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Material Safety Data Sheet

Organisms

Organisms this product works with:

Dehydrated Culture Media

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ALKALINE SALINE PEPTONE WATER (ISO)

Code: CM1117

a broth medium for the enrichment of Vibrio species from food and water samples; this formulation conform to ISO/TS 21872-1&2:20071,2

Typical Formula*g/litre
 Peptone 20.0
 Sodium chloride20.0
 pH 8.6 ± 0.2 @ 25°C 

* Adjusted as required to meet performance standards

Directions
Dissolve 40g in 1 litre of distilled water. Mix well and distribute into final containers. Sterilize by autoclaving at 121°C for 15 minutes.

Description
Alkaline Saline Peptone Water (ASPW) was first formulated by Shread, Donovan and Lee3 to be used as a non-selective enrichment broth for the cultivation of Aeromonas species. Cruickshank reported that the raised pH of the medium could be used to effectively cultivate Vibrio species4. The medium exploits the ability of Vibrio species to tolerate alkaline conditions and high sodium chloride levels.

Technique
For the detection of Vibrio parahaemolyticus and Vibrio cholerae
The following is an overview of ISO/TS 21872-1:20071 – Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 1: Detection of Vibrio parahaemolyticus and Vibrio cholerae. Prepare samples in accordance with ISO 6887 (appropriate part)5, ISO 72186 and ISO 82617.

  1. Add xg of sample to 9xml or 9xg of ASPW (ISO) at ambient temperature.
  2. Incubate for 6 ± 1 hour, at 41.5 ± 1°C for fresh products or 37 ± 1°C for deep-frozen/salted products.
  3. Remove 1ml of incubated medium and inoculate into a fresh 10ml of Alkaline Saline Peptone Water (ISO) and incubate for 18 ± 1 hour at 41.5°C.
  4. From the cultures obtained in the ASPW (ISO), inoculate with a microbiological loop the surface of a TCBS Agar plate (CM0333) and second optional selective isolation medium in order to obtain single colonies.
  5. Incubate the plates at 37°C for 24 ± 3 hours (or as appropriate for the second optional medium).
  6. Examine the plates for the presence of typical colonies* of presumptive pathogenic Vibrio spp. Select 5 typical colonies for subculture onto Saline Nutrient Agar followed by biochemical confirmation.

For the detection of species other than Vibrio parahaemolyticus and Vibrio cholerae
The following is an overview of ISO/TS 21872-2:20072 – Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 2: Detection of species other than Vibrio parahaemolyticus and Vibrio cholerae. Prepare samples in accordance with ISO 6887 (appropriate part)5, ISO 72186 and ISO 82617.

  1. Add xg of sample to 9xml or 9xg of ASPW (ISO) at ambient temperature.
  2. Incubate for 6 ± 1 hour, at 41.5- ± 1°C for fresh products or 37 ± 1°C for deep-frozen/salted products.
  3. Remove 1 ml of incubated medium and inoculate into a fresh 10ml of Alkaline Saline Peptone Water (ISO) and incubate for 18 ± 1 hour at 37 ± 1°C.
  4. From the cultures obtained in the ASPW (ISO), inoculate with a microbiological loop the surface of a TCBS Agar plate (CM0333) and second optional selective isolation medium in order to obtain single colonies.
  5. Incubate the plates at 37°C for 24 ± 3 hours (or as appropriate for the second optional medium).
  6. Examine the plates for the presence of typical colonies of presumptive pathogenic Vibrio spp. Select 5 typical colonies for subculture onto Saline Nutrient Agar followed by biochemical confirmation.

Refer to ISO/TS 21872-1&2:20071 or Oxoid manual entries for CM0333 (and entries for other optional second media as appropriate) for interpretation of colony type.

Quality control

Positive controls:Expected results
Vibrio parahaemolyticus NCTC 10885Turbid growth
Vibrio furnissii NCTC 11218Turbid growth
Vibrio vulnificus ATCC® 29307Turbid growth
Vibrio cholerae non-O1/non-O139 ATCC® 14733*Turbid growth
Negative control: 
Uninoculated mediumNo change

* This organism is available as a Culti-Loop® 

References
1. ISO/TS 21872-1:20071, Microbiology of food and animal feeding stuffs – Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 1: Detection of Vibrio parahaemolyticus and Vibrio cholerae.
2. ISO/TS 21872-2:20072, Microbiology of food and animal feeding stuffs – Horizontal method for the detection of potentially enteropathogenic Vibrio spp. Part 2: Detection of species other than Vibrio parahaemolyticus and Vibrio cholerae.
3. Shread P., Donovan T. J., and Lee J. V. (1991) Soc. Gen. Microbiol. Q. 8:184.
4. Cruickshank R. (1968) Medical Microbiology. 11th ed. Livingstone Ltd, London, UK.
5. ISO 6887 (all parts), Microbiology of food and animal feeding stuffs – Preparation of test samples, initial suspension and decimal dilutions for microbiological examination.
6. ISO 7218 Microbiology of food and animal feeding stuffs – General requirements and guidance for microbiological examinations.
7. ISO 8261 Microbiology of food and animal feeding stuffs – General guidance for the preparation of test samples, initial suspensions and decimal dilutions for microbiological examination.

 
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