller microbes to pass through.
    • Can be used to sterilze such heat-sensitive materials such as opthalmic solutions, antibiotics, vaccines, liquid vitamins, enzymes, and culture media.
  80. Over the years, what have filters been constructed out of?
    • Porcelain, glass, cotton, asbestos, adn diatomaceous earth, a substance composed of the innumerable glasslike cell walls of single-celled algae called diatoms.
    • Scientists today typically use thin (0.1mm), circular membrane filters manufactured of nitrocellulose or plastic and containing specific pore sizes ranging from 25um to < 0.01um in diameter.
  81. What do microbiologists also use filtration for?
    To estimate the number of microbes in a fluid by counting the number deposited on the filter after passing a given volume through a filter.
  82. HEPA Filter
    • High-efficiency particular air (HEPA) filters are crucial parts of biological safety.
    • Are mounted on air ducts of some operating rooms, rooms occupied by patients with airborne diseases such as tuberculosis, and rooms of immunocompromised patients such as burn victims and AIDS patients.
  83. How is osmotic pressure used in microbial control?
    High concentrations of salt and sugar in foods help to inhibit microbial growth.
  84. What is osmosis?
    The net movement of water across a semipermeable membrane (such as a cytoplasmic membrane) from an area of higher water concentration to an area of lower water concentration.
  85. What happens to cells in a hypertonic solution of salt or sugar?
    Cells lose water and then desiccates. The removal of water inhibits cellular metabolism because enzymes are fully functional only in aqueous enviornments.

    Thus, osmosis preserves honey, jerky, jams, jellies, salted fish, and some types of pickles from most microbial attacks.
  86. Which microbes have a greater ability than bacteria to tolerate hypertonic enviornments with little moisture?
    Fungi
  87. What two types of radiation are used in physical methods of microbial control?
    Particulate radiation and electromagnetic radiation.
  88. What is particulate radiation?
    It consists of high-speed subatomic particles, such as protons, that have been freed from their atoms.
  89. What is electromagnetic radiation?
    Can be defined as energy without mass traveling in waves at the speed of light. Energy is released from atoms that have undergone internal changes.
  90. What are wavelengths of radiation?
    The distance between two crests of a wave, ranging from very short waves (gamma, xrays, UV light, and visible light) to long waves (infrared) to very long waves (radio).

    The shorter the wavelength, the more energy, therefore, shorter-wavelength radiation is more suitable for microbial control than longer-wavelength radiation, which carries less energy and is less penetrating.
  91. What are the types of radiation described by scientists when determining its effects on chemicals within cells?
    Ionizing or nonionizing, according to its effects on the chemicals within cells.
  92. What is ionizing radiation?
    When radiation strike molecules, they have sufficient energy to eject electrons from atoms, creating ions. Such ions disrupt hydrogen bonding, oxidize double covalent bonds, and create highly reactive hydroxyl radicals. These ions in turn denature other molecules, particularly DNA, causing fatal mutations and cell death.

    Examples: electron beams, gamma rays, xrays, all of which have wavelengths shorter than 1nm.
  93. What is nonionizing radiation?
    Electromagnetic radiation with a wavelength greater than 1nm that does not have enough energy to force electrons out of orbit. However, it does contain enough energy to excite electrons and cause them to make new covalent bonds, which can affect the three-dimensional structure of proteins and nucleic acids.
  94. How are electron beams effective in microbial control?
    Electron beams are highly energetic in killing microbes in just a few seconds, but cannot sterilize thick objects or objects coated with large amounts of organic matter.
  95. How are gamma rays used in microbial control?
    Gamma rays are emitted by some radioactive elements such as radioactive cobalt, penetrating much farther than electron beams but require hours to kill microbes.

    Irradiation with gamma rays kills not only microbes but also the larvae and eggs of insects; it also kills the cells of fruit and vegetables, preventing both microbial spoilage and overripening.
  96. How are xrays used in microbial control?
    Xrays travel the farthest through matter, but have less energy than gamma rays and require a prohibitive amount of time to make them practival for microbial control.
  97. What are some examples of nonionizing radiation?
    Electromagnetic, UV light, visible light, infrared, and radio waves.

