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Fig. per05.01
V. Definitions: Control of microbial growth.
A. Biocide - general term describing a chemical agent with broad
spectrum activity that inactivates microorganisms - can be
antiseptic, disinfectant or sterilant.
B. -cidal / -cide - irreversible, meaning killer or killing
germicide: germ-killing agent
bactericide: kills bacteria
sporicide: kills bacterial endospores
fungicide: kills fungi
virucide: kills viruses.
C. -static / -stasis: growth is halted, when the agent is removed
microorganisms can grow.
D. Sterilization - the killing or removal of all microorganisms, including bacterial
spores. Does not destroy all bacterial products (LPS = exogenous pyrogen).
E. Disinfection - the killing of many but not all microorganisms. For adequate
disinfection pathogenic microorganisms must be killed. Disinfectants are toxic to human
tissues and should be used only on inanimate objects. Can be sporostatic or sporocidal.
F. Pasteurization - the use of heat at a temperature sufficient to kill important pathogens
in liquids, e.g. milk, egg products, but at a temperature below that needed for sterilization
(milk 740C for 3-5 sec). Spores are not killed.
G. Sanitization - clean and no pathogens. Does not imply sterilization or complete
disinfection.
H. Asepsis - literally, without infection. Refers to techniques that prevent entry of living
organisms.
I. Antiseptic - chemicals used to kill microorganisms on the surface of skin and mucous
membranes.
VI. Control of microbial growth concepts - Control of
microorganism numbers is critical, not always necessary to kill
everything. It is relatively easy to control microorganisms, yet it is
difficult to eliminate microbes or their products completely from
most environments (e.g. LPS).
A. Organisms differ in susceptibility to agents used in controlling
microbial growth.
What are you trying to destroy?
Prions (infectious proteins) - the most difficult to destroy
Endospores - Bacillus and Clostridium - only extreme
heat or strong chemical treatment will destroy them.
Mycobacterium - waxy cell wall make them resistant to
chemical treatments - stonger more toxic disinfectants needed.
Pseudomonas - very common found everywhere, but can grow
in phenols and are pathogenic.
Viruses - enveloped viruses (membrane and nucleocapsid)
more susceptible than naked viruses (nucleocapsid)
Factors affecting the killing of microorganisms: application time; number of organisms
and concentration of the agent.
A. If the concentration of the agent is increased, the length of time needed will
decrease for most agents (alcohol is the exception - in that case 70% is better than 100%).
Concentration/time is not a linear relationship. For instance, If phenol is diluted by 1/2, the
length of time to kill increases 64 times. It is imperative to follow label instructions
carefully.
B. Temperature - depending on the antimicrobial agent used for every 10 oC increase in
temperature the chemical activity doubles and killing time deceases.
C. pH (e.g. glutaraldehyde more active in alkaline environment) and presence of organic
matter (e.g. chlorine effectiveness decreases in the presence of organic matter).
VIII. Aspects of biocide selection or disinfection/sterilization technique: agent effective
against organism(s) in question; presence or absence of organic matter; cost; toxicity or
corrosiveness (e.g. I2 is both corrosive and toxic); time; smell; taste and esthetics
How many microbes present?
Decimal reduction time - D- value - time required for
killing 90% of a population of microbes under a certain
disinfection process.
Classification of medical devices in relation to infectious risk
Critical items are those which must be sterilized. These enter sterile
tissue or the blood stream (e.g. surgical instruments, cardiac and urinary
catheters and IV fluids).
Semi-critical items are those that come in contact with broken skin or mucus
membranes. These materials require treatment with disinfectants that have
high level of activity: kills Mycobacterium; endospores; vegetative bacteria;
fungi and viruses (e.g. endoscopes, reusable thermometers, respiratory therapy
equipment).
Non-critical may be cleaned and treated with low-level disinfectants:
kills Mycobacterium; some fungi and some viruses (e.g. blood pressure cuff,
electrocardiagram electrodes).
Medical equipment. Low level disinfection with at least anti-HIV activity
(e.g. examination table).
