CONTROL OF MICROBIAL GROWTH WITH

DISINFECTANTS AND ANTSEPTICS

© Health Protection Agency / Science Photo Library

Microbiology: A Clinical Approach © Garland Science

WHY IS THIS
IMPORTANT?

•Infection control
using disinfectants
and antiseptics is
essential to keep
infections from
spreading,
particularly in
hospitals.
OVERVIEW
IMPORTANT
TERMINOLOGY

• Some treatments are used

for both disinfection and
antisepsis.
• Disinfection is
associated with
inanimate objects.
• Antisepsis is associated
with human tissue and
skin.
..IMPORTANT
TERMINOLOGY

• Sterilization – the removal

of all microbes
• Sterilization does not
destroy prions.
• Aseptic – an environment or
procedure free from
contamination
..IMPORTANT
TERMINOLOGY

• Disinfection – the use

of chemical or physical
agents to kill or inhibit
the growth of
microorganisms
• Heat
• Alcohol
• Ultraviolet radiation
• Some disinfecting
agents do not affect
spores.
…IMPORTANT
TERMINOLOGY

•A chemical used on
inanimate object is
called a
disinfectant.
•The same chemical
used on skin and
tissue is called an
antiseptic.
…IMPORTANT
TERMINOLOGY

• De-germing – the
removal of microbes
from a surface by
mechanical means
• Sanitization –
disinfection of places
or things used by the
public
• Used to reduce the
number of
pathogens to meet
accepted public
health standards
…IMPORTANT
TERMINOLOGY

• Sanitization is not
sterilization but the same
techniques can be used.
• Steaming
• High-pressure
• High-temperature
• Washing and scrubbing
…IMPORTANT
TERMINOLOGY

• Pasteurization – using
heat to kill pathogens
• Does not sterilize but
is used to reduce
number of
pathogens
• Also reduces the
number of organisms
that can cause
spoilage
..IMPORTANT
TERMINOLOGY

-static – an agent that

inhibits growth
• In a bacteriostatic
environment, numbers do
not multiply but
organisms are not dead.

-cidal – an agent that

kills
TARGETS FOR
DISINFECTANTS
AND ANTISEPTICS

Bacteria are single-celled

organisms with a simple anatomy.

They have several targets:

• Cell wall
• Plasma membrane
• Responsible for enclosing the
cytoplasm
• Involved with DNA replication
• Involved in ATP Production
….TARGETS

When cell wall is damaged, it the makes

a cell susceptible to lysis.

When plasma membrane is disrupted,

the cell loses its selective permeability
& leads to the death of the cell.

When a protein’s shape is changed it is

called denaturing & loses its function;
inhibition or elimination of function can
result in cell death.
…TARGETS
Plasma Membrane
 Nucleic acids are required
for cell survival.

…TARGETS Some agents can disturb this

Nucleic Acid synthesis by binding
Synthesis irreversibly to DNA.
• This prevents gene expression.

Others are mutagenic and

cause lethal mutations.
…TARGETS
Nucleic Acid Synthesis

• Radiation can interfere with

DNA and RNA function.
• Irradiation with gamma
rays, ultraviolet radiation
and X-rays causes
mutations.
• These can result in
permanent inactivation of
nucleic acids.
 There are special requirements to
define microbial death.

The most efficient is to determine

whether an organism can
reproduce.
MICROBIAL
• Move it from the antimicrobial environment
DEATH to fresh media.
• If there is no growth, the organism is dead.

