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Sterilization
Destruction of all microorganisms in or on an object
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Antisepsis and Disinfection
- reduction in the number of microorganisms and viruses
- same thing except that antisepsis is for living objects and disinfectin is for inanimate objects and can be much stronger
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Degerming
- removing microbes by physical and mechanical means
- handwashing
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Sanitization
removal of pathogens from objects to meet public health standards
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Action of antimicrobial agents
- alteration of cell walls and membranes
- Damage to proteins and nucleic acids
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Ideally agents should be:
- Inexpensive
- Fast-acting
- Stable durnig storage
- Capable of controlling growth while being harmless to humans, animals, and objects
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Factors affecting the efficiency of Antimicrobial Methods
- Site to be treated
- Relative susceptability of microorganisms
- the higher the temperature, the higher the efficiacy of an antimicrobial agent
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Germicide Classification
- High-Level: kills all pathogens, including endospores, not prions
- Intermediate: kills fungal spores, protozoan cysts, viruses and pathogenic bacteria
- Low-Level: kill vegetative bacteria, fungi, protozoa, and some viruses
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Methods for Evaluating Disinfectants and Antiseptics (4)
- Phenol Coefficient
- Use-dilution test
- Kelsey Sykes Capacity test
- In-Use test
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Phenol Coefficient
- evaluates the efficiacy of agents by comparing it's ability to control microbes to phenol
- greater than 1.0 indicates agent is more effective
- has been replaced by newer methods
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Use-dilutin test
- metal cylinders dipped into broth cultures of bacteria
- cylinder immersed in dilution of disinfectant
- cylinders removed, washed, placed in medium
- effective agents prevent growth
- current standard test in U.S.
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Kelsey Sykes Capacity Test
- bacterial suspensions added to the chemical being tested
- samples removed at certain times and incubated
- lack of bacterial reproduction reveals minimum time requierd for the disinfectant to be effective
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In-Use Test
- swabs taken from objects before and after use of disinfectant
- swabs innoculated into medium and incubated
- medium monitored for growth
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Physical Methods of Microbial Control
- Heat-Related Methods
- Radiation
- Bio-Safety Levels
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Heat Related Methods of Microbial Control
- Effects of high temperature-denatures proteins, interferes with integrity of cytoplasmic membrane and cell wall
- Moist Heat
- Dry Heat
- Refrigeration and Freezing
- Dessication and Lyophilization
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Thermal Death Point
lowest temperature that kills all cells in broth in ten minutes
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Thermal death time
time to sterilize volume of liquid at certain temperature
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Decimal Reduction Time
Time required to destroy 90% of the microbes in a sample
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Moist Heat (Four methods)
- more effective than dry heat because water is a better conductor of heat than air
- boiling
- autoclaving
- pasteurization
- ultra high-temperature sterilization
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Moist Heat: Boiling
- kills vegetative cells of bacteria and fungi, protozoa, and most viruses
- boiling time is critical
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Moist Heat: Autoclaving
- pressure applied to boiling water prevents steam from escaping
- boiling temp increaes as pressure increases
- autoclave conditions: 121*C, 15 psi, 15 min
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Moist Heat: Pasteurization
- kills microbes by heating without raising the temp so much that the taste is ruined
- used for milk, yogurt, ice cream, and juices
- not sterilization: heat tolerant microbes survive
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Moist Heat: Ultrahigh-temperature sterilization
- 140*C for one second, then rapid cooling
- treated liquids can be stored at room temperature for months (dairy creamer)
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Dry Heat
- used for materials that cannot be sterilized with moist heat (powders and oils)
- denatures proteins and oxidezies metabolic and structural chemicals
- requires higher temps for longer times than moist heat
- incineration is ultimate means of sterilization
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Refrigeration and Freezing
- decrease microbial metabolism, growth, and reproduction
- chemical reactions occur slower
- slow freezing more effective than quick freezing due to ice crystals
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Dessication and Lyophilization
- drying inhibits growth due to removal of water
- Lyophilization- technique that combines freezing and drying (used for long-term preservation)
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Method of Microbial Control: Radiation (two kinds)
