biology 120 lecture 5 2011 2012
TRANSCRIPT
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CONTROL OF MICROORGANISMS
Lecture 5Thursday, September 15, 2011
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LECTURE OUTLINEDefinition of Terms in Microbial Control
Pattern of Microbial Death
Conditions Influencing the Effectiveness of Antimicrobial Agents
Use of Physical Agents
Use of Chemical Agents
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Thursday, September 15, 2011
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The Control of Microbial Growth SEPSIS
microbial contamination
ASEPSIS absence of significant
contamination
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The Control of Microbial Growth SEPSIS
microbial contamination
ASEPSIS absence of significant
contamination
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The Control of Microbial Growth STERILIZATION
Removal of all microbial life
COMMERCIAL STERILIZATION Killing C. botulinum endospores
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The Control of Microbial Growth STERILIZATION
Removal of all microbial life
COMMERCIAL STERILIZATION Killing C. botulinum endospores
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The Control of Microbial Growth DISINFECTION
Removal of pathogens
ANTISEPSIS from living tissue
DEGERMING from a limited area
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The Control of Microbial Growth DISINFECTION
Removal of pathogens
ANTISEPSIS from living tissue
DEGERMING from a limited area
Thursday, September 15, 2011
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The Control of Microbial Growth SANITATION
Lower microbial counts on eating utensils
BIOCIDE/GERMICIDE: Kills microbes
BACTERIOSTATS: Inhibiting, not killing, microbes
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The Control of Microbial Growth SANITATION
Lower microbial counts on eating utensils
BIOCIDE/GERMICIDE: Kills microbes
BACTERIOSTATS: Inhibiting, not killing, microbes
Thursday, September 15, 2011
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Bacterial populations
die at a constant
logarithmic rate
PATTERN OF MICROBIAL DEATH
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Bacterial populations
die at a constant
logarithmic rate
PATTERN OF MICROBIAL DEATH
Thursday, September 15, 2011
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The Control of Microbial Growth
HOW DO WE DECIDE WHETHER THEY ARE ACTUALLY DEAD?
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The Control of Microbial Growth
HOW DO WE DECIDE WHETHER THEY ARE ACTUALLY DEAD?
Thursday, September 15, 2011
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The Control of Microbial Growth
HOW DO WE DECIDE WHETHER THEY ARE ACTUALLY DEAD?
“a microbe is defined DEAD if it does not grow when inoculated into culture medium
that would normally support its growth”Thursday, September 15, 2011
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Conditions Influencing Effectiveness of
Antimicrobial Agent Activity1. Population size
Larger population requires a longer time to die
2. Population compositionMicroorganisms vary markedly on susceptibilityVegetative versus SporesYoung versus Mature cells
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Conditions Influencing Effectiveness of
Antimicrobial Agent Activity1. Population size
Larger population requires a longer time to die
2. Population compositionMicroorganisms vary markedly on susceptibilityVegetative versus SporesYoung versus Mature cells
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3. Concentration or Intensity of an Antimicrobial Agent
The more concentrated an agent the more rapidly microbes can be destroyedBUT sometimes an agent may be more effective at lower concentrations (e.g. 70% alcohol)
4. Duration of ExposureThe longer the exposure to an agent the more they will be killed
Conditions Influencing Effectiveness of
Antimicrobial Agent Activity
Thursday, September 15, 2011
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3. Concentration or Intensity of an Antimicrobial Agent
The more concentrated an agent the more rapidly microbes can be destroyedBUT sometimes an agent may be more effective at lower concentrations (e.g. 70% alcohol)
4. Duration of ExposureThe longer the exposure to an agent the more they will be killed
Conditions Influencing Effectiveness of
Antimicrobial Agent Activity
Thursday, September 15, 2011
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5. TemperatureAn increase in temperature at which a chemical acts often enhances it activityExample: acids used in high T = more effective
6. Local environmentpH, organic matter, etcControls or Protects the pathogen
Conditions Influencing Effectiveness of
Antimicrobial Agent Activity
Thursday, September 15, 2011
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5. TemperatureAn increase in temperature at which a chemical acts often enhances it activityExample: acids used in high T = more effective
6. Local environmentpH, organic matter, etcControls or Protects the pathogen
Conditions Influencing Effectiveness of
Antimicrobial Agent Activity
Thursday, September 15, 2011
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Thursday, September 15, 2011
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PHYSICAL METHODS: HEAT
Fire and boiling Sufficient to destroy vegetative