    Only UV light has sufficient energy to be a practical antimicrobial agent, suitable primarily for disinfecting air, transparent fluids, and surfaces of objects such as barber's shears and operating tables.
  98. Who created four levels of safety in microbiological laboratories dealing with pathogens?
    The Centers for Disease Control and Prevention (CDC)
  99. What are the four levels of biosafety?
    Each level raises personnel and enviornmental safety by specifying increasingly strict laborabory techniques, use of safety equipment, and design of facilities.

    • Biosafety Level 1 (BSL-1)
    • Biosafety Level 2 (BSL-2)
    • Biosafety Level 3 (BSL-3)
    • Biosafety Level 4 (BSL-4)
  100. Biosafety Level 1 (BSL-1)
    Suitable for handling microbes not known to cause disease in healthy humans. Precautions are minimal and include handwashing with antibacterial soap and washing surfaces with disinfectants.
  101. Biosafety Level 2 (BSL-2)
    Are designed for handling moderately hazardous agents such as hepatitis, influenza, and MRSA. Access is limited when work is being conducted, extreme precautions are taken with contaminated sharp objects and procedures that might produce aerosols are conducted within safety cabinets.
  102. Biosafety Level 3 (BSL-3)
    A stricter level, requiring that all manipulations be done within HEPA safety cabinets and specifying special design features for the laboratory for experimentation on microbes such as TB, anthrax bacteria, viruses of yellow fever, and Rocky Mountain spotted fever.

    Include double sets of doors and ventilation such that air only moves into the room through an open door.  Air leaving the room is HEPA-filtered before being discharged outside the room.
  103.  Biosafety Level 4 (BSL-4)
    Most secure laboratories designated for working with dangerous or exotic microbes that cause severe or fatal diseases in humans, such as Ebola, smallpox, and Lassa fever viruses.

    Are either separate buildings or completely isolated from all other areas of their buildings. Entrance and exits are strictly controlled through electronically sealed airlocks with multiple showers, a vacuum room, a UV light room, and other safety precautions designed to destroy all traces of the biohazard. All air and water entering/leaving facility are filtered to prevent accidental release. Personnel wear "space suits" supplied with air hoses are pressurized so microbes are swept away from workers.
  104. What are chemical agents used for in microbial control?
    They act to adversely affect microbes' cell walls, cytoplasmic membranes, proteins, or DNA. Effect of agent varies with temp, length of exposure, and amt of contaminating organic matter in the enviornment. The effect also varies with the pH, concentration, and freshness of the chemical.
  105. What microbes do chemical agents work best/worse with?
    They tend to destroy or inhibit the growth of enveloped viruses and the vegetative cells of bacteria, fungi, and protozoa more than fungal spores, protozoan cysts, or bacterial endospores. (i.e. numerous failed attempts in US Senate office building of anthrax endosports in 2001).
  106. What are the nine categories of antimicrobial chemicals used as antiseptics and disinfectants?
    • Phenols
    • Alcohols
    • Halogens
    • Oxidizing Agents
    • Surfactants
    • Heavy Metals
    • Aldehydes
    • Gaseous Agents
    • Enzymes
  107. Phenol and Phenolics
    Intermediate to low-level disinfectants that denature proteins and disrupt cell membranes in a wide variety of pathogens.

    • Phenol (carbonic acid)
    • Phenolics (compounds synthesized from phenol, have greater antimicrobial efficacy with fewer side effects)
    • Bisphenols (paried, cavalently linked phenolics)
  108. Phenolics
    Compounds derived from phenol molecules that have been chemically modified by the addition of halogens or organic functional groups.
  109. What natural oils can be used as phenolic antiseptics?
    Pine and clove oils.
  110. What methods are scientists turning to with the development of resistance to common disinfectants and antiseptics?
    Natural parasites of bacteria (bacteriophages or "phages") to control bacterial contamination.
  111. Bacteriophages
    "Bacteria eaters", the term for viruses that specifically attack particular strains of bacteria.

    Unlike disinfectants and most antimicrobial drugs, phages attack specific bacterial strains and leave neighboring bacteria unharmed.

    A phage injects its genetic material into a bacterial cell, causing the bacterial cells to produce hundreds of new phages before bursting out of the bacterium and killing it.
  112. Where are phage solutions that researchers have developed being used?
    Medical settings, food, and in patients.
  113. Bisphenolics
    Composed of two covalently linked phenolics.

