C. BOTULINUM ISOLATION AGAR BASE

Clostridium botulinum is an anaerobic, spore forming bacteria that produces a neurotoxin protein botulin. Severe food poisoning results from the consumption of this protein (toxin), which may be produced in foods contaminated with Clostridium botulinum. C. botulinum Isolation Agar Base is formulated as per the recommendation of APHA for the selective isolation of C. botulinum from food samples.
The antibiotic supplement containing the broad spectrum antibiotics namely cycloserine, sulphamethoxazole and trimethoprim makes the medium very selective. Egg yolk emulsion helps in detecting lecithinase, lipase and proteolytic activity. Lecithinase degrades lecithin present in the egg yolk producing an insoluble, opaque precipitate in the medium surrounding the growth. Lipase break down free fats present in the egg yolk causing an iridescent (oil on water) sheen to form on the surface of the colonies.
Botulinal toxin is heat-labile. Therefore, the test samples and cultures should be maintained at refrigeration temperatures. The pH of the toxic material should also be maintained at a slightly acidic pH since botulinal toxin is less stable at alkaline pH. Inoculate 1-2 grams of solid or 1-2 ml of liquid food per 15 ml of enrichment broth. The enrichment broth employed is Cooked Meat Medium. After an incubation at 35?C for 7 days, observe for turbidity, gas production and meat digestion. Carry out gram staining and spore staining. To isolate C.botulinum mix enrichment broth with equal amount of sterile ethanol (alcohol treatment). The alcohol treated culture is further streaked on C.botulinum Isolation Agar Base. Alternatively untreated enrichment cultures or stool can be streaked directly on C.botulinum Isolation Agar Base.

for isolation of Clostridium botulinum from food and clinical samples