cells (10 minutes) Not high for killing endospores Disinfection but not
sterilization!Thursday, September 15, 2011
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PHYSICAL METHODS: HEAT
Fire and boiling Sufficient to destroy vegetative
cells (10 minutes) Not high for killing endospores Disinfection but not
sterilization!Thursday, September 15, 2011
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Thermal Death Point (TDP)The lowest temperature at which a
microbial suspension in killed in 10 minutes
Thermal Death Time (TDT)The shortest time needed to kill all
organisms in a microbial suspension at a specific temperature and under defined conditions
PHYSICAL METHODS: HEAT
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Thermal Death Point (TDP)The lowest temperature at which a
microbial suspension in killed in 10 minutes
Thermal Death Time (TDT)The shortest time needed to kill all
organisms in a microbial suspension at a specific temperature and under defined conditions
PHYSICAL METHODS: HEAT
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However, such a destruction is logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy” microbes in a sample
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However, such a destruction is logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy” microbes in a sample
Thursday, September 15, 2011
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However, such a destruction is logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy” microbes in a sample
Decimal Reduction Time or D valueTime required to kill 90% of the
microorganisms or spores in a sample at a specified temperatureTime required for the line to drop by one
log cycle or tenfoldUsed to estimate the relative resistance
of a microbe to different temperaturesThursday, September 15, 2011
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PHYSICAL METHODS: HEAT
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PHYSICAL METHODS: HEAT
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z ValueThe increase in temperature required to
reduce D to 1/10 its value or to reduce it by one log cycle
PHYSICAL METHODS: HEAT
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z ValueThe increase in temperature required to
reduce D to 1/10 its value or to reduce it by one log cycle
F valueTime in minutes at a specific temperature
needed to kill a population of cells or sporesUsually 121°C
PHYSICAL METHODS: HEAT
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APPLICATION: FOOD INDUSTRY
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APPLICATION: FOOD INDUSTRY
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APPLICATION: FOOD INDUSTRY
After a food have been canned, it must be heated to eliminate the risk of botulism arising from the presence of Clostridium spores
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APPLICATION: FOOD INDUSTRY
After a food have been canned, it must be heated to eliminate the risk of botulism arising from the presence of Clostridium spores
Example: (1012 to 100 spores) IF the D value = 0.204 minutes It would take 12D or 2.5 minutes to
reduce the spore number by heating at the specified temperature
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APPLICATION: FOOD INDUSTRY
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APPLICATION: FOOD INDUSTRY
Thursday, September 15, 2011
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If the z value for Clostridium spores is 10°C
APPLICATION: FOOD INDUSTRY
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If the z value for Clostridium spores is 10°C It takes a 10°C change in temperature to
alter the D value tenfold
APPLICATION: FOOD INDUSTRY
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If the z value for Clostridium spores is 10°C It takes a 10°C change in temperature to
alter the D value tenfold
APPLICATION: FOOD INDUSTRY
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If the z value for Clostridium spores is 10°C It takes a 10°C change in temperature to
alter the D value tenfold
THUS: if the cans are to be processed at 111°C rather than 121°C, the D value would increase by tenfold t 2.04 minutes
APPLICATION: FOOD INDUSTRY
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If the z value for Clostridium spores is 10°C It takes a 10°C change in temperature to
alter the D value tenfold
THUS: if the cans are to be processed at 111°C rather than 121°C, the D value would increase by tenfold t 2.04 minutes The 12D value = 24.5 minutes
APPLICATION: FOOD INDUSTRY
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PHYSICAL METHODS: HEAT (MOIST HEAT
STERILIZATION)
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PHYSICAL METHODS: HEAT (MOIST HEAT
STERILIZATION)
Thursday, September 15, 2011
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Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
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Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
Thursday, September 15, 2011
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Flaming
Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
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Flaming
Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
Thursday, September 15, 2011
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Flaming
Incineration
Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
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Flaming
Incineration
Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
Thursday, September 15, 2011
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Flaming
Incineration
Hot-air sterilization
Hot-air Autoclave
Equivalent treatments 170˚C, 2 hr 121˚C, 15 min
PHYSICAL METHODS: HEAT (Dry Heat Sterlization)
Thursday, September 15, 2011
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HOW DOES HEAT KILL MICROBES
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HOW DOES HEAT KILL MICROBES
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HOW DOES HEAT KILL MICROBES
MOIST HEAT Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and other essential proteins May also disrupt cell membranes
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HOW DOES HEAT KILL MICROBES
MOIST HEAT Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and other essential proteins May also disrupt cell membranes
Thursday, September 15, 2011
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HOW DOES HEAT KILL MICROBES
MOIST HEAT Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and other essential proteins May also disrupt cell membranes
DRY HEAT Microbial death results from the oxidation of
cell constituents and denaturation of proteins
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PHYSICAL METHODS: FILTRATION
Applicable for heat-sensitive materials that needs sterilization
Types of filters Depth filters: consist of fibrous or granular materials that have been bonded into a thick layer filled with twisting channels of small diameter
Membrane filters: porous membranes; 0.2 µm pore sizes
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PHYSICAL METHODS: FILTRATION
Applicable for heat-sensitive materials that needs sterilization
Types of filters Depth filters: consist of fibrous or granular materials that have been bonded into a thick layer filled with twisting channels of small diameter
Membrane filters: porous membranes; 0.2 µm pore sizes
Thursday, September 15, 2011
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PHYSICAL METHODS: FILTRATION
Laminar flow hood versus biological safety cabinets (HEPA filters)
High Efficiency Particulate Air Remove 99.97% particles 0.02 µm for sterilizing AIR
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PHYSICAL METHODS: FILTRATION
Laminar flow hood versus biological safety cabinets (HEPA filters)
High Efficiency Particulate Air Remove 99.97% particles 0.02 µm for sterilizing AIR
Thursday, September 15, 2011
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BIOSAFETY CABINETS
Class1 (from room=outside) protection: person and environment
Class 2 (Type A and B) ) Type A and B: product, person and
environment difference: type A air is recirculated back
to room, type B exhausted outside the building
Class 3: contained facility, higher level of protection and containment
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BIOSAFETY CABINETS
Class1 (from room=outside) protection: person and environment
Class 2 (Type A and B) ) Type A and B: product, person and
environment difference: type A air is recirculated back
to room, type B exhausted outside the building
Class 3: contained facility, higher level of protection and containment
Thursday, September 15, 2011
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PHYSICAL METHODS: RADIATION
IONIZING RADIATION X rays, gamma rays, electron beamsExcellent as a sterilizing agent and penetrates deep into objects
NON-IONIZING RADIATION UV (about 260nm) Quite lethal but does not penetrate glass, dirt films, water and other substances very effectively
Microwaves: kill by heat not usually antimicrobial
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PHYSICAL METHODS: RADIATION
IONIZING RADIATION X rays, gamma rays, electron beamsExcellent as a sterilizing agent and penetrates deep into objects
NON-IONIZING RADIATION UV (about 260nm) Quite lethal but does not penetrate glass, dirt films, water and other substances very effectively
Microwaves: kill by heat not usually antimicrobial
Thursday, September 15, 2011
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Thursday, September 15, 2011
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CHEMICAL AGENTS
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PHENOLICS
ALCOHOLS
HALOGENS
HEAVY METALS
QUATERNARY AMMONIUM COMPOUNDS
ALDEHYDES
STERILIZING GASES
CHEMICAL METHODS
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PHENOLICS
ALCOHOLS
HALOGENS
HEAVY METALS
QUATERNARY AMMONIUM COMPOUNDS
ALDEHYDES
STERILIZING GASES
CHEMICAL METHODS
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CHEMICAL METHODS
Chemical agent Effectiveness againstEffectiveness againstEndospores Mycobacteria
Phenolics Poor GoodQuats None NoneChlorines Fair FairAlcohols Poor GoodGlutaraldehyde Fair Good
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CHEMICAL METHODS
Chemical agent Effectiveness againstEffectiveness againstEndospores Mycobacteria
Phenolics Poor GoodQuats None NoneChlorines Fair FairAlcohols Poor GoodGlutaraldehyde Fair Good
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First widely used antiseptic and disinfectant
Joseph Lister (1867): reduced the risk of infection during operations
Example: LYSOLR
Act by denaturing proteins and disrupting cell membranes
