Complete HE Agar Lab - lab PDF

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SARAH GILDENBERG

LONE STAR COLLEGE - MONTGOMERY

BIOL 2420 - SPRING 2020

Microbiology Lab Notebook Report

Hektoen Enteric Agar - Exercise 3-5 March 24, 2020

Introduction Hektoen enteric agar is currently used as direct and indirect plating medium to recover gastrointestinal pathogens, such as Salmonella and Shigella, from food, water, and fecal samples suspected of containing these organisms.

Purpose Hektoen enteric agar is a selective as well as differential media for the isolation and differentiation of enteric pathogens from clinical specimens.The presence of the bile salts and dyes inhibit most gram-positive organisms allowing only gram-negative rods to grow on HE agar. The high concentration of bile salts partially or fully inhibits most of the nonpathogenic coliform flora of the intestinal tract. Since the enteric pathogens Salmonella and Shigella can tolerate these inhibitory substances they generally grow faster and larger than the coliforms.

Materials • • •

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Proteose, peptone and Yeast extracts : Animal peptones and yeast extract provide the nutritive base. Sodium chloride Bile salts: The high bile salt concentration inhibits growth of all gram positive bacteria and retards the growth of many strains of coliforms. Sodium thiosulfate (Sulfur source) and Ferric ammonium citrate (to detect production of H2S gas) Bromthymol blue and Acid fuchsin

Procedure 1. Mark the bottom of an HE plate into quarters, label each sector with the name of one organism 2. Aseptically streak each organism onto its sector. 2. Repeat for a control on a nutrient agar plate. 3. Incubate 24-48 hours at 37C 5. Observe plates, record results.

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SARAH GILDENBERG

LONE STAR COLLEGE - MONTGOMERY

BIOL 2420 - SPRING 2020

Results

- Rapid lactose fermenters (such as coli) are moderately inhibited and produce bright-orange to salmon pink colonies. - Salmonella colonies are blue-green typically with black centers from hydrogen sulphide gas. - Shigella appear more green than Salmonella, with the color fading to the periphery of the colony. - Proteus strains are somewhat inhibited; colonies that develop are small transparent, and more glistening or watery in appearance than species of Salmonella or Shigella.

Conclusion Most enterics can ferment at least one of the sugars in this medium. Salmonella and Shigella are not able to ferment any of them, making the colonies colorless or bluish. Salmonella is able to reduce sulfur; its colonies will be black or will have black edges.

Follow-Up Questions 1. What is the purpose of a control plate? -Purpose of control plates: to compare it to the transformation plates 2. Which ingredients in the agar differentiate between Salmonella and Shigella? -Salmonella reduces sulfur to H2S, so the colonies also contain FeS, which makes them partially or completely black.

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