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Hektoen enteric agar is used to recover gastrointestinal pathogens, such as Salmonella and Shigella, from food, water and fecal samples suspected of containing these organisms. Because of its selective nature, it inhibits most non-pathogenic enteric organisms and thus is used in clinical microbiology to recover Salmonella and Shigella from feces. It is also a differential medium that allows microbiologists to note visual differences in colony morphology and quickly eliminate non-pathogenic, gram-negative rods from pathogenic, gram-negative rods with minimal additional testing.

HE agar can be used for the primary plating of fecal specimens. 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. 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.

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