CRITERION™ SKIM MILK AGAR

Cat. no. C7930 CRITERION™ Skim Milk Agar 142gm
Cat. no. C7931 CRITERION™ Skim Milk Agar 500gm
Cat. no. C7932 CRITERION™ Skim Milk Agar 2kg
Cat. no. C7933 CRITERION™ Skim Milk Agar 10kg
Cat. no. C7934 CRITERION™ Skim Milk Agar 50kg

INTENDED USE

Hardy Diagnostics CRITERION™ Skim Milk Agar is used for the cultivation and differentiation of microorganisms based on proteolytic activity.

Dehydrated culture media is a raw material not intended for use in the diagnosis of human disease. For proper use, the product requires additional processing and supplementation of ingredients before use.

SUMMARY

Skim Milk Agar is commonly used to demonstrate proteolysis by organisms capable of hydrolyzing casein. Proteolytic bacteria use the enzyme caseinase to hydrolyze casein and form soluble nitrogenous compounds displayed as a clear zone around colonies. This clearing is much more pronounced on agar containing milk if the bacteria are able to produce acid from fermentable carbohydrates in the medium. Moreover, the hydrolysis of casein is often used to evaluate proteolytic activity of psychrotrophic microorganisms of significance to food, water, and dairy industries.(1,3,5)

Psychrotrophs, such as Pseudomonas aeruginosa, are strongly proteolytic and often responsible for spoilage of meat and dairy foods; this spoilage can result in a stale, bitter or rancid taste and smell.(3,4) It is generally known that Pseudomonas species are most often responsible for the spoilage of fish.(5) In addition, pseudomonads are often isolated from large bodies of water, and some species have been linked to eye, ear, and skin infections from recreational water use. Thus, pseudomonads may serve as an indicator of recreational water quality. P. aeruginosa is also commonly found in drinking water and is known to be very resistant to ozonation and chemical disinfection in swimming pools.(2)

Many methods have been used to enumerate P. aeruginosa from water, but the most-probable-number (MPN) method results in satisfactory recovery of the organism.(1) However, this method is not suitable for large-volume water testing and lacks precision. The membrane filtration (MF) technique eliminates these deficiencies.

CRITERION™ Skim Milk Agar is an improved formulation from standard skim milk formulas and provides greater sensitivity.(11) CRITERION™ Skim Milk Agar contains casein peptone and glucose to support growth. Yeast extract is added as a vitamin source. The medium is a modification of Brown and Foster's formulation and can be used as a differential and confirmatory test for the identification of P. aeruginosa in water.(1) P. aeruginosa hydrolyzes casein as indicated by a zone of clearing around the colonies. There may also be a yellow to green pigment diffused into the medium.(1)

FORMULA*

Ingredients per liter of deionized water:*

Gram weight per liter: 71.0gm/L
Dry Milk, Instant Nonfat 50.0gm
Pancreatic Digest of Casein 5.0gm
Yeast Extract 2.5gm
Glucose 1.0gm
Agar 12.5gm

Final pH 6.8 +/- 0.2 at 25ºC.

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

STORAGE AND SHELF LIFE

Store the sealed bottle(s) containing dehydrated culture medium at 2-30ºC. Dehydrated culture medium is very hygroscopic. Keep lid tightly sealed. Protect dehydrated culture media from moisture and light. The dehydrated culture media should be discarded if it is not free-flowing or if the color has changed from its original white to off white.

Store the prepared culture media at 2-8ºC.

PRECAUTIONS

METHOD OF PREPARATION FOR DEHYDRATED CULTURE MEDIA

1. Suspend 71.0gm of the dehydrated culture medium in 1 liter of deionized water. Stir to mix thoroughly.

2. Gently heat to dissolve completely.

3. This is a heat sensitive product. Based on individual autoclaves, time or temperature adjustments may be necessary.
Sterilize in the autoclave at:

4. Cool to 45-50ºC. and dispense as desired.

PROCEDURE AND INTERPRETATION OF RESULTS

For information on procedures and interpretation of results, consult listed references.(1-5)

For the presumptive detection of P. aeruginosa from recreational water, a 200-500ml sample of water is filtered through a sterile membrane filter. The membrane is then placed on a plate of mPA Agar (Cat. no. G133). Inoculated plates are inverted and incubated at 41.5ºC. for 72 hours. Colonies are isolated from mPA Agar and subcultured using a direct inoculum to Skim Milk Agar by placing a single streak 2 to 4cm long down the center of the plate. Incubate Skim Milk Agar plates at 35ºC. for 24 to 48 hours and observe for characteristic clearing and pigment formation.(1)

For food and dairy testing of P. aeruginosa using CRITERION™ Skim Milk Agar, see listed references.(3,5)

LIMITATIONS

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as autoclaves, incinerators, and incubators, etc., are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Pseudomonas aeruginosa
ATCC ® 27853
* 24-48hr 35°C Aerobic Growth; clear zone surrounding the colonies may have a yellowish to green pigment
Escherichia coli
ATCC ® 25922
* 24-48hr 35°C Aerobic Growth; no clear zone

User Quality Control:

Physical Appearance:

CRITERION™ Skim Milk Agar powder should appear homogeneous, free-flowing, and white to off-white in color. The prepared media should appear opaque, and white in color.

REFERENCES

1. American Public Health Association. Standard Methods for the Examination of Water and Wastewater, APHA, Washington, D.C.

2. Hurst, C.J., Crawford, R.L., Knudsen, G.R., McInerney, M.J., Stetzenbach, L.D. 1996-2005. Manual of Environmental Microbiology. 2nd ed. ASM Press, Washington, D.C.

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

4. Hui, Y.H., Pierson, M.D., Gorham, J.R. 2001. Foodborne Disease Handbook.& 2nd ed. Marcel Dekker, Inc., New York, NY.

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

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. MacFaddin, J.F. 1985. Media for Isolation, Cultivation, Identification, Maintenance of Bacteria, Vol. I. Williams & Wilkins, Baltimore, MD.

11. Martley, F.G., et al. 1969. J. Appl. Bact.; 33:363-370.


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

062116gr