The salt and mannitol agar or salty mannitol is a solid, selective and differential culture medium. It was created by Chapman for the isolation of pathogenic Gram positive cocci, especially Staphylococcus aureus.
However, it is also useful for isolating Staphylococus epidermidis, which can sometimes be present as an opportunistic pathogen, and Staphylococcus saprophyticus, recognized urinary pathogen, among other species.
Some Enterococcus are capable of growing on this medium, as well as certain Gram positive spore-forming rods..
This medium is very useful in the analysis of clinical samples, but it is also used in the microbiological study of food and in the quality control of industrial products, such as cosmetics, medicines, among others..
Salty Mannitol Agar is composed of extracts and peptones from beef, triptein, mannitol, sodium chloride, phenol red and agar.
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Mannitol agar is selective thanks to its high concentration of salt. Salinity acts as an inhibitory substance and prevents the growth of Gram negative bacteria.
It is also differential due to the presence of the carbohydrate mannitol and the phenol red pH indicator. From this, the bacteria capable of fermenting mannitol produce acids, acidifying the medium, turning the colonies and the medium yellow..
On the other hand, colonies that do not ferment mannitol grow in the medium taking the nutrients provided by meat extracts and peptones and triptein. From there the bacteria extract the carbon, nitrogen, vitamins and minerals necessary for their growth..
The colonies in this case can be weak or strong pink, and the medium remains the same color or changes to fuchsia.
Agar is the substance that provides the consistency to the medium.
To prepare one liter of salty mannitol agar, 111 g of the dehydrated medium from the preferred commercial company are weighed and dissolved in 1000 ml of distilled water, using a flask..
Heat is applied with frequent stirring of the medium to improve the dissolution process. Boil for a minute.
Place the flask in the autoclave at 121 ° C for 15 minutes.
At the end of the time, remove the flask from the autoclave, let it rest and serve between 15 to 20 ml on sterile Petri dishes when the temperature is approximately 50 to 55 ° C..
It is allowed to solidify, ordering in an inverted way in plaqueros and keeping in the refrigerator until use. Before sowing a sample, wait for the plate to reach room temperature.
The plates are seeded by streaking or by surface seeding with a drigalski spatula. The final pH of the prepared medium must be 7.4 ± 0.2
The color of the dehydrated medium is light beige and the color of the prepared medium is red-orange.
Due to its high selectivity, this medium is ideal for sowing samples with mixed flora in which you want to look for the presence of Staphylococus aureus, as the main pathogen of this genus.
In this sense, one of its most frequent uses is in the microbiological analysis of pharyngeal exudate and nasal discharge samples, especially to detect asymptomatic carriers of S. aureus.
Some countries have implemented this analysis as a mandatory requirement for people who want to work as food vendors..
This control prevents the hiring of carriers of S. aureus, thus avoiding massive food poisoning, due to consumption of food contaminated with staphylococcal enterotoxin.
It can also be included in the seeding of wound infections, blood cultures, CSF, bronchoalveolar lavage, among others.
Salted mannitol agar is useful for re-isolating colonies from urine cultures from CLED agar or blood agar whose Gram has revealed Gram positive cocci in clusters..
It is also valid in the microbiological analysis of food, drinking water, soils, among other applications..
Once a batch of plates with salty mannitol agar has been prepared, it is advisable to perform a quality control. Control strains are sown to show whether or not there is growth of the same.
Known strains of Staphylococcus aureus. It should grow satisfactorily developing yellow colonies, and the medium also turns the same color.
Likewise, it is convenient to include a known strain of Staphylococcus epidermidis. Should grow satisfactorily developing pink colonies, and the medium stays the same color or darkens to a stronger pink.
As a negative control, strains are used that should not grow on this medium. For example, a known strain of Escherichia coli or Klebsiella pneumoniae. The expected result is complete inhibition, that is, no growth.
Additionally, an uninoculated plate must be incubated. In it there should be no growth or color change.
It is important that the plate is not used if there are signs of deterioration, such as contamination, dehydration, discoloration, among others.
When using salty mannitol agar medium, there are some important aspects to consider:
-Obtaining a growth of yellow colonies does not indicate that it is Staphylococcus aureus. It should be remembered that some strains of Enterococcus are capable of growing in this medium and fermenting mannitol, as well as certain Gram positive spore-forming rods..
Therefore, it is important to perform a Gram on the colony and a catalase test..
-On the other hand, it must be considered that other Staphylococcus species other than aureus they are also capable of fermenting mannitol. Therefore, it is important to subculture the colony to a nutrient broth to take from there and perform the coagulase test..
Among the Staphylococcus species of clinical importance to man that ferment mannitol are: S. aureus, S. simulans, S. capitis ssp capitis, S. capitis ssp urealyticus, S. xylosus, S. cohnii ssp urealyticum, among others.
Others can give a variable reaction, that is, sometimes positive and sometimes negative. Some are S. saprophyticus, S. haemolyticus, S. warneri, S. intermedius, among other.
-It is not recommended to take colonies directly from mannitol agar to perform the coagulase test, as the high salt concentration of the medium can interfere with the result..
-Finally, it is recommended to incubate the plates seeded with salty mannitol for up to 48 hours, because some strains of S. aureus can slowly ferment mannitol, although this is rare.
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