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Culturing Microbes
Learning intention
• To describe the main factors that are
required to culture microbes
Factors affecting microbial
growth
Factors affecting microbial
growth
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Contamination
Growth media
Availability of oxygen
Temperature
pH
Preventing contamination
• Agents that destroy or kill bacteria are
called bactericidal agents.
• However, killing bacteria is not always
possible so agents that inhibit bacterial
growth are often used. These are called
bacteriostatic agents.
Why control microbial growth?
• This is done in the food industry to
preserve food and prevent spoilage.
• It can prevent the spread of infectious
disease, given the rapid growth rate of
microbes.
How to prevent contamination
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Heat sterilisation
Pasteurisation
Radiation sterilisation
Filter sterilisation
Using chemical agents
Using disinfectants and antiseptics
Heat sterilisation
• This involves using an autoclave.
• This sealed device allows the entrance of
steam under pressure.
• The temperature used is 121ºC for 10–15
minutes.
• It is not the pressure in the autoclave that
causes the death of the microorganisms but
the high temperature achieved when the
steam is placed under pressure.
• This procedure also ensures that bacterial
spores are eliminated.
Pasteurisation
• The process used to reduce the microbial
population in milk and other heat-sensitive
products.
• Named after Louis Pasteur, who first used
heat for controlling the spoilage of wine.
• Pasteurisation involves passing milk through
a heat exchanger. The temperature of the
milk is raised to 71°C for 15 seconds. The
milk is then rapidly cooled.
Radiation sterilisation
• The number of microbes can be
significantly reduced by using
electromagnetic radiation.
– UV waves, for example, break down the
DNA molecules in the microbe, thus
causing death.
– Microwaves use thermal effects to kill
bacteria.
Filter sterilisation
• Membrane and depth filters can be
used. This method of sterilisation is
used for media that are sensitive to
heat.
Chemical agents
• Chlorine is used in air-conditioning
systems to prevent the growth of
bacteria such as Legionella.
Disinfectants and antiseptics
• Disinfectants are chemicals that kill
microorganisms.
– They should be used on inanimate
objects.
• Antiseptics are chemical agents that kill
or inhibit the growth of microorganisms
but are non-toxic enough to be applied
to living tissue.
Nutrients
• Microbes are grown on culture media.
• This can be a solid or liquid preparation
made to ensure the microbe has all the
nutrients it needs for growth.
• The media is sterilised before use.
Types of media
• Complex media contain one or more
crude sources of nutrients. Often their
exact chemical composition and
components are unknown.
• Defined media are also called synthetic
media. The components of these media
are known and are relatively pure.
Type of
medium
Example
Extra
constituents
Bacteria grown
Complex
Nutrient agar
Meat extracts,
yeast extract
Many bacteria will
grow on this
Defined
M9
Enriched
Blood agar
Blood
Streptococcus
pyogenes
Selective
MacConkey
agar
Bile salts and
crystal violet dye
Gram –ve bacteria
Escherichia coli
Oxygen
• Not all microbes need oxygen, in fact for some microbes
exposure to oxygen can be fatal!
• A microbe that must have oxygen present in order to extract
energy from foodstuffs is called an obligate aerobe. It needs
oxygen for respiration.
• An organism that would die in the presence of oxygen
is an obligate anaerobe. This is because these organisms lack
the enzymes needed to break down the dangerous chemicals
(peroxides) made in aerobic reactions.
• Some microbes (especially yeast) can grow
in the presence or absence of oxygen. These
are called facultative anaerobes. They grow
better in the presence of oxygen.
Getting oxygen
• In fermenters the actual process of
delivering oxygen can be quite tricky.
– Piping may need to be used.
– Stirrers may be placed in the fermenter to
ensure that the oxygen is available to all
the microbes.
Temperature
• All microbes have a range of temperatures
within which they are able to grow.
• However, within this range there will be a
temperature at which the organism grows
best: its optimum temperature.
• Growth occurs due to chemical
reactions and since enzymes are often
involved, temperature will have to be
tightly controlled.
• Some species of microbes live in
extremes of temperature and can
survive and grow despite the
temperature.
Psychrophiles
• Found in oceans, at the north or south
poles.
• Adapted to live in very cold
temperatures.
Mesophiles
• Microbes that live at ‘normal’
temperatures, eg room temperature
• They live on the human body.
Thermophiles
• Microbes that can live in very hot
environments.
• Examples: volcanic vents, compost
heaps, hot springs.
pH
• As with temperature, microbes have an
optimum pH.
• Fungi tend to prefer a more acidic
environment (pH 4–6).
• Bacteria like a neutral pH.
• Vinegar, citric acid and other chemicals can
be used to preserve food because they stop
microbes from growing.
• Buffers can be used to keep the pH relatively
constant.