BLUECOLI™ URINE BIPLATE

Cat. no. J123 BluEcoli™ Urine Biplate, 15x100mm Biplate, 10ml/10ml 10 plates/bag

INTENDED USE

Hardy Diagnostics BluEcoli™ Urine Biplate is a urine culture media, consisting of Blood Agar on one side and BluEcoli™ Agar on the other side, which is used for the isolation of urinary pathogens and for the identification of E. coli .

SUMMARY

Diagnosis of urinary tract infections contributes significantly to the daily workload in a microbiology laboratory, therefore any attempts of innovation by reducing the work load and cost are always welcome while a high quality standard is still maintained. For several years, development of culture media containing chromogens and fluorogens has led to the development of a great number of methods for the identification of microorganisms in primary isolation media.

The BluEcoli™ Agar side of the biplate uses a MacConkey Agar base with beta-D-glucoronidase (GUD) as indicator for E. coli since this enzyme is present in 94-96% of members of this species. (1) Presence of GUD can be measured by using different chromogenic and fluorogenic substrates. Fluorogens are hydrolyzed by GUD yielding fluorescent colonies of E. coli under a long-wave (366nm) UV light source, whereas chromogens release chromophores resulting in easy to read purple-blue colored colonies of E. coli . (1)

E. coli is known as the most common pathogen responsible for great majority (75-90%) of urinary tract infections. (11) Therefore rapid identification and reporting is useful in the direction of therapy, and also streamlines the overall turn-around-time and allows microbiologists to devote more time to problematic cultures. (2,3)

Several chromogenic media have been compared to traditional urine culture media (i.e., Blood and MacConkey Agars) and were found to be at least as good as traditional media for the isolation of urinary pathogens. (2-5) Overall, studies agree that rapid detection and identification of microorganisms is of high importance in a diverse array of clinical and research settings. By incorporating synthetic enzyme substrates into primary isolation media, the enumeration and detection by color reactions can be performed directly on the isolation plate. This enhances the accuracy and performance of the microbiologist, allowing them to quickly recognize E. coli isolates and rule out clinically insignificant mixed cultures. (3)

Since very limited information is documented, in terms of accuracy, concerning usage of colonies from chromogenic media for susceptibility testing in automated systems, it is recommended that colonies from the Blood Agar side of the BluEcoli™ Urine Biplate be used for susceptibility testing.

FORMULA

Ingredients per liter of deionized water:*

Blood Agar (Side I):
Pancreatic Digest of Casein 15.0gm
Pancreatic Digest of Soybean Meal 5.0gm
Sodium Chloride 5.0gm
Sheep Blood 50.0ml
Agar 12.0gm

Final pH 7.3 +/- 0.2 at 25ºC.

BluEcoli™ Agar (Side II):
Pancreatic Digest of Gelatin 17.0gm
Lactose 10.0gm
Sodium Chloride 5.0gm
Pancreatic Digest of Casein 1.5gm
Peptic Digest of Animal Tissue 1.5gm
Bile Salts 1.5gm
Neutral Red 30.0mg
Chromogenic Substances 90.0mg
Crystal Violet 1.0mg
Agar 13.5gm

Final pH 7.1 +/- 0.3 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 (shrinking, cracking, or 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: Specimens should be submitted directly to the laboratory without delay and protected from excessive heat and cold. If there is to be a delay in processing, the specimen should be contained within an air-tight transport vial and refrigerated until inoculation. Consult listed references for information on specimen collection. (6-10)

Prepared media should be inoculated, incubated, and results recorded according to accepted procedures described in the listed reference texts. (6-10)

INTERPRETATION OF RESULTS

E. coli appears as purple-blue colored colonies on the BluEcoli™ Agar side of the biplate. Other gram-negative rods that grow on the BluEcoli™ Agar side may appear pink to slight pink for lactose-positive colonies, and colorless for lactose-negative colonies, as is typical of MacConkey Agar.

The Blood Agar side of the biplate will exhibit typical colony morphology of urinary pathogens and contaminating organisms.

