Microbial control

Dr. Bhavesh Patel ([Link]., Ph.D.)

Principal
V.P. and R.P.T.P. Science College
Vallabh Vidyanagar, Gujarat
(bhavesh1968@[Link])
Microbial control
What is control ?
 Restricting the growth and normal activities of
microorganisms.
Microbial control
Why control ?
 To prevent transmission of diseases.
 To prevent contamination at various levels.

To prevent spoilage of materials.
Microbial control
How control ?
 By killing the microorganisms (cidal)
 By inhibiting the microorganisms (static)
 By removing the microorganisms
Microbial control
Mechanism of antimicrobial action
 Affecting the cell wall
 Affecting the cell membrane
 Interfering with protein structure and function
 Affecting nucleic acids
Microbial control
Factors affecting antimicrobial
activities
 Type and state of microorganisms
 Initial microbial load
 Concentration or dose of agent
 Time of exposure
 Environmental conditions
 Microbial control - Practical Concern

 Does the application require sterilization ?

 Is the item to be reused ?
 Can the item withstand heat, pressure, radiation
or chemical ?
 Will the agent penetrate to the necessary
extent ?
 Is the method cost and labor efficient and is it
safe ?
 Microbial control - Some useful
terms

 Sterilization :killing or removal of all life form

i.e. free from life.
 Disinfection :surface removal of organisms.
 Sanitation :reduction in number of organism.
 Antiseptic :agent that prevent sepsis.
 Pasteurization :killing of organism without
affecting quality
of material.
 Kinetics of heat sterilization
 Death from heating is an exponential (first order)
function and occurs more rapidly as the temp
raised.
 Thus if we practically wants to sterilize a microbial
population , it will take longer at low temp than at
higher temp.
 It is thus required to adjust time and temp to
achieve sterilization for each specific set of
conditions.
 The nature of heat is also important – moist heat is
more penetrating than dry heat.
Kinetics of heat sterilization
 Kinetics of heat sterilization
Death rate-Decimal reduction time (D-value)
 It is the time in minute to kill 90% of the cells in a
population. OR
 It is the time to reduce the population by one decimal
Death rate- Thermal Death Point (TDP)
 Lowest temperature required to kill all the
microorganisms in the liquid suspension in 10 min.
Death rate- Thermal Death Time (TDT)
 Minimum time required to kill all microorganisms in
liquid suspension at given temperature
 Physical control

 Heat (low temp, high temp)

 Desiccation
 Osmotic pressure
 Radiation(ionizing & non ionizing)
 Filtration(porcelain, barkfeld, sintered
glass)
 High temperature

Moist heat Dry heat

o
 o
Boiling (100 C)

Incineration(>500 C)
 Tyndalization
 Hot air oven
o
o
(100 C,3days) (160 C,2hrs. or
o
 Autoclave 170 C,1 h.)
o  o
(15 lbs/[Link].,121 C- Flaming (>500 C)
20min.)
 Pasteurization(LTHT-
o
63 C,30min.,HTST-
o
72 C,15sec.)
 Moist vs Dry heat

o
Temp( C) Time(min)
Moist heat 121 15
125 10
134 3
Dry heat 121 600
140 180
160 120
170 60
Autoclave
 Heat
Low temp.(Static) High temp.
 o
Freezing (below 4 C) (Cidal)
o o

Chilling (4-6 C) 
Moist heat (>100 C)
o

Dry heat (>100 C)
 Desiccation

Control by drying - vacuum, heat etc.

Susceptibility to dessication varies widely:
 Neisseria gonnorrhea: Only survives about one hour.
 Mycobacterium tuberculosis: May survive several
months.
 Viruses are fairly resistant to dessication.
 Clostridium spp. and Bacillus spp.: May survive
decades.
 Filtration
 Depth filters-particulate material (porcelain,
glass, earth, asbestos)
 Membrane filters (cellulose acetate,
polycarbonate, etc.) different pore sizes; 0.2 or
0.4 micrometer remove bacteria
 HEPA (high-efficiency particulate air) filters
used for hospitals, laminar flow hoods in
laboratories, being marketed or home use to
reduce allergens
 Filtration
Laminar air flow Membrane filtration
 Osmotic pressure

 High sugar and salt conc. Can be used to

restrict the growth (Bacteriostatic).
 Many microbes can tolerate sugar conc. Up to
10% (saccharophilic or saccharolytic).
 Some can tolerate salt conc. Up to 30%
(halophiles).
Radiation
 Radiation
Radiation: Three types of radiation kill microbes:
1. Ionizing Radiation: Gamma rays, X rays,
electron beams, or higher energy rays. Have short
wavelengths (less than 1 nanometer).
Dislodge electrons from atoms and form ions.
Cause mutations in DNA and produce peroxides.
Used to sterilize pharmaceuticals and disposable
medical supplies. Food industry is interested in using
ionizing radiation.
Disadvantages: Penetrates human tissues. May
cause genetic mutations in humans.
 Radiation
2. Ultraviolet light (Non ionizing Radiation): Wavelength is
longer than 1 nanometer. Damages DNA by producing
thymine dimers, which cause mutations.
Used to disinfect operating rooms, nurseries, cafeterias.
Disadvantages: Damages skin, eyes. Doesn’t penetrate
paper, glass, and cloth.

