What is Mannitol Salt Agar
A mannitol salt agar test is used to isolate and identify the presence of Staphylococcus aureus in a clinical specimen, which makes it both a selective, differential, and indicator medium.
Picture 1: Mannitol salt agar test as a part of a selective medium. On the left side grows Staphylococcus aureus and no growth took side on the right side.
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Picture 2: Mannitol salt agar test on three plates with corresponding results.
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Components of mannitol salt agar
- 1% Mannitol ( a type of sugar)
- 7% salt
- Agar (solidifying agent)
- Enzymatic digest of casein
- Enzymatic digest of animal tissue
- Beef extract
- Phenol (1, 2)
- Prepare the medium as directed by the manufacturer.
- It is best to use a ready to use dehydrated powder (the one readily available on most suppliers of culture media).
- The medium has a concentration of 11.1 grams in every 100 ml of distilled water. Sterilize through autoclaving at a temperature of 121 degree Celsius for 15 minutes.
- Allow the medium to cool down.
- Mix well before putting in a sterile petri dish. Put a label on the medium.
- Place the plate at 2 to 8 degree Celsius in a plastic bag in order to prevent loss of moisture.
- The medium can last for a few weeks provided no abnormalities in the medium’s appearance. Do not use the medium if there are any signs of abnormalities as they could indicate a possible contamination, alteration, and deterioration.
- The pH of the medium ranges between 7.3 and 7.7 at a room temperature. (2, 3, 4, and 5)
Picture 3: Staphylococcus epidermidis on mannitol salt agar.
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Picture 4: Mannitol salt agar; the plate on the left tests negative while the one on the right tests positive.
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Purpose of Mannitol salt agar
The primary purpose of mannitol salt agar is to determine or select bacteria that have the ability to grow in a high salt environment. If there is no growth after adding salt, it means that the microorganism is intolerant to salt.
Usually, Staphylococci have the ability to tolerate salt concentration. Mannitol salt sugar usually inhibits the growth of gram-positive and gram-negative bacteria. (5, 6, and 7)
Uses of Mannitol salt agar
- It differentiates and isolates Staphylococcus aureus in a clinical sample.
- It is used to identify Staphylococci in foods and dairy products.
- It is a part of the bacteriological examination of drinking water, spas, and swimming pool water.
- Mannitol salt agar is a part of the bacteriological analytical manual for cosmetic testing. (7, 8, and 9)
Mannitol salt agar principle
The peptones and beef extract present in mannitol salt agar supply all the essential nutrients for growth of the medium such as vitamins, nitrogen, minerals, and amino acids. A 7.5% sodium chloride concentration will result in the inhibition of species other than the Staphylococci. The addition of salt supplies the important electrolytes needed for transport and osmotic balance.
Mannitol is a fermentable carbohydrate. The process of fermentation produces acid which is detected by a phenol red indicator. The acid helps in the fermentation of Staphylococcal species. (1, 6, and 10)
Why Mannitol is used in mannitol salt agar?
There are different types of sugar but why mannitol is used in mannitol salt agar? The addition of sugar in the medium is used to find out if a particular microorganism can ferment sugar that is present in the medium.
For Staphylococcus, mannitol is the right sugar to use. Other organisms have a corresponding type of sugar too such as TCBS where sucrose is used. Differentiation using mannitol salt agar test is a must because not all Staphylococci are pathologic to humans.
Pathogenic Staphylococci, the ones that cause a disease to humans have the ability to ferment mannitol. If a particular specimen contains Staphylococcus aureus, you can expect that it can easily ferment mannitol. The fermentation of sugar leads to the production of acid, which changes the pH of the medium to acidic.
Mannitol salt agar has a pH indicator in the form of phenol red. If the pH level is 6.9 and above, the color of the medium is yellow. However, if the microorganism being tested has the ability to ferment mannitol, the pH level becomes acidic changing the color from yellow to pink. (2, 5, 8, and 9)
- Yellow colonies with yellow zones – It indicates the presence of Staphylococcus aureus.
- Red colonies with red zones/Colorless – it indicates the presence of organisms other than Staphylococcus aureus, usually the ones found naturally occurring in the human body such as Staphylococcus epidermidis (found on human skin).
- No growth/traces of growth – It indicates the presence of Streptococci and gram-negative bacteria.
- Large white to orange – It indicates the presence of micrococci. (3, 6, 9, and 10)
What to keep in mind?
Staphylococcus saprophyticus may have the ability to ferment mannitol. It produces yellow hallow around the colonies during mannitol salt agar test resembling with Staphylococcus aureus. Escherichia coli does not grow in the medium while Staphylococcus epidermidis has colorless to pink colonies. It is normally found on the skin of humans and does not cause any danger at all.
After performing a mannitol salt agar test, a confirmatory diagnosis is also made in the form of a coagulase test. Mannitol salt agar is a selective and differential medium. It isolates and identifies Staphylococcus aureus. At the same time, it also encourages the growth of some types of bacteria while inhibiting the growth of others. (2, 5, 8, and 9)