Permanent loss of reproductive

capability is the accepted definition
of microbial death.
• The death rate is logarithmic.
 Factors Affecting Rate

…MICROBIAL
DEATH

Several factors affect the rate of

microbial death in a clinical setting:

Duration of Endospore
Numbers Temperature Environment
exposure formation
…MICROBIAL
DEATH
Factors Affecting Rate

¡ Numbers – the greater the

number of organisms, the
longer it will take to kill.
§ If large numbers of
organisms are present, it
takes time for the agent to
reach them all.
¡ Duration of exposure can vary
depending on: Type of
microbe , Accessibility &
types and nature of radiation.
¡ Temperature – the lower the
temperature, the longer it will
take to kill.
 ¡ The environment – particularly
important in health care
…MICROBIAL environments

DEATH § Many pathogens are

associated with organic
Factors materials.
Blood
Affecting Rate Saliva
Bodily fluids
Fecal material
§ These inhibit accessibility of
the antimicrobial agent to the
organism.
 • Endospore formation –
may be the most
…MICROBIAL important factor
DEATH • Spore-forming
organisms are not
Factors susceptible to most
Affecting Rate chemical agents.
• This is especially
important for
nosocomial
infections.
METHODS FOR
CONTROLLING
MICROBIAL GROWTH
• There are three major
methods for controlling
microbial growth:
• Chemical
• Physical
• Mechanical removal
 Many chemicals can kill
microbes.
CHEMICAL
METHODS FOR
CONTROLLING Chemicals can also be
MICROBIAL harmful to humans.
GROWTH

If they are to be useful as

disinfectants, they must
also be safe to use.
POTENCY OF
DISINFECTANTS AND
ANTISEPTICS
• Factors to consider when
evaluating effectiveness
are:
• Time
• Temperature
• Concentration
 Death rate using a
chemical agent is
accelerated by increasing
the temperature.
..POTENCY OF
DISINFECTANTS • An increase of 10˚C doubles
AND the rate of chemical reaction.
ANTISEPTICS

Changes in pH can
increase or decrease an
agent’s potency.
 For most chemical agents,
increase in concentration
= increase in its potency.
…POTENCY OF
DISINFECTANTS
AND
ANTISEPTICS

This is not true for alcohol.

Alcohol must have some
Increase in alcohol water associated with it.
concentration actually •This makes for better penetration
hinders killing. and denaturation of proteins.
 There is no completely
satisfactory method for
evaluating antimicrobial
chemical agents.
EVALUATION
OF
DISINFECTANTS
AND
ANTISEPTICS
There are several tests
including:
Comparison of the
agent with phenol Use dilution
– the phenol Disk method
method
coefficient
 Phenol was first used as a
disinfectant by Joseph Lister in 1867.

It is still considered the benchmark

disinfectant that others are
…EVALUATION compared with.
METHODS:
The Phenol
Coefficient Comparison is reported as the
phenol coefficient.

• Phenol coefficient of 1.0 = same

effectiveness as phenol
• Greater than 1.0 = efficiency greater than
phenol
• Less than 1.0 = efficiency less than phenol
…EVALUATION
METHODS:
The Disk Method

¡ The disk method uses tiny disks

of filter paper soaked in the
agent.
¡ An agar plate is inoculated and
the disks are placed at various
positions.
¡ Inhibition of growth around the
disk is called the zone of
inhibition.
§ Sizes of the zones are not
comparable.
§ Sizes may reflect differences
in concentration and
diffusion rates.
¡ This method cannot distinguish
between microbicidal and
microbistatic.
…EVALUATION
METHODS:
The Use Dilution
Method
• The use dilution method is
time-consuming.
• It can tell whether the
agent is microbistatic or
microbicidal.
• A series of solutions of
different concentrations of
the disinfectant are
prepared.
SELECTING AN ANTIMICROBIAL
AGENT

Some chemical
agents are better for Qualities to be
certain uses than considered include:
others.

Is it reactive against
Is it effective in the Is it stable and, if
all types of
presence of organic possible,
organisms without
material. inexpensive.
destroying tissue.
TYPES OF CHEMICAL AGENTS

Chemical agents are used more than physical means for

disinfection, antisepsis, and preservation.

Chemical agents affect cell walls, plasma membranes, proteins,

or nucleic acids.