    • Orthophenylphenol (active ingredient in Lysol)
    • Triclosan (garbage bags, diapers, cutting boards)
  114. Where are phenolics commonly used?
    Health care settings, laboratories, and households.
  115. What are some negative aspects of phenolics?
    Their disagreeable odor and possible side effects (irritation of skin).
  116. What bisphenolic was once a popular household, when was found to cause brain damage in infants?
    Hexachlorophene, now only available by prescription and used in nurseries only in response to severe staphylococcal contamination.
  117. Alcohols
    Are intermediate-level disinfectants.

    Denature proteins and disrupt cytoplasmic membranes.

    Are bactericidal, fungicidal, and virucidal against enveloped viruses, but are not effective against fungal spores or bacterial endospores.
  118. What is slightly superior to ethanol as a disinfectant and antiseptic?
    Isopropanol
  119. Tinctures
    Are solutions of other antimicrobial chemicals in alcohol that often are more effective than the same chemicals dissolved in water.
  120. Is pure alcohol an effective chemical agent in microbial control?
    No, because the denaturation of proteins requires water, so solutions of 70%-90% are typically used to control microbes.
  121. What are the advantages and disadvantages of alcohol use in microbial control?
    • Advantages:
    • evaporate quickly and leave no residue
    • more effective than soap in removing bacteria from hands

    • Disadvantages:
    • may not contact microbes long enough to be effective
  122. What are the four very reactive, nonmetallic chemical elements in halogens?
    Iodine, chlorine, bromine, and fluorine.
  123. Halogens
    • Intermediate-level antimicrobial chemical.
    • Effective against vegetative bacterial and fungal cells, fungal spores, some bacterial endospores and protozoan cysts, and many viruses.
    • Used both alone and combined with other elements in organic and inorganic compounds.
    • Thought to damage enzymes via oxidation or by unfolding them, denaturing them.
  124. Iodine
    • Well-known antiseptic.
    • Used either as a tincture or as an iodophor.
    • Used to be used as iodine tablets in disinfecting water for backpackers and campers, but protozoan cysts still survived unless the iodine concentration was so high that the water would be undrinkable.
  125. Iodophor
    • An iodine-containing organic compound that slowly releases iodine.
    • Have advantage of being long lasting and nonirritating to the skin.
  126. Example of iodine use.
    Betadine - used in medical institutions to prepare skin for surgery, injections, and to treat burns.
  127. Examples of chlorine (Cl2).
    Used in its elemental form to treat drinking water, swimming pools, and wastewater from sewage treatment plants.

    Sodium Hypochlorite (NaOCl) (calcium hypochlorite): household chlorine bleach, but does not kill all protozoan cysts, bacterial spores, or viruses. 

    Dairy industry and resturants: disinfect utensils.

    Medical field: to disinfect hemodialysis systems.

