and Citrobacter freundii, also produce H2S, but these organisms are usually inhibited by the bile salts in the HE agar. Other nonpathogenic enteric organisms, such as Proteus spp. The H2S gas reacts with the ferric ammonium citrate, forming a precipitate, resulting in colonies that are black or have a black center. This sulfide molecule then couples with a hydrogen ion to form H2S gas. Salmonella produces bacterial enzymes that cause a sulfide molecule to be released from the thiosulfate present in the media. The production of H2S by certain enteric gram-negative rods, such as Salmonella, can be detected on HE agar due to the addition of thiosulfate and ferric ammonium citrate to the formula. Stool culture on Hektoen enteric agar: mixed flora including Escherichia coli (red arrow), Salmonella (blue arrow), and Proteus vulgaris (yellow arrow). Sodium thiosulfate (sulfur source) and ferric ammonium citrate (to detect the production of H2S gas).Bile salts: The high bile salt concentration inhibits the growth of all gram-positive bacteria and retards the growth of many strains of coliforms.Protease, peptone, and yeast extracts: Animal peptones and yeast extract provide the nutritive base.Since the enteric pathogens, Salmonella and Shigella can tolerate these inhibitory substances they generally grow faster and larger than the coliforms.
The high concentration of bile salts partially or fully inhibits most of the nonpathogenic coliform flora of the intestinal tract. The presence of the bile salts and dyes inhibits most gram-positive organisms allowing only gram-negative rods to grow on HE agar. Hektoen enteric agar is a selective as well as differential media for the isolation and differentiation of enteric pathogens from clinical specimens. Direct inoculation of colonies from agar plates may produce sufficient growth of organisms that would otherwise be inhibited in a more dilute inoculum from diarrheal feces or broth culture. It may also be used to subculture the overnight growth from enrichment broths (such as gram-negative broth or selenite broth) inoculated with fecal specimens suspected of containing low numbers of Salmonella. HE agar can be used for the primary plating of fecal specimens. Differential medium: It allows microbiologists to note visual differences in colony morphology and quickly eliminate nonpathogenic gram-negative rods from pathogenic gram-negative rods with minimal additional testing.Selective medium: It inhibits most nonpathogenic (normal flora) enteric organisms and assists in selective recovery of Salmonella and Shigella from feces.Hektoen enteric agar is currently used as a 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.
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