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Further Information

Organisms

Organisms this product works with:

Dehydrated Culture Media

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BOLTON SELECTIVE ENRICHMENT BROTH   

BOLTON BROTH

CODE: CM0983

A medium for the selective pre-enrichment of Campylobacter organisms in food samples.

Typical Formula*

gm/litre

Meat peptone

10.0

Lactalbumin hydrolysate

5.0

Yeast Extract

5.0

Sodium chloride

5.0

Alpha-ketoglutaric acid

1.0

Sodium pyruvate

0.5

Sodium metabisulphite

0.5

Sodium carbonate

0.6

Haemin

0.01

pH 7.4 ± 0.2 @ 25°C

 
* Adjusted as required to meet performance standards 

BOLTON BROTH SELECTIVE SUPPLEMENT 

CODE: SR0183

Vial contents (each vial is sufficient for 500ml of medium)

per vial
per litre

Cefoperazone

10.0mg

20.0mg

Vancomycin

10.0mg

20.0mg

Trimethoprim.

10.0mg

20.0mg

Cycloheximide

25.0mg

50.0mg

Directions
Add 13.8g of Bolton Broth to 500ml of distilled water. Sterilise by autoclaving at 121°C for 15 minutes. Cool to 50°C. Aseptically add 25ml Laked Horse Blood SR0048 and 1 vial of Bolton Broth Selective Supplement SR0183, reconstituted as directed. Mix well and distribute into sterile screw top containers.

Description
Bolton Selective Enrichment Broth is intended for the pre-enrichment of Campylobacter in food samples. Campylobacter are Gram-negative, spirally shaped microaerophilic organisms which may be present in raw milk, untreated water, improperly handled food and undercooked meats, poultry and shellfish. Human consumption of these organisms can result in a range of clinical illnesses from transient asymptomatic colonisation to severe dysentery. The symptoms of Campylobacter enteritis include diarrhoea, stomach pain, nausea, fever, headache and muscle pain. Complications of infection by Campylobacter jejuni may include unnecessary appendectomies as a result of abdominal pain, reactive arthritis or Guillian-Barré syndrome1. Campylobacter infection is recognised as one of the most common causes of bacterial gastroenteritis in humans, and the minimum infective dose may be as low as 500-800 cells1.

Since awareness of the apparent role of Campylobacter in human disease was heightened by Skirrow in 19772, a great number of culture media have evolved in response to the need to optimise performance. There was early recognition of the need for enrichment culture when examining food samples to overcome the damaging effects that food processing and preservation techniques can have on Campylobacter cells. Use of lower incubation temperatures in the early stages of enrichment is now widely established as an aid to cell recovery3. This principle was employed by Bolton in the development of his enrichment broth4.

Campylobacter can be injured by food processing and preservation procedures3. This makes them susceptible to selective agents which are tolerated by undamaged cells. False negative results are avoided through use of recovery medium such as Bolton Selective Enrichment Broth which increases the number of cells available for culture, first by resuscitating injured organisms and then encouraging them to multiply.
Bolton Selective Enrichment Broth contains nutrients to aid resuscitation of sublethally injured cells, and is formulated to avoid the need for a microaerobic atmosphere. Initial incubation is carried out at 30-37°C, depending on the type of food to be examined. After the pre-enrichment, the incubation temperature is raised to 42°C to increase the selective pressures on competing organisms.

Inclusion of sodium metabisulphite and sodium pyruvate in Bolton Broth quenches toxic compounds that may form in the culture medium. These additions also increase the aero-tolerance of the culture. The antibiotics contained in Bolton Broth Selective Supplement SR0183 optimise selectivity for Campylobacter spp. Vancomycin - active against Gram- positives. Cefoperazone - predominantly active against Gram- negatives. Trimethoprim - active against a wide variety of Gram-negative and Gram-positive organisms. Cycloheximide - active against yeasts.

Technique
One method of use is as follows:
Place 25g of food sample in 225ml Bolton Selective Enrichment Broth (prepared as described above) and homogenise the mixture using a Stomacher (or similar device). Bolton Selective Enrichment Broth does not require incubation in a microaerobic environment, but must be used in screw topped containers which are filled to within 20mm of the top4. Incubate for 4 hours at 37°C, followed by further incubation at 42°C . The broth can be subcultured after 24 hours and 48 hours on to either Modified CCDA (CM0739 + SR0155) or Preston Agar (CM0689 + SR0117 + SR0048)4.
For other methods please refer to BAM5.

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 medium at 2-8°C for up to 2 weeks.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Broth: Straw coloured solution containing small black particles

Quality control

Positive control:

Expected results: when subcultured on modified CCDA

Campylobacter jejuni ATCC® 29428*

Good growth; grey coloured colonies

Negative control:

 
Escherichia coli ATCC® 25922*No growth
* This organism is available as a Culti-Loop®.
References
1.
National Advisory Committee on Microbiological Criteria for Foods (1994). Journal of Food Protection 57 (12): 1101-1121.
2. Skirrow, M.B. (1977). British Medical Journal 2: 9-11.
3. Post, D. E. (1995). Food-Borne Pathogens Monograph Number 3 Campylobacter .
4. Bolton, F.J. (1995) Personal communication.
5. Hunt, J.M. (1998) Campylobacter. In: F.D.A. Bacteriological Analytical Manual, 8th Edition (Revision A) 7.01-7.27. AOAC, Arlington Va.
 
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