Methods for controlling microorganisms by sterilization. Monitoring of all
sterilization procedures is advised.
A. Temperature - kills by denaturation, burning.
1. Incineration - dressings and public health hazards. Dry heat - (oven;
180oC, 3 hrs.) glassware, bulk powders.
2. Steam under pressure - Autoclaving - 15 lb psi, 121 oC, 15 min. Easy,
safe, clean, least expensive for repeat use, rapid. Can't use for protein
containing solutions.
B. Radiation - extensive changes to DNA and ionizes other components
of cell (free radical formation).
1. Ionizing radiation - Cobalt 60. Bone grafts and food.
2. Non-ionizing radiation. Ultraviolet light (won't penetrate surfaces) and
microwave.
Table 05.02
Fig. 05.08
Chemical agents - many heat sensitive medical devices and surgical
supplies can be effectively sterilized with liquid/vapor sterilants.
Important to monitor effectiveness of these agents, since with time, some
need to be replaced. These agents are also used in mechanical sterilizers.
Good mechanical sterilizers should include a washing step before
sterilization, especially for endoscopes.
1. Ethylene oxide - causes macromolecule crosslinking. Ethylene oxide is
an explosive gas and mutagenic. Most commonly used biocide for
sterilization of temperature sensitive medical devices and supplies (plastics,
surgical materials) in U.S. Toxic residues can be removed by aeration.
2. Aldehydes - causes macromolecule crosslinking
a. Glutaraldehyde - less toxic and more effective than formaldehyde.
Important for low temperature disinfection and sterilization of endoscopes
and surgical equipment. Irritant.
b. Formaldehyde (formalin = 37% formaldehyde in solution). Solution
vaccine preservative (0.1%), 8% surfaces. Vapor can be used to sterilizefilters
and rooms. Exposure monitored - carcinogen. Use inhospitals is limited.
c. o-Pthalaldehyde - suggested as a replacement for glutaraldehyde in
endoscope disinfection. Does not require exposure monitoring, barely
perceptible odor, lasts longer than glutaraldehyde (but more expensive) in
endoscope reprocessor. Stains proteins gray (including unprotected skin).
3. Hydrogen peroxide vapor/gas plasma (gas consisting
of ions, electrons or neutral particles, commonly
produced in vacuum with electromagnetic field) - free
hydroxyl radicals cause DNA strand breakage and
oxidize and disrupt thiol groups in proteins. Used as low-
temperature liquid sterilant for medical devices (Sterrad 50
and 100). Is as effective as ethylene oxide and leaves no
toxic residue.
4. Peracetic acid - liquid sterilant (STERIS SYSTEM I),
mechanism of action similar to hydrogen peroxide. More
potent and greater activity in the presence of organic
contaminants, compared to hydrogen peroxide. Decomposes
to acetic acid and oxygen (non-toxic).
STERILIZATION METHODS
PHYSICAL Applications and notes Associated hazards
Incineration/heat Loops / dressings / public health hazards Destroys
Dry Heat/ heat Glassware / bulk powders Heat labile (unstable) substances
180C, 3 hrs
Autoclaving/ 15 lb. Broad;easy,safe ,inexpensive,rapid Protein containing solutions; heat
psi pressure, 1210C, labile substances destroyed
15 min.
Ionizing Bone grafts/ fo od/extensive changes to Radiation
Radiation/Cobalt 60 DNA and ionizes other components of
cell
UV light Surfaces/ air/extensive changes to DNA Won't penetrate surfa ces or glass
and ionizes other components of cell
Microwave/heat Must contain water Heat labile / utility not fu lly
electromagnetic explored
energy.
CHEMICAL
Ethylene oxide/gas Plastics, surgical materials; extensively Flamm able & mutagenic and
used in hospitals/crosslinks mo lecules toxic if not completely
eliminated, special equipment
needed.
