UREASE TEST Urea is a nitrogen containing compound that
is produced by the decarboxylation of the amino acid arginine during the urea cycle.
Some bacteria produce the enzyme urease as part of its metabolism to break down urea
to ammonia and carbon dioxide. The purpose of Urease test is to determine the ability
of microorganisms to degrade urea by means of the enzyme urease. The presence of urease is detectable when
the organisms are grown in a medium containing urea and the pH indicator Phenol red. Phenol
red turns yellow in an acidic environment and deep pink in an alkaline environment.
If the urea in the medium is degraded and ammonia is produced, an alkaline environment
is created, and the media turns pink. This is a positive reaction for the presence of
urease. Failure of deep pink color to develop is evidence of a negative reaction.
Rapid urease test, also known as the CLO test (Campylobacter-like organism test), is a rapid
test for diagnosis of Helicobacter pylori. The basis of the test is the ability of Helicobacter
pylori to secrete the urease enzyme, which catalyzes the conversion of urea to ammonia
and bicarbonate. 24-48 hours tryptic soy broth cultures
Urea slant Bunsen burner
Inoculating loop Test tubes
PROCEDURE Arrange the materials required for the test
on LAF. Sterilize the inoculating loop in the blue
flame of the bunsen burner till red hot and then allowed to cool.
Take the Tryptic soy broth tube from the rack containing 24hour culture; Gently tap it,
remove the cap and flame the neck of the tube Using aseptic technique, take a loopful of
culture from the TSB (tryptic soy broth) tube with the needle.
Again flame the neck of the tube ,cap it back and replace the tube in the test tube rack.
Take a sterile urease slant tube from the rack, remove the cap and flame the neck of
the tube. Insert the inoculation loop and streak the
slanted surface of the tube in a zigzag manner Again flame the neck of the urease tube, cap
it back and place it in the test tube rack. Incubate at 37oc for 18 to 24 hours.
Tighten the cap and incubate at 37°C for 24-48 hours
After 24 hours, take out the slants from the incubator and observe the results
RESULTS OBSERVED: Positive reaction will yield deep pink coloration
in the media. Negative results will yield yellow coloration
in the media.