Hektoen Enteric Agar : Principle | Preparation | Interpretation

☰ Sommaire :

Ⅰ. Overview

Ⅱ. Preparation / Composition

Suspend the components, dried powder, in water (72.66 grams in 1000 ml purified/distilled water). The medium is boiled for a few seconds until the ingredients are completely dissolved. Do not autoclave or overheat .

Cool to 47°C and pour into Petri dishes. Final pH should be 7.5 + 0.2.

Hoben and al added 15 mg / liter of novobiocin to improve the selectivity of the medium by inhibiting the Citrobacter and Proteus species.

Ⅲ. Principle of Hektoen Agar

Hektoen Enteric agar is a selective and differential medium for the isolation and differentiation of pathogens from clinical samples.

◈ Selectivity : The presence of bile salts and dyes inhibits most Gram-positive organisms, which only allows Gram-negative bacilli to develop on agar.

The high concentration of bile salts partially or totally inhibits most of the non-pathogenic coliform flora of the intestinal tract. Because Salmonella and Shigella can tolerate these inhibitory substances, they generally develop faster and larger than coliforms.

◈ Differentiation : The fermentation of carbohydrates such as lactose, sucrose and salicin is one of the differentiating characteristics used to identify coliforms. Bromothymol blue turns yellow in the presence of acid and fuchsin turns red in the presence of aldehyde.

Salmonella and Shigella are unable to use these three specific carbohydrates (translucent colonies, light green or greenish blue ), while most non-pathogenic coliforms can use at least one (orange, yellow or salmon coloured colonies ).

Additional differentiation based on hydrogen sulphide production is possible due to the presence of sodium thiosulphate and iron citrate. It results in black colonies or black center, staining due to the formation of iron sulphide .

Hektoen Enteric Agar

Salmonella and E.coli on Hektoen Enteric agar

Note :
- Bile salts can precipitate out of the medium and appear as a cloudy area around colonies.
- High peptone content compensates for the inhibitory effect of bile salts on Shigella species in particular


◈ After 24 hours at 37°C aerobically, generally:

  • Fermentation of at least one of the sugars results in a "salmon / yellow / orange" coloration of the colonies.
  • The absence of fermentation results in a blue or green coloration of the colonies.
  • The production of hydrogen sulfide (H2S) is characterized by black or black center colonies.
species Growth
E. coli Large size, yellow to salmon colour, possible inhibition of some strains
Enterobacter/Klebsiella Large size, yellow to salmon
Proteus Variable size, from blue-green to blue or salmon, most strains being black in the centre or on their entire surface
Salmonella Blue-green to blue in colour, most strains being black in the centre or over their entire surface. All Salmonella serotypes have this aspect on HE agar with the exception of the Typhi serotype, which is a low producer of H2S, and rare lactose-fermenting strains of Salmonella.
Shigella Green color, moist and convex
Pseudomonas Irregular, green to brown in color
Gram-positive bacteria No / low growth


  1. Handbook of Culture Media for Food Microbiology, J.E.L. Corry et al : Hektoen Enteric (HE) agar 2003
  2. ASM : Hektoen Enteric Agar Protocol
  3. Color Atlas of Medical Bacteriology
  4. BD Hektoen Enteric Agar (HE Agar)
  5. ASM MicrobeLibrery
  6. Oxoid™ Hektoen Enteric Agar
  7. Austin Community College District
  8. HOBEN : Some Observations on the Incorporation of Novobiocin into Hektoen Enteric Agar for Improved Salmonella Isolation