Formaldehyde Vaccine preservative (0.1%) / surfaces Carcinogenic/ volatile / toxic to
(forma lin 37% (8%) / filters and rooms (12 hrs) /limited tissue in high concentrations
aqueous soln.) use/ crosslinks macromolecules
Glutaraldehyde/o- Heart valves / respiratory therapy Irritant/toxic/OPA less irritable
phthalaldehyde equipme nt, endoscopes etc. / less toxic then glutaraldehyde
(OPA) and more eff ective than formaldehyde;
activity not affec ted by the presence of
organic ma tter/crosslinks proteins
Peracetic acid (P AA) Medical devices , endoscopes, etc. U sed HP- nontoxic/ P AA- decomposes
/liquid with mechanical sterilizers/ denatures to safe b y-products (acetic acid
proteins and attacks sulfhydryl groups, and oxygen).
Hydrogen peroxide
disulfide bonds and DNA.
(HP)/gas plasma
PAA - Environmental surfa ce sterilant,
potency greater than HP
Methods for controlling microbial growth by disinfection or antisepsis.
A. Halogen releasing agents- oxidize thiol groups, disrupt oxidative
phophorylation and/or inhibit DNA synthesis.
1. Chlorine - lethal at highest concentrations, but is rapidly inactivated by
organic material. Used in potable water supply but protozoan cysts are not
inactivated, 5-10% works as a sanitizing agent, concentration must be
adjusted for the organic load.
2. Iodine. Iodophores - complex of I2 with detergent, most effective skin
antiseptic in use. Highly corrosive.
B. Hydrogen peroxide - liquid used as an antiseptic for wounds and to
disinfect contact lens.
C. Heavy metals - can cause DNA strand breakage and interact with
thiol groups in membrane bound enzymes. Silver is the most widely used
and new long-acting silver compounds are being tested. Crede procedure. -
AgNO3, 1% in eyes. 0.5% with antibiotic - can also be used on extensive
burns.
D. Membrane active compounds - disrupt membrane structure, proton
motive force and cause leakage of cytoplasmic contents.
1. Phenol - as a disinfectant, good when dealing with fecal contamination or soil.
Mouth sprays, tablets and antiseptics. Very toxic.
Triclosan (Bis-phenol) - inhibits bacterial fatty acid synthesis. Found in
majority of all U.S. household soap products sold, even if not mentioned
as an ingredient. Shown to select for bacteria with elevated levels of
antibiotic resistance.
2. Anionic detergents. Tide - sodium lauryl sulfate.
3. Alcohol. 70 - 95%. Requires 10-15 min. to kill. Evaporation is a problem.
Used to spot clean - alcohol wipes. Important in hand antisepsis for hospital
staff in alcohol gels and hand lotions.
4. Chlorhexidine - most widely used in antiseptic products (worldwide).
Irritant, but may be formulation dependent.
E. Heat.
1. Boiling. (10 min.) Doesn't kill endospores.
2. Pasteurization. (74oC, 3-5 sec.) - used for wine, milk and
vaccines and kills pathogens rapidly.
3. Refrigeration, freezing and freeze-drying (lyophilization).
F. Filtration (0.2 microns). For heat-labile solutions; removes
most bacteria.
Fig. 05.07
DECONTAMINATION AND DISINFECTION/ANTISEPSIS MEASURES
ANTISEPTICS Applications and notes Associated hazards.
Silver Crede Procedure: 1% in eyes / Heavy me tal
topical (0.5%) fo r burns/
interaction with thiol groups in
membrane proteins
Iodine/Iodophores Most eff ective skin antiseptic in Highly corrosive
use/inhibition of DNA
synthesis, oxidises thiol groups
and disrupts oxidative
phosphorylation
Hydrogen peroxide Wounds / contact lens/ Toxic in high concentrations.
denatures proteins and attacks
sulfhydryl groups, disulfi de
bonds and DNA
Anionic Detergents (Tide / Disrupt me mbranes/ Drying
Sodium Lauryl Sulfa te) clean wounds
Alcohol Spot clean/in gels or lotions- Requires 10-15 min to kill, thus
70-90% hand decontamination, increases evaporation is a problem.
