Are you familiar with Mannitol salt agar (MSA)? Have you ever used Mannitol salt agar in your laboratory? If you have used it, but don’t remember why and when, than you are in right place. In this post I am discussing about MSA, its composition, uses and the colony of characteristics of organisms that grows on MSA.
Remember this statement: Mannitol Salt Agar (MSA) is a selective, differential and indicator medium which is used to isolate and identify S. aureus from the clinical specimen.
Mannitol Salt Agar (MSA) contains:
- Mannitol (1%): Mannitol is one of the major ingredients. Do you remember what is Mannitol? It’s a sugar.
- Salt (7.5%): Salt is the common ingredient but see the percentage, which is very high compared with other media.
- Agar: the medium contains agar (a solidifying agent).
Composition of Mannitol Salt Agar
- Enzymatic Digest of Casein (Source of nitrogen, vitamin and carbon)
- Enzymatic Digest of Animal Tissue (Source of nitrogen, vitamin and carbon)
- Beef Extract (Source of nitrogen, vitamin and carbon)
- D-Mannitol : Only Carbohydrate source present in the medium
- Sodium Chloride (Why?? see below)
- Phenol Red (Indicator)
Final pH: 7.4 ± 0.2 at 25oC
(Remember the ingredients in Bold letters)
You can purchase prepared Mannitol Salt Agar from the commercial suppliers or get the Mannitol Salt Agar Powder and prepare the media in your laboratory.
So what’s the purpose of using salt in Mannitol Salt Agar?
Incorporation of 7.5% sodium chloride in the medium helps to select only those bacteria which can tolerate high salt concentrations. MSA helps to demonstrate the ability of a bacterium to grow in a 7.5% salt environment (growth indicates tolerance for high salt environment – no growth means intolerance). Species of staphylococci are able to tolerate this salt concentration but other pathogenic bacteria may not. This concentration inhibits the growth of most other gram-positive and gram-negative bacteria
Thus MSA selectively isolates Staphylococcus spp i.e. Selective media for Staphylococcus spp
Why Mannitol (Sugar) is used in Mannitol Salt Agar?
Whenever there is the use of any sugar in any media used in Microbiology, we try to find does a particular microorganism is able to ferment sugar present in this media or not?
For example, in MacConkey Agar we try to find, does a gram negative rod is lactose fermenter or not? In TCBS, we try to identify the isolate on the basis of sucrose fermentation. Vibrio cholerae is able to ferment Sucrose and gives yellow color in TCBS.
Similarly in MSA, we try to find does a particular organism ferments Mannitol or not?
You must be aware by now; all the staphylococci spp are not pathogenic to human. So, next task of microbiology laboratory (microbiologist) is differentiating S. aureus from other Staphylococcus spp.
Pathogenic staphylococci, i.e. Staphylococcus aureus is able to ferment Mannitol (It is coagulase test positive) but others (coagulase negative Staphylococcus) are not. So, if that particular specimen contains S. aureus, it ferments Mannitol (whenever sugar is fermented acid is produced) and changes the pH of medium to acidic. As MSA contains phenol red as a pH indicator, at pH levels below 6.9, the medium is a yellow color. But if Coagulase negative staphylococcus (CONS) grow, they cant ferment Mannitol, so the color of the media around the bacterial colony does not change to yellow, it appear pink.
# Note: Staphylococcus saprophyticus (coagulse negative Staphylococcus) may ferment mannitol, producing yellow halo around colonies in MSA thus resembling with S. aureus.
So, MSA is also a differential medium.
Remember that in the neutral pH (6.9 to 8.4) the color of phenol red is red; while above pH 8.4, the color of phenol red is pink.
Expected colony characteristics of organism in Mannitol Salt Agar
- Escherichia coli: Does not grow
- Staphylococcus epidermidis: Colorless to pink colonies
- Staphylococcus aureus: Yellow colonies; may have yellow halo around colonies.
- When grown on mannitol salt agar some species of Micrococcus (Micrococcus is a normal flora of human skin, mucosa, and oropharynx), such as M. luteus (yellow) can produce yellow colonies. M. roseus (red) produces pink colonies on MSA. Find out difference between Micrococcus and Staphylococcus here
- Enterococcus faecalis and Enterococcus faecium (most common enterococcal species that has been isolated from human infections) are salt tolerant bacteria. They can ferment mannitol and produce lactic acid, producing yellow colored colonies on MSA. Catalase test can help to differentiate between Enterococcus (-ve) and Staphylococcus (+ve).