Chemical agents destroy or inhibit They are ineffective against

growth of enveloped viruses, protozoan cysts and bacterial
bacteria, fungi, and protozoans. endospores.
…TYPES OF
CHEMICAL
AGENTS

• The effect of chemical

agents varies with:
• Temperature
• Length of exposure
• Amount of
contaminating
organic material
• pH
• Concentration
• Stability
…TYPES OF
CHEMICAL
AGENTS
¡ There are eight major
categories of chemical
agents:
§ Phenol and phenolic
compounds
§ Alcohols
§ Halogens
§ Oxidizing agents
§ Surfactants
§ Heavy metals
§ Aldehydes
§ Gaseous agents
PHENOL AND PHENOLIC COMPOUNDS

Many have greater efficacy

Phenolic compounds are
and fewer side effects than
derived from phenol. phenol.

Phenols and phenolic compounds are low-level to

intermediate-level disinfectants and antiseptics.

They are commonly used as disinfectants in health

care settings and laboratories.
 ALCOHOLS

Alcohols are
bacteriocidal, They have no effect on
They are intermediate-
fungicidal, and fungal spores and
level disinfectants.
virucidal for enveloped bacterial endospores.
viruses.

It is routinely used as a
Alcohol denatures
de-germing agent to
proteins and disrupts
prepare sites for
the plasma membrane.
injection.
 Four members of this chemical
family have antimicrobial activity:

Iodine Chlorine Bromine Fluorine

HALOGENS

They are all intermediate-level

antimicrobial chemical agents.
..HALOGENS

Halogens are effective against: Bacterial and fungal

cells, Fungal spores, Some bacterial endospores,
Protozoan cysts & many viruses

Iodine is a well-known antiseptic.

…HALOGENS
Chlorine

Chlorine is found in drinking water, swimming

pools, and waste-water.

It is major ingredient in disinfectants such as

chlorine bleach.

It is used to disinfect kidney-dialysis

equipment.
..HALOGENS
Chlorine

• Chloramines are
combinations of chlorine
and ammonia.
• Used in wound
dressings, skin
antiseptics, water
supplies.
• Less effective than
chlorine as
disinfectants/antiseptic
s.
 Oxidizing agents are high-level
disinfectants and antiseptics that
prohibit bacterial metabolism.

They release hydroxyl radicals

OXIDIZING which kill anaerobic organisms.
AGENTS
They are very effective against
infections of deep tissues.

• They are routinely used in deep puncture

wounds.
 The three most
commonly used Hydrogen
are: peroxide is a
• Hydrogen peroxide common
• Ozone household
• Peracetic acid antiseptic.
..OXIDIZING
AGENTS
Ozone is a very Peracetic acid is
reactive form of the peroxide form
oxygen & some of acetic acid and
cities use ozone an extremely
for water effective
treatment sporicide.
 There are two common
surfactants:

• Soap
• Detergents
SURFACTANTS Soaps are good de-germing
agents but poor antimicrobial
agents.

Detergents are positively charged

organic surfactants that are more
soluble in water than soap.
 QUATS contain ammonium cations.

They are low-level

…SURFACTANTS disinfectants/antiseptics.
Detergents –
QUATS
Their advantage is being odorless,
tasteless, and harmless to humans.

They are used in many industrial and

medicinal applications including
mouthwash.
 QUATS disrupt the plasma membrane
and are:

• Bactericidal (especially for Gram-positive

bacteria)
• Fungicidal
..SURFACTANTS
• Virucidal against enveloped viruses
Detergents –
QUATS They are not useful for non-enveloped
viruses, mycobacteria, or bacterial
endospores.

They are inhibited by the presence of

organic contaminants.
 The ions of heavy metals are
inherently antimicrobial- The
mechanism of action is
through protein denaturation.
HEAVY
METALS

Heavy metals include:

Arsenic Zinc Mercury Silver Copper

..HEAVY
METALS

• Mercury and silver were formerly used in

clinical situations.
• Mercury has been abandoned.
• Silver is still occasionally used in:
• Surgical dressings
• Burn creams
• Catheters
 Aldehydes are compounds
containing a terminal –CHO
group- denature proteins and
inactivate nucleic acids.

