CONTROLLING MICROBIAL GROWTH IN VITRO
, medium) that are used in
Availability of Nutrients
- All living organisms require nutrient the
various chemical compounds that
organisms use to sustain life. Therefore, to
survive in a particular environment,
appropriate nutrients must be available.
Moisture
- Water is essential for life, as we know it.
Cells are composed of between 70% and
95% water.
Temperature
- Every microorganism has an optimal, a
minimum, and a maximum growth
temperature.
- Thermophiles
- Mesophiles.
- Psychrophiles
- Psychroduric organisms
pH ( potential of hydrogen)
- The term “pH” refers to the hydrogen ion
concentration of a solution and, thus, the
acidity or alkalinity of the solution.
- Acidophiles
- Alkaliphiles
Osmotic Pressure and Salinity
Osmotic pressure is the pressure that is exerted on a
cell membrane by solutions both inside and outside
the cell.
Hypertonic, hypotonic and isotonic..
Osmosis is defined as the movement of a solvent
(e.g., water), through a permeable membrane, from
a solution having a lower concentration of solute to
a solution having a higher concentration of solute.
- Crenation
- Plasmolysis and plasmoptysis.
- Halophilic and haloduric organisms.
Barometric Pressure
- Most bacteria are not affected by minor
changes in barometric pressure.
- Piezophiles,
Gaseous Atmosphere -microorganisms vary with
respect to the type of gaseous atmosphere that they
require.
The media (sing.
microbiology laboratories to culture
bacteria are referred to as artificial media or
synthetic media, because they do
not occur naturally; rather, they are
prepared in the laboratory.
Thioglycollate broth (THIO) is a very
popular liquid medium for use in the
bacteriology laboratory.
IMPORTANCE OF USING “ASEPTIC TECHNIQUE”
- to maintain the sterility of the media before
inoculation and to avoid touching the agar surface
with fingertips or other nonsterile objects.
INCUBATION
After media are inoculated, they must be incubated
(i.e., they must be placed into a chamber [called an
incubator] that contains the appropriate
atmosphere and moisture level and is set to
maintain the appropriate temperature).
Once a particular species of bacteria has been
isolated from a clinical specimen, it can be separated
from any other organisms that were present in the
specimen and can be grown as a pure culture.
Three types of incubators
A CO2 (carbon dioxide) incubator is an incubator
to which a cylinder of CO2 is attached. CO2 is
periodically introduced into the incubator to
maintain a CO2 concentration of about 5% to 10%.
A non-CO2 incubator is an incubator containing
room air; thus, it contains about 20% to 21% oxygen.
An anaerobic incubator is an incubator
containing an atmosphere devoid of
oxygen.
viable plate count is used to determine the number
of viable bacteria in a liquid sample, such as milk,
water, ground food diluted in water, or a broth
culture. In this procedure, serial dilutions of the
sample are prepared, and then 0.1-mL or 1-mL
aliquots (portions) are inoculated onto plates of
nutrient agar.
Sterilization- involves the destruction or elimination
of all microbes, including cells, spores, and viruses.
Disinfection - involves the elimination of most or all
pathogens (except bacterial spores) from nonliving
objects.
Pasteurization - is a method of disinfecting liquids.
Disinfectants - used to disinfect inanimate objects,
such as bedside equipment and operating rooms
Sanitization - the reduction of microbial populations
to levels considered safe by public health standards
Microbicidal Agents - are disinfectants or antiseptics
that kill microbes.
Microbistatic Agents- a drug or chemical that
inhibits reproduction of microorganisms, but
does not necessarily kill them
Lyophilization - a good method of preserving
microorganisms for future use.
Sepsis - refers to the presence of pathogens in
blood or tissues, whereas asepsis means the
absence of pathogens.
Aseptic techniques - are used to eliminate and
exclude pathogens.
Antisepsis - is the prevention of infection.
Antiseptic technique - developed by Joseph
Lister in 1867, refers to the use of antiseptics.
Sterile technique - is practiced when it is necessary
to exclude all microorganisms from a particular area,
so that the area will be sterile.
Using Physical Methods to Inhibit Microbial Growth
- The methods used to destroy or inhibit microbial
life are either physical or chemical, and sometimes
both types are used.
Heat is the most common type of sterilization for
inanimate objects able to withstand high
temperatures.
Two factors—temperature and time determine the
effectiveness of heat for sterilization.
The thermal death point (TDP) of any particular
species of microorganism is the lowest temperature
that will kill all the organisms in a standardized pure
culture within a specified period.
The thermal death time (TDT) is the length of time
necessary to sterilize a pure culture at a specified
temperature.
Dry-heat baking in a thermostatically controlled
oven provides effective sterilization of metals,
glassware, some powders, oils, and waxes.
Incineration (burning) is an effective means of
destroying contaminated disposable materials.
Moist Heat
- Heat applied in the presence of moisture, as
in boiling or steaming, is faster and more
effective than dry heat, and can be
accomplished at a lower temperature.
Autoclave is like a large metal pressure cooker that
uses steam under pressure to completely destroy all
microbial life.
Cold-Most microorganisms are not killed by cold
temperatures and freezing, but their metabolic
activities are slowed, greatly inhibiting their growth.
Refrigeration merely slows the growth of most
microorganisms; it does not completely inhibit
growth.
Desiccation -For many centuries, foods have been
preserved by drying. However, even when moisture
and nutrients are lacking, many dried
microorganisms remain viable, although they cannot
reproduce.
Radiation - Studies performed in radiation research
laboratories have demonstrated that these
radiations (X-rays, gamma and beta rays) can be
used for the prevention of food spoilage, sterilization
of heat-sensitive surgical equipment, preparation of
vaccines, and treatment of some chronic diseases
such as cancer, all of which are very practical
applications for laboratory research.
Ultrasonic Waves - In hospitals, medical clinics, and
dental clinics, ultrasonic waves are a frequently used
means of cleaning delicate equipment.
Filtration-Filters of various pore sizes are used to
filter or separate cells, larger viruses, bacteria, and
certain other microorganisms from the liquids or
gases in which they are suspended.
Gaseous Atmosphere -In limited situations, it is
possible to inhibit growth of microorganisms by
altering the atmosphere in which they are located.
Because aerobes and microaerophiles require
oxygen, they can be killed by placing them into
an atmosphere devoid of oxygen or by removing
oxygen from the environment in which they are
living.

USING CHEMICAL AGENTS TO INHIBIT MICROBIAL

GROWTH

Disinfectants-Chemical disinfection refers to the use

of chemical agents to inhibit the growth of
pathogens, either temporarily or permanently.
Various factors affect the efficiency or effectiveness
of a disinfectant (Fig. 8-15), and these factors must
be taken into consideration whenever a disinfectant
is used.

Antiseptics -Antimicrobial chemical agents that can

safely be applied to skin are called antiseptics. An
antiseptic merely reduces the number of organisms
on a surface; it does not penetrate pores and hair
follicles to destroy microorganisms residing there.