compliance
DISINFECTION
Chlorine Lethal at highest concentrations Rapidly inactivated by organic
/ potable water/sanitizing agent/ material, thus concentration must
inhibition of DNA synthesis, be adjusted for organic load
oxidises thiol groups and /toxic
disrupts oxidative
phosphorylation
Good when dealing with fecal
Phenol Very toxic
contamination, soil/ damages
membranes
Boiling (10 min) Inactivation of toxins and some Doesn't kill endospores
vegetative bacteria and viruses
Pasteurization Kills pathogens / Does not render sterile
(740C, 3-5 sec) wine / milk / vaccines
Refr igeration, Freezing, Slows/halts growth of Some organisms grow in cold
fre eze-drying organisms /preserves organisms (psychrophiles)
(lyophilization)
Filtration (0.2 microns) For heat-labile Does not render sterile
solutions;removes mo st bacteria
F. Biocide resistance - has been reported on extensively. It is
unclear what impact evolution of biocide resistance has had on
human disease. Some biocide resistance mechanisms in bacteria
are linked to increases in antibiotic resistance levels. Slight but
measurable increases in biocide resistance, aids in survival of
potential pathogen in toxic environments?
1. Intrinsic resistance - mediated by unaltered chromosomally
located genes, can be dependent on structural state of
organism (spores and cysts). Some organisms can use biocides as
a carbon and energy source (e.g. Pseudomonas spp. - phenol),
others have permeability barriers (e.g. Mycobacterium - cell wall).
2. Plasmid-mediated and chromosomal mutations leading to
resistance.
a. qac - plasmid encoded - chlorhexidine resistance in
staphylococci. Heavy metal resistance also expressed on
plasmids.
b. Mar (multiple antibiotic resistance) mutants (e.g. E. coli)-
low level multiple antibiotic resistant mutants also resistant to
detergents, terpenes, triclosan and organic solvents.
Chromosomal-encoded multidrug efflux pumps play an important
role.
c. Triclosan resistance - chromosomal mutation (e.g. E. coli) in
enoyl-reductase (Fatty acid synthesis). Triclosan can select for
elevated antibiotic resistance in bacteria (e.g. Pseudomonas)
which is clinically relevant. Multidrug efflux pumps also play a
role in the latter mechanism.
Fig. pg.128b
3. Prions - infectious proteins that can cause disease - highly
resistant to many biocides. Chlorine releasing agents, sodium
hydroxide, some phenols and other agents are most effective
against these infectious proteins.
Barrier technique.
A. Hand hygiene - single most important factor in
preventing nosocomial infections.
1. Compliance with handwashing by health care workers is
never over 40%.
2. Adequate facilities with good antiseptic agents in the soap
are required.
3. Soaps containing chlorhexidine or triclosan can be used in
increase disinfection (preventing infections with common
hospital pathogens).
4. Alcohol-based rub-in cleanser for hand disinfection -
biologically more or as effective as handwashing with soaps
containing biocides; presence of dispensers near patients
beds and nurses desks, increases compliance.
B. Centers for Disease Control handwashing
guidelines.
1. Before performing invasive procedures.
2. Before taking care of particularly susceptible
patient.
3. Before and after touching wounds.
4. After situations where microbial contamination
of hands is likely to occur.
5. After touching inanimate objects that are likely
to be contaminated.
6. After taking care of an infected patient.
7. Between contacts with different patients in high
risk units.
C. Gloves prevent contamination of hands with organisms. Must be
changed when going from one patient to another, and also when
handling a contaminated area (wound) and then a clean area (vascular
catheter). Hand washing is still required upon glove removal
D. Gowns. Indicated when soiling is likely and should be disposed of
or sterilized
between patients.
E. Masks, Glasses, Face Shields. Necessary when caring for patients
under stringent types ofisolation. Used to protect personnel, isolated
patient and other patients.
F. Personal Hygiene other than handwashing to include keeping
fingernails short, neat and clean; hair kept clean, in an bundled or
short style or under a covering.
Table 05.03
Table 05.04a
Table 05.04b
