This document discusses different types of culture media used for growing microbes. It describes media based on consistency (solid, semisolid, liquid), composition (synthetic vs non-synthetic), and application (basic, selective, differential, etc.). Key aspects covered include the use of agar as a solidifying agent, nutrients needed for microbial growth, and raw materials used in media preparation like water, carbohydrates, minerals, and buffering agents. Selective agents are also discussed which suppress unwanted microbial growth. The major nutritional requirements of microbes are carbon, nitrogen, sulfur and trace elements.

1. Culture media

3. Types of Microbial Culture Media:
(A)Culture Media Based on Consistency: 1. Solid Media
2. Semisolid media
3. Liquid Media
(B)Culture Media Based on Composition:
1. Synthetic (chemically defined) media: Known chemical composition
2. Non-synthetic/ Complex (chemically not defined) media: Unknown chemical
composition
(C)Culture Media Based on Application:
1. Basic Media: nutrient broth, nutrient agar 2. Anaerobic media
3. Enriched Media 4. Enrichment Media 5. Differential Media

4. 6. Transport Media
7. Assay Media
8. Selective Media:
i. Thayer-Martin Media ii. Mannitol- Salt Agar Media
iii. Mac-Conkeys Agar Media
iv.Wilson and Blair Agar Media
v.Crystal violet Blood Agar Media
vi. Pseudosel Agar Media

5. (A) Culture Media Based on Consistency:
1. Liquid Media or nutrient broth: Liquid consistency, Fast growth
2. Solid Media or nutrient agar:
solidifying agent (1.5-2% agar): plates, slants
• The most common solidifier is agar, first used by Robert Koch.
3. Semisolid Media: To study the motility; Agar: 0.5-0.7%
Properties of Agar:
Melts above 95℃.
• Once melted, does not solidify until it reaches 40℃., Cannot be degraded
by most bacteria.
• Polysaccharide made by red algae., Originally used as food thickener

6. (B) Culture Media Based on Composition:
Nutrient material whose exact chemical composition is known
(Defined Media):
•For chemo heterotrophs, must contain organic source of carbon
and energy (e.g.: glucose, starch etc.).
May also contain amino acids, vitamins, and other important building
blocks required by microbes.
• Expensive and not widely used.

7. Nutrient material whose exact chemical composition is unknown
(Complex Media):
• Widely used for heterotrophic bacteria and fungi.
• Made of extracts from yeast, meat, plants, protein digests etc.
• Composition may vary slightly from batch to batch.
• Vitamins and organic growth factors provided by meat and yeast extracts.
Example: Solid media (Nutrient agar), Liquid media (Nutrient broth)

8. (C)Culture Media Based on Application:
Anaerobic Growth Media:
• Used to grow anaerobes that might be killed by oxygen (Reducing media)
• Contain ingredients that chemically combine with oxygen and remove it from
the medium.
• Example: Sodium thioglycolate
• Tubes are heated shortly before use to drive off oxygen.
• Plates must be grown in oxygen free containers (anaerobic chambers).

9. Selective Media:
Used to suppress the growth of unwanted bacteria and encourage the growth of desired
microbes.
• Sabouraud’s Dextrose Agar: contains peptones, pH of 5.6 discourages bacterial growth.
Used to isolate fungi, filamentous bacteria like Nocardia
• Brilliant Green Agar: Green dye selectively inhibits gram-positive bacteria. Used to
isolate gram negative Salmonella species.
• Bismuth Sulfite Agar: Used to isolate Salmonella typhi. Inhibits growth of most other
bacteria uses glucose as a primary source of cabon.
• Differential/ Indicator Media: Used to distinguish colonies of a desired organism.

10. • Blood Agar: Used to distinguish bacteria which destroy red blood cells (hemolysis).
Hemolysis appears as an area of clearing around colony. Example: Streptococcus
pyogenes.
• Mannitol Salt Agar:
• Used to distinguish and select for Staphylococcus aureus.
• High salt (7.5% NaCl) discourages growth of other organisms.
• pH indicator changes color when mannitol is fermented to acid.

11. • MacConkey Agar:
• Used to distinguish and select for Salmonella
• Bile salts and crystal violet discourage growth of gram positive bacteria.
• Lactose plus pH indicator: Lactose fermenters produce pink or red colonies, non
fermenters are colorless.
• Enriched Media
• Enriched media contain the nutrients required to support the growth of a wide
variety of organisms, including some fastidious ones.
• They are commonly used to grow as many different types of microbes as are present in
the specimen.

