LECTURE 7:
CULTURE MEDIA
Microbiology and Virology; 3 Credit hours
Atta-ur-Rahman School of Applied Biosciences (ASAB)
National University of Sciences and Technology (NUST)
CULTURE MEDIA
• A culture medium is a solid or liquid
preparation used to grow, transport, and store
microorganisms.
• Much of microbiology depends on the ability
to grow and maintain microorganisms in the
laboratory
• Microorganism’s normal habitat reflects its
nutritional requirements
Types of Culture Media
Culture media can be classified on the basis of
several parameters
Defined medium
A medium in which all chemical components are
known is a defined or synthetic medium.
Complex Media
• Media that contain some ingredients of unknown
chemical composition are complex media.
• Such media are very useful
• Nutritional requirements of a particular
microorganism are unknown, and thus a defined
medium cannot be constructed
• Complex media contain undefined components
like peptones, meat extract, and yeast extract
Components of Complex Media
• Peptones are protein hydrolysates prepared
by partial proteolytic digestion of meat,
casein, soya meal, gelatin, and other protein
sources.
• They serve as sources of carbon, energy, and
nitrogen.
Components of Complex Media
• Beef extract and yeast extract are aqueous
extracts of lean beef and brewer’s yeast,
respectively.
• Beef extract contains amino acids, peptides,
nucleotides, organic acids, vitamins, and
minerals.
• Yeast extract is an excellent source of B
vitamins as well as nitrogen and carbon
compounds.
Components of Complex Media
• Agar is a sulfated polymer composed mainly of D-galactose,
-L-galactose and D-glucuronic acid.
• It usually is extracted from red algae.
• It melts at about 90°C but once melted does not harden
until it reaches about 45°C.
• Thus after being melted in boiling water, it can be cooled to
a temperature that is tolerated by human hands as well as
microbes.
• Microbes growing on agar medium can be incubated at a
wide range of temperatures.
• Most microorganisms cannot degrade it.
Components of Complex Media
Components of Complex Media
Functional Types of Media
• Media such as tryptic soy broth and tryptic soy agar are
called general purpose media or supportive media
because they sustain the growth of many
microorganisms
• Blood and other special nutrients may be added to
general purpose media to encourage the growth of
fastidious microbes.
• Neisseria gonorrhoeae : requires blood or hemoglobin
and several amino acids and vitamins in order to grow.
• These specially fortified media (blood agar) are called
enriched media
Functional Types of Media
• Selective media favor the growth of particular
microorganisms.
• inhibit the growth of some organisms but allow others to
grow.
• Mannitol Salt Agar contains a high concentration of sodium
chloride that inhibits the growth of most organisms but
permits staphylococci to grow
• Bile salts or dyes like basic fuchsin and crystal violet favor
the growth of gram negative bacteria by inhibiting the
growth of gram-positive bacteria; the dyes have no effect
on gram negative organisms.
• Endo agar, eosin methylene blue agar, and MacConkey agar
Functional Types of Media
• Differential media
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contain compounds that allow groups of microorganisms to be
visually distinguished i: appearance of the colony
ii: on the basis of some biochemical difference between the
two groups.
Blood agar is one type of differential medium, allowing
bacteria to be distinguished by the type of hemolysis produced
Some differential media are also selective, i.e., most of the
standard enteric agars such as MacConkey and EMB agars
selective for
gram-negative coliforms
Differentiate lactose-fermenting and non-lactose-fermenting
bacteria.
• Hemolysis
• Observation of the hemolytic reactions on blood agar is a very
useful tool in the identification of bacteria, particularly streptococci.
The types of hemolysis are defined as follows:
• Alpha (α) hemolysis:
An indistinct zone of partial destruction of red blood cells (RBCs)
appears around the colony coupled with a greenish to brownish
discoloration of the medium. Streptococcus pneumoniae and many
oral streptococci are α hemolytic.
• Beta (β) hemolysis:
A clear, colorless zone appears around the colonies, in which the
RBCs have undergone complete lysis. S. pyogenes, S. agalactiae,
and several other species of streptococci are β hemolytic.
• Many other bacteria besides streptococci can be β hemolytic,
including Staphylococcus aureus, Pseudomonas aeruginosa.
• Hemolytic reactions can also be a useful diagnostic tool for these
organisms.
• (γ) hemolysis: No apparent hemolytic activity or discoloration is
produced by the colony
Functional Types of Media
• It is used to distinguish pathogenic bacteria
based on the effect of bacterial enzymes
known as hemolysins which lyse red blood
cells.
• Blood agar is mainly used clinically to detect
the presence of Streptococcus pyogenes
Functional Types of Media
• MacConkey agar is both
differential and selective.
• Bile salts to support the
growth of Gram negative
microbes
Crystal violet to inhibit
the growth of Gram
positive microbes
• Since it contains lactose
and neutral red dye,
lactose fermenting
colonies appear pink to
red in color and are easily
distinguished from
colonies of non
fermenters.
Lac+ bacteria such as Escherichia coli,
Enterobacter and Klebsiella will produce acid,
which lowers the pH of the agar below 6.8 and
results in the appearance of red/pink colonies.
Non-Lactose fermenting bacteria such as
Salmonella, Proteus species, Pseudomonas
aeruginosa and Shigella cannot utilize lactose,
and will use peptone instead
Colony Morphology Characteristics