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Dehydrated Culture Media

AMIES TRANSPORT MEDIUM

Code: CM0425

An improved transport medium, containing charcoal to prolong the viability of pathogenic organisms.

Typical Formula*

gm/litre

Charcoal pharmaceutical

10.0

Sodium chloride

3.0

Sodium hydrogen phosphate

1.15

Potassium dihydrogen phosphate

0.2

Potassium chloride

0.2

Sodium thioglycollate

1.0

Calcium chloride

0.1

Magnesium chloride

0.1

Agar

4.0

pH 7.2 + 0.2 @ 25°C

* Adjusted as required to meet performance standards 

Directions
Suspend 20g in 1 litre of distilled water. Bring to the boil to dissolve the agar completely. Distribute into small, screwcap bottles, stirring meanwhile to keep the charcoal evenly suspended. Screw down the caps firmly on the completely filled bottles. Sterilise by autoclaving at 121°C for 15 minutes. Invert the bottles whilst cooling to distribute the charcoal uniformly. Store in a cool place.

Description
Amies1 modified Stuart’s Transport Medium2,3,4 by replacing glycerophosphate with an inorganic phosphate buffer and adding charcoal to the medium.

The metabolism of glycerophosphate by coliform organisms and other Gram-negative rods in Stuart’s original formulation resulted in the proliferation of these organisms from wound swabs and faecal specimens.

A concentration of NaCl at 0.3% w/v was discovered by Amies to be optimal for the preservation of Neisseria gonorrhoeae.

Calcium and magnesium salts were added in the belief that these ions were of importance in controlling the permeability of the bacterial cells and so contributing to their survival.

Stuart3 showed that the survival of Neisseria gonorrhoeae was increased by the use of charcoal swabs, but because they were black and dusty, they proved unpopular with the patients. Amies1 overcame this problem by incorporating charcoal in this medium.

Survival of N. gonorrhoeae at 22°C

85 strains

With charcoal

Without charcoal

Time

No. of strains surviving

No. of strains surviving

24 hours

82

20

48 hours

70

0

72 hours

38

0

 

100 strains

Stuart’s Medium

Amies Medium

Time

No. of strains surviving

No. of strains surviving

24 hours

91

98

48 hours

79

87

72 hours

56

77

( Tables taken from Amies1 ).
The agar concentration was increased from that proposed by Stuart because the presence of charcoal particles interferes with the gelling properties of the agar.

Amies removed the methylene blue indicator from Stuart’s formulation considering it superfluous because of the presence of charcoal in the medium. Care should be taken to ensure that the prepared bottles of transport medium are not stored longer than 9 months from the date of preparation, or freshly steamed and the charcoal resuspended before use.

The value of these modifications was shown in two studies which tested the efficiency of various transport media5,6 . Amies Transport Medium is recommended for the transport of specimens to be cultured for Bacteroides ureolyticus.7

Technique
Use sterile, cotton-tipped swabs on wooden sticks to collect the specimen. Push the swab down one third of the medium depth and cut the stick so that when the cap is screwed down, the swab is forced to the bottom of the medium.

Make sure the cap is screwed firmly on the bottle and keep cool during the transport period.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
The prepared medium, held in tightly screw-capped bottles, can be stored at room temperature.

Appearance
Dehydrated medium: Black coloured, free-flowing powder
Prepared medium: Straw coloured semi-solid gel

Quality control

Positive controls:

Expected result at 35°C

Staphylococcus aureus ATCC® 25923 *Good growth

Escherichia coli ATCC® 25922 *

Good growth

Negative control:

 

Uninoculated medium

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

Precautions
It is important that the charcoal is properly suspended in the medium, invert the bottles when the bottles are cool but the agar still liquid.
During preparation of the medium, avoid prolonged heating in open flasks because thioglycollate is volatile.
Old medium should be freshly steamed and the charcoal resuspended before use.
Keep medium cool during transport but do not freeze.

References
1. Amies C.R. (1967) Can. J. Pub. Hlth. 58. 296-300.
2. Stuart R.D. (1946) J. Path. Bact. 58. 343-345.
3. Stuart R.D. (1959) Pub. Hlth. Rep. 74. 431-435.
4. Stuart R.D., Toshach Sheila R. and Patsula Teresa M. (1954) Acta. Pathol. Microbiol. Scand. 74. 371-374.
5. Gastrin L., Kallings O. and Marcetic A. (1968) Acta. Pathol. Microbiol. Scand. 74. 371-374.
6. Barry A.L., Fay G.D. and Sauer R.L. (1972) Appl. Microbiol. 24. 31-33.
7. Bennett K.W., Eley A. and Woolley P.D. (1990) Eur. J. Clin. Microbiol. Inf. Dis. 9. 237-238.

 
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