    Chlorine dioxide (ClO2): gas that can be used to disinfect large spaces. (i.e. federal office contaminated with anthrax spores in 2001 bioterrorism attack)
  128. Chloramines
    • Chemical combination of chlorine and ammonia.
    • Are less effective antimicrobial agents than other forms of chlorine, but release chlorine slowly and are thus longer lasting.
    • Used in wound dsgs, skin antiseptics, and in some municipal water supplies.
  129. Uses of bromine.
    • Effective disinfectant in hot tubs.
    • Evaporates more slowly than chlorine at high temps.
    • Also used as an alternative to chlorine in the disinfection of swimming pools, cooling towers, and other water containers.
  130. Uses of fluorine.
    • Works in part by disrupting metabolism in the biofilm of dental plaque.
    • In form of fluoride is antibacterial in drinking water and toothpastes, which can help reduce the incidence of dental cavities.
  131. What are three types of oxidizing agents?
    Peroxides, ozone, and peracetic acid.
  132. Oxidizing Agents
    • High-level disinfectants and antiseptics.
    • Work by releasing oxygen radicals by oxidizing their enzymes preventing metabolism.
    • Particularly effective against anaerobic microorganisms.
    • Health care workers use oxidizing agents to kill anaerobes in deep puncture wounds.
  133. Hydrogen Peroxide
    • Common household chemical that can disinfect and even sterilize surfaces of inanimate objects such as contact lenses.
    • Does not make a good antiseptic for open wounds because catalase (an enzyme released from damaged human cells) quickly neutralizes the hydrogen peroxide by breaking it down into water and oxygen gas, which can be seen as escaping bubbles.
    • Although aerobes and facultative anaerobes on inanimate surfaces also contain catalase, the volume of peroxide used as a disinfectant overwhelms the enzyme, making hydrogen peroxide a useful disinfectant.
  134. Uses of ozone (O3).
    • A reactive form of oxygen that is generated when molecular oxygen (O2) is subjected to electrical discharge.
    • Gives "fresh air" smell after a thunderstorm.
    • More effective antimicrobial agent than chlorine, but more expensive and difficult to maintain an effective concentration of ozone in water.
  135. Uses of peracetic acid.
    • Extremely effective sporicide used to sterilize surfaces.
    • Food processors and medical personnel use to sterilize equipment because it is not adversely affected by organic contaminants, and leaves no toxic residue.
  136. Surfactants
    "Surface active" chemicals that reduce the surface tension of solvents such as water by decreasing the attraction among molecules.
  137. What are two common surfactants?
    Soaps and detergents
  138. How is soap an effective surfactant?
    • One end of a soap molecule is hydrophobic (composed of fatty acids) and the other is hydrophilic and negatively charged. The hydrophobic ends are effective at breaking oily deposits into tiny droplets and the hydrophilic ends attract water molecules. The oily material and any bacteria they harbor are more easily dissolved in and washed away by water.
    • Good degerming agents, though poor antimicrobial agents (only antiseptic when they contain antimicrobial chemicals).
  139. Detergents
    Synthetically positively charged organic surfactants that are more soluble in water then soaps.
  140. Quaternary Ammonium Compounds or QUATS
    • Low-level disinfectants.
    • Most popular and are composed of an ammonium cation (NH4+) where the hydrogen atoms are replaced by other functional groups or hydrocarbon chains.
    • Function by disrupting cellular membranes so that affected cells lose essential internal ions, such as potassium.
    • Are not only antimicrobial, are colorless, tasteless, and harmless to humans (except at high concentrations).
    • Are bactericidal (particularly against Gram-positive bacteria), fungicial, and virucidal against enveloped viruses, but are not effective against nonenveloped viruses, mycobacteria, or endospores.
    • Action of QUATS are retarded by organic contaminants and deactivated by soaps.
    • Ideal for industrial and medical applications.
  141. Heavy Metal Ions
    • Ions of arsenic, zinc, mercury, silver, and copper.
    • Antimicrobial because they combine with sulfur atoms in molecules of cysteine, and amino acid.
    • Such bonding denatures proteins, inhibiting or eliminating their function.
    • Low-level bacteriostatic and fungistatic agents, with few exceptions their use has been superseded by more effective antimicrobial agents.
  142. What heavy metal ion was once used to treat the eyes of newborns to prevent blindless caused by Neisseria gonorrhoeae?
    • 1% silver nitrate (AgNO3)
    • Today silver still plays an antimicrobial role in some surgical dsgs, burn creams, and catheters.
  143. What heavy metal ion did drug companies use for over 70 years?
    • Thimerosal, a mercury-containing compound, to preserve vaccines.
    • In 1999, US Public Health Services recommended alternatives to be used because mercury is a metabolic poison, though the small amt of mercury in vaccines is considered safe.
    • Today only a few adult vaccines contain thimerosal (whole-cell pertussis and some against tetanus, flu, and meningococcal meningitis).
  144. Uses of copper.
    • Interferes with chlorophyll.
