Terminology of Microbial Control
I. Sterilization (all microbes)
sterilization
 removal or destruction of all microbes,
including viruses and bacterial
endospores, in or on an object
 the eradication of harmful
microorganisms and viruses
aseptic
 environment or procedure that is free of
contamination of pathogens
disinfection
 destruction of most microorganisms and
viruses on nonliving tissue (inanimate
objects)
 use of disinfectants that are more
concentrated than antiseptics and can be
left on surface for longer periods of time
antisepsis
 reduction in the number of
microorganisms and viruses, particularly
potential pathogens, on living tissues
 use of antiseptic
degerming
 removal of microbes from a surface by
scrubbing, such as when you wash your
hands, or a nurse prepares an area of skin
for an injection
 chemicals such as soap or alcohol are
commonly used
sanitization
 process of disinfecting places and utensils
used by the public to reduce the number of
pathogenic microbes to meet accepted
public health standard
pasteurization
 use of heat to kill pathogens and reduce
the number of spoilage microorganisms in
food or beverages
-stasis or -static
 indicate that a chemical or physical agent
inhibits microbial metabolism and growth
but does not necessarily kill microbes
-cide or -cidal
 indicate the agents that destroy or
permanently inactivate a particular
microbe
2 Major Types of Microbial Control
II. Disinfection or antisepsis (vegetative cells and
many viruses)
a. Degerming
b. Sanitization
c. Pasteurization
microbial death
 permanent loss of reproductive ability
under ideal environmental conditions
microbial death rate
 calculating technique for evaluating the
efficacy of an antimicrobial agent which is
usually found to be constant over time for
any particular microorganisms under a
particular set of conditions
Action of Antimicrobial Agents
Alteration of Cell Walls and Membranes
 a cell wall maintains cellular integrity by
counteracting the effects of osmosis when
the cell is in a hypotonic solution
 when damaged, effects of osmosis
cause cells to burst
 cytoplasmic membrane acts as a bag that
contains the cytoplasm and controls the
passage of chemicals into and out of the
cell
 when damaged, cellular contents
leak out
 enveloped viruses have envelope that acts
as a membrane that is responsible for the
attachment of the virus to its target cell
 when damaged, viral replication is
prevented
 nonenveloped viruses can better tolerate
harsh conditions
Damage to Proteins and Nucleic Acids
 proteins regulate cellular metabolism,
function as enzymes in most metabolic
reactions, and form structural
components in membranes and
cytoplasm
 denatured proteins cease to
function, bringing about cellular
death
 genes of a cell or virus are composed of
nucleic acids
 chemicals, radiation, and heat can
alter or destroy nucleic acids
 disruption can produce fatal
mutations
 halt protein synthesis through
action on RNA
The Selection of Microbial Control
Methods
Ideal agents are:
 inexpensive
 fast-acting
 stable during storage
 capable of controlling microbial growth
while being harmless to humans, animals,
and objects
Factors affecting the efficacy of Antimicrobial
Methods:
Site to be treated
 harsh chemicals and extreme heat cannot
be used on humans, animals, and fragile
objects
 when performing medical procedures,
medical personnel must choose a method
and level of microbial control based on
the site of the procedure
Relative Susceptibility of Microorganisms
 most resistant microbes:
a. Bacterial endospores
o Endospores of Bacillus and
Clostridium are the most resilient
forms of life
o Can survive environmental
extremes of temperature, acidity,
and dryness and can withstand
many chemical disinfectants
b. Species of Mycobacteria
o Cell walls of members of the
genus Mycobacterium such as
Mycobacterium tuberculosis,
contain a large amount of a waxy
lipid
o The wax allows the bacteria to
survive drying and protects them
from most water-based
chemicals
o Must use strong disinfectants or
heat
c. Cysts of Protozoa
o Protozoan cyst’s wall prevents entry
of most disinfectants, protects
against drying, and shields against