CHEMICAL METHODS: Phenolics
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First widely used antiseptic and disinfectant
Joseph Lister (1867): reduced the risk of infection during operations
Example: LYSOLR
Act by denaturing proteins and disrupting cell membranes
CHEMICAL METHODS: Phenolics
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Disruption of Cell Membranes
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ADVANTAGES: effective in the presence of organic material and remain active on surfaces long after application
DISADVANTAGE: disagreeable odor and can cause skin irritation and in some instances brain damage (hexachlorophene)
CHEMICAL METHODS: Phenolics
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ADVANTAGES: effective in the presence of organic material and remain active on surfaces long after application
DISADVANTAGE: disagreeable odor and can cause skin irritation and in some instances brain damage (hexachlorophene)
CHEMICAL METHODS: Phenolics
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Widely used disinfectant and antiseptics
Bactericidal and fungicidal but not sporicidal
May not destroy lipid-containing viruses
CHEMICAL METHODS: Alcohols
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Widely used disinfectant and antiseptics
Bactericidal and fungicidal but not sporicidal
May not destroy lipid-containing viruses
CHEMICAL METHODS: Alcohols
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DENATURES PROTEINS, DISSOLVES
LIPIDS
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Example: ethanol and isopropanol (70-80% concentration)
Act by denaturing proteins and possibly by dissolving membrane lipids
10-15 soaking in alcohol is sufficient to disinfect thermometers and small instruments
CHEMICAL METHODS: Alcohols
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Example: ethanol and isopropanol (70-80% concentration)
Act by denaturing proteins and possibly by dissolving membrane lipids
10-15 soaking in alcohol is sufficient to disinfect thermometers and small instruments
CHEMICAL METHODS: Alcohols
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Iodine Kills by oxidizing cell constituents
and iodinating cell proteins
Kill spores at high concentrations
DISADVANTAGE: a stain may be left (answer = iodophor)
CHEMICAL METHODS: Halogens
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Iodine Kills by oxidizing cell constituents
and iodinating cell proteins
Kill spores at high concentrations
DISADVANTAGE: a stain may be left (answer = iodophor)
CHEMICAL METHODS: Halogens
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Chlorine Usually for water supply
Kills by oxidation of cellular materials and destruction of vegetative bacteria, fungi
Will not kill spores
Death within 30 minutes
CHEMICAL METHODS: Halogens
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Chlorine Usually for water supply
Kills by oxidation of cellular materials and destruction of vegetative bacteria, fungi
Will not kill spores
Death within 30 minutes
CHEMICAL METHODS: Halogens
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Mercury, Arsenic, Zinc, Copper Used as germicides How do they Kill:
Heavy metals combine with proteins, often with their sulfhydryl groups and inactivate them May also precipitate cell proteins
CHEMICAL METHODS: Heavy Metals
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Mercury, Arsenic, Zinc, Copper Used as germicides How do they Kill:
Heavy metals combine with proteins, often with their sulfhydryl groups and inactivate them May also precipitate cell proteins
CHEMICAL METHODS: Heavy Metals
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DETERGENTSAmphipathic (both polar and non-polar
ends)Kill by disrupting microbial membranes and
denature proteins
ADVANTAGE: stable, non-toxicDISADVANTAGE: inactivated by hard
CHEMICAL METHODS: Quats
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DETERGENTSAmphipathic (both polar and non-polar
ends)Kill by disrupting microbial membranes and
denature proteins
ADVANTAGE: stable, non-toxicDISADVANTAGE: inactivated by hard
CHEMICAL METHODS: Quats
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Surface-Active Agents or SurfactantsSoap Degerming
Acid-anionic detergents
Sanitizing
Quarternary ammonium compoundsCationic detergents
Bactericidal, Denature proteins, disrupt plasma membrane
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FORMALDEHYDES Very reactive molecules that
combine with proteins and inactivate them
Sporicidal and can be used as sterilants
CHEMICAL METHODS: Aldehydes
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FORMALDEHYDES Very reactive molecules that
combine with proteins and inactivate them
Sporicidal and can be used as sterilants
CHEMICAL METHODS: Aldehydes
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EVALUATION OF ANTIMICROBIAL AGENT
EFFECTIVENESS PHENOL COEFFICIENT TEST
Best-known disinfectant screening test Potency of a disinfectant is compared
with that of phenol The highest dilution that killed bacteria
after a 10 minutes exposure are used to calculate phenol coefficient
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EVALUATION OF ANTIMICROBIAL AGENT
EFFECTIVENESS PHENOL COEFFICIENT TEST
Best-known disinfectant screening test Potency of a disinfectant is compared
with that of phenol The highest dilution that killed bacteria