    • Are antibacterial.
    • Used to control algal growth in reservoirs, fish tanks, swimming pools, and water storage tanks.
    • In the absence of organic contaminants, copper is an effective algicide.
    • Copper, zinc, and mercury are also used to control mildew in paint.
  145. Aldehydes
    • Compounds containing terminal - CHO groups.
    • Function by cross-linking amino, hydroxyl, sulfhydryl, and carboxyl organic functional groups, denaturing proteins and inactivating nucleic acids.
  146. Aldehyde - Glutaraldehyde
    • A highly reactive chemical liquid compound.
    • Used by hospital personnel and scientists with 2% solutions to kill bacteria viruses, and fungi. A 10 minute treatment effectively disinfects most objects, including medical and dental equipment. A 10 hour treatment sterilizes.
    • Less irritating and more effective than formaldehyde, but more expensive.
  147. Aldehyde - Formaldehyde
    • A highly reactive chemical gas compound.
    • Morticians and health care workers use dissolved in water to make a 37% solution called formalin.
    • Used for embalming and to disinfect isolation rooms, exhaust cabinets, surgical instruments, and reusable kidney dialysis machines.
    • Must be handled with care because it irritates mucous membranes and is carcinogenic.
  148. Gaseous Agents
    • Highly reactive microbicidal and sporicidal gases, such as ethylene oxide, propylene oxide, and beta-propiolactone.
    • These gases rapidly penetrate paper and plastic wraps and diffuse into every crack.
    • Over time (usually 4-18 hours), they denature proteins and DNA by cross-linking organic functional groups, killing everything they contact without harming inanimate objects.
  149. What are the advantages of gaseous agents?
    Objects and equipment (such as heart-lung machine components, sutures, plastic lab ware, mattresses, pillows, artificial heart valves, catheters, electronic equipment, dried or powdered foods) that cannot be sterilized easily with heat or water-soluble chemicals; nor irradiation for it is not practical for large or bulky items can be sterilized within a closed camber with these highly reactive microbicidal and sporicidal gases.
  150. Gaseous Agent - Ethylene oxide
    • Frequently used as a gaseous sterilizing agent in hospitals and dental offices.
    • Large hospitals chambers, similar to autoclaves, to sterilize instruments and equipment sensitive to heat.
    • NASA uses the gas to sterilize spacecraft designed to land on other worlds, lest they accidentally export earthly microbes.
  151. What are some disadvantages of gaseous agents?
    • Can be extremely hazardous to the people using them.
    • Must be administered by combining them with 10%-20% nitrogen gas or carbon dioxide, because they are often highly explosive.
    • Extremely poisonous, so workers must extensively flush sterilized objects with air to remove every trace of the gas.
    • Potentially carcinogenic (especially beta-propiolactone)
  152. Antimicrobial Enzymes
    • Many organisms produce chemicals that inhibit or destroy a variety of fungi, bacteria, or viruses, these are enzymes that act against MO.
    • i.e. Human tears contain an enzyme lysozyme, which is a protein that digests the peptidoglycan cell wall of bacteria, causing the cell to rupture due to osmotic pressure and thus protecting the eye from most bacterial infections.
  153. Uses of antimicrobial enzymes.
    • Food processors: lysozyme to reduce number of bacteria in cheese.
    • Vintners: lysozyme instead of poisonous sulfur dioxide (SO2) to remove bacteria that would spoil wine.
  154. Why are scientists, food processors, and medical personnel researching ways to use natural and chemically modified antimicrobial enzymes?
    To control microbes in the enviornment, inhibit microbial decay on foods and beverages, and reduce the number and kids of microbes on medical equipment.
  155. In 2006, the European Union approved of what enzyme to safely and completely remove prions on medical instruments?
    • Prionzyme, the first certified, noncaustic chemical to target prions.
    • Prions, the highly infectious and deadly proteins may remain infectious even after normal autoclaving, boiling, exposure to aldehydes, ethylene oxide, or even 24 hours of dry heat at 160 degrees.
    • Until the development of Prionzyme, harsh methods such as autoclaving in sodium hydroxide for 30 minutes or complete incineration were required to eliminate prions.
  156. Antimicrobials
    • Include antibiotics, semisynthetics, and synthetics.
    • Typically used for treatment of disease, not for enviornmental control of microbes. Although, some are used for antimicrobial control outside the body. (i.e. Nisin and natamycin are used to reduce the growth of bacteria and fungi in cheese)
  157. Antibiotics
    Are antimicrobial chemicals produced naturally by MO.
  158. Semisynthetics
    When scientists chemically modify an antibiotic.
  159. Synthetic
    When scientists have developed wholly antimicrobial drugs.
  160. Resistant Microbes
    • Development of strains of microbes may become resistant to antimicrobial chemicals because, while susceptible cells die, resistant cells remain to proliferate.
    • i.e. TB, Ecoli, P. aeruginosa, and S. aureus, are less susceptible to common disinfectants and antiseptics.

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