radiation and heat
 Effectiveness of germicides: (depends on
the proficiency in inactivating or
destroying microorganisms that cannot
be sterilized with heat)
a. High-level germicides
o Kill all pathogens, including
bacterial endospores
o Used to sterilize invasive
instruments
b. Intermediate-level germicides
o Kill fungal spores, protozoan
cysts, viruses, and pathogenic
bacteria, but not bacterial
endospores
o used to disinfect instruments
that come in contact with
mucous membranes but are
noninvasive
c. Low-level germicides
o Eliminate vegetative bacteria,
fungi, protozoa, and some
viruses
o Used to disinfect items that
contact only the skin
Environmental Conditions
 Temperature and pH affect microbial
death rates and the efficacy of
antimicrobial methods
 Acidic conditions enhance the
antimicrobial effect of heat
 Clean objects before sterilization or
disinfection so that antimicrobial agents
can thoroughly contact all the object’s
surfaces
Biosafety Levels:
Biosafety Level 1 (BSL-1)
 Handling microbes that do not cause
disease in healthy humans
 Precautions are minimal and include
handwashing with antibacterial soap and
disinfecting surfaces
Biosafety Level 2 (BSL-2)
 Handling moderately hazardous agents
 Procedures that may produce aerosols
are conducted within safety cabinets
Biosafety Level 3 (BSL-3)
 Stricter, requiring manipulations be done
within safety cabinets containing high-
efficiency particulate air (HEPA) filters
 Entry through double sets of doors and
lower air pressure in the laboratory
Biosafety Level 4 (BSL-4)
 Handling dangerous or exotic microbes
that cause severe or fatal diseases in
humans
  Lab space is isolated and personnel wear
protective suits
Physical Methods of Microbial Control
Heat-Related Methods
 Heat – older and more common means of
microbial control
 Effects of high temperature:
 Denature proteins
 Interfere with the integrity of
cytoplasmic membranes and cell
walls
 Disrupt the function and structure
of nucleic acids
 Thermal death point – lowest
temperature that kills all cells in a broth
in 10 minutes
 Thermal death time – time it takes to
completely sterilize a particular volume of
liquid at a set temperature
 Decimal reduction time (D) – time
required to destroy 90% of the microbes
in a sample
Moist Heat
 Commonly used to disinfect, sanitize,
sterilize, and pasteurize
 Kills cells by denaturing proteins and
destroying cytoplasmic membranes
 More effective in microbial control than
dry heat because water is a better
conductor of heat than air
 Methods of moist heat:
a. Boiling
 Kills the vegetative cells of bacteria
and fungi, the trophozoites of
protozoa, and most viruses within
10 minutes at sea level
 Boiling time is the critical factor –
different elevations require
different boiling time
 Bacterial endospores, protozoan
cysts, and some viruses can
survive boiling
 Not recommended when true
sterilization is required
 Effective for sanitizing restaurant
tableware or disinfecting baby
bottles
b. Autoclaving
 True sterilization using heat
requires higher temperatures than
that of boiling water
 Pressure is applied to boiling
water to prevent escape of heat
 Temperature of 121°C, which
requires the addition of 15 psi of
pressure above that of normal
pressure, destroys all microbes in
a small volume in about 15
minutes
 Autoclave
o used to sterilize chemicals and
objects that can tolerate moist
heat
o consists of pressure chamber,
pipes to introduce and evacuate
steam, valves to remove air and
control pressure, and pressure
and temperature gauges to
monitor the procedure
c. Pasteurization
 Developed by Louis Pasteur which
is a method of heating beer and
wine just enough to destroy the
microorganisms that cause
spoilage without ruining the taste
 Used to kill pathogens in milk, ice
cream, yogurt, and fruit juices
 It is not sterilization because heat-
tolerant and heat-loving microbes
survive
 Pasteurization of milk:
o Batch method – 30 minutes at
63°C
o Flash pasteurization – high-
temperature, short-time method
wherein milk flows through
heated tubes that raise its
temperature to 72°C for only 15
seconds
o Ultra-high-temperature
pasteurization – heats the milk
to at least 135°C for only 1
