Published: 25th Jun 2026, 17:38

In their daily life, a microbiologist works with a variety of samples. And the one who works with Stool Samples is familiar with the common challenge of stool samples.

How to find the hidden pathogen from a pool of harmless intestinal bacteria?

In the clinical laboratory settings, when a faecal sample comes with the suspicion of salmonellosis or shigellosis, the main aim is now to identify the enteric pathogens Salmonella and Shigella. These target organisms are probably contributing a small fraction of the total microbial population present in the sample. Hence, it is difficult to find them without a Selective Culture Media.

Hope you know about Selective Culture Media if not, then click to know: https://www.tmmedia.in/selective-media-the-gatekeeper-of-microbiology-labs/

So now back to our topic, so this is the situation when hektoen enteric agar (HE Agar) comes into the frame or we can say on the LAF (Laminar Air Flow).

Why Hektoen Enteric Agar?

HE Agar is a medium that performs double duty. It is both a selective and a differential agar but it is not a general-purpose agar. This medium contains bile salts and its speciality is that it inhibits many Gram-positive organisms and reduces the interference arising from them, as our target microorganisms are gram-negative in nature. Other than bile salt, this medium also contains lactose, sucrose, and salicin all of these ingredients help distinguish organisms based on their carbohydrate fermentation patterns.

And the most impressive thing is HE Agar can also detect whether an organism is producing hydrogen sulphide or not. This characteristic helps in rapid identification of Salmonella because its colonies often develop distinctive black centers due to hydrogen sulphide production. Hence, microbiologists can presume the presence of salmonella before biochemical tests.

But Biochemical confirmatory tests are always mandatory because some species of Proteus can also produce hydrogen sulphide and similar black centers colonies. Also, atypical strains may not always exhibit textbook morphology.

So the conclusion is never skip Biochemical Confirmatory Tests.

Composition of Hektoen Enteric Agar (TM 121)

Let’s Know About The Colonies of Other Gram-Negative Organisms.

The HE Agar has the ability to translate metabolism reactions into visible clues. Organisms such as E. coli ferment the carbohydrates present in the medium, which results in acid production and orange or salmon-pink-coloured colonies. These bright colonies generally indicate normal enteric flora rather than pathogens.

On the other hand, Shigella species cannot ferment these carbohydrates and appear as green-to-blue-green colonies.

And as previously mentioned, Salmonella species often produce the most recognizable appearance due to hydrogen sulphide production. They appear as blue-green colonies with characteristic black centers.

Suppressing competing flora while simultaneously differentiating pathogens based on their biochemical behaviour, this ability of HE Agar has made it a staple in clinical, food, environmental, and pharmaceutical microbiology laboratories worldwide.

Role of HE Agar Outside Clinical Lab

The HE Agar is not only associated with Stool Samples, Salmonellosis or Shigellosis. Its applications extend much further.

In Food laboratories,

Foodborne outbreaks are one of the major public health concerns globally. Food laboratories routinely use HE Agar to investigate poultry, meat products, dairy products, spices, processed foods, and ready-to-eat meals.

In Pharmaceutical manufacturing,

Pharmaceutical manufacturing is one of the industries where sterility and asepsis are most important. Any contamination by enteric pathogens can have serious consequences. Pharmaceutical industries frequently use HE Agar during microbiological quality control programmes to screen raw materials and finished products for Salmonella contamination.

In Environmental Monitoring,

Enteric bacterial contamination in water is a serious concern. Water quality laboratories use HE Agar during monitoring of drinking water, wastewater, and other environmental samples.

With the modernization of the world, microbiology labs are also stepping up and we are in the Molecular Era. 

So why are the traditional culture-based methods still relevant?

Modern diagnostic laboratories rely on PCR, MALDI-TOF MS, and next-generation sequencing technologies that may indicate the presence of Salmonella DNA, but they cannot provide a viable isolate for further analysis. And regulatory agencies, food safety laboratories, and pharmaceutical quality control facilities still require viable isolates for confirmation, antimicrobial susceptibility testing, epidemiological investigations, and strain characterization.

Culture Media (HE Agar) bridges this gap. Because microbiologists can recover the actual organism through colonies. For this reason, culture media continue to play a crucial role alongside molecular diagnostics rather than being replaced by them.

Tips to get reliable results with HE Agar

If you are using Dehydrated Hektoen Enteric Agar

1.      Accurately prepared the media.

2.      Avoid overheating during preparation.

3.      Adhere to the recommended incubation periods for reading plates.

4.      Follow up with biochemical and serological tests for confirmation.

If you are using a Ready-to-Use Hektoen Enteric Agar Plate

1.      Open plates aseptically.

2.      Adhere to the recommended incubation periods for reading plates.

3.      Follow up with biochemical and serological tests for confirmation.

Conclusion

HE Agar is more than a selective agar; it is a practical example of how microbiological principles can be translated into a powerful diagnostic tool. By combining selective inhibition, carbohydrate fermentation analysis, and hydrogen sulphide detection, it enables microbiologists to rapidly identify potential enteric pathogens from a pool of unwanted microorganisms.

With the modernization of labs, traditional culture methods are still the gold standard for identifying, studying and isolating the microorganisms.

Frequently Asked Questions (FAQs)

1. Are there any other organisms, besides Salmonella, which give distinctive black centers colonies due to hydrogen sulphide production?

A. Yes, some species of Proteus can also produce hydrogen sulphide and similar black centers colonies.

2. What are the differences in salmonellosis and shigellosis?

A. Both of them are bacterial infections of the digestive system. Salmonellosis is caused by Salmonella bacteria and shows symptoms such as watery, self-limiting diarrhoea. It is spread through contaminated food. On the other hand, Shigellosis is caused by Shigella bacteria, which cause severe bloody dysentery and it spreads from person to person through the faecal-oral route.

3. Is Hektoen Enteric Agar a selective culture media or a differential culture media?

A. It is both a selective and a differential agar.

4. Hektoen Enteric Agar is used for which species of bacteria?

A. Hektoen Enteric Agar is used to isolate and identify enteric pathogens, Salmonella & Shigella.

5. What is the role of lactose, sucrose, and salicin in Hektoen Enteric Agar?

A. HE Agar contains lactose, sucrose, and salicin; all of these ingredients help distinguish organisms based on their carbohydrate fermentation patterns.