ALDEHYDES
Two highly reactive aldehydes
are used as antimicrobials.
Formaldehyde – used
Glutaraldehyde –
in both liquid form
used in liquid form and gaseous form.
 Many items cannot be sterilized with heat or
chemicals.

GASEOUS
AGENTS

They can be sterilized using highly reactive

antimicrobial and sporicidal gases.

Propylene β-
Ethylene oxide
oxide propiolactone
 Gases rapidly penetrate and diffuse
into any space.

Over time, they can denature proteins

and DNA.

…GASEOUS
AGENTS They kill everything they come in
contact with and cause no damage to
inanimate objects.

E.g Ethylene oxide is used in hospitals

and dentists’ offices for sterilizing
instruments and equipment
..GASEOUS
AGENTS

• Gaseous agents have

several disadvantages:
• They are explosive,
poisonous, and
potentially carcinogenic.
• Disinfection with
gaseous agents takes
considerable time.
• There is a need for
continuous cleanup.
PHYSICAL METHODS
FOR CONTROLLING
MICROBIAL
GROWTH
• Physical methods for
controlling microbial growth
include:
• Drying
• Heating
• Cold
• Filtration
• Osmotic pressure
• Radiation
HEAT

• Heat is usually lethal to most

pathogenic microbes.
• Two types of heat can be used:
• Moist heat – from hot water,
boiling water, or steam- it
denatures proteins quicker
than dry heat & halts microbe
metabolism and causes
death.

• Dry heat – from hot air with

low moisture e.g. from ovens-
Dry heat dehydrates microbial
cells & protein denaturation
…HEAT
• Hot air ovens are used for
glassware, metallic
instruments, powders, and
oils.
• Temperatures between 150-
180˚C for two to four hours
ensure the destruction of
spores as well as vegetative
cells.
• Exposure to very high
temperature dry heat reduces
microbes to ash and gases.
 Adequate sterilization with heat depends on:

Temperature Length of time

Higher temperatures require shorter treatment

times.

..HEAT Thermal death time (TDT) is the shortest length of

time needed to kill all organisms at a specific
temperature.

Thermal death point (TDP) is the lowest

temperature needed to kill all organisms in 10
minutes.
 ..HEAT
• There are three
ways of using
moist heat:
• Pressurized
steam
• Boiling
• Pasteurization
…HEAT

1. Pressurized steam gives

the highest temperature
which results from
increased pressure,
example an autoclave.
• Sterilization occurs when
steam condenses to
liquid water on the object.
• Autoclaves are superior
for sterilizing heat-
resistant materials.
..HEAT

2. Boiling is easy but

does not kill heat-
resistant cells.
• It is effective for
disinfection but
not sterilization.
• Boiling for 30
minutes kills most
non spore-forming
pathogens.
..HEAT

3. Pasteurization is
used to reduce
microbial load.
•It destroys
pathogens.
•It preserves flavor
and nutritive value
in foods.
•It does not sterilize.
… HEAT

• Pasteurization is
accomplished in two ways:
• Flash method –
temperature of 71.6˚C for
15 seconds
• Batch method –
temperature of 63-66˚C
for 30 minutes
• It does not affect
endospores, non-
pathogenic lactobacilli,
micrococci, or yeasts.
 Cold temperatures retard the
growth of microorganisms.
• They slow the rate of enzymatic
reactions.
REFRIGERATION,
FREEZING, AND • They do not kill.
FREEZE-DRYING
Refrigeration is used to delay
the spoilage of food.
• Bacteria and molds will continue to
grow.
• It is useful only for a limited period.
…REFRIGERATION,
FREEZING, AND
FREEZE-DRYING

