Fill in the blanks in this diagram using: Escherichia coli Staphylococcus aureus

Question

Fill in the blanks in this diagram using:

Escherichia coli
Staphylococcus aureus
Pseudomonas aeruginosa
Staphylococcus epidermis

For each of the answers, indicate if the bacterium is Gram-positive or Gram-negative.

[If unable to see diagram picture use table]

Growth on EMB

Color +

Growth on mannitol salt agar -

Metallic green

colorless

Yellow colonies

White colonies

Result:

Result:

Result:

Result:

Growth on EMB

Color +

Growth on mannitol salt agar -

Metallic green

colorless

Yellow colonies

White colonies

Result:

Result:

Result:

Result:

St.

Explanation / Answer

The flowchart designates whether there is growth of bacteria in Eosin-methylene blue (EMB) agar medium (which is shown on the left side of the chart) and if not, their growth is observed on mannitol salt agar medium (shown on right side of the chart).

EMB is a selective stain for Gram-negative bacteria. It is a blend of the stains eosin and methylene blue in 6:1 ratio. This stain is used in preparation of EMB agar, a growth medium which inhibits the growth of Gram-positive bacteria but allows many Gram-negative organisms to grow. Metallic green and colorless colonies indicates there is growth in the medium, hence they belong to Gram-negative group.

EMB agar medium serves as a color indicator that helps in distinguishing lactose-fermenting Gram-negative bacteria and those that do not ferment lactose. Gram-negative bacteria that ferment lactose will be seen as purple colored colonies since the acid produced from lactose act upon the dyes and lactose non-fermenters will be seen as colorless. But in the case of most strains of Escherichia coli, colonies show characteristic green metallic sheen in EMB agar. This is due to the fact that E. coli produces strong acids by fermenting lactose quite rapidly. This results in the rapid reduction of the pH of EMB agar which leads to the formation of green metallic sheen. So the metallic green colored colonies are E. coli and colorless ones are Pseudomonas aeruginosa.

Since EMB agar inhibits the growth Gram-positive bacteria, they are allowed to grow in mannitol salt agar. This medium contains high concentration of salt (NaCl) and helps in distinguishing pathogenic microbes in a short period of time. Owing to high concentration of salt, growth of most bacteria are inhibited but allows the growth of certain Gram-positive bacteria like Staphylococcus. Therefore, yellow and white colonies in the medium suggests they belong to Gram-positive group.

Mannitol salt agar is used as a selective and differential medium for mannitol-fermenting staphylococci, containing carbohydrate mannitol. This medium distinguishes coagulase (an enzyme that causes blood clot formation)-negative and coagulase-positive species of Staphylococcus. Coagulase-positive species (most Staphylococcus aureus strains) produce yellow colonies with yellow zones and coagulase-negative species (Staphylococcus epidermis) produce white colonies.

So the answers would be

(a) Escherichia coli (Gram negative)

(b) Pseudomonas aeruginosa (Gram negative)

(c) Staphylococcus aureus (Gram positive)

(d) Staphylococcus epidermis (Gram positive)

Growth on EMB

Color +

Growth on mannitol salt agar -

Metallic green

colorless

Yellow colonies

White colonies

Result: Escherichia coli (Gram negative)

Result: Pseudomonas aeruginosa (Gram negative)

Result: Staphylococcus aureus (Gram positive)

Result: Staphylococcus epidermis (Gram positive)

Growth on EMB

Color +

Growth on mannitol salt agar -

Metallic green

colorless

Yellow colonies

White colonies

Result: Escherichia coli (Gram negative)

Result: Pseudomonas aeruginosa (Gram negative)

Result: Staphylococcus aureus (Gram positive)

Result: Staphylococcus epidermis (Gram positive)

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