MacConkey agar media was first introduced by a scientist named Alfred Theodore MacConkey in the year 1890s. It is primarily exercised for the isolation and differentiation of the non-fastidious, gram-negative bacilli belonging to the family Enterobacteriaceae. Previously, it was first considered as “Solid differential media”. But now, it is practised as both liquid broth and solid culture media that can function as a selective as well as differential media.
MacConkey agar media comprises an indicator system that includes bile salt, lactose and neutral red that helps us to study and identify the growth pattern of the bacterial isolate based on bile salt precipitation, lactose fermentation and formation of acid.
Content: MacConkey Agar Media
Definition of MacConkey Agar Medium
MacConkey agar media can define as the indicator, selective and differential media, which determines the growth or presence of the gram-negative enteric bacilli from the different samples that are important for the sanitary purpose. It can function as a selective media as it ensures the growth of gram-negative enteric bacilli by restricting the growth of other organisms.
MacConkey agar media can also work as a differential media by which one can easily distinguish between the closely related gram-negative bacteria depending upon the lactose fermentation and acid production. It acts as an indicator media that indicate the presence or absence of lactose fermenters by a definite colour change in the developing colonies as well as the media.
- In dehydrated form: The media appears beige-pink and homogenous.
- In prepared form: The media appears slightly lustrous and pinkish-red in colour.
The ingredients of the MacConkey agar media in 1L of distilled water, are as follows:
- Peptone: 7g
- Proteose peptone: 3g
- Lactose: 10g
- Bile salts: 1.5g
- Sodium chloride: 5g
- Neutral red: 0.03g
- Crystal violet: 0.001g
- Agar: 13.5g
Role of media components
The proteoses and peptones provide vital elements and the sole N-source for the growth of gram-negative rods. Lactose is a fermentable carbohydrate that provides a sole C-source to promote bacterial growth. Bile salts function as a selective agent that particularly suppress the gram-positive organisms to grow. Gram-negative enteric bacteria are resistant to bile-salts by having a bile-resistant outer membrane.
Sodium chloride functions to supply essential electrolytes for the cell growth of gram-negative organisms and also maintains the osmotic concentration. Neutral red is a pH indicator dye, which can help us to discriminate the organism based on the colour change.
When the bacteria produce a sufficient amount of acid, the pH of the media falls, resulting in the colour change of the neutral red pH indicator into pink. Crystal violet is also a selective agent that mainly restricts the growth of gram-positive bacteria. Agar acts as a solidifying agent of the media.
Preparation of the Media
Weigh all the contents accurately and dissolve it by adding 1L of sterile water into a sterilized conical flask. Then put a cotton plug into the mouth of the conical flask to avoid any contamination. Then mix the contents thoroughly over the hot plate by shaking the flask in a clockwise and anticlockwise direction.
After getting a homogenous solution, place the flask in an autoclave for 30 minutes at 121 degrees Celsius, under 15lps pressure. Then, take out the flask from the autoclave directly to the laminar airflow chamber for the plating and the remaining media can be kept in a refrigerator for further use.
The MacConkey agar media principally helps in the isolation and differentiation of the gram-negative bacterial isolates. The property of differentiating bacteria depends upon the capability of the organism to ferment lactose. The lactose fermenting organisms will produce a certain amount of acid as a result of fermentation, and dropdowns the pH of the media.
As the pH drops below 6.8, the neutral red indicator will turn into pink that will be absorbed by the test organism. Strong lactose fermenters will utilize lactose to form enough acids, resulting in bile precipitation around the bacterial growth. The precipitation of bile salt gives a hazy appearance around the growth and appears as a pink halo. In contrast, the weaker lactose fermenting bacteria will not form a pink halo, and the non-lactose fermenting bacteria will look colourless.
The result can be indicated by the growth and colour of the bacteria in the culture media.
Strong lactose fermenters: Its presence is indicated by the development of pink colonies encircled by a pink halo (zone of precipitated bile-salt). Strong lactose fermentors include Klebsiella species, Escherichia species etc.
Weak lactose fermenters: Its presence can be visualized by the growth of pink colonies after 48 hours of the incubation period. These bacteria do not form a zone of precipitated bile. Weak lactose fermenters include Citrobacter species, Serratia species etc.
Non-lactose fermentors: Its presence is indicated by the growth of colourless colonies over the transparent MacConkey agar surface. Non-lactose fermentors include Salmonella species, Proteus species etc.
Modifications in the MacConkey Agar Media
MacConkey media without CV: The MacConkey agar media without crystal violet is less selective as it allows the development of Staphylococcus and Enterococcus species. It also helps in determining the Mycobacterium fortuitum and M. chelonae from other fast-growing mycobacteria.
Controlled swarming MacConkey agar media: It lacks either crystal violet or salt in order to control the flocking of Proteus species.
Sorbitol MacConkey agar media: It is an alternative of MacConkey agar media that uses sorbitol as a sole carbohydrate source in place of lactose.
MacConkey agar works as a selective medium that isolates the gram-negative microorganisms from different sources like urine, stool, food samples etc. It also works like a differential media that discriminates between the closely related gram-negative enteric bacilli.
MacConkey agar medium has a drawback that it gives presumptive test results of the isolated organism. For the confirmatory results, the test organism needs further subculturing and biochemical tests. There is another limitation where some strains of the Proteus may flock on this medium.