Culture Media

Question 1

Use of culture media?

Answer 1

Culture media are required to grow the organisms from infected material to identified the causative agent. After identification of organism, culture media are also used for antibiotic susceptibility testing.

Question 2

Basic constituents of culture media?

Answer 2

1. Water : source of hydrogen and oxygen
2. Electrolyte : sodium chloride or other electrolytes
3. Peptone : it is a complex mixture of partially digested proteins. It is obtained from lean meat or other protein materials such as heart muscle, casein or fibrin, usually by digestion with proteolytic enzymes.
4. Meat extract : it is available commercially as ‘ Lablemco’. It contain protein degradation products, inorganic salts,carbohydrates and growth factors
5. Blood and serum : these are used for enriching culture media. Usually 5-10 defibrillated sheep blood is used. In certain media serum is used.
6. Agar : it is prepared from sea weed ( algae -geladium species). It contains mainly long chain polysaccharide, a smal amount of protein-like material and a variety of inorganic salts. It is available either in long shreds or powder form. It is used in concentration of 2-3 %. It melts at 98 C and usually solidifies at 42 C. Agar does not provide any nutrition to the bacteria but acts as a solidifying agent only.

Question 3

Difference between New Zealand agar and Japanese agar?

Answer 3

New Zealand agar has twice the jellifying capacity than that of Japanese agar.

Question 4

Brain- heart infusion?

Answer 4

It is used for the culture of non-fastidious and moderately fastidious microorganisms.

This is also recommended for the culture of Streptococcus pneumoniae for the bile solubility test

It contains beef heart, peptones, glucose, sodium chloride and buffers.

It is often used as blood culture medium

This broth can also be used to prepare inoculate for antibiotic sensitivity testing.

Question 5

Classification of media?

Answer 5

1. Based on physical state
   A. Liquid media
   B. Semisolid media
   C. Solid media
2. On the basis of presence of molecular oxygen and reducing substances in the media
    A. Aerobic media
    B. Anaerobic media
3. Based on nutritional factors
    A. Simple media
    B. Complex media
    C. Synthetic media
    D. Special media
         I. Enriched media 
        II. Enrichment media
        III. Selective media
        IV. Differential media
         V. Indicator media
        VI. Transport media
        VII. Sugar media

Question 6

Simple media?

Answer 6

• nutrient broth is an example of simple medium ( basal medium)
• it contains peptone water and meat extract 1%.
• when glucose (0.5%) is added to nutrient broth, it become glucose broth.
• when 2-3 % agar is added to nutrient broth, it becomes nutrient agar

Question 7

Commonly employed medium in the laboratory?

Answer 7

Nutrient agar for diagnostic purposes

Question 8

How to enables motile bacteria to spread?

Answer 8

If conc of agar is reduced ( 0.2-0.4 % ), semisolid medium is obtained which enable motile bacteria to spread.

Question 9

Complex media?

Answer 9

• All media other than simple media are complex.
• complex media have added ingredients for bringing out certain properties or providing special nutrient required for the growth of the bacterium.

Question 10

Synthetics media ?

Answer 10

• These are prepared from pure chemicals and the exact composition of the medium is known.
• these are used for special studies such as metabolic requirement
  eg. Dubo’s medium with tween 80

Question 11

Enriched media?

Answer 11

When basal medium is added with some nutrient such as blood,serum or egg,it is called enriched medium.

Eg: blood agar, chocolate agar, Loeffler’s serum slope

Question 12

Enrichment media?

Answer 12

-Some substances are incorporated in the liquid medium which have a stimulating effect on the bacteria to be grown or inhibits its competitors. This results in an absolute increase in the number of wanted bacteria related to other bacteria. Such media are called enrichment media.

Question 13

Uses of enrichment media?

Answer 13

Useful for culture of faeces where the non-pathogenic or commensalism bacteria tend to overgrow the pathogenic ones. Eg- salmonella being overgrown by E. coli.

Question 14

Example of enrichment media?

Answer 14

1. Tetrathionate broth : tetrathionate is added which inhibit coliforms while allows typhoid-paratyphoid bacilli to grow
2. Selenite F broth : selenite has similar action as that of tetrathionate in tetrathionate broth
3. Alkaline peptone water : it is used to grow Vibrio cholerae.

Question 15

Selective bacteria?

Answer 15

Selective media contain substances that inhibit all but a few types of bacteria and facilitate the isolation of a particular species.

Question 16

Difference between selective media and enrichment media?

Answer 16

Selective media are solid in contrast to enrichment media which are liquid.

Question 17

Examples of selective media?

Answer 17

1. Deoxycholate citrate agar (DCA): addition of deoxycholate acts as a selective agent for enteric bacilli(salmonella, shigella)
2. Bile salt agar (BSA) : bile salt is a selective agent. It favours the growth of only vibrio cholerae whereas inhibits the growth of other intestinal organisms.

Question 18

Differential media?

Answer 18

When a medium contains substances which help to distinguish differing characteristics of bacteria, it is called differential medium e.g. MacConkey’s medium which contain peptone, lactose, agar, sodium taurocholate and neural red. The lactose fermenters form pink coloured colonies whereas non-lactose fermenters produce colourless or pale colonies.

Question 19

Indicator media?

Answer 19

These media contain an indicator which change colour when a bacterium grows in them.

Question 20

Indicator media for salmonella typhi?

Answer 20

Salmonella typhi grow as black colonies on Wilson and Blair medium containing sulphite.

Question 21

MacConkey’s agar?

Answer 21

It is also an indicator medium. Due to fermentation of lactose, there is acidic pH which forms the pink colonies in the presence of neutral red indicator. The blood agar is an enriched medium but bacteria losing red blood cells give clearing (haemolysis) around their colonies, thus, it is also an indicator medium

Question 22

Indicator for corynebacterium diphtheria?

Answer 22

Potassium tellurite in potassium tellurite blood agar is reduced to metallic tellurium by corynebacterium diphtheria to produce black colonies.

Question 23

Transport media?

Answer 23

These are used in the case of delicate organism( eg gonococci) which may not survive the time taken for transit or may be overgrown by non pathogenic of specimen to the laboratory, special media are devised and these are termed transport media.

Question 24

Eg of transport media?

Answer 24

1. Stuart’s transport media : is a non nutrient soft agar gel containing a reducing agent to prevent oxidation and charcoal to neutralise bacterial inhibitors. It may be used for organism such as gonococci
2. buffered glycerol saline transport medium: for enteric bacilli

Question 25

Sugar media?

Answer 25

Sugar media helps in identification of bacteria. The term sugar in microbiology denotes any fermentable substances. Glucose, lactose, sucrose and mannitol are routinely employed for fermentation tests.

Question 26

Anaerobic media?

Answer 26

1. Cooked meat broth (CMB)

2. Thioglycollate broth

Question 27

Media for growing fungi?

Answer 27

Sabouraud dextrose agar (SDA)