second
d. Ultra-High-Temperature Sterilization
 Involves flash heating milk or
other liquids to rid them of all
living microbes
 Involves passing the liquid through
superheated steam at about 140°C
for 1 to 3 seconds
 Treated liquids can be stored at
room temperature
Dry Heat
 Used for materials that cannot be
sterilized with moist heat
 Hot air is an effective sterilizing agent
because it denatures proteins and fosters
the oxidation of metabolic and structural
chemicals
 Requires higher temperatures for longer
times than moist heat because dry heat
penetrates more slowly
Refrigeration and Freezing
 Decrease microbial metabolism, growth,
and reproduction because chemical
 reactions occur more slowly at low
temperatures because liquid water is not
available at subzero temperatures
 Refrigeration halts the growth of most
pathogens, which are predominantly
mesophiles
 Slow freezing, during which ice crystals
have time to form and puncture cell
membranes, is more effective than quick
freezing in inhibiting microbial
metabolism
Desiccation and Lyophilization
 Desiccation (drying)
 Preserve foods such as fruits, peas,
beans, grain, nuts, and yeast
 Inhibits microbial growth because
metabolism requires liquid water
 Lyophilization (freeze-drying)
 A technique that combines freezing
and drying, to preserve microbes and
other cells for many years
 Used for long-term preservation of
microbial cultures
 Prevents formation of damaging ice
crystals
Filtration
 Passage of a fluid through a sieve
designed to trap particles and separate
them from the fluid
 Viruses (filterable viruses) – pathogens
that are too small to be trapped in the
pores of filters
 Use filtration to estimate the number of
microbes in a fluid by counting the
number deposited on the filter after
passing a given volume through the filter
 Health care and laboratory workers
routinely use filtration to prevent
airborne contamination by microbes
Osmotic Pressure
 Use of high concentrations of salt and
sugar in foods to inhibit microbial growth
by osmotic pressure
 Removal of water inhibits cellular
metabolism because enzymes are fully
functional only in aqueous environments
 Fungi have greater ability than bacteria to
survive hypertonic environments
Radiation
 2 types of radiation:
a. Particulate radiation
 Consists of high-speed subatomic
particles, such as protons, that
have been freed from their atoms
b. Electromagnetic radiation
 Energy without mass traveling in
waves at the speed of light
 The shorter the wavelength, the
more energy it carries
 All types of radiation are described as
either ionizing or nonionizing based on its
effects on the chemicals within cells
Ionizing Radiation
 Wavelengths shorter than 1nm
 When they strike molecules, they have
sufficient energy to eject electrons from
atoms, creating ions
 Ions disrupt hydrogen bonding, oxidize
double covalent bonds, and create highly
reactive hydroxy radicals which in turn
denature other molecules, particularly
DNA, causing fatal mutations and cell
death
 Nonionizing radiations:
a. Electron beams
 Produced by cathode ray
machines
 Highly energetic and very
effective in killing microbes in just
few seconds
 Cannot sterilize thick objects or
objects coated with large amount
of organic matter
b. Gamma rays
 Emitted by some radioactive
elements
 Penetrate much farther than
electron beams but require hours
to kill microbes
 Kills not only microbes but also
larvae and eggs of insects
 Prevents both microbial spoilage
and overripening
c. X-rays
 Travel the fastest through matter
but have less energy than gamma
rays
 Require long time to kill microbes
 Not practical for microbial control
Nonionizing Radiation
 Wavelength greater than 1nm
 Does not have enough energy to force
electrons out of orbit but contains enough
energy to excite electrons and cause them
to make new covalent bonds, which can
affect the 3-D structure of proteins and
nucleic acids
 Ultraviolet (UV) light
 Has sufficient energy to be a practical
antimicrobial agent
 Causes pyrimidine dimers in DNA
 Does not penetrate well
 Suitable for disinfecting air,
transparent fluids, and surface of
objects
Chemical Methods of Microbial Control