Control of Microorqanisms

Methods used to control the growth of microorganisms and their

transmission of infectious disease involve stopping the growth of
the microorganism for a period of time, reducing the number of
microorganisms to a safe level, or destroying the microorganisms.
The degree of effectiveness depends on the number of
microorganisms, the type of microorganism, their physiological
state, such as the stage of growth or formation of endospores, and
the ~nvironment in which they are growing (glassware, instruments,
tissue, food).
Terms Related to Destruction of Microorqanisms
I.
sterilization - the destruction of all microorganisms,
including endospores, on an object or in a material.
2.
disinfection
the destruction of pathogens, but not
endospores, on an object or in a material. The number of pathogens
is reduced or growth is inhibited to a level that does not produce
disease.
3.
antisepsis - chemical disinfection of the skin, mucosal
membranes, or other living tissues.
4. germicide ("cide" = kill) - a chemical agent
kills microorganisms.
Specific germicides include:
a.
b.
c.
d.

that

rapidly

sporicide - kills spores

bactericide - kills bacteria
viricide - kills viruses
funqicide - kills fungi

Terms Related to Suppression of Microorqanisms

i. asepsis ("without infection")
the absence of pathogens from
an object or area.
Aseptic techniques prevent the entry of
pathogens into the body.
There are two types of asepsis:
a.
surqical asepsis - techniques designed to exclude all
microorganisms.
It prevents infectious agents from reaching a
wound. Prevention begins before the patient enters the operating
room and includes removal of hair and cleansing and disinfecting
the skin with an antiseptic that removes skin oils and
microorganisms from the area of the skin to be opened. Substances
include Betadine, Isodine, Ioprep, and Surgidine.

b.
medical asepsis - techniques designed to exclude
microorganisms associated with communicable diseases. It includes
dust control, hand-washing, use of individualized equipment and
instruments, waste disposal, care of instruments, syringes,
needles,~ thermometers, and dressings.
2.
sanitization - the reduction or removal of pathogens on
inanimate objects by chemical or mechanical cleansing.
3.
bacteriostasis -("static" = halt) - bacterial growth and
multiplication are inhibited, but the bacteria are not killed.

Terms for Destruction or Suppression o__f Microorqanisms

i.
antimicrobial
microorganism.

agents that destroy or suppress any

2. antibiotic - a product formed by microorganisms that destroy or

suppress microorganisms.

Physical Aqents
I.

Heat

Heat is the most common, inexpensive, simplest, reliable, and

effective method used to destroy microorganisms. Heat denatures
the proteins and enzymes of the microorganisms. Most pathogens
will be destroyed at temperatures between 50 and 70C for a
duration of i0 minutes, except endospores which may survive 1 to 2
hours at 100C.
The heat used in sterilization is either moist or dry heat.
a. moist heat

(i). boilinq - Bacteria, fungi, and many viruses are

destroyed by boiling at 100C for I0 to 30 minutes. Some viruses
and endospores may require boiling for up to 20 hours.
Sterilization is accomplished by denaturing the proteins and
enzymes of the microorganisms.
(2).
steam under pressure - Water is heated under
pressure which raises the temperature above 100C which denatures
proteins and amino acids.
The most common device used is an
autoclave for sterilizing surgical bandages, instruments, media,
and contaminated material. At a pressure of 15 pounds/square inch
and a temperature of 121C for 15 to 20 minutes it will destroy
microorganisms and endospores.

5O

(3). pasteurization - It is mainly used in the food and

dairy industries and it involves raising the temperature high
enough to destroy pathogens or inhibit their growth without
affecting the quality of the product.
b.

dz-Z heat

Dry heat penetrates substances more slowly than moist heat.

(I). incineration - The burning of disposable substances
in a chamber.
(2).
direct flaminq - Sterilization of inoculating
loops, needles, and rims of test tubes with a bunsen burner. Wire
heated to a red glow is 100% effective.
(3). hot-air oven - Sterilization of glassware, test
tubes, petri dishes, instruments, syringes, and needles requires
higher temperatures for a longer
period of time than other
methods.
Most endospores will be destroyed at 160-165"C for a
period of 2 hours.
2.

cold

The effect of low temperature on microorganisms depends on the

type of microorganism and the intensity of the application.
Temperature in a refrigerator ranges from 0-8"C and has a
bacteriostatic effect which reduces the metabolic rate of most
organisms so that they cannot reproduce or synthesize toxins.
Freezing at -20"C kills most bacteria, but some may survive in
a frozen state.
3.

dryinq or desiccation

To grow and multiple, microorganisms require water.

The
removal of water by evaporation or freeze-drying (solid to gas)
inhibits growth and reproduction of microorganisms by inhibiting
enzymes.
They may be viable for years so when water is made
available they resume growth and reproduction.
ultra-violet radiation and ionizinq radiation (x-rays and
~amma rays)
Both types of radiation damage the DNA of microorganisms and
denatures their proteins.
U-V radiation cannot penetrate
materials such as glass, clothing, dirt, paper, or pus and is used
to kill microorganisms on surfaces. Germicidal lamps in operating
rooms, nurseries, and communicable disease wards reduce the number
of bacteria in the air, but they do not sterilize.

