THE CULTURE MEDIA
(Microbiology and Parasitology)
Prepared by:
NESTLEE S. CABACCAN, RN, MSN-MCN,
[Link].
Associate professor III
What is Culture Media?
• Culture media- Any
materials where
microbes may tribe
for their nourishment
for medium their
reproduction.
History of Culture Media
• Robert Koch (1884-1910)-Could be
conisidered the father of the
culture media.
• His first success in bacteriology was
in the isolation of Bacillus anthracis
which at the time was causing the
disease in the cattle.
• This was the first time that any
pathogenic organism had been
isolated and studie outside of the
host body.
Koch’s Postulate
1. A particular microbe must be
found in all cases of the disease
and must not be present in
healthy animals or humans.
2. The microbe must be isolated
from the diseased animal or
human and grown in pure culture
in the laboratory.
3. The same disease must be
produced when microbes from the
pure culture are inoculated into
healthy susceptiblelaboratory
animals.
4. The same microbe must be
recovered from the experimentally
infected animals and grown again
in pure culture.
Exceptions:
• Obligate intracellular pathogens (viruses, rickettsias and chlamydias)
• Fastidious microorganisms
• Species-specific pathogens
• Synergistic infection (ex. ANUG and bacterial vaginosis)
• Effects of in vitro culture
• Not all diseases are caused by microorganism s
• Koch developed methods of cultivating bacteria
on solid media. One of Koch’s colleagues, R.J.
Petri, invented a flat glass dish (now known as a
Petri dish) in which to culture bacteria on solid
media. It was Frau Hesse—the wife of another
of Koch’s colleagues—who suggested the use
of agar (a polysaccharide obtained from
seaweed) as a solidifying agent. These methods
enabled Koch to obtain pure cultures of
bacteria.
• The term pure culture refers to a condition in
which only one type of organism is growing on
a solid culture medium or in a liquid culture
medium in the laboratory; no other types of
organisms are present.
• Koch discovered the bacterium (M.
tuberculosis) that causes tuberculosis and the
bacterium (Vibrio cholerae) that causes
cholera.
• Koch’s work on tuberculin (a protein derived
from M. tuberculosis) ultimately led to the
development of a skintest valuable in
diagnosing tuberculosis.
• Previously, other early
scientist had experimented
with various substrate for
growing bacteria; among
them are bread, potato,
coagulated egg albumin,
starch paste and meat.
• Koch observed that few
pathogenic organism would
grow on potato and other
media such as egg, prevented
adequate inspection of
colonies for growth.
• Credit must also given to Italian,
Bartolomeo Bizio, whose work
preceded Koch’s by 50 years.
• He was possibly the first to
attempt to grow an organism on
the solid medium. In 1832 there
were reports of the miraculous
appearance of “Blood spots” on
bread and common wafers.
• Bizio was able to subculture a
chromogenic bacteria onto other
bread surfaces in pure culture
and named Serratia marcenscens.
• Partial success was reached
when Koch discover the use
of gelatin that could mixed
with broths and bouillons.
In 1881, he presented this
technique at the
International Medical
Congress in London, which
was attended by Louis
Pasture and Joseph Lister.
• However, there were two discouraging problems associated
with the use of this new “Nutrient gelatin” medium:
1. It changed from solid to liquid at about 25 degrees C,
preventing it from being incubated at 37 degree C, which
is optimal for most bacteria.
2. Gelatin is liquified by the enzyme gelatinase, which is
produced by most proteolytic bacteria.
• Agar is a polysaccharide that is
derived from red seaweed. Its
ability to melt at 85 to 90 degrees C.
and solidifying at 34 to 42 degrees
C. make it ideal for culture media.
• Another clear benefits it that it is
clear for ease of observing colonies
and was found not to be toxic to
bacteria.
• When Dr. Hesse visited Koch’s lab
for several weeks, he introduced
him to the agar base media.
Unfortunately, Fanny Hesse was not
recognized for her contribution to
microbiology until 1939, five years
after her death.
Petri’s Contribution
• When agar was discovered, it was common
practice at that time to pour the media onto a
flat glass plate, then cover it with a bell jar
while it cooled.
• In 1987, another associate of Koch’s, by the
name of Petri decided to modify this method
by putting the media into glass dishes, with
sides that were about 15 mm tall and diameter
of about 100mm.
• The orginal design of this “ Petri Dish” has not
changed up to this day, except for being made
of disposable plastic.
The Introduction of Peptone
• Koch and his associates continued to experiment
with bacteria using meat infusion or extracts as their
nutrient base.
• Although this is an adequate sources of most growth
factors for bacteria, it lacks sufficient concentration
of amino-nitrogen for optimal growth.
• It was Frederick Loeffler, who was investigating
Corynebacterium diptheriae who first had the idea
of adding peptones to the nutrient broth.
• Peptone are short chain amino acids that ate
prepared by digesting meat with enzymes.
• Loeffler also added sodium chroride to the media,
which created the proper osmotic level for optimal
growth.
Requirements
• Cultivation of microorganism depends on a number
of important factors:
1. Proper nutrients must be available.
2. Oxygen nutrients must be available as required.
3. Moisture is necessary
4. The medium must have an appropriate pH.
5. Proper temperature relations must prevail.
6. The medium must be free of interfering bioburden.
7. Contamination must be prevented.
A satisfactory culture medium must contain
available sources of:
a) Carbon
b) Nitrogen
c) Inorganic phosphate and sulfur
d) Trace metals
e) Water
f) Vitamins
• These were originally supplied in the form of meat infusion.
• Beef or yeast extracts frequently replace meat infusion in
culture media.
• The addtion of peptones, which are digest of proteins,
provides readily available sources of nitrogen and carbon.
• The pH of the culture medium is important for
microorganism growth.
• Temperature is another important parameter:
Mesophilic bacteria and fungi have optimal
growth temperatures of 25-40 degree C;
Psychrophulic (cold loving) organism require
temperature below 20 degree C.
• Human pathogenic organism ate generally
mesophiles.