LIMITATIONS

Speciation and susceptibility tests should be done with pure cultures taken from the Blood Agar side of the biplate. Erroneous antimicrobial susceptibility results may occur from colonies taken from the BluEcoli™ Agar side.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as calibrated loops, specimen cups, other culture media, 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
Blood Agar (Side I):
Streptococcus pneumoniae

ATCC ® 6305
A 24hr 35°C Aerobic Growth; alpha-hemolysis
Streptococcus pyogenes
ATCC ® 19615**
A 24hr 35°C Aerobic Growth; beta-hemolysis
Staphylococcus aureus
ATCC ® 25923
A 24hr 35°C Aerobic Growth
Escherichia coli
ATCC ® 25922
A 24hr 35°C Aerobic Growth
CAMP Test:
Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Staphylococcus aureus
ATCC ® 33862
H 18-24hr 35°C Aerobic Growth
Streptococcus agalactiae
ATCC ® 12386
H 18-24hr 35°C Aerobic Growth; positive (enhanced arrowhead hemolysis)
Streptococcus pyogenes
ATCC ® 19615
H 18-24hr 35°C Aerobic Growth; negative (no enhanced arrowhead hemolysis)
BluEcoli™ Agar (Side II):
Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 25922**
A 24hr 35°C Aerobic Growth; purple-blue colonies
Enterobacter cloacae
ATCC ® 23355
A 24hr 35°C Aerobic Growth; pink colonies
Klebsiella pneumoniae
ATCC ® 13883**
A 24hr 35°C Aerobic Growth; pink colonies
Proteus mirabilis
ATCC ® 12453
A 24hr 35°C Aerobic Growth; clear colonies
Staphylococcus aureus
ATCC ® 25923
B 24hr 35°C Aerobic Inhibited
Enterococcus faecalis
ATCC ® 29212
B 24hr 35°C Aerobic Inhibited

** Recommended QC strains for User Quality Control according to the CLSI document M22 when applicable.

User Quality Control

PHYSICAL APPEARANCE

BluEcoli™ Urine Biplate should appear as follows:

Blood Agar (Side I) should appear opaque, and cherry red in color.
BluEcoli™ Agar (Side II) should appear translucent, and pink in color.

E. coli growing on a BluEcoli Urine Biplate

Escherichia coli (ATCC ® 25922) colonies growing on a BluEcoli™ Urine Biplate (Cat no. J123). Incubated aerobically for 24 hours at 35ºC.

Mixed culture growing on BluEcoli Agar

Mixed culture with Enterobacter cloacae (ATCC ® 23355) colonies (pink) and Escherichia coli (ATCC ® 25922) colonies (blue) growing on BluEcoli™ Agar (Cat no. J123). Incubated aerobically for 24 hours at 35ºC.

REFERENCES

1. Manafi, M. 2000. New developments in chromogenic and fluorogenic culture media. Int. J. of Food Microbiol.; 60:205-218.

2. Samra, Z., et al. 1998. Evaluation of use of a new chromogenic agar in detection of urinary tract pathogens. J. Clin. Microbiol.; 36:990-994.

3. Hengstler, K.A., et al. 1997. Evaluation of BBL CHROMagar™ Orientation medium for detection and presumptive identification of urinary tract pathogens. J. Clin. Microbiol.; 35:2773-2777.

4. D'Souza, H., et al. 2004. Practical bench comparison of BBL CHROMagar™ Orientation and standard two-plate media for urine cultures. J. Clin. Microbiol.; 42:60-64.

5. Aspevall, O., et al. 1996-2005. Performance of four chromogenic urine culture media after one or two days of incubation compared with reference media. J. Clin. Microbiol.; 40:1500-1503.

6. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

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

8. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

9. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

10. Koneman, E.W., et al. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott Company, Philadelphia, PA.

11. Gupta, K., et al. 2001. Increasing Antimicrobial Resistance and the Management of Uncomplicated Community-Acquired Urinary Tract Infections. Ann. Intern. Med.; 135:41-50.


ATCC is a registered trademark of the American Type Culture Collection.
CHROMagar is a trademark of Dr. Alain Rambach, France.

121415hh