Mannitol Salt Agar (MSA) is a differential and selective growth medium devised by Chapman for the isolation and identification of coagulase-positive staphylococci (e.g., Staphylococcus aureus) from coagulase-negative staphylococci from the clinical and non-clinical specimens. MSA medium is important in medical labs by distinguishing pathogenic microbes
Composition of Mannitol Salt Agar
1. Peptic Digest of Casein: Supply of nitrogen, vitamins and essential amino acids
2. Peptic Digest of Animal Tissue: Supply nitrogen, vitamin and carbon
3. Beef Extract: Source of nitrogen, vitamin and amino acids.
4. D-Mannitol: Fermentable Carbohydrate source present in the medium
5. Sodium Chloride :
• Inhibit the growth of certain bacteria other than staphylococci
• Supply electrolyte for transport and osmotic balance
6. Phenol Red (Indicator): helps in differentiating staphylococcal species
7. Agar: Solidifying agent.
Selective media: MSA selectively isolates Staphylococcus spp.Incorporation of 7.5% sodium chloride in the medium results in partial or complete inhibition of bacterial organisms other than staphylococci. It helps to select only those bacteria which can tolerate high concentrations of salt. Staphylococci spp. are capable of tolerating higher salt concentration but other pathogenic bacteria may or may not survive under high salt concentrations.
Differential media: MSA is also a differential medium for mannitol-fermenting staphylococci, containing carbohydrate mannitol and the indicator phenol red indicator. Since all the staphylococci spp are not pathogenic to human. So, it is necessary to differentiate S. aureus from other Staphylococcus spp. Pathogenic staphylococci, i.e. Staphylococcus aureus can ferment Mannitol (It is coagulase test positive) but others (coagulase-negative Staphylococcus) are not. So, if that particular sample contains S. aureus, it ferments Mannitol (whenever sugar is fermented acid is produced) and changes the pH of the medium to acidic. MSA contains phenol red as a pH indicator which is used to detect acid produced by mannitol-fermenting staphylococci. at pH levels below 6.9, Staphylococcus aureus produces yellow colonies with yellow zones, whereas other coagulase-negative staphylococci produce small pink or red colonies with no color change to the medium. If an organism can ferment mannitol, an acidic byproduct is formed that causes the phenol red in the agar to turn yellow. It is used for the selective isolation of presumptive pathogenic (pp) Staphylococcus spp.
Preparation of Mannitol salt agar
Mannitol salt agar is best prepared from ready to use dehydrated powder, available from most suppliers of culture media. The medium is usually used at a concentration of 111.0.25 gm in 1000 ml distilled water.
1. Prepare the medium as instructed by the manufacturer. Sterilize by autoclaving at 121oC for 15 minutes.
2. When the medium has cooled to 50-55oC, mix well, and dispense it aseptically in sterile Petri dishes. Date the medium and give it a batch number.
3. Store the plates at 2-8oC preferably in plastic bags to prevent loss of moisture.
Shelf life: Several weeks providing there is no change in the appearance of the medium to suggest contamination, deterioration, or alteration of pH.
pH of the medium: pH 7.3- 7.7 at room temperature.
Expected results on Mannitol Salt Agar
1. Escherichia coli: Does not grow
2. Staphylococcus epidermidis: Colorless to pink colonies
3. Staphylococcus aureus: Yellow colonies; may have a yellow halo around colonies.
Uses of Mannitol Salt Agar
• For the selective isolation and differentiation of Staphylococcus aureus from clinical and nonclinical samples.
• For detection of staphylococci in food and dairy products.
• MSA is included in the Bacteriological Analytical Manual for cosmetics testing.
• For bacteriological examination of swimming pool water, spas and drinking water using membrane filtration
Limitations of Mannitol Salt Agar
• Several Staphylococcus species other than aureus are mannitol positive and produce yellow colonies surrounded by yellow zones on this medium (e.g. S. capitis, S. xylosus, S. cohnii, S. sciuri, S. simulans, and other species) therefore, further biochemical tests are necessary for the identification of Staphylococcus aureus or other species.
• Most organisms other than staphylococci are inhibited by the high salt concentration found in Mannitol Salt Agar except for some halophilic marine organisms.
• Certain strains of S.aureus may show a delayed fermentation of mannitol. Negative plates should be re-incubated overnight before discarding.
• Presumptive Staphylococcus aureus must be confirmed with a coagulase test.