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5. filtration - Filtration is the passage of material or liquids

through a filter containing small pores that retain microorganisms.
Membrane filters of cellulose acetate are the most common, but
gauze, cotton, and paper serve as filters for air. Vaccines that
require the presence of live viruses, such as polio, are passed
through filters which prevent bacteria from going through the
pores. Air filters are used in operating rooms to lower the number
of airborne microorganisms.
Chemical Aqents

Chemical agents are used to control the growth of

microorganisms on inanimate objects and on living tissue. Most
chemical agents reduce the number of microorganisms (disinfect),
but do not achieve sterility. Selection of a disinfectant depends
on the mode of action, concentration necessary, the type of
microorganism, the number of microorganisms present, the type of
material to be disinfected, temperature, and pH.
Disinfectants are classified based on their chemical structure
and activity.
I.

phenols (carbolic acid)

Phenol and derivatives called phenolics disrupt the plasma

membrane, denature proteins, and inactivate enzymes.
Phenol is
rarely used any more because it causes skin irritation and has a
disagreeable odor.
Common phenolics are cresols such as that foLuud in L_~I, and
hexachlorophene which is used as an antiseptic.

2.

alcohols

Ethanol and isopropanol are widely used as skin antiseptics,

but they are not effective against enveloped viruses or bacterial
endospores.
They effectively destroy bacteria and fungi by
disrupting the lipids in the plasma membrane resulting in lysis,
and denatures proteins.
3.

surface-active agents (surfactants)

They include soaps and detergents that lower the surfacetension of liquid molecules and make microorganisms accessible to
other agents. The pH of soaps is usually alkaline which destroys
some bacteria. Detergents are more effective against gram-positive
than gram-negative bacteria and they disrupt the plasma membrane.

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4.

haloqens

The major halogens are iodine and chlorine.

Iodine is an
effective antiseptic for the skin and superficial wounds.
It
destroys many kinds of bacteria, some endospores, fungi, and some
viruses. Iodine inhibits protein function in the microorganism.
Chlorine is used as a disinfectant and it is a strong oxidizing
agent which affects functions of the microorganismss enzymes.
5.

io___ns of heav~ metals

The major metal ions are mercury and silver, and they function
in denaturing proteins and enzymes. Both are used in combination
with other substances as antiseptics.
6.

chlorhexidine

Chlorhexidine is used as an antiseptic on skin and mucosal

membranes and functions in disruption of the plasma membrane of
gram-positive and gram-negative bacteria resulting in lysis.
7.

alkylatinq awents

Alky!ating agents disrupt the structure of proteins and

nucleic acids.
Formaldehyde (.2 - .4%) is used to inactivate
viruses and toxins used in vaccines, it also destroys spores in
bacteria and fungi. Glutaraldehyde is less irritating and more
effective than formaldehyde.
It kills many microorganisms,
viruses, and endospores. Ethylene oxide is a gas used in a closed
chamber to sterilize materials. It kills all bacteria, including
endospores. Betapropiolactone kills spores in concentrations not
much more than required for killing cells. The effect is rapid and
it disappears in a few hours. It is used to sterilize bone,
cartilage, and artery grafts.
Antimicrobial Chemotherapy
Chemotherapy is the use of chemical substances to treat
various aspects of disease.
Chemotherapeutic aqents (drugs) are any chemical substances
used in medical practice to treat disease.
Antimicrobial aqents are a special group of chemotherapeutic
agents used to treat infection caused by microorganisms. Many are
produced by microorganismsand are specifically called antibiotics.
Some antimicrobial agents are synthetic druq~which are produced in
a laboratory.
Others may be semis~nthetic dru~s which are
synthesized by chemically modifying a substance from a
microorganism.

Properties of Antimicrobial Aqents

!. solubility in bod~ fluids - They must dissolve in body fluids
to be transported and reach the area of infection or
microorganisms.
2. selective toxicit~ - They must harm the microorganisms without
causing significant damage to the host.
3. spectrum of activity - The range of different microorganisms on
which an antimicrobial agent acts is its spectrum of activity.
There are two major types of spectrum activity:
a. broad-spectrum activity - Agents are effective against a
great number and types of microorganisms, including gram-positive
and gram-negative bacteria. They are useful when an infection is
caused by an unidentified microorganism and it increases the
chances that the microorganism will be susceptible to the drug.
A disadvantage is that many normal flora of the host are
destroyed. Normal flora compete with and help destroy pathogens
and other microorganisms. Some normal flora may flourish when the
competition is diminished and they become opportunistic pathogens.
b. narrow-spectrum activity - Agents are effective against
only a small number or type of microorganism. They reduce the
chance of normal flora being destroyed and decrease the chance the
microorganisms will develop drug resistance.
4. nonallerqenic - The drug should not cause an allergic reaction,
hypersensitivity, or anaphy!actic shock in the host.
5.
stability - Toxicity should not be altered by food, other
drugs, or conditions of the host and the concentration in the blood
and tissue fluids should be stable over a period of hours (degraded
and excreted slowly).
6. resistance - Few microorganisms should be resistant or have the
ability to develop resistance to the effects of the drug. Two
drugs with different modes of action on the microorganism may be
administered. The drugs do not interfere with each other, have an
additive or synerqistic effect and reduce the likelihood of
resistance.

