Detection of toxins in human cases or outbreaks of foodborne botulism 

Clostridium botulinum is a strict, gram-positive, anaerobic bacillus of the genus Clostridium, family Clostridiaceae, spore-forming that produces a neurotoxic toxin. This bacterium is usually found in the earth, in waters and its untreated sediments (oceans, lakes), with a worldwide distribution. In some circumstances this organism can contaminate food and grow in them producing its toxin/s. Botulism, a serious form of food poisoning, results mainly from the ingestion of food containing the toxin. Although this foodborne disease is rare, its mortality rate is high. When the type of toxin was determined among the 1036 cases detected in the USA. between 1899 and 1990, 384 were due to toxin type A, 106 for toxin B, 105 for toxin E and 3 for toxin F. Sometimes cases can occur due to 2 toxins, for example, A and B.

The detection of the preformed botulinum toxin can be carried out in a liquid like the serum obtained from blood. It can also be detected from the remains of an ingested food that has caused a case/outbreak of botulism, preparing an extract filtered from it. In cases of infant botulism or intestinal botulism, the presence of botulinum toxin can be detected in the faeces of patients, but we consider it more advisable to perform a previous culture of the faeces. To detect the toxin, inoculation of experimental animals (mice) is carried out, which will develop a paralytic symptomatology, followed by death, if they were inoculated with botulinum toxin. To corroborate that the mice have died from the inoculation of botulinum toxin, it is necessary to perform a neutralization test, which confirms that botulinum toxin exists. 

Tests available and recommended 

• Inoculation test to mice with serum or food extract ingested.
• In case of being positive, confirmation test by neutralization.
• Alternative: Culture test with stool sample, followed by molecular detection by real-time PCR of the presence of botulinum toxin-producing Clostridium botulinum, without specifying type.
• Optional in case of positive culture: If the presence of Clostridium botulinum is obtained in stool culture, identification of the type of toxin by real-time PCR for each of the genes producing the different types of botulinum toxin.

Required sample 

• Serum of the patient, at least 5 mL obtained as recently as possible, for inoculation in mice and confirmation of toxin, in case the mice were affected. The presence of botulinum toxin can be detected in patients´ faeces, but we consider it more advisable to perform a prior faeces culture. 

Schedule for results 

• We cannot give exact times. If only the detection of the toxin in mice is carried out and is negative, the report would be in a maximum of one week. If a culture of the sample is carried out, and subsequently the detection of the toxin by inoculating culture filtrates into batches of mice, the time would be two weeks. If any of these mouse inoculation tests, with a sample extract or with the enrichment culture filtrate, are positive, the neutralization test must be performed on mice to confirm that it is botulinum toxin, which takes 15 days to be completed.

Sample storage and shipping conditions 

• Samples must be stored and shipped in the conditions at least under conditions that guarantee refrigeration during transport (packaging the sample inside an expanded polystyrene box – white cork – with frozen cold accumulators – frozen packs). Note: this microorganism does not pose a biological risk of contamination by exposure. 

Form with product characteristics and chosen test(s) 

• If you request that test be carried out, you must send us with the product the specific request form that we have attached, indicating the characteristics of the product and the chosen tests that you wish to be carried out on the sample sent.