Hardy Diagnostics HUGO

MODIFIED LISTERIA ENRICHMENT BROTH

Cat. no. U167 Modified Listeria Enrichment Broth, 500ml Polycarbonate Bottle, 225ml 10 bottles/box

INTENDED USE

Hardy Diagnostics Modified Listeria Broth is used for selectively enriching Listeria monocytogenes from raw and pasteurized milk and milk products according to the International Dairy Federation (IDF) and the U.S. FDA Bacteriological Analytical Manual (BAM). (5,6)

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

SUMMARY

First described by Murray, Webb, and Swann, Listeria monocytogenes is a widespread problem in public health and the food industries. (7) This organism can cause human illness and death, particularly in immunocompromised individuals and pregnant women. The first reported food-borne outbreak of listeriosis was in 1985. (5) Since then, microbiological and epidemiological evidence from both sporadic and epidemic cases of listeriosis has shown that the principal route of transmission is via the consumption of foodstuffs contaminated with Listeria monocytogenes . (9)

Implicated vehicles of transmission include turkey frankfurters, coleslaw, pasteurized milk, Mexican-style and other soft cheeses, pate and pickled pork tongue. (5) The organism has been isolated from commercial dairy and other food processing plants. (6) It is ubiquitous in nature, being present in a wide range of unprocessed foods and in soil, sewage, silage and river water.

Listeria spp. grow over a pH range of 5.0-9.6 and survive in food products with pH levels outside these parameters. (11)

Listeria Enrichment Broth is based on the formula developed by Lovett, et al. in which Tryptic Soy Broth was supplemented with yeast extract for optimum growth of Listeria spp. (12) Modified Listeria Enrichment Broth is an adjusted version of Listeria Enrichment Broth in which the acriflavine content has been reduced from 15.0mg to 10.0mg per liter. This modification reflects the formula specified by the FDA and the International Dairy Federation for isolation of L. monocytogenes from milk and milk products. (5,6)

Modified Listeria Enrichment Broth contains pancreatic digest of casein, papaic digest of soybean meal, and yeast extract as nitrogen and vitamin sources. Dextrose provides an energy source. Sodium chloride maintains the osmotic balance of the medium. Dipotassium phosphate is a buffering agent. Nalidixic acid is incorporated to inhibit growth of gram-negative organisms, while acriflavine is added to suppress growth of gram-positive bacteria. The antifungal cycloheximide is used to inhibit saprophytic fungi.

Identification of Listeria spp. is based on successful isolation of the organism, biochemical characterization and serological confirmation.

FORMULA

Ingredients per liter of deionized water:*

Pancreatic Digest of Casein 17.0gm
Yeast Extract 6.0gm
Sodium Chloride 5.0gm
Papaic Digest of Soybean Meal 3.0gm
Dextrose 2.5gm
Dipotassium Phosphate 2.5gm
Cycloheximide 0.05gm
Nalidixic Acid 0.04gm
Acriflavine 0.01gm

Final pH 7.3 +/- 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), contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.

PRECAUTIONS

PROCEDURE

Consult listed references for information regarding sample preparation and processing. (1,2,5-12)

For dairy samples:

1. Add 25ml of liquid or 25.0gm of solid test material to 225ml of Modified Listeria Enrichment Broth (Cat. no. U167) and mix or blend thoroughly.

2. Incubate for 48 hours at 30ºC.

3. At 48 hours, streak the Modified Listeria Enrichment Broth culture onto plates of Oxford Medium (Cat. no. G46) or PALCAM Medium (Cat. no. G149).

4. Incubate the agar plates at 37ºC. for 48 +/- 2.0 hours.

INTERPRETATION OF RESULTS

Examine plates for typical Listeria spp. colonies.

Listeria monocytogenes will show growth with black colonies upon Oxford Medium. L. monocytogenes will also display growth with gray-green colonies and black precipitate on PALCAM Medium.

LIMITATIONS

Since the nutritional requirements of organisms vary, some strains of Listeria spp. may be encountered that fail to grow or grow poorly on this medium.

Modified Listeria Enrichment Broth is a partially selective medium. Growth of some contaminating strains will be markedly but not totally inhibited.

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, swabs, applicator sticks, 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
Listeria monocytogenes
ATCC ® 7644
A 24-48hr 35°C Aerobic Growth
Escherichia coli
ATCC ® 25922
B 24-48hr 35°C Aerobic Partial to complete inhibition
Staphylococcus aureus
ATCC ® 25923
B 24-48hr 35°C Aerobic Partial to complete inhibition

USER QUALITY CONTROL

Physical Appearance

Modified Listeria Enrichment Broth should appear clear with slight opalescence, and light amber in color.

REFERENCES

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

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

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

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

5. U.S. Food and Drug Administration. Bacteriological Analytical Manual. AOAC, Arlington, VA. www.fda.gov/Food/ScienceResearch/LaboratoryMethods/BacteriologicalAnalyticalManualBAM/default.htm


6. International Dairy Federation. 1990. Milk and milk products - detection of Listeria monocytogenes . IDF Provisional International Standard No. 143. International Dairy Federation, Brussels.

7. Murray, E.G, et al. 1926. A disease of rabbits characterized by large mononuclear leucocytosis caused by a hitherto undescribed bacillus Listeria monocytogenes . J. Path. Bact. ; 19:407-439.

8. Monk, J.D, et al. Irradiation inactivation of Listeria monocytogenes and Staphylococcus aureus in law and high-fat frozen and refrigerated ground beef. J. Food Prot. ; 57:769-772.

9. Wehr, H.M. 1987. Listeria monocytogenes - a current dilemma Special Report. J. Assoc. Anal . Chem. ; 80:769-7762.

10. Grau, F.H., et al. 1995. Occurrence, numbers, and growth of Listeria monocytogenes on some vacuum-packaged processed meats. J. Food Prot. ; 55:4-4.

11. Kramer, P.A., et al. 1969. Media selective for Listeria monocytogenes . J. Appl. Bacteriology ; 32:381-394.

12. Lovett, J.D., et al. 1987. Listeria monocytogenes . J. Food Prot. ; 50:188-192.


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

030216sw