Mannitol Salt Agar MSA is used as a selective and differential medium for the isolation and identification of Staphylococcus aureus from clinical and non-clinical specimens. It encourages the growth of a group of certain bacteria while inhibiting the growth of others. It is a selective medium prepared according to the recommendations of Chapman for the isolation of presumptive pathogenic staphylococci. Mannitol Salt Agar contains peptones and beef extract, which supply nitrogen, vitamins, minerals and amino acids essential for growth.
The 7. Sodium chloride also supplies essential electrolytes for transport and osmotic balance. Mannitol is the fermentable carbohydrate, fermentation of which leads to acid production, detected by phenol red indicator, aids in the differentiation of staphylococcal species.
Coagulase positive staphylococci e. Agar is the solidifying agent. The salt clears the egg yolk emulsion and lipase production is detected as yellow opaque zone around the colonies. Hallo, My respected sir, You please tell me… How process can I measurements the antimicrobial sensitivity test of Staphylococcus aureus???
Which process or method can I following for the sensitivity test of Staphylococcus aureus?????? I kindly requested to you, please you response me. With my respect. If the organism in MSA ferments mannitol and forms yellow colonies, can we directly assume it as S.
What is the ideal method? When the electron donor is oxidized by cytochrome oxidase it turns a dark purple.
In the picture below the organism on the right Pseudomonas aeruginosa is oxidase positive. Coagulase test Coagulase is an enzyme that clots blood plasma by catalyzing the conversion of a soluble protein fibrinogen to an insoluble protein fibrin.
This test is performed on Gram-positive, catalase positive species to identify the coagulase positive Staphylococcus aureus. Coagulase is a virulence factor of S. The formation of clot around an infection caused by this bacteria likely protects it from phagocytosis. Taxos A bacitracin sensitivity testing This is a differential test used to distinguish between organisms sensitive to the antibiotic bacitracin and those not. Bacitracin is a peptide antibiotic produced by Bacillus subtilis.
It inhibits cell wall synthesis and disrupts the cell membrane. This test is commonly used to distinguish between the b -hemolytic streptococci: Streptococcus agalactiae bacitracin resistant and Streptococcus pyogenes bacitracin sensitive.
The plate below was streaked with Streptococcus pyogenes ; notice the large zone of inhibition surrounding the disk. MacConkey agar This medium is both selective and differential. The selective ingredients are the bile salts and the dye, crystal violet which inhibit the growth of Gram-positive bacteria. The differential ingredient is lactose.
Fermentation of this sugar results in a pH of less than 6. Thus organisms capable of lactose fermentation such as Escherichia coli , form bright pinky-red colonies plate pictured on the left here.
MacConkey agar is commonly used to differentiate between the Enterobacteriaceae. Spirit Blue agar This agar is used to identify organisms that are capable of producing the enzyme lipase. This enzyme is secreted and hydrolyzes triglycerides to glycerol and three long chain fatty acids. These compounds are small enough to pass through the bacterial cell wall. Glycerol can be converted into a glycolysis intermediate.
Spirit blue agar contains an emulsion of olive oil and spirit blue dye. Bacteria that produce lipase will hydrolyze the olive oil and produce a halo around the bacterial growth. The Gram-positive rod, Bacillus subtilis is lipase positive pictured on the left The plate pictured on the right is lipase negative. Starch hydrolysis test This test is used to identify bacteria that can hydrolyze starch amylose and amylopectin using the enzymes a -amylase and oligo-1,6-glucosidase.
Often used to differentiate species from the genera Clostridium and Bacillus. Because of the large size of amylose and amylopectin molecules, these organisms can not pass through the bacterial cell wall. In order to use these starches as a carbon source, bacteria must secrete a -amylase and oligo-1,6-glucosidase into the extracellular space. These enzymes break the starch molecules into smaller glucose subunits which can then enter directly into the glycolytic pathway.
In order to interpret the results of the starch hydrolysis test, iodine must be added to the agar. The iodine reacts with the starch to form a dark brown color. Thus, hydrolysis of the starch will create a clear zone around the bacterial growth. Bacillus subtilis is positive for starch hydrolysis pictured below on the left.
The organism shown on the right is negative for starch hydrolysis. This is a synergistic test between Staphylococcus aureus and Streptococcus agalactiae. The two bacteria are streaked at 90 o angles of one another. They do NOT touch. As a result, an arrow of beta-hemolysis is produced between the two streaks.
The test is presumptive for S. In the picture here, Streptococcus agalactiae was streaked throughout the top region of the plate and brought down toward the center of the plate. Staphylococcus aureus was streaked in a straight line across the center of the plate. Rings of hemolysis are evident all around S. Motility agar is a differential medium used to determine whether an organism is equipped with flagella and thus capable of swimming away from a stab mark. The results of motility agar are often difficult to interpret.
Generally, if the entire tube is turbid, this indicates that the bacteria have moved away from the stab mark are motile. The organisms in the two tubes pictured on the right are motile. If, however, the stab mark is clearly visible and the rest of the tube is not turbid, the organism is likely nonmotile tube pictured on the left. Rachel Watson, M. AG Cell: rwatson uwyo. Taxos A bacitracin sensitivity testing. It is often used to differentiate between members of Enterobacteriaceae.
In organisms capable of utilizing citrate as a carbon source, the enzyme citrase hydrolyzes citrate into oxaoloacetic acid and acetic acid. The oxaloacetic acid is then hydrolyzed into pyruvic acid and CO 2. If CO 2 is produced, it reacts with components of the medium to produce an alkaline compound e. The alkaline pH turns the pH indicator bromthymol blue from green to blue. This is a positive result the tube on the right is citrate positive.
Klebsiella pneumoniae and Proteus mirabilis are examples of citrate positive organisms. Escherichia coli and Shigella dysenteria e are citrate negative. TOP Spirit Blue agar This agar is used to identify organisms that are capable of producing the enzyme lipase. TOP Starch hydrolysis test This test is used to identify bacteria that can hydrolyze starch amylose and amylopectin using the enzymes a -amylase and oligo-1,6-glucosidase.
In the mixed acid fermentation pathway, glucose is fermented and produces several organic acids lactic, acetic, succinic, and formic acids. The stable production of enough acid to overcome the phosphate buffer will result in a pH of below 4.
If the pH indicator methyl red is added to an aliquot of the culture broth and the pH is below 4. If the MR turns yellow, the pH is above 6. Technique Streak or smear the Staphylococcus Medium No. Pigmented colonies are a deep orange colour, whilst non-pigmented colonies are white. Acid production from mannitol is best demonstrated by adding a drop of 0.
The above reactions may be conveniently performed using short sleeves, 5mm long and 10mm diameter, cut from polythene tubing. Coagulase tests should not be carried out without first sub-culturing in Nutrient Broth No. Appearance Dehydrated medium: Light straw coloured, free-flowing powder.
Prepared medium: Straw coloured gel. Due to the high slat content of this medium a slight haze may be visable in the prepared medium. This in no way affects the performance.
Precautions Enterococcus faecalis may grow on this medium as tiny colonies with slight mannitol fermentation.
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