Dichloran Rose Bengal Chloramphenicol (DRBC) Agar

Cat. no. G389 Dichloran Rose Bengal Chloramphenicol (DRBC) Agar, 15x100mm Plate, 18ml 10 plates/bag
Cat. no. W89 Dichloran Rose Bengal Chloramphenicol (DRBC) Agar, 15x100mm Plate, 26ml 10 plates/bag

INTENDED USE

Hardy Diagnostics Dichloran Rose Bengal Chloramphenicol (DRBC) Agar is recommended for the enumeration of yeasts and molds in food and dietary supplements.

This product is not intended to be used for the diagnosis of human disease.

SUMMARY

Fungi are recovered from air, soil, lakes, ponds, rivers, streams, wastewaters, and well waters. Due to their heterotrophic nature and their ability to adapt to a wide range of environmental conditions, fungi are also frequently encountered as contaminants in various commodities, including foods, inadequately cleaned food processing equipment, and food storage facilities. Since yeasts and molds can initiate growth over a wide pH and temperature range, growth can occur on almost any type of food, including processed foods and food ingredients. (1,3)

Smith and Dawson found that rose bengal added to a near-neutral medium (pH of 6.8) allowed for more colonies to develop than did an acidified medium such as Sabouraud Dextrose Agar. (5) Traditionally, low pH media are used to enumerate yeasts and molds from water, soil, and food. Such media are now believed to be inferior to selective media with antibiotics. The use of antibiotics for suppressing bacteria, rather than acid, results in improved recovery of injured (acid-sensitive) fungal cells, better control of bacteria, and less interference during counting from precipitated food particles. (2,6)

Hardy Diagnostics DRBC Agar contains peptone as a source of carbon and nitrogen, dextrose as an energy source, and magnesium sulfate to provide trace elements. (5) The medium contains chloramphenicol, which is added to inhibit most bacterial growth. (1) In addition to chloramphenicol, rose bengal is added to increase the selectivity and to help control overgrowth of rapidly growing molds such as Neurospora and Rhizopus species. Dichloran is added to the media to ihibit the spreading of molds by reducing colony diameters. (7) DRBC Agar conforms to the APHA guidelines for the mycological examination of foods. (1)

FORMULA

Ingredients per liter of deionized water:*

Dextrose 10.0gm
Peptone 5.0gm
Monopotassium Phosphate 1.0gm
Magnesium Sulfate 0.5gm
Chloramphenicol 0.1gm
Rose Bengal 0.025gm
Dichloran 0.002gm
Agar 15.0gm

Final pH 5.6 +/- 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 (shrinking, cracking, or discoloration), hemolysis - for media w/blood, contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.

PRECAUTIONS

PROCEDURE

1. If using the dilution method, add 40ml of the food sample to 200ml of 0.1% Peptone Water (Cat no. U201) and homogenize in a stomacher for 2 minutes. (3)

2. Inoculate 0.1ml of sample onto the agar surface.

3. Spread the inoculum evenly over the entire surface using a sterile bent glass rod or disposable spreader (Cat No. 174CS200).

4. Incubate plates at 22 to 25ºC. and examine plates after 3, 4, and 5 days of incubation. (1,3) Record results as colony forming units per gram of food.

INTERPRETATION OF RESULTS

Colonies should be apparent within five days of incubation. Yeast colonies will appear pink due to the uptake of rose bengal. Report counts as colony forming units (CFU) per gram or ml of sample.

LIMITATIONS

It is important to protect this medium from light since photodegradation of rose bengal produces compounds that are toxic to fungi. (1,4)

Due to the selective nature, some strains may grown poorly or fail to grow at all on this medium.

Chloramphenicol may not be sufficient to inhibit all bacterial flora.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, disposable spreaders (Cat No. 174CS200), swabs, applicator sticks, other culture media, such as Peptone Water 0.1% (Cat No. U201), stomacher, 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
Aspergillus brasiliensis
ATCC® 16404
A 3-5 days 15-30°C Aerobic Growth; white and filamentous, black specks on colonies
Candida albicans
ATCC® 10231
A 48-96hr 15-30°C Aerobic Growth; pink smooth raised colonies
Escherichia coli
ATCC® 25922
B 24 hr 35°C Aerobic Inhibited
Bacillus subtilis
ATCC® 6633
B 24 hr 35°C Aerobic Inhibited

USER QUALITY CONTROL

Physical Appearance

Dichloran Rose Bengal Chloramphenicol (DRBC) Agar should appear slightly opalescent, and bright pink in color.

REFERENCES

1. APHA Technical Committee on Microbiological Methods for Foods. Compendium of Methods for the Microbiological Examination of Foods, APHA, Washington, D.C.

2. American Public Health Association. Standard Methods for the Examination of Dairy Products , APHA, Washington, D.C.

3. U.S. Food and Drug Administration. Bacteriological Analytical Manual . AOAC, Arlington, VA. www.fda.gov/Food/FoodScienceResearch/LaboratoryMethods/ucm2006949.htm

4. Banks, J.G., R.G. Board. 1985. Preservation by the lactoperoxidase system (LP-S) of a contaminated infant formula. Letters in Applied Microbiology. 1:81–85.

5. Smith, N.R., V.T. Dawson. 1944. The bacteriostatic action of Rose Bengal in media used for plate counts of soil fungi. Soil Sci .; 58: 467-471.

6. King, Hocking and Pitt. 1979. Appl. Environ. Microbiol. 37:959.

7. Henson, OE. 1981. Dichloran as an Inhibitor of Mold Spreading in Fungal Plating Media: Effects on Colony Diameter and Enumeration. Appl Environ Microbiol . 42(4): 656–660.

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

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