1. CULTURE MEDIA LECTURE 7: Microbiology and Virology; 3 Credit hours
Atta-ur-Rahman School of Applied Biosciences (ASAB)
National University of Sciences and Technology (NUST)

2. 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

3. Types of Culture Media
Culture media can be classified on the basis of several parameters

4. Defined medium
A medium in which all chemical components are known is a defined or synthetic medium.

5. 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

6. 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.

7. 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.

8. 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.

9. Components of Complex Media

10. Components of Complex Media

11. 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
A fastidious organism is any organism that has a complex nutritional requirement

12. 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

13. Functional Types of Media
Differential media
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.

14. 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

15. 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

16. 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

17. Colony Morphology Characteristics