after a 10 minutes exposure are used to calculate phenol coefficient
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The reciprocal of the appropriate test disinfectant dilution is divided by that for phenol to obtain the coefficient
Example: phenol dilution = 1/90 and the maximum effective dilution for disinfectant X = 1/450 Phenol coefficient = 5
CALCULATING PHENOL COEFFICIENTS
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The reciprocal of the appropriate test disinfectant dilution is divided by that for phenol to obtain the coefficient
Example: phenol dilution = 1/90 and the maximum effective dilution for disinfectant X = 1/450 Phenol coefficient = 5
CALCULATING PHENOL COEFFICIENTS
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CALCULATING PHENOL COEFFICIENTS
The higher the phenol coefficient value, the more effective the disinfectant under this conditions
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CALCULATING PHENOL COEFFICIENTS
The higher the phenol coefficient value, the more effective the disinfectant under this conditions
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DILUTION TESTS Metal rings dipped in test bacteria are dried
Dried cultures placed in disinfectant for 10 min at 20°C
Rings transferred to culture media to determine whether bacteria survived treatment
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DILUTION TESTS Metal rings dipped in test bacteria are dried
Dried cultures placed in disinfectant for 10 min at 20°C
Rings transferred to culture media to determine whether bacteria survived treatment
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DISK-DIFFUSION METHOD
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DISK-DIFFUSION METHOD
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CHEMOTHERAPEUTIC AGENTS
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CHEMOTHERAPEUTIC AGENTS
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Antibiotics are medicines used to treat infections caused by bacteria only
Infections are usually caused by bacteria or viruses
Antibiotics, therefore, do not cure all infections
Many infections like the common cold, flu, mild sore throat or diarrhea are caused by viruses
CHEMOTHERAPEUTIC AGENTS
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Antibiotics are medicines used to treat infections caused by bacteria only
Infections are usually caused by bacteria or viruses
Antibiotics, therefore, do not cure all infections
Many infections like the common cold, flu, mild sore throat or diarrhea are caused by viruses
CHEMOTHERAPEUTIC AGENTS
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WHAT IF ANTIBIOTICS WERE USED INCORRECTLY?
No healing effect - If antibiotics are used for viral infections, there will be no effect on the illness
Antibiotic resistance - This occurs when one antibiotic no longer works on a specific type of bacteria
A stronger antibiotic will be needed to treat the infection caused by this resistant strain of bacteria
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WHAT IF ANTIBIOTICS WERE USED INCORRECTLY?
No healing effect - If antibiotics are used for viral infections, there will be no effect on the illness
Antibiotic resistance - This occurs when one antibiotic no longer works on a specific type of bacteria
A stronger antibiotic will be needed to treat the infection caused by this resistant strain of bacteria
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ANTIBIOTIC MECHANISMS
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ANTIBIOTIC MECHANISMS
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RESISTANCE
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RESISTANCE
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RESISTANCEThursday, September 15, 2011
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RESISTANCEThursday, September 15, 2011
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DO YOU CONTRIBUTE TO RESISTANCE?
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DO YOU CONTRIBUTE TO RESISTANCE?
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DO YOU CONTRIBUTE TO RESISTANCE?
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DO YOU CONTRIBUTE TO RESISTANCE?
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Another factor that contributes to resistance is that when patients are prescribed antibiotics for a just cause, many do not finish their medication
This allows resistant bacteria to survive more easily
The practice of saving unused medication to treat themselves or others at a later date can also lead to resistant strains
DO YOU CONTRIBUTE TO RESISTANCE?
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DO YOU CONTRIBUTE TO RESISTANCE?
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DO YOU CONTRIBUTE TO RESISTANCE?
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Also contributing to antibiotic resistance is the widespread use of antibiotics to promote weight gain and to control disease in cattle, pigs, and chickens
Forty to fifty percent of antibiotics produced are used in livestock feed
This leads to an increase of resistant bacteria in these animals, which is then spread to humans
DO YOU CONTRIBUTE TO RESISTANCE?
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ANY QUESTIONS???
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NEXT MEETING: INTERACTIVE LECTURE/QUIZ ON METABOLISM
Thursday, September 15, 2011