3.4: Use of Specialized Media

To supplement mechanical techniques of isolation such as the streak plate method, many special-purpose media are available to the microbiologist to aid in the isolation and identification of specific microorganisms. These special purpose media fall into four groups: selective media, differential media, enrichment media, and combination selective and differential media.

1. Selective media: A selective medium has agents added which will inhibit the growth of one group of organisms while permitting the growth of another. For example, Columbia CNA agar has the antibiotics colistin and nalidixic acid added which inhibit the growth of Gram-negative bacteria but not the growth of Gram-positives. It is, therefore, said to be selective for Gram-positive organisms, and would be useful in separating a mixture of Gram-positive and Gram-negative bacteria.

2. Differential media: A differential medium contains additives that cause an observable color change in the medium when a particular chemical reaction occurs. They are useful in differentiating bacteria according to some biochemical characteristic. In other words, they indicate whether or not a certain organism can carry out a specific biochemical reaction during its normal metabolism. Many such media will be used in future labs to aid in the identification of microorganisms.

3. Enrichment media: An enrichment medium contains additives that enhance the growth of certain organisms. This is useful when the organism you wish to culture is present in relatively small numbers compared to the other organisms growing in the mixture.

4. Combination selective and differential media: A combination selective and differential medium permits the growth of one group of organisms while inhibiting the growth of another. In addition, it differentiates those organisms that grow based on whether they can carry out particular chemical reactions.

For example, MacConkey agar (see Fig. \(\PageIndex{1}\)) is a selective medium used for the isolation of non-fastidious Gram-negative rods, particularly members of the family Enterobacteriaceae and the genus Pseudomonas, and the differentiation of lactose fermenting from lactose non-fermenting Gram-negative bacilli. MacConkey agar contains the dye crystal violet well as bile salts that inhibit the growth of most Gram-positive bacteria but do not affect the growth of most Gram-negatives. If the Gram-negative bacterium ferments the sugar lactose in the medium, the acid end products lower the pH of the medium. The neutral red in the agar turns red in color once the pH drops below 6.8. As the pH drops, the neutral red is absorbed by the bacteria, causing the colonies to appear bright pink to red.

Fig (\PageIndex{1A}\): Uninoculated Plate of MacConkey Agar	Fig. (\PageIndex{1B}\): Escherichia coli Growing on MacConkey Agar	Fig. (\PageIndex{1C}\): Klebsiella aerogenes (formerly known as Enterobacter aerogenes) Growing on MacConkey Agar	Fig. (\PageIndex{1D}\): Proteus mirabilis Growing on MacConkey Agar
MacConkey agar is a selective medium used for the isolation of non-fastidious Gram-negative rods, particularly members of the family Enterobacteriaceae and the genus Pseudomonas, and the differentiation of lactose fermenting from lactose non-fermenting Gram-negative bacilli. MacConkey agar contains the dye crystal violet well as bile salts that inhibit the growth of most Gram-positive bacteria but do not affect the growth of most Gram-negatives.	MacConkey agar is a selective medium used for the isolation of non-fastidious Gram-negative rods, particularly members of the family Enterobacteriaceae and the genus Pseudomonas, and the differentiation of lactose fermenting from lactose non-fermenting Gram-negative bacilli. MacConkey agar contains the dye crystal violet well as bile salts that inhibit the growth of most Gram-positive bacteria but do not affect the growth of most Gram-negatives. Strong fementation of lactose with high levels of acid production by the bacteria causes the colonies and confluent growth to appear bright pink to red. The resulting acid, at high enough concentrations, can also causes the bile salts in the medium to precipitate out of solution causing a pink precipitate (cloudiness) o appear around the the growth (arrows).	MacConkey agar is a selective medium used for the isolation of non-fastidious Gram-negative rods, particularly members of the family Enterobacteriaceae and the genus Pseudomonas, and the differentiation of lactose fermenting from lactose non-fermenting Gram-negative bacilli. MacConkey agar contains the dye crystal violet well as bile salts that inhibit the growth of most Gram-positive bacteria but do not affect the growth of most Gram-negatives. Weak fermentation of lactose by the bacteria causes the colonies and confluent growth to appear pink or pink with red centers (arrows), but without the precipitation of bile salts there is no pink halo around the growth.	MacConkey agar is a selective medium used for the isolation of non-fastidious Gram-negative rods, particularly members of the family Enterobacteriaceae and the genus Pseudomonas, and the differentiation of lactose fermenting from lactose non-fermenting Gram-negative bacilli. MacConkey agar contains the dye crystal violet well as bile salts that inhibit the growth of most Gram-positive bacteria but do not affect the growth of most Gram-negatives. If the bacteria do not ferment lactose, the colonies and confluent growth appear colorless and the agar surrounding the bacteria remains relatively transparent (arrow).
Copyright; Gary E. Kaiser, Ph.D. The Community College of Baltimore County, Catonsville Campus CC-BY-3.0

Typical colony morphology on MacConkey agar is as follows:

Escherichia coli: colonies and confluent growth appear bright pink to red and surrounded by a pink precipitate (cloudiness) in the agar surrounding the growth. (See Fig. (\PageIndex{1B}\).)

Enterobacter and Klebsiella: colonies and confluent growth appear bright pink to red but are not surrounded by a pink precipitate (cloudiness) in the agar surrounding the growth. (See Fig. (\PageIndex{1C}\).)

Pseudomonas, Salmonella, Serratia, Proteus, and Shigella: colorless colonies; agar relatively transparent. (See Fig. (\PageIndex{1D}\).)

There are literally hundreds of special-purpose media available to the microbiologist. Today we will combine both a mechanical isolation technique (the streak plate) with selective and selective-differential media to obtain pure cultures from a mixture of bacteria. In future labs, such as 12 - 16, which deal with the isolation and identification of pathogenic bacteria, we will use many additional special-purpose media.

Contributors and Attributions

• Dr. Gary Kaiser (COMMUNITY COLLEGE OF BALTIMORE COUNTY, CATONSVILLE CAMPUS)