MacConkey Broth: Principle, Composition, Preparation, and Uses in Microbiology

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MacConkey Broth: Principle, Composition, Preparation, and Uses in Microbiology

MacConkey Broth is a selective and differential medium used for the detection and differentiation of coliform organisms (especially Escherichia coli) from water, food, dairy, and clinical samples. It is an essential medium in microbiological testing to identify lactose-fermenting and non-lactose-fermenting Gram-negative bacteria.

📘 Introduction

The MacConkey medium was originally developed by Alfred Theodore MacConkey in the early 20th century. It exists in both solid (agar) and liquid (broth) forms. MacConkey Broth serves as a selective enrichment medium that supports the growth of enteric Gram-negative bacteria while inhibiting Gram-positive organisms through bile salts and crystal violet.

🧪 Principle of MacConkey Broth

MacConkey Broth works on two key principles — selectivity and differentiation:

  • Selectivity: Bile salts and crystal violet inhibit the growth of Gram-positive bacteria, allowing only Gram-negative enteric bacilli to grow.
  • Differentiation: The presence of lactose and the pH indicator neutral red helps differentiate bacteria based on lactose fermentation ability.

Lactose-fermenting bacteria (e.g., E. coli) produce acid, lowering the pH and changing the color of the medium to yellow or pink, whereas non-lactose fermenters (e.g., Salmonella, Shigella) do not cause a color change.

⚗️ Composition of MacConkey Broth (per liter)

Ingredients Quantity (g/L) Function
Peptone 20.0 Provides nitrogen, vitamins, and amino acids for bacterial growth
Lactose Monohydrate 10.0 Fermentable carbohydrate for differentiation
Bile Salts 5.0 Inhibits Gram-positive bacteria
Sodium Chloride (NaCl) 5.0 Maintains osmotic balance
Neutral Red 0.01 pH indicator (turns red below pH 6.8)
Final pH (at 25°C) 7.4 ± 0.2

⚙️ Preparation of MacConkey Broth

  1. Weigh the required amount of dehydrated medium as per the manufacturer’s instructions.
  2. Dissolve it in 1000 ml of distilled water with gentle heating and stirring.
  3. Ensure complete dissolution and adjust the pH to 7.4 ± 0.2 if necessary.
  4. Dispense the medium into test tubes or bottles with Durham tubes inserted upside down for gas collection.
  5. Sterilize by autoclaving at 121°C for 15 minutes.
  6. Cool the medium to room temperature before use.

🧫 Inoculation and Incubation

  • Inoculate the sample (e.g., water, food, or environmental sample) into the MacConkey Broth tube.
  • Incubate at 35–37°C for 24–48 hours.
  • Observe for acid and gas production in the Durham tube.

🔍 Interpretation of Results

Observation Interpretation
Yellow color with gas in Durham tube Positive for lactose fermentation (Coliform present)
Yellow color without gas Partial lactose fermentation
No color change (remains red or purple) Negative for lactose fermentation (No coliforms)

🧍‍♂️ Quality Control Strains

  • Escherichia coli ATCC 25922 – Positive (acid and gas formation)
  • Enterobacter aerogenes ATCC 13048 – Positive (acid and gas formation)
  • Salmonella typhimurium ATCC 14028 – Negative (no acid or gas)

💡 Applications of MacConkey Broth

  • Used in Most Probable Number (MPN) tests for coliform detection in water analysis.
  • Applied in food and dairy microbiology for hygiene monitoring.
  • Used in pharmaceutical microbiology for microbial limit testing.
  • Helps differentiate lactose-fermenting enteric bacilli from non-fermenters.
  • Serves as an enrichment medium for further plating on MacConkey Agar or EMB Agar.

⚖️ Advantages

  • Simple and reliable medium for detecting coliform bacteria.
  • Provides both qualitative and semi-quantitative results.
  • Supports further biochemical identification and confirmation steps.

⚠️ Limitations

  • Some non-coliform bacteria may ferment lactose and give false-positive results.
  • Further biochemical or serological tests are required for definitive identification.
  • Over-incubation may alter color reactions due to peptone utilization.

📚 References

  • MacConkey, A.T. (1905). “Lactose-fermenting bacteria in feces.” Journal of Hygiene.
  • ISO 4832: Microbiology of Food and Animal Feeding Stuffs – Enumeration of Coliforms.
  • USP & IP Microbiology Guidelines.
  • HiMedia and Oxoid Product Manuals for MacConkey Broth.

💬 About the Author

Siva Sankar is a Pharmaceutical Microbiology Consultant and Auditor with extensive experience in sterility testing, validation, and GMP compliance. He provides consultancy, training, and documentation services for pharmaceutical microbiology and cleanroom practices.

📧 Contact: siva17092@gmail.com
Mobile: 09505626106

📱 Disclaimer: This article is for educational purposes and does not replace your laboratory’s SOPs or regulatory guidance. Always follow validated methods and manufacturer instructions.

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