• Freezing can
preserve food.
• It does not
sterilize.
• It slows
metabolic
rate.
• There is no
growth or
spoilage.
…REFRIGERATION,
FREEZING, AND
FREEZE-DRYING

• Freezing can also be

used to preserve
microorganisms.
• Organisms to be
preserved are frozen
in glycerol.
• This prevents the
formation of ice
crystals.
..REFRIGERATION,
FREEZING, AND
FREEZE-DRYING

• Freeze-drying (lyophilization)
preserves cells by removal of
water.
• Organisms are frozen in
liquid nitrogen and subjected
to high vacuum.
• Containers are then sealed
under vacuum.
• Organisms are viable in this
state for years.
• It is used for long-term
storage.
• Addition of water restarts the
growth process.
FILTRATION
• Filtration is useful for sterilizing liquids.
• It involves passing the liquid through
membrane filters.
• Pores in the filter are too small to
allow for the passage of
microorganisms.
• Filters are made with specific pore
sizes.
• Filtration is used to sample and test
water samples for fecal coliform
contamination.
…FILTRATION

• Filters can also purify air.

• High-Efficiency Particulate
Air filters are called HEPA
filters.
• These are seen in operating
rooms, burn units, clean
rooms of laboratories.
• They are used in laboratory
facilities such as the Centers
for Disease Control (CDC) to
keep organisms from
escaping.
• Filters are soaked in
formalin before disposal.
OSMOTIC
PRESSURE

• Osmotic pressure has been used

in food preservation for many
decades.
• High concentrations of salt or
sugar or other substances are
used in food preservation
because:
• It creates a hypertonic
medium.
• It draws water from the
organisms.
• It leads to plasmolysis and
death.
RADIATION
• Radiation is energy emitted from atomic
activities & it is dispersed at high velocity.
• The cell’s molecules absorb some of the
energy which changes DNA structure.
• Two types of radiation:
• Ionization:
• Includes gamma rays, X-rays, and
high-speed electron beams
• DNA is very sensitive to this type of
radiation.
• When DNA is exposed to this type this
causes the breakdown of
chromosomes.
 Non-ionizing radiation – excites atoms but does not ionize them

It leads to abnormal bonds within

Best seen with ultraviolet radiation molecules such as the formation of
thymine dimmers.

Non-ionizing radiation can penetrate liquids and most solid

materials.

…RADIATION
Gamma rays are the most penetrating.

Flour, meat, fruits, and vegetables are routinely irradiated to kill

microorganisms, parasites, and insects.
 Sterilization of medical products
by ionizing radiation is rapidly
expanding and now includes:
• Drugs
• Vaccines
..RADIATION • Plastics
..Ionizing • Syringes
Radiation • Gloves
• Tissue used in grafting
• Heart valves

The main drawback is the

potential radiation poisoning of
the operators.
 Ultraviolet radiation disrupts cells by generating
free radicals and is a powerful killer of:

Fungal Bacterial
Spores Protozoans Viruses
cells cells

..RADIATION
Ultraviolet It is used for disinfection but not sterilization.
Radiation

It is used in germicidal lamps in hospital rooms,

operating rooms, food preparation areas, and
dental offices.
 • The major disadvantages
are:
• Poor penetration
• Damaging effects seen
over long exposure to
..RADIATION human tissues
• Retinal damage
..Ultraviolet Radiation • Cancer
• Skin wrinkling
HAND WASHING
¡ Hand washing is one of the
most important historical
discoveries in medicine.
¡ The simple act of washing
your hands can inhibit the
spread of pathogens.
¡ The effectiveness of hand
washing is related to:
§ The type of soap used
§ The time taken
¡ Hospitals use bacteriocidal
soaps because they are very
effective at preventing
pathogen transmission.
¡ Household soap can be
effective if enough time is
taken to do the job
thoroughly.
References

• Microbiology, A clinical Approach -Danielle Moszyk-

Strelkauskas-Garland Science 2010
• http://en.wikipedia.org/wiki/Scientific_method