Pseudomonas Selective Agar Base


Cat. No. 1.07620.0500
 (500 g)
Product available March 2005


For the detection and enumeration of Pseudomonas.



Pseudomonas C-F-C Selective Agar
When supplemented with Pseudomonas C-F-C Selective Supplement (Cat.No.1.07627) the medium complies with the recommendations of ISO 13720 for the detection and enumeration of Pseudomonas spp. in foods and animal feed.

Pseudomonas C-N Selective Agar
When supplemented with Pseudomonas C-N Selective Supplement (Cat.No. 1.07624) the medium complies with the recommendations of DIN/EN 12780 for the detection and enumeration of Pseudomonas aeruginosa in water using the membrane filtration technique.

Mode of Action Quality Control
Typical Composition (g/liter) Additives/Reagents
Preparation Literature
Experimental Procedure and Evaluation  


Mode of Action
 The peptone mixture in Pseudomonas Selective Agar Base allows growth of a broad spectrum of Pseudomonades. The amount of potassium sulfate and manganese chloride supports forming of pigments.

By use of the appropriate selective supplement and the incubation temperature the medium becomes selective for Pseudomonas spp. including Burkholderia cepacia, formerly known as Pseudomonas cepacia (C-F-C Agar), or Pseudomonas aeruginosa (C-N Agar).


Typical Composition (g/liter)
 Peptone from gelatine 16.0; Casein hydrolysate 10.0; potassium sulfate 10.0; manganese chloride 1.4; agar-agar 11.0.

Preparation
 Suspend 24.2 g in 500 ml of purified water, add 5 ml glycerol and heat to boiling until completely dissolved.

Autoclave for 15 minutes at 121°C.

Cool the medium to 45-50°C and aseptically add the contents of one vial of Pseudomonas C-F-C Selective Supplement (Cat. No. 1.07627) or Pseudomonas C-N Selective Supplement (Cat. No. 1.07624). Mix thoroughly and pour plates.

pH 7.1 ± 0.2 at 25°C.

The prepared plates are clear and colorless and can be stored for up to 4 weeks at 2-8°C in the refrigerator.

Protect from light and drying.

Do not keep the liquid medium (45-50°C) longer than 4 hours.

Do not remelt the medium several times.

Experimental Procedure and Evaluation
Pseudomonas C-F-C Selective Agar
 Inoculate the medium using the surface spread method.

Incubation 44 ± 4 hours at 25 ± 1°C.

All grown colonies are suspect Pseudomonas spp. and counted as such.

The suspect colonies must be confirmed. Colonies which show a positive oxidase reaction but no glucose fermentation are confirmed Pseudomonas spp. colonies.

Pseudomonas C-N Selective Agar
Inoculate the medium using the membrane filtration technique.

The filter material impacts results. Good results were achieved using Cellulose-Mixed Ester membranes (e.g. Pall GN-6).

Incubation: 44 ± 4 hours at 36 ± 2°C.

Check the membrane filters for growth after 22 ± 2 h and 44 ±4 h.

All grown colonies with a blue-green pigmentation are considered confirmed Pseudomonas aeruginosa colonies and counted as such.
Check the membrane filters under UV-light. All colonies not showing the blue-green pigmentation but fluoresce are suspect P. aeruginosa colonies and confirmed by use of acetamide solution.
All other reddish-brown pigmenting colonies, which do not fluoresce are considered suspect P. aeruginosa colonies and confirmed by the oxidase test, acetamide solution and King's B Medium.


Quality control of Test Agar pH 6.0 (1.10663)

Test strain
 Inhibition zones in mm Ø

Gentamicin
         10 µg                  30 µg
 		Penicillin
 10 IU
 		Streptomycin
 10 µg
Bacillus subtilis strain BGA (DSM 618)
 		20-28
 		22-30
 		36-48
 		19-27


Quality Control
Pseudomonas C-F-C Selective Agar 44 h ± h at 25 ± 1°C

Test strain
 		Recovery Rate
Pseudomonas aeruginosa ATCC 27853
 > 70%
Pseudomonas putida ATCC 12633
 > 70%
Pseudomonas fluorescens ATCC 13525
 > 70%
Pseudomonas fragi ATCC 27362
 > 70%
Burkholderia cepacia ATCC 17759
 > 70%
Proteus mirabilis ATCC 14153
 < 0.01%
Staphylococcus aureus ATCC 25923
 < 0.01%


Quality Control
Pseudomonas C-N Selective Agar 44 h ± 4 h at 36 ± 2°C
Test strain
 		Recovery Rate
Pseudomonas aeruginosa ATCC 27853
 > 70%
Pseudomonas fluorescens ATCC 13525
 < 0.01%
Aeromonas hydrophila ATCC 7966
 < 0.01%
Klebsiella pneumoniae ATCC 13883
 < 0.01%
Proteus mirabilis ATCC 14153
 < 0.01%
Providencia rustigianii ATCC 13159
 < 0.01%


Additives


EMD Cat. No. Product Pack Size
1.07627.0001 Pseudomonas C-F-C Selective Supplement 16 vials
1.07624.0001 Pseudomonas C-N Selective Supplement 16 vials
     


Literature

Goto, S., and S. Enomoto. 1970. Nalidixic Acid Cetrimide Agar. A New Selective Plating Medium for the Selective Isolation of Pseudomonas aeruginosa. Japan. J. Microbiol. 14: 65 - 72.
Mead, G.C., and B.W. Adams. 1977. A selective medium for the rapid isolation of Pseudomonas associated with poultry meat spoilage. Br. Poult. Sci. 18: 661 - 670.
ISO INTERNATIONAL STANDARDISATION ORGANISATION. Microbiology of food and animal feeding stuffs - Horizontal method for the enumeration of Pseudomonas spp. ISO/WD 13720:2000.
EN EUROPEAN STANDARD. Water Quality - Detection and enumeration of Pseudomonas aeruginosa by membrane filtration. DIN/EN 12780:2002.



© 2002 Merck KGaA, Darmstadt, Germany