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Dehydrated Culture Media

CZAPEK DOX AGAR (MODIFIED)

Code: CM0097

a solid defined medium for the cultivation of those fungi and bacteria which are able to utilise sodium nitrate as the sole source of nitrogen. The acidity of the medium may be increased for the cultivation of acidophilic organisms such as yeasts.

Typical Formula*

gm/litre

 Sodium nitrate

2.0

 Potassium chloride

0.5

 Magnesium glycerophosphate

0.5

 Ferrous sulphate

0.01

 Potassium sulphate

0.35

 Sucrose

30.0

 Agar

12.0

 pH 6.8 ± 0.2 @ 25°C

 

* Adjusted as required to meet performance standards

Directions
Suspend 45.4g in 1 litre of distilled water. Bring to the boil to dissolve completely. Sterilise by autoclaving at 121°C for 15 minutes. Mix well before pouring. If required, adjust the reaction to pH 3.5 ± 0.2 by adding 10ml of Lactic Acid 10% (SR0021) per litre after sterilisation.

Description
Czapek Dox Agar (Modified) is a medium containing sodium nitrate as the sole source of nitrogen, it is one of the most useful solid media for the general cultivation of fungi.

In the Oxoid medium magnesium glycerophosphate and potassium sulphate replace the magnesium sulphate and potassium phosphate of the original. This modification prevents the precipitation of magnesium phosphate. The medium is also a highly satisfactory substrate for chlamydospore production by Candida albicans 1.

Dawson1 employed Oxoid Czapek Dox Agar (Modified) in her technique for the identification of Candida albicans by chlamydospore formation in primary culture, using swabs taken from the mouth and from the vagina. Identification was usually possible within 24 hours. The Oxoid medium showed good chlamydospore production whereas the original formulation did not. After 24 hours incubation 23 out of 27 Candida albicans strains had formed chlamydospores on Oxoid Czapek Dox Agar (Modified), 21 on rice infusion agar, 10 on Oxoid Corn Meal Agar (CM0103) and 10 on a corn meal agar made in the laboratory. After 48 hours 25 strains had formed chlamydospores on both the Oxoid medium and the rice agar, 24 on Oxoid Corn Meal Agar and 20 on the laboratory medium. Dawson concluded that the Oxoid Czapek Dox medium and the rice infusion agar were the most satisfactory media. None of 14 strains of unidentified yeasts formed chlamydospores on any medium.

Smith2 cited the following recommendations for the use of Czapek Dox Agar for taxonomic studies: by Thom and Church3 for Aspergillus; by Thom4 and by Raper and Thom5 for Penicillium; and by Wakesman6 for actinomycetes.

Technique
General Cultivation
To avoid excessive condensation cool the molten medium to 50°C before pouring approximately 12ml into each 9cm diameter Petri dish. Store the poured plates in an inverted position and inoculate using a needle or wire, with the plate still inverted in order to avoid scattering stray fungal spores over the surface of the medium. Time and temperature of incubation vary considerably according to the species being cultivated. As a general guide, incubate for 1-2 weeks at 25°C. Most Penicillium species have an optimum growth temperature between 20° and 25°C, whilst many Aspergillus species grow best at about 30°C. However, different fungi grow over a wide range of temperatures; Aspergillus fumigatus grows well at 50°C (Smith2) and Cladosporium herbarum will grow on meat at -6°C 7,8.

Identification of Candida albicans 1

  1. Using an inoculating needle (previously flamed, cooled and rubbed against the swab) cut across and through the medium in a Czapek Dox Agar plate to the base of the petri dish. With the same needle, raise the medium along the whole of one side of the cut - so that the inoculum is spread between the agar and the base of the dish.
  2. Incubate the inoculated plates for 24 hours at 28°C.
  3. Using a low-power objective, microscopically examine the unopened plates for chlamydospores through the base of each dish. Alternatively, remove the tops of the dishes, and examine through the top of the medium.
  4. If no chlamydospores are seen, incubate for a further 24 hours and re-examine.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared agar plates at 2-8°C.

Appearance
Dehydrated medium: White coloured, free-flowing powder
Prepared medium: Off-white coloured gel

Quality control

Positive control:

Expected results

Aspergillus niger ATCC® 9642 * White / yellow mycelium, black spores
Candida albicans ATCC® 10231 *Good growth; cream coloured colonies
Negative control: 

Uninoculated medium

No change
* This organism is available as a Culti-Loop®

References
1. Dawson Christine O. (1962) Saboutaudia 1. 214-219
2. Smith G. (1960) An Introduction to Industrial Mycoloogy 5th ed., Edward Arnold Ltd., London.
3. Thom C. and Church M. B. (1926) ’The Aspergilli’ Williams and Wilkins Co. Baltimore.
4. Thom C. (1930) ’The Aspergilli ’Williams and Wilkins Co. Baltimore.
5. Raper K. B. and Thom C. (1949) ’Manual of the Penicillia’ Williams and Wilkins Co. Baltimore.
6. Wakesman S. A. (1931) ’Principals of soil Microbiology’ Bailliere Tindall and Cox, London.
7.
Brooks F. T. and Kidd M. N. (1921) Specia. Report No 6, Food Invest. Board, DSIR, London.
8. Brooks F. T. and Handsford C. G. (1922) Trans. Brit. Mycol. Soc. 8. 113-142.

 
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