Exam 2 Physical and Chem Control 10/12 Flashcards
sterilization
destruction or removal of all viable organisms
Sterilants
disinfection
killing, inhibition, or removal of disease causing organisms
Disinfectants
sanitization
reduction of microbial population
antisepsis
Destruction or inhibition of microorganisms on living tissue
Antiseptics
chemotherapy
use of chemicals to kill or inhibit growth of microorganisms within host tissue
agents that kill microorganisms or inhibit their growth
cidal agents kill
static agents inhibit growth
The Pattern of Microbial Death
microorganisms are not killed instantly
population death usually occurs exponential
measure of agent’s killing efficiency
decimal reduction time (D-value) – time to kill 90% of microorganisms and spores in a sample under specific conditions
Conditions Influencing the Effectiveness of Antimicrobial Agent Activity
1) population size
larger populations take longer to kill than smaller populations
2) population composition
microorganisms differ markedly in their sensitivity to antimicrobial agents
3) concentration or intensity of an antimicrobial agent
usually higher concentrations or intensities kill more rapidly (not always, e.g., EtOH)
relationship is not linear
4) Contact time - duration of exposure
longer exposure more organisms killed
5) temperature
higher temperatures usually increase amount of killing
6) local environment
many factors (e.g., pH, viscosity, and concentration of organic matter) can profoundly impact effectiveness
Physical Control Methods
Filtration
Heat
Radiation
Filtration
Physical removal of microbes by filtration
Filtration has been a method of purifying liquids for centuries
Lab applications use membrane nylon/Teflon filters with a pore size of 0.2 or 0.45 μm
Viruses can be removed from liquids by ultrafiltration methods (reducing pore size 10 to 100 nm)
Problems can result, though…
Large particles clog filters.
Ultrafiltration requires high pressure.
Viscous liquids don’t filter well.
Membrane Filter Sterilization
-Removes microorganisms from heat sensitive liquids
Biological Safety Cabinet and other filtration devices
N-95 masks removes 95% of particles 0.3um or larger
high-efficiency particulate air (HEPA) filters
used in laminar flow biological safety cabinets
Removes 99.97% of particles 0.3 um or larger
Temperature
Heat
-proteins and nucleic acids disrupted
Freezing
-Can damage cells by forming ice crystals
-Can stop biochemical reactions in microbes
Steam Sterilization
must be carried out above 100oC which requires saturated steam under pressure
carried out using an autoclave (Developed by Chamberland, 1884)
(121 C at 15 psi for 10-15 min)
effective against all types of microorganisms including spores
*^ spores usually have a harder time living in high pressure vs just high temperature, so to sterilize we need to apply pressure
quality control – indicator tape or ampules of Geobacillus stearothermophilus
Tyndallization
Developed by John Tyndall
Intermittent sterilization
30-60 min of steam repeated 3 times with 23-24 hr incubations between
Used to kill spores – spores germinate and are then killed by the steam treatment
Pasteurization
controlled heating at temperatures well below boiling (55 - 60 C)
used for milk, beer, and other beverages
process does not sterilize but does kill pathogens present and slows spoilage by reducing the total load of organisms present
Introduced to US in 1889
Dry Heat Sterilization
less effective than moist heat sterilization, requiring higher temperatures and longer exposure times
items subjected to 160–170oC for 2 to 3 hours
E.g., C. botulium – 5 min at 121C or 2 hrs at 160C
oxidizes cell constituents and denatures proteins
Dry Heat Incineration
bench top incinerators are used to sterilize inoculating loops used in microbiology laboratories
Radiation-UV
Using electromagnetic radiation to control microbes
UV radiation of 260 to 280 nm wavelengths can damage DNA, forming thymine dimers.
Ionizing Radiation
gamma 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
Chemical Control Agents
Disinfection
Antisepsis
Sterilization
Chemical Methods:
Chemical methods of controlling microbes
Disinfectants: Chemicals used on non-living surfaces to kill pathogenic microbes.
Antiseptics: Chemicals that can be used on living tissue to kill pathogenic
What makes a good chemical agent?
Should kill a wide-range of microbes
Shouldn’t be corrosive or overly toxic
Shouldn’t leave a residue
Shouldn’t emit fumes or smell TOO bad
Should be cheap
Should be temperature stable
Which Method to use?
How do you pick a method?
What microbes are present? What about endospores?
How many microbes are present? Do they all need to die?
What kind of object needs to be treated?
If using a physical method, how long/intense does it need to be?
If using a chemical method, how powerful must it be, and how long does it need to be applied?
Do we need to worry about toxicity to humans or other life?
Phenolics
Phenol was 1st widely used antiseptic and disinfectant. Used in 1867 by Joseph Lister
commonly used as laboratory and hospital disinfectants (e.g., Lysol)
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
E.g., triclosan is used in hand sanitizers
Aldehydes
commonly used agents are formaldehyde and glutaraldehyde
highly reactive molecules
sporicidal and can be used as chemical sterilants
combine with and inactivate nucleic acids and proteins
Alcohols
among the most widely used disinfectants and antiseptics
two most common are ethanol and isopropanol (60 – 80 %)
not sporicidal
inactivate some viruses
denature proteins and possibly dissolve membrane lipids
Halogens - Iodine
skin antiseptic
oxidizes cell constituents and iodinates proteins
at high concentrations kills spores
skin damage, staining, and allergies can be a problem
iodophore
Halogens - Chlorine
oxidizes cell constituents
destroys vegetative bacteria and fungi,
chlorine gas is sporicidal
can react with organic matter to form carcinogenic compounds
E.g., Bleach is highly effective (10% with a little vinegar)
Heavy Metals
e.g., ions of mercury, silver, arsenic, zinc, and copper
effective but usually toxic
combine with and inactivate proteins; may also precipitate proteins
Quaternary Ammonium Compounds
amphipathic organic cleansing agents
act as wetting agents and emulsifiers
Disrupt membranes and denature proteins
kill most bacteria, but not Mycobacterium tuberculosis or endospores
safe and easy to use, but inactivated by hard water and soap
Evaluating a disinfectant
Disk-diffusion method
Sterilizing Gases
used to sterilize heat-sensitive materials like plastics
microbiocidal and sporicidal
ethylene oxide sterilization is carried out in equipment resembling an autoclave
EtO is a very strong alkylating agent
betapropiolactone and vaporized hydrogen peroxide
Which physical or chemical agents would be the best choice for sterilizing the following items: glass pipettes, tryptic soy broth tubes, antibiotic solution, plastic-wrapped Petri plates?
