MUG (beta-Glucuronidase) test for rapid identification of E. coli

MUG is an acronym for 4-methylumbelliferyl-beta-D-glucuronide. Escherichia coli produces the enzyme beta-D-glucuronidase. Beta-D-glucuronidase is a enzyme which hydrolyzes the beta-D-glucopyranosid-uronic derivatives to aglycons and D-glucuronic acid.

E. coli use MUG as a substrate, releasing a fluorogenic compound
E. coli use MUG as a substrate, releasing a fluorogenic compound

The substrate 4-methylumbelliferyl-beta-D-glucuronide is impregnated in the disc and is hydrolyzed by the enzyme (beta-D-glucuronidase) to yield the 4-methylumbelliferyl moiety, which fluoresces blue under long wavelength ultraviolet light.

Significance of MUG Test: 

U.S. Environmental Protection Agency has approved 4-methylumbelliferyl-β-D-glucuronide Escherichia coli broth medium (EC-MUG) is an effective and rapid method for detection and verification of E. coli in food, water, and environmental samples.

Standard analysis of water includes the most probable number (MPN) for the presumptive and quantitative detection of coliform and fecal coliform bacteria  in water samples. According to ASM EC broth and agar media with MUG is best suited for confirmatory testing of the presence of E. coli after a presumptive positive result for fecal coliform bacteria.

Procedure for MUG Test (b-Glucuronidase test)

  1. Prepare a dense milky suspension of the organism to be tested in a small tube containing 0.25 ml of saline. The suspension should be prepared from colonies growing on MacConkey agar.
  2. Add one tablet of 4-nitrophenyl-β-D-glucopyranosiduronic acid (PGUA) for detection of β-glucuronidase activity.
  3. Place a stopper in the tube and agitate vigorously for a few seconds.
  4. Incubate the tube at 35-37 °C for 4 hours.
  5. The development of a yellow colour in the supernatant indicates a positive test for Ecoli.

Expected results of MUG Test

  1. Positive: Electric blue fluorescence
  2. Negative: Lack of fluorescence

Quality control of MUG Test

  1. Positive: Escherichia coli
  2. Negative: Pseudomonas aeruginosa

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