Eosin Methylene Blue (EMB) agar is both selective and differential culture medium. It is a selective culture medium for Gram-negative bacteria (selects against Gram-positive bacteria) and is commonly used for the isolation and differentiation of coliforms and fecal coliforms.
EMB media assists in visual distinction Escherichia coli, other nonpathogenic lactose-fermenting enteric gram-negative rods, and the Salmonella and Shigella genera.
Differentiation between these gram-negative bacilli is based on the colony color.
- Colored colonies in EMB agar: Lactose fermenter
- Colorless colonies in EMB agar: Non-lactose fermenter
The aniline dyes (eosin and methylene blue) inhibit Gram-positive and fastidious Gram-negative bacteria.
They combine to form a precipitate at acid pH, thus also serving as indicators of acid production.
Gram-negative bacteria that ferment the lactose produce acid which turns the colonies dark purple as the acid acts upon the dyes. In addition, certain lactose-fermenting bacteria produce flat, dark colonies with a green metallic sheen. Other lactose fermenters produce larger, mucoid colonies, often purple only in their center. In EMB agar, most of the strains of E.coli colonies have a characteristic green sheen. Rapid fermentation of lactose & production of strong acids, thus a rapid reduction in the pH of the EMB agar the critical factor in the formation of the green metallic sheen observed with E.coli, rapid fermentation of lactose and formation of strong acids. Lactose non-fermenters are either colorless or light lavender.
Composition of EMB agar
|Ingredient||Eosin methylene blue (EMB) Agar (g/L)|
|Dipotassium, PO4||2 g|
|Eosin Y||0.4 g|
|Methylene blue||0.065 g|
|Distilled water||1 L|
- Enzymatic Digest of gelatin
- Lactose: Sugar, helps to differentiate lactose fermenter from the non-lactose fermenter
- Dipotassium Phosphate
- Eosin Y: Indicator
- Methylene blue: pH indicator
Expected colony characteristics of organisms in EMB agar (Quality control)
- Escherichia coli ATCC 25922: Blue-black bull’s eye; may have green metallic sheen
- Pseudomonas aeruginosa ATCC 27853: Colorless
Uses of EMB agar
Primary purpose: Isolation and differentiation of lactose fermenting and non-lactose fermenting enteric bacilli.
- EMB agar is used in water quality tests to distinguish coliforms and fecal coliforms that signal possible pathogenic microorganism contamination in water samples (presence of E.coli in the river/water sample indicates the possibility of fecal contamination of water so does the presence of other pathogenic enterics).
- EMB agar is also used to differentiate the organisms in the colon-typhoid-dysentery group. For culture of Salmonella and Shigella, selective medium such as MacConkey agar and EMB agar is commonly used.
Escherichia coli colonies grow with a metallic sheen with a dark center, Aerobacter aerogenes colonies have a brown center, and nonlactose-fermenting gram-negative bacteria appear pink.