12. • Examples are:
• Blood agar is an enriched medium in which nutritionally-rich whole blood supplements
the basic nutrients.
• Chocolate agar is enriched with heat-treated blood (40-45°C), which turns brown and
gives the medium that chocolate color.
• Enrichment media: These media promotes the growth of a particular organism by
providing it with the essential nutrients and rarely contains certain inhibitory substance to
prevent the growth of normal competitors.
• Used to favor the growth of a microbe that may be found in very small numbers.
• Unlike selective medium, does not necessarily suppress the growth of other microbes.

13. After incubation in enrichment medium, greater numbers of the organisms,
increase the likelihood of positive identification.
ex: Selenite F broth favors the growth of Salmonella also prevents the growth of
normal competitors like E. coli.
E. coli does not die in the medium but they do not flourish like Salmonella does

14. Raw materials for culture Media
Raw materials play an important role in media preparation. The quality of media
depends on the quality of raw materials. The most important raw materials are used for
preparations of media are water, agar, peptone, casein hydrolysate, meat extract, yeast
extract, and malt extract.
1. Water as solvent:
2. Petri dishes:
3. Energy source: The most common substance added to culture media is glucose that
acts as a source of energy and also increases the rate of growth of organisms.
4. Nutrients: The nutrients of culture media are selected to recover the required
spectrum of organisms in the sample e.g. coliforms or anaerobes. Proper nutrients
help for bacterial growth.

15. 5. Essential Metals and Minerals:
Macro-components (gm/liter): Na, K, Cl, P, S, Ca, Mg, Fe. Micro-components (mgm-
microgram/liter): Zn, Mn, Br, B, Cu, Co, Mo, V, Sr, etc.
6. Buffering Agents: The pH of a culture medium is poised around the optimum
necessary for the growth of the desired micro-organisms. examples of buffering agents
are phosphates, acetates, citrates, zwitterion compounds, and specific amino acids
7. Indicator Substances: The addition of colored indicator substances in the medium is
very effective in detecting the fermentation of specific carbohydrates.
They change color distinctly and rapidly at critical pH values.
Examples: phenol red, Bromo-cresol purple, etc., are toxic and it is essential to use in
low concentrations.
Known sensitive strains of micro-organisms are also used in the screening tests.

16. 8. Selective Agents: Chemicals or antimicrobials are added to culture media to make
them selective for certain micro-organisms. They are added at specific concentrations
to suppress the growth of unwanted organisms in a poly microbial sample. Examples:
bile salts, dye-stuffs, selenite, tetrathionate, tellurite, and azide.
9.Gelling Agents:
Gelatin is still used for a few specific media
carrageenans, alginates, silica gel, and polyacrylamides are sometimes used
as gelling agents

17. Reasons for using agar:
The most important gel-forming substance used in culture media is agar.
It is inert to microbial action and melting temperatures (38°C and 84°C
respectively)
the high gel strength which allows low concentrations of agar to be used.
It has low toxicity
It is also not hydrolyzed by agarase enzyme at high temperatures.
Microbiological agar is specially processed to yield low toxicity, high clarity, low
mineral, and high diffusion gel.

18. 10. Other Components: Many other substances are added to culture media for specific
purposes like growth factors for fastidious organisms, pH-reducing compounds for
anaerobic organisms (thioglycollate and cysteine)
11. Meat extract: It is obtained by hot water extraction of lean beef and then
concentrated by evaporation. It contains gelatin, peptones, proteases, amino acids,
creatinine, purines, and accessory growth factors.
12. Yeast extract: It is prepared from washed cells of bakers’ yeast and contains a wide
range of amino acids, growth factors, and inorganic salts.
13. Malt extract: It is prepared by extracting soluble materials from sprouted barley in
water at 55o C and concentrated by evaporation. It contains maltose, starch, dextrin,
glucose, and small amounts of protein and protein breakdown products and growth
factors.

19. Nutritional Requirements for Culture Media
Major macronutrients: C, H, O, N, S, P, K, Mg, Fe, Co, and Mn.
Major micronutrients: Zn, Co, Cu, and Mo.
Carbon and energy sources.
Growth factors.
Vitamins.

20. Nutritional Type Energy Source Carbon Source Examples
Photoautotrophs Light CO2
Cyanobacteria, some
Purple and Green
Bacteria
Photoheterotrophs Light Organic compounds
Some Purple and
Green Bacteria
Chemoautotrophs or
Lithotrophs
Inorganic
compounds, e.g. H2,
NH3, NO2, H2S
CO2
A few Bacteria and
many Archaea
Chemoheterotrophs
or Heterotrophs
Organic compounds Organic compounds
Most Bacteria, some
Archaea
Prokaryotes as per nutritional requirement