3. Microwave Radiation: Wavelength ranges from 1 mm to 1m

Heat is absorbed by water molecules.
May kill vegetative cells in moist foods.
Bacterial endospores, which do not contain water, are not
damaged by microwave radiation.
Radiation
Chemical Agents
 Phenolics
 Alcohols
 Halogens
 Heavy metals
 Quaternary Ammonium Compounds
 Aldehydes
 Sterilizing Gases
Chemical Agents: Phenolics
 Aromatic organic compounds with attached -OH
 Denature protein & disrupt membranes
 Phenol, orthocresol, orthophenylphenol,
hexachlorophene
 Commonly used as disinfectants (e.g. “Lysol”);
are tuberculocidal, effective in presence of
organic matter, remain on surfaces long after
application
 Disagreeable odor & skin irritation;
hexachlorophene once used as an antiseptic but
its use is limited as it causes brain damage
Chemical Agents: Alcohols
 Ethanol; isopropanol; used at concentrations
between 70 – 95%
 Denature proteins; disrupt membranes
 Kills vegetative cells of bacteria & fungi but
not spores
 Used in disinfecting surfaces; thermometers;
“ethanol-flaming” technique used to sterilize
glass plate spreaders or dissecting instruments
at the lab bench
Chemical Agents: Halogens
 Act as oxidizing agents; oxidize proteins
& other cellular components
 Chlorine compounds
 Used in disinfecting municipal water supplies
(as sodium hypochlorite, calcium
hypochlorite, or chlorine gas)
 Sodium Hypochlorite (Chlorine Bleach) used
at 10 - 20% dilution as bench top disinfectant
 Halazone tablets (parasulfone dichloro amido
benzoic acid) used by campers to disinfect
water for drinking
Chemical Agents: Halogens
 Iodine Compounds
 Tincture of iodine (iodine solution in alcohol)
 Potassium iodide in aqueous solution
 Iodophors: Iodine complexed to an organic
carrier; e.g. Wescodyne, Betadyne
 Used as antiseptics for cleansing skin
surfaces and wounds
Chemical Agents: Heavy
Metals
 Mercury, silver, zinc, arsenic, copper ions
 Form precipitates with cell proteins
 At one time were frequently used medically as
antiseptics but much of their use has been
replaced by less toxic alternatives
 Examples: 1% silver nitrate was used as
opthalmic drops in newborn infants to prevent
gonorrhea; has been replaced by erythromycin
or other antibiotics; copper sulfate used as
algicide in swimming pools
Chemical Agents: Quaternary
Ammonium Compounds
 Quaternary ammonium compounds are
cationic detergents
 Amphipathic molecules that act as
emulsifying agents
 Denature proteins and disrupt membranes
 Used as disinfectants and skin antiseptics
 Examples: cetylpyridinium chloride,
benzalkonium chloride
Chemical Agents: Aldehydes
 Formaldehyde and gluteraldehyde
 React chemically with nucleic acid and protein,
inactivating them
 Aqueous solutions can be used as disinfectants
Chemical Agents:Sterilizing
Gases
 Ethylene oxide (EtO)
 Used to sterilize heat-sensitive equipment and
plastic ware
 Explosive; supplied as a 10 – 20% mixture
with either CO2 or dichlorofluoromethane
 Its use requires a special EtO sterilizer to
carefully control sterilization conditions as well
as extensive ventilation after sterilation
because of toxicity of EtO
 Much of the commercial use of EtO (for
example, plastic petri dishes) has in recent
years been replaced by gamma irradiation
Chemical Agents:Sterilizing
Gases
 Betapropiolactone (BPL)
 In its liquid form has been used to sterilize
vaccines and sera
 Decomposes after several hours and is not
as difficult to eliminate as EtO, but it doesn’t
penetrate as well as EtO and may also be
carcinogenic
 Has not been used as extensively as EtO
 Vapor-phase hydrogen peroxide
 Has been used recently to decontaminate
biological safety cabinets