How does emb agar work

The green metallic sheen was not observed in the rest of the five species. Two out of the five species include the bacteria Aerobacter aerogenes and Aerobacter cloacae which are lactose fermenters. The authors reasoned that green metallic sheen formation is triggered only when the pH of the medium drops to a certain threshold. In this case, only E. Formation of this new molecule called eosin-methylene blue complex is associated with the green metallic sheen.

Like eosin and methylene blue, this complex can be taken into the bacterial cells Figure 4. The pH threshold was determined to be about 4. In other words, if any micro-organism can drop the pH of the media to 4.

In one study conducted by R. S Horvath and M. When E. Although the E. Similarly, Enterobacter aerogenes now Klebsiella aerogenes , a lactose fermenter which generally does not produce green sheen, produced green sheen on an unbuffered EMB medium. These findings further confirm that pH alone is responsible for the green metallic sheen formation.

This research group also confirmed that the Eosin — Methylene blue complex is made up of one molecule of Eosin and one molecule of Methylene blue. In the complex, Eosin and Methylene blue connected by an amide bond Figure 5. When the bacteria lactose fermenters failed to drop the pH below 4. The colour of the colony differs from the centre to edge.

The centre part of the colony being coloured, and the edge stays colourless. The reason could be, due to the difference in the density of the bacteria which in turn reflects on the amount of acid production.

On the other hand, if the bacteria increase the pH of the medium turning to alkaline would end up taking methylene blue into the cells results in blue colonies. They were capable of an increase in the pH by deamination of proteins.

One can observe the increase in blue colonies, even by increasing the concentration of methylene blue in the media to ratio or by making the EMB media initial pH between In summary, how a colony stains on EMB agar, is an interplay between the pH changes the bacteria brings in the medium, relative ratio of dyes and buffering capacity of the media.

Although the reasons mentioned above for the colony colour are relatively straightforward, it can be more complex in reality. For example, strong fermenters can form EMB complex and takes up the dye. Since the Eosin is in excess, and shown to be taken into cells under acidic conditions, plenty of chance to take up Eosin along with EMB complex.

This situation becomes more complicated when pH changes are linked to change in the colour of these dyes. Alternatively, the commercially available EMB agar media powders can be used. Weigh the mixture of content as prescribed by the manufacturer. The test sample can be directly streaked on the medium plates and then the plates should be incubated, protected from light. However, in order to obtain isolated colonies, standard procedures should be followed.

A non-selective medium should be inoculated along with conjunction with EMB Agar. Identification of isolates should be done with further confirmatory tests. The pH of the media plays a critical role in the colony colour. Care to be taken to keep the pH to the recommended value. Save my name, email, and website in this browser for the next time I comment. Table of Contents. Add a header to begin generating the table of contents.

Introduction Eosin-methylene blue EMB agar was initially formulated in by Holt-Harris and Teague, to visibly differentiate between the lactose fermenting and non-fermenting microorganisms through the use of eosin and methylene blue dyes.

Principle Eosin Methylene Blue agar EMB agar is both selective and differential culture medium ideally used for isolating faecal coliforms. Neither Eosin Y red , methylene blue blue nor a mixture of the of them is green coloured. Where does the green sheen come from?

Why some fermenting bacteria develop green sheen while others do not? Sometimes, I see blue colour colonies appear in the medium why? Figure 1. Levine modification contains 10g of lactose twice as in EMB agar and contains no sucrose.

Any growth on the media should be regarded as contamination and the whole lot should be discarded. Performance testing of prepared EMB agar plates can be done by inoculating known strains of bacteria into the medium and observing growth and colonial characteristics. News Ticker. About Acharya Tankeshwar Articles.

Hello, thank you for visiting my blog. I am Tankeshwar Acharya. Blogging is my passion. I am working as an Asst. Colonies are mm in diameter, and have greenish metallic sheen in reflected light, dark or even black centre in transmitted light. Colonies are mm in diameter, raised and mucoid, tending to become confluent.