1. Control of Microorganisms by Physical and Chemical Agents
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Chapter 7
Control of Microorganisms by Physical and Chemical Agents

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3. Definition of Frequently Used Terms
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Definition of Frequently Used Terms
sterilization
destruction or removal of all viable organisms
used on inanimate objects
disinfection
killing, inhibition, or removal of pathogenic organisms
disinfectants
agents, usually chemical, used for disinfection
usually used on inanimate objects

4. More definitions… sanitization antisepsis
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More definitions…
sanitization
reduction of microbial population to levels deemed safe (based on public health standards)
antisepsis
prevention of infection of living tissue by microorganisms
antiseptics
chemical agents that kill or inhibit growth of microorganisms when applied to tissue

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Antimicrobial agents
agents that kill microorganisms or inhibit their growth
-cidal agents kill
-static agents inhibit growth

6. -cidal agents -cide suffix indicating that agent kills germicide
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-cidal agents
-cide
suffix indicating that agent kills
germicide
kills pathogens and many nonpathogens but not necessarily endospores
include bactericides, fungicides, algicides, and viricides

7. -static agents -static suffix indicating that agent inhibits growth
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-static agents
-static
suffix indicating that agent inhibits growth
include bacteriostatic and fungistatic

8. The Pattern of Microbial Death
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The Pattern of Microbial Death
microorganisms are not killed instantly
population death usually occurs exponentially
microorganisms were previously considered to be dead when they did not reproduce in conditions that normally supported their reproduction
however we now know that organisms can be in a viable but nonculturable (VBNC) condition
once they recover they may regain the ability to reproduce and cause infection

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10. The pattern of microbial death
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The pattern of microbial death
D = decimal reduction time

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Conditions Influencing the Effectiveness of Antimicrobial Agent Activity
population size
larger populations take longer to kill than smaller populations
population composition
microorganisms differ markedly in their sensitivity to antimicrobial agents

12. More conditions… concentration or intensity of an antimicrobial agent
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More conditions…
concentration or intensity of an antimicrobial agent
usually higher concentrations or intensities kill more rapidly
relationship is not linear
duration of exposure
longer exposure  more organisms killed

13. More conditions… temperature
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More conditions…
temperature
higher temperatures usually increase amount of killing
local environment
many factors (e.g., pH, viscosity and concentration of organic matter) can profoundly impact effectiveness
organisms in biofilms are physiologically altered and less susceptible to many antimicrobial agents

14. The Use of Physical Methods in Control
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The Use of Physical Methods in Control
heat
low temperatures
filtration
radiation

15. Moist heat sterilization
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Moist heat sterilization
must be carried out above 100oC which requires saturated steam under pressure
carried out using an autoclave
effective against all types of microorganisms including spores
degrades nucleic acids, denatures proteins, and disrupts membranes

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17. The autoclave or steam sterilizer
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The autoclave or steam sterilizer

18. Pasteurization controlled heating at temperatures well below boiling
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Pasteurization
controlled heating at temperatures well below boiling
used for milk, beer and other beverages
process does not sterilize but does kill pathogens present and slow spoilage by reducing the total load of organisms present

19. Dry heat sterilization
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Dry heat sterilization
less effective than moist heat sterilization, requiring higher temperatures and longer exposure times
items subjected to oC for 2 to 3 hours
oxidizes cell constituents and denatures proteins

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Dry heat incineration
bench top incinerators are used to sterilize inoculating loops used in microbiology laboratories

21. Measuring Heat-Killing Efficiency
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Measuring Heat-Killing Efficiency
thermal death time (TDT)
shortest time needed to kill all microorganisms in a suspension at a specific temperature and under defined conditions
decimal reduction time (D or D value)
time required to kill 90% of microorganisms or spores in a sample at a specific temperature

22. Z value increase in temperature required to reduce D by 1/10
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Z value
increase in temperature required to reduce D by 1/10

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24. Low temperatures freezing refrigeration
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Low temperatures
freezing
stops microbial reproduction due to lack of liquid water
some microorganisms killed by ice crystal disruption of cell membranes
refrigeration
slows microbial growth and reproduction

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Filtration
reduces microbial population or sterilizes solutions of heat-sensitive materials by removing microorganisms
also used to reduce microbial populations in air

26. Filtering liquids depth filters membrane filters
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Filtering liquids
depth filters
thick fibrous or granular filters that remove microorganisms by physical screening, entrapment, and/or adsorption
membrane filters
porous membranes with defined pore sizes that remove microorganisms primarily by physical screening

27. Membrane filter sterilization
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Membrane filter sterilization

28. Nylon membrane with 0.2 μm pores
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Nylon membrane with 0.2 μm pores
Polycarbonate membrane with 0.4 μm pores