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Ionizing Radiation
- wavelength shorter than one nmelectron beams and gamma rays
- eject electrons from atoms to create ions, ions disrupt hyrdrogen bonding, oxidized double covalent bonds, and create hyrdroxide ions which denature DNA
- Electron beams- effective at killing, but doesnt penetrate well
- Gamma Rays-penetrate well but requires hours to kill
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Nonionizing Radiation
- wavelengths greater than one nmExcites electrons, causing them to make new covalent bonds (affects 3D structure of proteinds and nucleic acids)
- UV light causes pyrimidine dimers in DNA
- Suitable for disinfecting air, transparent fluids, and surfaces of objects
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Biosafety Levels
- Biosafetly Level 1:handling pathogens that don't cause disease in healthy humans (E.coli)
- Biosafety Level 2:handling of moderately hazardous agents, has potential to cause disease (our Micro Lab)
- Biosafety Level 3:handling of microbes in safety cabinets (space suits)
- Biosafety Level 4:handling of microbes that cause severe or fatal disease, there is no cure for these (isolated labs, islands)
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Chemical Methods of Microbial Control
- Affect microbes cell walls, membranes, proteins, or DNA
- often more effective against enveloped viruses and vegetative cells of bacteria, fungi, and protozoa
- Phenol and phenolics
- Alcohols
- Halogens
- Oxidizing Agents
- Surfactants
- Heavy Metals
- Aldehydes
- Gaseous Agents
- Enzymes
- Antimicrobials
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Chemical Methods of Microbial Control: Phenols and Phenolics
- intermediate to low level disinfectants
- denature proteins and disrupt cell membranes
- effective in presence of organic matter
- reamin active for prolonged time
- commonly used in health care settings,labs, and homes
- disagreeable odor and possible side effects
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Chemical Methods of Microbial Control: Alcohols
- intermediate level disinfectants
- denature proteins and disrupt cytoplasmic membrane
- more effective than soap in removing bacteria from hands
- swabbing skin with 70% before injecting (70% because microbes need water to metabolize)
- Advantage: doesn't leave residue
- Disadvantage: may not contact microbe long enough to be effective (high evaporation rate)
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Chemical Methods of Microbial Control: Halogens
- intermediate level anitmicrobial, non-metallic chemicals (iodine,chlorine, bromine, fluorine)
- effective against fungal spores and some bac. endospores
- damage enzymes via oxidation or by denaturation
- Widely used: water treatment, bleach, chlorine dioxide- buildings, anthrax)
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Chemical Methods of Microbial Control: Oxidizing Agents
- Peroxides, Ozone, and paracetic acid
- kill by oxidation of microbial enzymes
- high-level disinfectants and antiseptics that work by releasing oxygen radicals
- H2O2-sterilize surfaces, not wounds because of catalase
- Ozone-treatment of drinking water
- peracetic acid- effective sporocide used to sterilize equipment
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Chemical Methods of Microbial Control: Surfactants
- "surface active" chemicals
- reduce surface tension of solvents
- soaps and detergents
- Quartemery ammonium compounds (Quats)
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Quartemery ammonium compounds
- *Surfactants in which hydrogen atoms of an ammonium ion are replaced by other functional groups
- low level disinfectants that function by disrupting cellular membranes
- ideal for many medical and industrail applications
- not effective against non-enveloped viruses, mycobacterium, or endospores
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Chemical Methods of Microbial Control: Heavy Metals
- Heavy-Metal Ions denature proteins (arsenic, zinc, mercury, silver, copper)
- low-level bacteriostatic and fungaistatic agents
- thimerosal (mercury compound) used to preserve vaccines
- Copper controls algae growth by interfering with chlorophyll
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Chemical Methods of Microbial Control: Aldehydes
- Compounds containing terminal -CHO groups
- cross-link functional organic groups (like amino, hydroxyl,carboxyl) to dentaure proteins and inactivate nucleic acids
- Formalin- used in embalming and disinfecting of rooms and instruments
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Chemical Methods of Microbial Control: Gaseous Agents
- microbicidal and sporicidal gases used in closed chambers to sterilize items
- denature proteins and DNA by cross-linking functinoal groups
- used in hospitals and dentist offices
- Disadvantages: can be hazardous,highly explosive, poisonous, and potentially carcinogenic.
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Chemical Methods of Microbial Control: Enzymes
- antimicrobial enzymes act against microorganisms
- human tears contain lysozyme (digest peptid. cell wall in bacteria)
- Lysozyme used to decrease number of bacteria in cheese
- prionzyme- can remove prions on medical instruments
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Chemical Methods of Microbial Control: Antimicrobials
- antibiotics, semi-synthetic, and synthetic chemicals
- antibiotics-antimicrobial chemicals produced naturally by microorganisms
- semi-syntheic agents- chemically modified agents
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