Result Interpretation on Mannitol Salt Agar

sugar mannitol

Mannitol Salt is a selective bacterial growth medium because it has a very high concentration of NaCl (7.5%). Most bacteria cannot survive in this highly saline, hypertonic environment. But the genus Staphylococcus has a protective slime layerthat protects it in a harsh, salty environment.

Mannitol Salt Agar (MSA) is a selective and differential medium. The high concentration of salt (7.5%) selects for members of the genus Staphylococcus, since they can tolerate high saline levels. Organisms from other genera may grow, but they typically grow very weakly.

Mannitol salt agar or MSA is a commonly used selective and differential growth medium in microbiology. It encourages the growth of a group of certain bacteria while inhibiting the growth of others.

What is the purpose of adding salt to mannitol salt agar? High concentrations of salt inhibit the growth of non-halophilic organisms but allow for the growth of bacteria from the genus Staphylococci.

What would be the likely consequences of omitting the NaCl in Mannitol Salt Agar? Why? Non-staphylococcus bacteria would be able to grow on the media. This may lead to false positives for Non-staphylococcus that can ferment mannitol.

The MSA agar will retain its initial red color and will not change to yellow. Gram-negative bacteria like E. coli and P. aeriginosa are not tolerant to salt (not halophilic) and will not grow colonies on MSA (see quadrants II and IV).

MacConkey agar is used for the isolation of gram-negative enteric bacteria and the differentiation of lactose fermenting from lactose non-fermenting gram-negative bacteria.

Altogether, MacConkey agar only grows gram-negative bacteria, and those bacteria will appear differently based on their lactose fermenting ability as well as the rate of fermentation and the presence of a capsule or not.

Escherichia coli (E. coli) is a Gram-negative, rod-shaped, facultative anaerobic bacterium.

Preparation of MacConkey Agar

1. Suspend 49.53 grams of dehydrated medium in 1000 ml of distilled water.
2. Heat to boiling to dissolve the medium completely.
3. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes.
4. Cool to 45°C -50°C.
5. Mix well before pouring into sterile Petri plates.

A differential and selective growth medium used to isolate and identify gram-negative bacilli, often enteric pathogens, based on fermentation or lack of fermentation of a sugar added to the media.

Sorbitol MacConkey agar is a variant of traditional MacConkey agar used in the detection of E. coli O157:H7. This is important because gut bacteria, such as Escherichia coli, can typically ferment lactose, while important gut pathogens, such as Salmonella enterica and most shigellas are unable to ferment lactose.

Nutrient broth, tryptic soy broth, and chocolate agar, are all examples of complex media. Selective media are used for the growth of only selected microorganisms. An example of a selective medium is MacConkey agar (Table 9.1 & Figure 9.26).

MacConkey agar not only selects for Gram-negative organisms by inhibiting Gram-positive organisms and yeast but also differentiates the Gram-negative organisms by lactose fermentation.

Blood agar contains many unspecified nutrients, supports the growth of a large number of bacteria, and allows differentiation of bacteria according to hemolysis (breakdown of blood). The medium is complex and differential.

A decolorizer such as ethyl alcohol or acetone is added to the sample, which dehydrates the peptidoglycan layer, shrinking and tightening it. The large crystal violet-iodine complex is not able to penetrate this tightened peptidoglycan layer, and is thus trapped in the cell in Gram positive bacteria.

The Crystal Violet assay is based on staining cells that are attached to cell culture plates. It relies on the detachment of adherent cells from cell culture plates during cell death. During the assay, dead detached cells are washed away. The remaining attached live cells are stained with Crystal violet.

Crystal violet is also a mitotic agent and mutagen. It has been used for topical treatment in creams against fungal and bacterial infections. Its been used against some pathogenic fungi such as Candida species and some gram-positive bacteria, e.g. Staphylococcus species.

In the Gram staining method, crystal violet is used to differentiate between Gram Positive and Gram Negative bacteria.

Harmful if inhaled. May be harmful if absorbed through the skin. May cause eye and skin irritation. May cause respiratory tract irritation.

When iodine is applied, decolorizing time for all cells is longer than without iodine. Thus, iodine penetrates the cell and serves to form a dye-iodine precipitate, and since all cells are less permeable for the dye-iodine toward decolorizing agents, slower removal of it results.

Crystal violet or gentian violet, also known as methyl violet 10B or hexamethyl pararosaniline chloride, is a triarylmethane dye used as a histological stain and in Gram’s method of classifying bacteria.

Crystal violet is a biohazardous substance, which by in-vitro investigations suggests it is poisonous and carcinogenic to some species of fish. In humans, it irritates the eyes moderately, can permanently injure the cornea and in extreme cases, cause respiratory and kidney failures [15] .

Gram-negative bacteria are classified by the color they turn after a chemical process called Gram staining is used on them. Gram-negative bacteria stain red when this process is used. Other bacteria stain blue.

What are 3 possible uses for Crystal Violet? Simple Stain. Gram Stain. Anti-fungal/anti-gram (+) bacteria.

The first step in gram staining is the use of crystal violet dye for the slide’s initial staining. The final step in gram staining is to use basic fuchsin stain to give decolorized gram-negative bacteria pink color for easier identification. It is also known as counterstain.

The purple, crystal-violet stained cells are referred to as gram-positive cells, while the red, safranin-dyed cells are gram-negative (Figure 3).

This is a simple assay useful for obtaining quantitative information about the relative density of cells adhering to multi-well cluster dishes. The dye in this assay, crystal violet, stains DNA.