29. Filtering air surgical masks cotton plugs on culture vessels
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Filtering air
surgical masks
cotton plugs on culture vessels
high-efficiency particulate air (HEPA) filters
used in laminar flow biological safety cabinets
laminar flow biological safety cabinet

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31. Ultraviolet (UV) Radiation
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Ultraviolet (UV) Radiation
limited to surface sterilization because it does not penetrate glass, dirt films, water, and other substances
has been used for water treatment
UV treatment system for disinfection of water

32. Ionizing Radiation penetrates deep into objects
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Ionizing Radiation
penetrates deep into objects
destroys bacterial endospores; not always effective against viruses
used for sterilization and pasteurization of antibiotics, hormones, sutures, plastic disposable supplies, and food

33. Sterilization with ionizing radiation Symbol on irradiated materials
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Sterilization with ionizing radiation
Irradiation machine that uses radioactive cobalt 60 as gamma radiation sources.
Fruits, vegetables, meats, fish, and spices.
Symbol on irradiated materials

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36. Chemical Control Agents - Disinfectants and Antiseptics
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Chemical Control Agents - Disinfectants and Antiseptics

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Phenolics

38. Phenolics commonly used as laboratory and hospital disinfectants
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Phenolics
commonly used as laboratory and hospital disinfectants
act by denaturing proteins and disrupting cell membranes
tuberculocidal, effective in presence of organic material, and long lasting
disagreeable odor and can cause skin irritation

39. Alcohols bactericidal, fungicidal, but not sporicidal
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Alcohols
bactericidal, fungicidal, but not sporicidal
inactivate some viruses
denature proteins and possibly dissolve membrane lipids

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Halogens
any of five elements: fluorine, chlorine, bromine, iodine, and astatine
iodine and chlorine are important antimicrobial agents

41. Halogens - iodine skin antiseptic
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Halogens - iodine
skin antiseptic
oxidizes cell constituents and iodinates proteins
at high concentrations may kill spores
skin damage, staining, and allergies can be a problem
Iodophore (e.g., Betadine)
iodine complexed with organic carrier
Water soluble, stable, nonstaining, and release iodine slowly
Minimize skin burns and irritations

42. Halogens - chlorine oxidizes cell constituents
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Halogens - chlorine
oxidizes cell constituents
important in disinfection of water supplies and swimming pools, used in dairy and food industries, effective household disinfectant
destroys vegetative bacteria and fungi, but not spores
can react with organic matter to form carcinogenic compounds

43. Heavy Metals ions of mercury, silver, arsenic, zinc, and copper
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Heavy Metals
ions of mercury, silver, arsenic, zinc, and copper
effective but usually toxic
combine with and inactivate proteins; may also precipitate proteins

44. Quaternary Ammonium Compounds
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Quaternary Ammonium Compounds

45. Quaternary Ammonium Compounds
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Quaternary Ammonium Compounds
detergents that have antimicrobial activity and are effective disinfectants
organic molecules with hydrophilic and hydrophobic ends (i.e. amphipathic)
act as wetting agents and emulsifiers
cationic detergents are effective disinfectants
kill most bacteria, but not Mycobacterium tuberculosis or endospores
safe and easy to use, but inactivated by hard water and soap

46. Aldehydes highly reactive molecules
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Aldehydes
highly reactive molecules
sporicidal and can be used as chemical sterilants
combine with and inactivate nucleic acids and proteins

47. Effects of glutaraldehyde
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Effects of glutaraldehyde
2% buffered solution is effective in disinfection

48. Sterilizing Gases used to sterilize heat-sensitive materials
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Sterilizing Gases
used to sterilize heat-sensitive materials
microbicidal and sporicidal
combine with and inactivate proteins

49. Ethylene oxide sterilizer
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Ethylene oxide sterilizer

50. Chemotherapeutic Agents
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Chemotherapeutic Agents
chemicals that can be used internally to kill or inhibit the growth of microbes within host cells
their selective toxicity allows them to target the microbe without harming the host
most are antibiotics, chemicals synthesized by microbes that are effective in controlling the growth of bacteria

51. Evaluation of Antimicrobial Agent Effectiveness
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Evaluation of Antimicrobial Agent Effectiveness
complex process regulated by US federal agencies
Environmental Protection Agency
Food and Drug Administration
phenol cofficient test
testing of antimicrobial agents to see if they are effective and at what concentrations
potency of a disinfectant is compared with that of phenol

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53. Other evaluation methods
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Other evaluation methods
use dilution test
determines dilution rate at which selected bacteria are destroyed by various chemical agents
use stainless steel cylinders, specific bacterial species, and appropriately diluted test disinfectants
determine disinfectant concentration that kills test organisms with a 95% level of confidence
in-use testing
testing done using conditions that approximate normal use of disinfectant