UREA R BROTH

Cat. no. Y141 Urea R Broth, 13x100mm Tube, 1ml 20 tubes/box

INTENDED USE

Hardy Diagnostics Urea R Broth is a highly sensitive medium used for the rapid detection of urease activity by microorganisms.

SUMMARY

The principle of the urease test is to determine whether an organism has the ability to hydrolyze urea into two molecules of ammonia by the enzyme urease (urea amidohydrolase) which results in increased alkalinity of the medium. This increase in alkalinity is detected by the use of phenol red as the pH indicator, changing from a orange-red to a bright pink. (1)

Urea R Broth is designed to be both rapid and sensitive for the detection of urease. Urea R Broth utilizes weak pH buffers monopotassium phosphate and disodium phosphate in very low concentrations in order to detect the smallest amount of alkali. Yeast extract acts as a source of nutrients, carbon, and nitrogen for those organisms incapable of using ammonia as a nitrogen source. (1)

There are several different formulas for determining the urease reaction of a microorganism. For the strong urease producers like Proteus spp. there is Rustigian and Stuart's Urea Broth. For the weak urease producers such as Klebsiella pneumoniae and Morganella morganii there is Urea R Broth. (1) Staphylococcus epidermidis , S. intermedius and most strains of S. saprophyticus are usually urease-positive using Urea R Broth. (5) In addition to these organisms, Urea R Broth aids in the identification of certain yeast and filamentous fungi. Cryptococcus spp. and Rhodotorula spp. are urease-positive. Most strains of Trichosporon spp. are urease-positive and Geotrichum spp. and Blastoschizomyces capitatus are urease-negative. (4,5) Almost all strains of Candida spp. found in clinical specimens are urease-negative, with the exception of C. lipolytica and some strains of C. krusei which are urease-positive. (5) Several species of Trichophyton are urease-positive when tested with Urea R Broth.

FORMULA

Ingredients per liter of deionized water:*

Urea 20.0gm
Yeast Extract 0.1gm
Disodium Phosphate 0.095gm
Monopotassium Phosphate 0.091gm
Phenol Red 0.01gm

Final pH 6.9 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE

Storage: Upon receipt store at 2-8ºC. away from direct light. Media should not be used if there are any signs of deterioration, discoloration, contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.

PRECAUTIONS

PROCEDURE

Specimen Collection: This product is not intended for primary isolation of patient specimens. It should be used only with cultures of isolated organism. This product is used in conjunction with other biochemical tests to identify cultures of isolated organism.

Method of Use:

Using a sterile loop, pick up three to four isolated colonies from a pure culture to inoculate the broth. Incubate aerobically at 35ºC. Check for color change at 6 to 8 and 12 to18 hours.

When testing Trichophyton spp. for urease, most reactions will become positive after 48-72 hours, but if still negative, hold for a maximum of 7 days before determining the test negative. (3)

INTERPRETATION OF RESULTS

A positive urease reaction is indicated by a color change from a reddish-orange to a bright pink. A negative reaction is indicated by no color change. It is often helpful to use an uninoculated tube as a negative control. (1) Consult listed references for the identification of colony morphology and further biochemical tests required for identification. (1,2,4,5)

LIMITATIONS

If not stored at 2-8ºC., urea has been shown to undergo autohydrolysis, which could interfere with the urease reaction. (1)

Plastic tubes or wells or tubes made of borosilicate type I glass is recommended to prevent non-specific alkaline shifts due to other glass types.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, other culture media, swabs, applicator sticks, incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Proteus hauseri
ATCC ® 13315
E 12-18hr 35°C Aerobic Positive; color change to bright pink
Escherichia coli
ATCC ® 25922
E 12-18hr 35°C Aerobic Negative; no color change

User Quality Control

Physical Appearance

Urea R Broth should appear clear, and reddish-orange in color.

P. vulgaris growing in Urea R Broth

Proteus hauseri (ATCC ® 13315) growing in Urea R Broth (Cat. no. Y141). The bright pink color change was indicative as positive for urease hydrolysis. Incubated aerobically for 18 hours at 35ºC.

E. coli growing in Urea R Broth

Escherichia coli (ATCC ® 25922) growing in Urea R Broth (Cat. no. Y141). The absence of a pink color change was indicative as negative for urease hydrolysis. Incubated aerobically for 18 hours at 35ºC.

REFERENCES

1. MacFaddin, J.F. Biochemical Tests for Identification of Medical Bacteria,, Lipincott Williams & Wilkins, Philadelphia, PA.

2. Baron, E.J, Peterson, L.R., and Finegold, S.M. 1994. Bailey and Scott's Diagnostic Microbiology, 9th ed. Mosby, St. Louis, MO.

3. Sutton, D.A., Fothergill, A.W., and Rinaldi, M.G. Guide to Clinically Significant Fungi. Williams & Wilkins.

4. St. Germain, Guy, et al. 1996. Identifying Filamentous Fungi. Star Publishing Company, Belmont, CA.

5. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

6. Larone, D.H. Medically Important Fungi: A Guide to Identification, American Society for Microbiology. Washington, D.C.

ATCC is a registered trademark of the American Type Culture Collection.

041816gr