MacConkey Agar is one of the most commonly used selective and differential media in microbiology. This media was first developed in the early 20th century by the British bacteriologist Alfred Theodore MacConkey, who originally designed to differentiate typhoid bacilli (Salmonella Typhi) from other coliform organisms in contaminated water supply.
It plays a very crucial role in identifying and differentiating gram-negative enteric bacteria particularly belonging to Enterobacteriaceae family and Pseudomonas species.
MacConkey agar is developed for the isolation of gram-negative enteric bacteria and differentiation of lactose fermenting from non-lactose fermenting gram-negative bacterias.
Lactose: Lactose acts as the fermentable carbohydrate source, lactose allows differentiation between gram-negative bacterias. Bacteria that can ferment produce acid (lactic acid, acetic acid, succinic acid, formic acid) as a byproduct which can be easily visualized in the medium. Depending on species of bacterias and specific fermentation pathway, bacterias might produce different organic acids.
Bile Salts & Crystal Violet: Bile salts and crystal violet selectively inhibit the growth of gram-positive bacterias by disrupting cell membranes, gram-negative bacteria's cell membrane blocks these chemicals, allowing gram-negative bacteria to grow. This is very crucial in clinical analysis, where gram-positive bacterias might overgrow and shadow the presence of gram-negative bacterial growth.
Neutral red: As we discussed earlier lactose fermenting bacterias produce organic acid, this drops the medium’s pH below 6.8. The neutral red in the medium absorbs hydrogen ions released by organic acid and undergoes structural change. This causes the indicator to turn bright red or pink, staining lactose fermenting bacterias, where non-lactose fermenting bacterias stays clear.
Peptones & sodium chloride: These components provide essential nutrients for bacterial growth, including amino acids and salts. This supports the fast growth of wide range gram-negative bacterias.
1. Dissolve the recommended quantity of MacConkey agar medium in purified water (quantity varies based on the manufacturer).
2. Heat the container with medium gently with frequent agitation until it is fully dissolved, make sure it doesn't have any clumps which might interfere with uniform growth.
3. Autoclave the prepared medium at 121°C for 15 minutes to eliminate any potential contaminants.
4. Allow the medium to cool to 45 - 50°C and pour the medium into sterile petri dishes.
5. After complete solidification of the petri dishes, label the media plates with their name and preparation date.
6. Use a sterile loop or swap to streak out the sample onto the surface of the plate.
7. Incubate the plates aerobically at 35 - 37°C for the primary result and read it again at 48 hours.
Lactose fermenting bacterias produce acid as a by-product, lowering the pH and turning colonies pink or red. Very strong fermenting bacterias such as E.coli may show pink halo around the colonies due to precipitation of bile salt.
Lactose non-fermentering bacteria cannot ferment lactose. These bacterial colonies remain colorless or appear pale. The medium surrounding the colonies stays relatively transparent.
MacConkey agar is used for the selective isolation of gram-negative bacterias, especially coliforms and enteric pathogens.
It is used in the differentiation of lactose fermenting bacterias from non-fermenting gram-negative bacterias.
In the microbiological examination MacConkey agar is widely used in clinical, food, water analysis to detect coliform count.
MacConkey agar can be only used for preliminary or probable identification only. It is necessary to subculture and carry out several other tests for final confirmation.
MacConkey agar is designed to support non-fastidious gram-negative rod bacterias; certain bacterias might be sensitive to the agents and exhibit poor growth.
Incubating MacConkey agar under increased CO2 forms carbonic acid which lowers the medium’s pH. This high acidic condition increases the toxicity of bile salts and crystal violet suppressing the growth of gram-negative bacterias.
It plays a very crucial role in identifying and differentiating gram-negative enteric bacteria particularly belonging to Enterobacteriaceae family and Pseudomonas species.
Principle of MacConkey Agar
MacConkey agar is developed for the isolation of gram-negative enteric bacteria and differentiation of lactose fermenting from non-lactose fermenting gram-negative bacterias.
Lactose: Lactose acts as the fermentable carbohydrate source, lactose allows differentiation between gram-negative bacterias. Bacteria that can ferment produce acid (lactic acid, acetic acid, succinic acid, formic acid) as a byproduct which can be easily visualized in the medium. Depending on species of bacterias and specific fermentation pathway, bacterias might produce different organic acids.
Bile Salts & Crystal Violet: Bile salts and crystal violet selectively inhibit the growth of gram-positive bacterias by disrupting cell membranes, gram-negative bacteria's cell membrane blocks these chemicals, allowing gram-negative bacteria to grow. This is very crucial in clinical analysis, where gram-positive bacterias might overgrow and shadow the presence of gram-negative bacterial growth.
Neutral red: As we discussed earlier lactose fermenting bacterias produce organic acid, this drops the medium’s pH below 6.8. The neutral red in the medium absorbs hydrogen ions released by organic acid and undergoes structural change. This causes the indicator to turn bright red or pink, staining lactose fermenting bacterias, where non-lactose fermenting bacterias stays clear.
Peptones & sodium chloride: These components provide essential nutrients for bacterial growth, including amino acids and salts. This supports the fast growth of wide range gram-negative bacterias.
Preparation of MacConkey Agar
1. Dissolve the recommended quantity of MacConkey agar medium in purified water (quantity varies based on the manufacturer).
2. Heat the container with medium gently with frequent agitation until it is fully dissolved, make sure it doesn't have any clumps which might interfere with uniform growth.
3. Autoclave the prepared medium at 121°C for 15 minutes to eliminate any potential contaminants.
4. Allow the medium to cool to 45 - 50°C and pour the medium into sterile petri dishes.
5. After complete solidification of the petri dishes, label the media plates with their name and preparation date.
6. Use a sterile loop or swap to streak out the sample onto the surface of the plate.
7. Incubate the plates aerobically at 35 - 37°C for the primary result and read it again at 48 hours.
Results on MacConkey Agar
Lactose fermenting bacterias produce acid as a by-product, lowering the pH and turning colonies pink or red. Very strong fermenting bacterias such as E.coli may show pink halo around the colonies due to precipitation of bile salt.
Lactose non-fermentering bacteria cannot ferment lactose. These bacterial colonies remain colorless or appear pale. The medium surrounding the colonies stays relatively transparent.
Applications of MacConkey Agar
MacConkey agar is used for the selective isolation of gram-negative bacterias, especially coliforms and enteric pathogens.
It is used in the differentiation of lactose fermenting bacterias from non-fermenting gram-negative bacterias.
In the microbiological examination MacConkey agar is widely used in clinical, food, water analysis to detect coliform count.
Limitations of MacConkey Agar
MacConkey agar can be only used for preliminary or probable identification only. It is necessary to subculture and carry out several other tests for final confirmation.
MacConkey agar is designed to support non-fastidious gram-negative rod bacterias; certain bacterias might be sensitive to the agents and exhibit poor growth.
Incubating MacConkey agar under increased CO2 forms carbonic acid which lowers the medium’s pH. This high acidic condition increases the toxicity of bile salts and crystal violet suppressing the growth of gram-negative bacterias.
