Question

1. Complete the following table to describe the characteristics for each media.

Medium	Selective component(s)	Differential component(s)	Application	Growth appearance and example (+)	Growth appearance and example (-)	Organism unable to grow on media
TSA (Tryptic Soy Agar )(TSA)
MAC(MacConkey Agar)
EMB(Eosin-methylene Blue Agar)
MSA (Mannitol Salt Agar)
TSI (Triple Sugar Iron Agar)
SBA(Sheep Blood Agar)

Answer 1

Answer:

Study of bacteria is incomplete without growing them or culturing them on different kinds of media and noting their reactions that help in their identification. Different kinds of growth media serve different purposes; selective media only allow growth of some organisms, differential media incorporate dyes and pH indicators to differentiate the growth of certain organisms from others; the nutrient media make base for many such media and supply abundant nutrients to let bacteria grow and biochemical media give different reactions with different organism. So, let us see a few commonly used media in the laboratory:

Medium - TSA - Trypticase Soya Agar, is a general purpose nutrient medium , supporting growth of all kinds of organisms. It is many a times used as a basal medium for making other differential media like Sheep Blood Agar or Chocolate Agar. However it is used mainly for maintenance of cultures of Salmonella in lab
Selective Components: There are no such selective components as it supports all growth.
Differential Components: not usually present though sometimes salt concentration is marginally increased.
Application: As a nutrient agar for subculturing / for colony counts / for basal medium for other agars
Growth Appearance (+): Staphylococcus aureus gives bright yellow colonies & Pseudomonas aeruginosa green pigmented colonies
Growth Appearance (-): Many colonies appear similar as this will not help any organism from growing
Organism inhibited: Not inhibitory to any organisms, but fastidious organisms like Streptococcus pyogenes will not grow as they are fastidious organisms and need more nutrients (adding of blood will support these organisms)

Medium - MAC - MacConkey's Agar is the most common differential as well as selective media used in Microbiology laboratory.
Selective Components: Bile salts and crystal violet dye make it a selective medium for non-fastidious gram negative organisms. Gram positive will not grow easily on this medium
Differential Components: Presence of a pH indicator Neutral Red along with lactose as the main source of Carbon in the medium helps it differentiate Lactose fermenters from lactose non-fermenters
Application: Used as selective and differential medium in clinical laboratories isolating Enterobacteriaceae from clinical specimens
Growth Appearance (+): Lactose fermenters like Escherichia coli and Klebsiella pneumoniae give Pink colored colonies
Growth Appearance (-): where as non-lactose fermenters give pale coloured colonies, like Salmonella typhi or Shigella dysenteriae
Organism inhibited: Staphylococci and streptococci will not grow on this media or will give very small pin-point colonies after longer incubation.

Medium - EMB - Eosin Methylene Blue Agar, as the name suggests has dyes Eosin and Methylene blue (ratio is normally 6 : 1) incorporated in its ingredients. These metachromatic dyes will inhibit gram positive bacteria as well as help identify between lactose fermenters and lactose non-fermenters.
Selective Components: This medium is specifically designed to discourage growth of Gram Positive organisms, making it a selective medium for Gram negatives
Differential Components: Presence of dyes also incorporate colour to colonies that can ferment lactose and leave colonies that can not ferment lactose, colourless.
Application: Just like MAC agar, it is commonly employed in clinical laboratories to pick up non-lactose fermenting pathogens and for identifying coliforms
Growth Appearance (+): Escherichia coli will give colonies with greenish black metallic sheen due to metachromatic nature of the dyes included, other lactose fermenters will give dark purple centered colonies
Growth Appearance (-): whereas non-lactose fermenters like Salmonella typhi or Shigella dysenteriae give pale, colourless colonies on EMB agar
Organism inhibited: Staphylococci and streptococci as well as Bacillus cereus and Bacillus subtilis are inhibited.

Medium - MSA - Mannitol Salt Agar is again another commonly used medium in quick identification of pathogenic bacteria. This too is selective only for certain groups of bacteria while inhibits others; at the same time help differentiate those who grow on the basis of mannitol fermentation
Selective Components: 7 - 10% NaCl
Differential Components: Mannitol and Phenol Red indicator
Application: Commonly used in medical laboratories for quick identification of certain pathogens
Growth Appearance (+): Staphylococcus aureus - Yellow colonies as it ferments Mannitol, producing acid
Growth Appearance (-): Staphylococcus epidermidis - Pink colonies as it does not ferment Mannitol and rather decreases pH by using amines and peptides in the medium
Organism inhibited: All Streptococci and most of the gram negative bacteria.

Medium - TSI - Triple Sugar Iron Agar, poured as slants in tubes is one of the rapid biochemical medium used for identifying organisms on the basis of fermentation of 3 sugars namely Glucose 0.1%, Lactose 1.0% and Sucrose 1.0% and ability to produce gas (H2) and also if they produce H2S.
Selective Components: It is not used as selective media as it does not select one organism over other
Differential Components: Three sugars / Phenol Red indicator / Sodium thiosulphate & ferrous sulphate
Application: as a biochemical test for identifying bacteria that will grow aerobically &/or facultatively as well as use three sugars and also produce H2S
Growth Appearance (+):
Escherichia coli - Acid Slant / Acid butt + gas but No H2S.
Salmonella typhi - Alk. Slant / Acid butt with black precipitation due to H2S production
Shigella flexneri - Alk. Slant / Acid butt without black precipitation as no H2S
Growth Appearance (-): Pseudomonas aeruginosa - no change in medium as only grows on slant (aerobic)
Organism inhibited: It allows growth of all organisms though strict aerobes will grow only on slant and will be non fermentative

Medium - SBA - Sheep Blood Agar is an enriched medium and due to presence of sheep blood it makes medium as good nutritive medium with lots of growth factors for bacteria to grow,
Selective Components: It does not include any selective components.
Differential Components: Presence of sheep blood make it differential by the way red blood cells are hemolysed by the organisms
Application: Used as a common differential media for gram positive organisms by identifying the haemolysis pattern
Growth Appearance (+):
Staphylococcus aureus = beta hemolysis- clear zone
Streptococcus viridans = alpha hemolysis - green zone around colony (partial hemolysis)
Growth Appearance (-): Staphylococcus epidermidis = gamma haemolysis (no hemolysis)
Organism inhibited: This is an enriched medium and not selective for any group. Hence it supports growth of most non-fastidious organisms