What is this media selective and differential for? – the isolation of staphylococci 2. What carbohydrate is in the media? – the sugar mannitol 3. What other molecule is important in the media? 4. What color is a negative and positive results? -Pos: Yellow -Neg:Red 5. Give one example of an organism that would be positive and one that would be negative. -Pos.: Staphylococcus aureus I -Neg.: Staphylococcus epidermidis Mannitol Salt Agar (MSA) This type of medium is both selective and differential. The MSA will select for organism: such as Staphylococcus species which can live in areas of high salt concentration (plate on the left in the picture below). This is in contrast to Streptococcus species, whose growth is selected against by this high salt agar (plate on the right in the picture below) The differential ingredient in MSA is the sugar mannitol. Organisms capable of using mannitol as a food source will produce acidic byproducts of fermentation that will lower the pH of the media. The acidity of the media will cause the pH indicator, phenol red, to turn yellow. Staphylococcus aureus is capable of fermenting mannitol (left side of loft plate) while Staphylococcus epidermidis is not right side of left plate) SA Mannitol Salt Agar History Early work indicated that the fermentation of mannitol could be used as a means of differentiating pathogenic staphylococci from nonpathogenic staphylococci. In 1942, Koch discovered that the presence of 7.5% sodium chloride in media inhibited the growth of most organisms except staphylococci. The agar then gets modified with the addition to the formula of the phenol-red mannitol agar and mannitol salt agar was the result. Purpose Mannitol salt agar (MSA) is both a selective and differential medium used in the isolation of staphylococci. It contains 7.5% sodium chloride and thus selects for those bacteria which can tolerate high salt concentrations. MSA also distinguishes bacteria based on the ability to ferment the sugar mannitol, the only carbohydrate in the medium. Theory Staphylococci can withstand the osmotic pressure created by 7.5% NaCl, while this concentration will inhibit the growth of most other gram-positive and gram-negative bacteria. Additionally, MSA contains mannitol and uses phenol red as a pH indicator (BK= 7.8) in the medium. At pH levels below 6.9, the medium is a yellow color. In the neutral pH ranges (6.9 to 8.4) the color is red; while above pH 8.4, the color of phenol red is pink. When mannitol is fermented by a bacterium, acid is produced, which lowers the pH and results in the formation of a yellow area surrounding an isolated colony on MSA. A nonfermenting bacterium that withstands the high salt concentration would display a red to pink area due to peptone breakdown. I PROTOCOL Streak a plate of mannitol salt agar with appropriate culture using the quadrant streak plate method to obtain isolated colonies. Well-isolated colonies will provide the best results in the biochemical differentiation bacteria using MSA. B

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