Can E. coli grow on MacConkey Agar?
For microorganisms to be grown in a laboratory, they have to be cultured in a man-controlled medium (plural: media), similar to the way animals are kept in a zoo. Such media are called growth media.
They are also called culture media or nutrient broth.
Culture media contain nutrients required for the growth of microorganisms such as bacteria and fungi (microbiological culture) or for specific cell types derived from plants or animals (cell culture).
When the microbiological culture is being used to grow a specific bacterium, the culture or medium is called a bacteria growth medium.
MacConkey agar is an example of a bacteria growth media.
What is MacConkey Agar (MAC)?
MacConkey agar (MAC) is a specialized bacterial growth medium that is selective for Gram-negative bacteria and can also differentiate for Gram-negative bacteria that are capable of fermenting lactose.
It inhibits the growth of Gram-positive bacteria due to the presence of crystal violet and bile salts.
The selective and differentiating properties of MacConkey agar makes it very useful for both research and clinical purposes. Clinical microbiologists find MAC very useful in diagnostic testing as part of their many important bacterial cultures.
MacConkey agar is still widely used in clinical laboratories to identify causal agents from a patient (i.e. stool sample).
MacConkey agar is named after Alfred Theodore MacConkey; the British bacteriologist who first developed the medium in the late 1890s.
Dr. MacConkey first developed a bile salt medium containing glycocholate, lactose, and litmus, to be incubated at 22°C. This formula was then altered by the replacement of glycocholate with taurocholate and the incubation temperature was raised to 42°C.
MacConkey was to later change the recipe again by substituting neutral red for litmus following a suggestion that neutral red is used as an indicator in bile salt lactose medium. The final media formulation was originally designed to support the growth of Shigella and is the one that is still most commonly used today
Components of MacConkey Agar.
MacConkey agar (MAC) has the following components:
- Peptone – 17 g
- Proteose peptone – 3 g
- Lactose – 10 g
- Bile salts – 1.5 g
- Sodium chloride – 5 g
- Neutral red – 0.03 g
- Crystal violet – 0.001 g
- Agar – 13.5 g
- Distilled Water – make it to 1 liter
- pH – adjust pH to 7.1 +/− 0.2.
Agar should be boiled to dissolve and sterilized at 121° C for 15 minutes.
There are many variations of MacConkey agar depending on the need. If the spreading or swarming of Proteus species is not required, sodium chloride is omitted. Crystal violet at a concentration of 0.0001% (0.001 g per liter) is included when needing to check if Gram-positive bacteria are inhibited.
MacConkey with sorbitol can also be used to isolate the enteric pathogen, E. coli O157.
The four key ingredients of MacConkey agar are:
- Lactose; lactose acts as a source of carbohydrates. Lactose-fermenting bacteria produce pink-red colonies, after fermenting the lactose to acids and dropping the pH of the indicator (neutral red) present in the medium. Since non-fermenters can’t utilize lactose, colonies appear colorless or transparent.
- bile salts; bile salts and crystal violet act as selective agents that inhibit the growth of Gram-positive organisms, and aid in the selective growth of non-fastidious gram-negative bacteria
- crystal violet; alongside the bile salts, crystal violet acts as a selective agent that inhibits the growth of Gram-positive organisms, and aid in the selective growth of non-fastidious gram-negative bacteria.
- neutral red is a pH indicator that turns red at a pH below 6.8 and is colorless at any pH greater than 6.8. Agar is the solidifying agent. This quality is vital to what makes MAC a selective and differential media.
Other ingredients such as enzymatic digestion of gelatin, casein, and animal tissue provide nitrogen, vitamins, minerals, and amino acids that are essential for growth. Sodium chloride provides osmotic balance and supplies essential electrolytes for transport. Agar is also incorporated as the solidifying agent.
Uses of MacConkey Agar Medium
- MacConkey agar is commonly used for the isolation of Gram-negative enteric bacteria.
- MacConkey agar is a commonly used media to differentiate members of Enterobacteriaceae. It differentiates between lactose-fermenting and non-fermenting gram-negative rods by the color of colonial growth.
- MacConkey agar is used for the selective isolation and identification of members of the family Enterobacteriaceae from feces, urine, wastewater, and foods.
E. coli Growth on MacConkey Agar Medium
Gram-negative enteric bacteria are a common cause of bacterial gastroenteritis, which is characterized by diarrhea, vomiting, and abdominal cramping. Escherichia coli is a common cause of bacterial gastroenteritis.
When bacterial gastroenteritis is suspected, the patient’s specimen can be sampled and cultured on a panel of bacterial cultures, which includes MacConkey agar medium.
MAC contributes to the identification of the causal agent by providing lactose-fermentation profiles in gram-negative species.
As a gram-negative bacterium, E. coli grows on MAC, and will by fermenting lactose, lower the pH and change the color from pink to dark pink, which is very useful in its identification. E. coli are, identified on the basis of color change in a MacConkey agar medium.
Like some other colonies that grow on MacConkey medium, besides differentiating on the basis of color, E.coli can further be presumptively identified based on their colonial appearances (shape, size, margin, time of growth, etc.).
The lactose fermenter, E.coli, gives flat, dry, pink, non-mucoid colonies with a surrounding darker pink area of precipitated bile salts.
Application of the Principle of Selectivity and Differentiation of MAC on E.coli
Selectivity: Crystal violet dye and bile salts stop the growth of gram-positive bacteria. This only allows gram-negative species, which have a relatively bile-resistant outer membrane, to form colonies on MacConkey agar.
Differentiation: Gram-negative bacteria that grow on MacConkey plates are differentiated by their ability to ferment lactose.
Bacteria that ferment lactose, like E. coli, decrease the pH of the medium, this decrease in pH is detected by neutral red (turns red if pH below 6.8).
When the pH drops, the neutral red is absorbed by the bacteria, which appears as bright pink to red colonies on the agar.
Gram-negative bacteria that thrive on MacConkey agar but do not ferment lactose appear colorless on the medium, and the agar surrounding the bacteria remains relatively transparent.
Such is the case with Salmonella, Proteus, Yersinia, and Pseudomonas.