| Red halo around H. pylori colony in MIU medium due to alkalinization |
Urease test for Mycobacteria
DESCRIPTION
The method used for determination of urease activity is a modification of one used by Toda and associates, which is based on
pH change associated with the large amount of ammonia released by urease-positive organisms.
PROCEDURE
Difco-Bacto Urea Agar Base (containing no agar) is diluted 1:10 in sterile water and dispensed in 3-ml amounts to sterile 13- by
100-mm screwcapped tubes. A generous loop or spadeful of growth from a fresh Lowenstein-Jensen culture is suspended in the
substrate solution and incubated at 36 to 37° C for 3 days.
Reaction is positive (urease activity) if a pink color appears within 3 days.
DESCRIPTION
The method used for determination of urease activity is a modification of one used by Toda and associates, which is based on
pH change associated with the large amount of ammonia released by urease-positive organisms.
PROCEDURE
Difco-Bacto Urea Agar Base (containing no agar) is diluted 1:10 in sterile water and dispensed in 3-ml amounts to sterile 13- by
100-mm screwcapped tubes. A generous loop or spadeful of growth from a fresh Lowenstein-Jensen culture is suspended in the
substrate solution and incubated at 36 to 37° C for 3 days.
Reaction is positive (urease activity) if a pink color appears within 3 days.
| Urea Hydrolysis Test (Urease Activity) |
DESCRIPTION
Urease is produced by numerous taxonomically diverse bacterial species, including normal flora and nonpathogens. Also, urease
has been demonstrated as a potent virulence factor for some species, including Proteus mirabilis, Helicobacter pylori.
Testing the ability to produce urease, an enzyme that hydrolyse urea providing a source of nitrogen for bacteria.
Urease is produced by numerous taxonomically diverse bacterial species, including normal flora and nonpathogens. Also, urease
has been demonstrated as a potent virulence factor for some species, including Proteus mirabilis, Helicobacter pylori.
Testing the ability to produce urease, an enzyme that hydrolyse urea providing a source of nitrogen for bacteria.
Urease test for Helicobacter
DESCRIPTION
Helicobacter pylori produces a urease that catalyses the hydrolysis of urea to yield
ammonia and carbonic acid. Urease aids in colonization of the host by neutralizing
gastric acid and providing ammonia for bacterial protein synthesis.
H. pylori produces such a high amount of urease that it can be detected quickly using
the MIU test without the need for incubation. The positive reaction can appear in 2-3
minutes.
PROCEDURE
Pick a well isolated colony and inoculate medium. Wait a few minutes until a red halo
appears around the colony.
DESCRIPTION
Helicobacter pylori produces a urease that catalyses the hydrolysis of urea to yield
ammonia and carbonic acid. Urease aids in colonization of the host by neutralizing
gastric acid and providing ammonia for bacterial protein synthesis.
H. pylori produces such a high amount of urease that it can be detected quickly using
the MIU test without the need for incubation. The positive reaction can appear in 2-3
minutes.
PROCEDURE
Pick a well isolated colony and inoculate medium. Wait a few minutes until a red halo
appears around the colony.
REFERENCES:
1. Helgomar Raducanescu, Valeria Bica-Popii,1986. Bacteriologie veterinara, Ed. Ceres, Bucuresti.
2. Murray, P.R., Baron, E. J., Jorgensen, J.J., Pfaller, M.A., and Yolken, R.H. Manual of Clinical Microbiology, 8th ed. ASM Press:
Washington, DC, 2003.
3. Lawrence G. Wayne 1973. Simple Pyrazinamidase and Urease Tests for Routine Identification of Mycobacteria. American Review of
Respiratory Disease, Volume 109, Issue 1, 147-151.
4. Mobley, H. (1996), The role of Helicobacter pylori urease in the pathogenesis of gastritis and peptic ulceration.. Aliment Pharmacol
Ther, 10: 57-64. https://doi.org/10.1046/j.1365-2036.1996.22164006.x
1. Helgomar Raducanescu, Valeria Bica-Popii,1986. Bacteriologie veterinara, Ed. Ceres, Bucuresti.
2. Murray, P.R., Baron, E. J., Jorgensen, J.J., Pfaller, M.A., and Yolken, R.H. Manual of Clinical Microbiology, 8th ed. ASM Press:
Washington, DC, 2003.
3. Lawrence G. Wayne 1973. Simple Pyrazinamidase and Urease Tests for Routine Identification of Mycobacteria. American Review of
Respiratory Disease, Volume 109, Issue 1, 147-151.
4. Mobley, H. (1996), The role of Helicobacter pylori urease in the pathogenesis of gastritis and peptic ulceration.. Aliment Pharmacol
Ther, 10: 57-64. https://doi.org/10.1046/j.1365-2036.1996.22164006.x
| Christensen's urea agar positive (left) and negative (right) |
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(c) Costin Stoica
MEDIUM PREPARATION
Composition of Christensen's urea agar medium:
Glucose ................................................................................1g
Peptone ................................................................................1g
KH2PO4 ...............................................................................2g
NaCl ......................................................................................5g
Phenol red ...................................................................0.012g
Agar ....................................................................................20g
H2O ad 1000 ml, pH = 6.8
To base medium add 20 ml urea solution 20% before solidification. Initial color of the
medium must be yellow.
Composition of Christensen's urea agar medium:
Glucose ................................................................................1g
Peptone ................................................................................1g
KH2PO4 ...............................................................................2g
NaCl ......................................................................................5g
Phenol red ...................................................................0.012g
Agar ....................................................................................20g
H2O ad 1000 ml, pH = 6.8
To base medium add 20 ml urea solution 20% before solidification. Initial color of the
medium must be yellow.
Urease test for Enterobacteria
DESCRIPTION
MIU medium may also be used for urease testing. Test checks simultaneously for the
motility, production of indole and urea hydrolysis. This medium, together with TSI and
Simmons agar, is most used for Enterobacteriaceae identification, especially
Escherichia and Salmonella.
PROCEDURE
Pick a well isolated colony and inoculate medium. Incubate 24 hours at 37 °C.
Reaction is positive if the medium become red (urea is decomposed resulting
ammonia which changes the pH).
NOTES
False positives may result due to protein hydrolysis when incubation is prolonged.
DESCRIPTION
MIU medium may also be used for urease testing. Test checks simultaneously for the
motility, production of indole and urea hydrolysis. This medium, together with TSI and
Simmons agar, is most used for Enterobacteriaceae identification, especially
Escherichia and Salmonella.
PROCEDURE
Pick a well isolated colony and inoculate medium. Incubate 24 hours at 37 °C.
Reaction is positive if the medium become red (urea is decomposed resulting
ammonia which changes the pH).
NOTES
False positives may result due to protein hydrolysis when incubation is prolonged.
| R E G N U M PROKARYOTAE |
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| Urea-positive result in the middle tube: MIU medium turning red (Proteus mirabilis) |