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Most antimicrobial agents are antibacterial agents that are

bactericidal (killing) or bacteriostatic (growth-inhibiting) while
having minimal effects on host cells. Antimicrobial agents may be
categorized by their mode of action.
There are five modes of action:
I. inhibition of cell wall s~nthesis - Peptidoglycan synthesis in
the cell wall of growing bacteria is affected. The cell wall is
weakened and lysis occurs. Human cells are not affected because
they do not have cell walls or peptidoglycan in their plasma
membrane. Drugs such as vancomTcin and bacitracin interfere with
Penicillin and
the synthesis of peptidoglycan chains.
cephalosporin prevent the linking between the peptidoglycan
chains.
2. disruption of the plasma membrane - The selective permeability
of the plasma membrane is affected and important substances used in
metabolic activities leave the bacterial cell. The drugs used are
selectively toxic. PolymTxin B causes damage by attaching to the
phospholipids of the membrane. It allows dissolved substances to
pass freely into and out of the cell eventually resulting in lysis.
Antifungal drugs, such as nTstatin, amphotericin ~, and miconazole,
combine with sterols in the funga! membrane causing disruption and
leakage of cytoplasmic contents.
3. inhibition of protein synthesis - Antimicrobial drugs affect
the bacterial ribosome which is made up of a large and small
protein subunit.
When ribosomal function is disturbed, the
synthesis of protein may slow down so that normal growth cannot
occur or protein synthesis may stop. Ribosomal function may be
affected in different ways Chloramphenicol and erythromycin react
with the large ribosomal subunit and prevent peptide bonds from
forming between the amino acids.
Streptomycin, neomycin,
kanamycin, and qentamicin bond with the small subunit and cause the
genetic code on mRNA to be improperly read and inhibit initiation
of protein synthesis by preventing the ribosome from moving.
Tetracycline bonds with the small subunit and interferes with the
attachment of the tRNA carrying the amino acid to the ribosome.
4. inhibition of nucleic acid sTnthesis - DNA replication and the
copying of the information from DNA by mRNA is affected
(transcription). Quinolones (ciprofloxacin) are synthetic drugs
that block the enzyme which unwinds the DNA double helix during
preparation for replication. Rifampin is a semisynthetic drug that
inhibits the enzyme RNA polymerase which is necessary for
transcription.

55

5.
inhibition o__f s~nthesis of essential metabolites - The
synthesis of folic acid (B vitamin), which is needed to make the
nitrogenous bases of DNA is inhibited. Sulfonamides (sulfa drugs)
are antimetabolites which function as competitive inhibitors of ~
enzyme in the pathway that forms folic acid. Sulfonamides resemble
the substrate para-aminobenzoic acid (PABA).
The enzyme that
normally converts PABA to fo!ic acid combines with the sulfonamide
instead. The combination prevents fo!ic acid synthesis and stops
the growth of the microorganism.

Antiviral Aqents
Chemical agents that inhibit the growth of viruses are not as
common as antibiotics. A few inhibit a wide variety of viruses,
while most are restricted to a single genus or group of viruses.
There are four modes of action:
i. antiviral proteins sT~thesized bv cells infected with a virus Interferons are small proteins that are produced in most cells when
they are infected by a virus. Once produced, interferon prevents
the subsequent infection of noninfect~d cells.
2. interfere with viral DNA synthesis - Most antiviral drugs are
structural analogues of the normal purines and pyrimidines that
form nucleic acids. Because they are structurally similar to the
normal nucleotides, they prevent viral reproduction by inhibiting
DNA synthesis. Iodoxuridine, Vidarabine, and AcTclovir are used to
treat herpesvirus eye infections, herpesvirus encephalitis,
cytomegalovirus infections in newborns, and herpes zoster.
3. acted on b~ viral enzTmes or that block the activit7 of viral
enzTmes - Retrovirus replication requires that the viral RNA first
be transcribed into DNA. The viral enzyme that catalyzes this step
is reverse transcriptase. Retroviruses cause leukemias and AIDS.
AzidothTmidine (AZT) inhibits the action of reverse transcriptase.
4. prevent the release o__f the viral nucleic acid into the
cytoplasm of the infected cell - Amantadine prevents the release of
viral RNA into the cytoplasm and is effective against influenza
virus type A.
Ribavirin inhibits nucleic acid synthesis by
interfering with the formation of guanine nucleotides.