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The presence of Crystal violet and bile salts incorporated in MacConkey agar help to inhibit the growth of Gram-positive bacteria making it a selective media and fastidious Gram-negative bacteria, such as Neisseria and Pasteurella. Gram-negative enteric bacteria can tolerate bile salts because of their bile-resistant outer membrane.
MacConkey agar is selective for Gram negative organisms, and differentiate lactose fermenting gram negative rods from non lactose fermenting gram negative rods. It is primarily used for detection and isolation of bacteries members of family enterobacteriaceae and Pseudomonas spp.
MacConkey agar is used for the selective isolation and identification of Enterobacteriaceae from samples such as feces, urine, wastewater and foods.
Principle of selective and differential ability of MacConkey agar
Those Gram-negative enteric bacteria that grow on MacConkey agar are differentiated by their ability to ferment lactose (Sugar). Fermentation of lactose by bacteria leads to the production of the acid that drops the pH of the media. The drop in pH is indicated by the change of neutral red indictor to pink (Neutral read appears pink at pH’s below 6.8).
Strongly lactose fermenting bacteria produce sufficient acid which causes precipitation of the bile salts around the growth. It appears as a pink halo surrounding individual colonies or areas of confluent growth. Pink halo in not seen around the colonies of weaker lactose fermenting bacteria.
As for the selective ability of MacConkey agr,Gram-negative bacteria that grow on MacConkey agar but do not ferment lactose appear colorless on the medium and the agar surrounding the bacteria remains relatively transparent.
Composition of MacConeky Agar and the functions:
- Enzymatic digest of Gelatin, Casein and Animal tissue: provides nitrogen, vitamins, minerals and amino acids essential for growth.
- Lactose: fermentable carbohydrate providing carbon and energy.
- Bile Salts: selective agents and inhibit Gram positive organisms.
- Crystal Violet: Gram positive bacteria are generally inhibited by crystal violet.
- Sodium Chloride: supplies essential electrolytes for transport and osmotic balance.
- Neutral Red: pH indicator. which is red in color at pH’s below 6.8.
- As a resulut of lactose fermentation, the pH of the medium decreases, changing the color of neutral red to pink
- Agar : Solidifying agent
How to Prepare MacConkey Agar
- According to manufacturer instruction,Suspend the measured amount of powder (See in the agar bottle and generally 50 gram) in 1 L of distilled water and mix thoroughly.
- Heat while constantly agitating and boil for 1 minute to completely dissolve the powder.
- Autoclave at 121°C for 15 minutes.
Colonies appearance on MacConkey Agar
- Lactose-fermenting organisms appears pink to brick red colonies with or without a zone of precipitated bile.
- Non-lactose fermenting organisms appears colorless or clear colonies
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| Identification of lactose fermenters and non lactose fermenters MacConkey Agar |
Lactose Fermenter Organisms as seen on MacConkey Agar:
- Citrobacter spp.: colonies are light pink after 48 hours.
- Klebsiella spp.: Mucoid lactose fermenter colonies
- Escherichia coli: Lactose fermenter; flat, dry, pink colonies with a surrounding darker pink area of precipitated bile salts.
- Serratia spp.: Late lactose fermenter; S. marcescens may be red pigmented, especially if the plate is left at 25°C
Non Lactose Fermenter Organisms as seen on MacConkey Agar
- Proteus spp.: may swarm depending on the amount of agar in the medium; characterize by a foul smell
- Shigella spp.: Shigella sonnei produces flat colonies with jagged edges.
- Yersinia spp.: may be colorless to peach.
- Salmonella spp.: Not a lactose fermenting
- Other organisms showing colorless colonies on MacConkey agar are; Edwardsiella spp, Hafnia spp., Morganella spp., Providencia spp.
As for gram positive bacteria like Staphylococcus aureus,no growth is observed
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