Table of Contents
In this Macconkey agar media composition and uses post we have briefly explained about Macconkey agar media principle, composition, appearances, storage, preparation, result, applications, and limitations of Macconkey agar media.
MacConkey Agar Media
MacConkey agar media was created by Alfred Theodore MacConkey, M.D. at the turn of the century. MacConkey agar media contains crystal violet and bile salts, which inhibit the growth of gram-positive bacteria. MacConkey agar media also inhibits the growth of mould. Gram-negative bacteria have a bile-resistant outer membrane; therefore they may be able to thrive in this environment.
Lactose non-fermenting Gram-negative colonies can grow on MacConkey agar media, although they appear as Colourless colonies. Gram-negative lactose fermenting bacteria include E.coli, Klebsiella pneumoniae, and Enterobacter cloacae. Gram-negative lactose non-fermenters include Shigella, Salmonella, Proteus, Serratia, and Pseudomas aeruginosa. Before checking for growth on MacConkey agar media, MacConkey agar media should be incubated at 35°C for 24 to 48 hours.
Principle
MacConkey agar media is used to isolate gram-negative enteric bacteria and distinguish lactose fermenting gram-negative bacteria from lactose non-fermenting gram-negative bacteria. The basic minerals, vitamins, and nitrogenous elements required for microorganism growth are provided by pancreatic digest of gelatin and peptones (meat and casein).
Lactose monohydrate is a carbohydrate that can be fermented. To suppress the growth of gram-positive bacteria, MacConkey agar media adds crystal violet and bile salts. Mold is also inhibited by this medium. Gram-negative bacteria have an outer membrane that is relatively resistant to bile. Lactose (Lac+) can be fermented by some gram-negative bacteria, but not by others (Lac-).
This is when MacConkey’s differential aspect comes into play. The pH of the media drops when Gram-negative bacteria that can ferment lactose are present, and this shift is recognized by another component, neutral red. Red that has a pH of less than 6.8 is known as neutral red. This dye is absorbed by bacteria at pH levels below 6.8, resulting in vivid pink to red colonies on the agar.
Composition
Typical Formula | (g/L) |
Pancreatic Digest of Gelatin | 17.0 |
Peptone from Meat | 1.5 |
Peptone from Casein | 1.5 |
Lactose | 10.0 |
Sodium Chloride | 5.0 |
Bile Salts | 1.5 |
Agar | 15.0 |
Neutral Red | 0.03 |
Crystal Violet | 0.001 |
Final pH 7.1 ± 0.2 at 25°C
Appearances, Storage
Dehydrated medium: free-flowing, homogeneous, beige-pink. Prepared medium: slightly opalescent, pinkish-red. Because the powder is quite hygroscopic, store it at 10-30°C in a dry atmosphere, tightly closed in its original container. Keep bottles and prepared plates away from light at 10-25°C. Use the product before the expiration date on the label, or if there is any evidence of contamination or degradation.
Preparation
Dehydrated medium
In 1 litre of distilled or deionized water, dissolve 51.5 g of the powder. Mix thoroughly. Bring to a boil for 1 minute, stirring constantly until the sugar is completely dissolved. Sterilize for 15 minutes in an autoclave at 121°C.
Medium in bottles
Melt the contents of the bottle in a water bath at 100°C until entirely dissolved (leaving the cap partially off). Then screw on the cap and turn the bottle upside down to check for homogeneity of the dissolved medium. Cool to 45-50°C, thoroughly mix to avoid foam formation, then distribute aseptically into Petri dishes.
Test Procedure
Inoculate the plates by strewing the specimen directly onto the agar surface or spreading a sample from an enrichment culture. Incubate aerobically for 18-24 hours at 35 2°C.
Test Interpretation
Lactose-fermenting
Lactose-fermenting strains produce red/pink growth that is surrounded by acid-precipitated bile. Acid generation due to lactose, neutral red absorption, and dye colour change if the pH level falls below 6.8 are all variables that contribute to the change in colour.
Non-Lactose-fermenting
Lactose strains that do not ferment the end result is colourless and transparent. The medium’s look has not changed significantly.
MacConkey agar media Composition and Uses: MacConkey agar media is a modern, commercially available agar. Lactose fermenter Escherichia coli is shown in Panel A. The colonies are surrounded by an opaque pink bile precipitation. Klebsiella pneumoniae, another lactose fermenter, is shown in panel B. The colonies are pink, indicating acid production, but there is no bile precipitation. Pseudomonas aeruginosa, a lactose non-fermenter, is shown in Panel C.
Bacterium | Gram – / + | Lactose Fermentation |
Bacillus subtilis | + | Not Determined |
Escherichia coli | – | yes |
Pseudomonas fluorescens | – | no |
Limitations
1. The isolated organisms only provide a speculative result. The ideal method is to execute a cultural operation and confirm the results using a confirmation test. Some bacterium strains grow slowly in the medium, while others fail to thrive at all.
2. The incubation of a MacConkey agar media plate exposes it to an increase in carbon dioxide, which can have a substantial impact on the growth/reduction of Gram-negative bacilli strains.
Applications
1. To suppress the growth of gram-positive bacteria, MacConkey agar media adds crystal violet and bile salts.
2. MacConkey agar media is used to distinguish lactose fermenting gram-negative bacteria from lactose non-fermenting bacteria.
3. Coliforms and intestinal pathogens are isolated from water, dairy products, and biological materials using this method.