Clostridium botulinum

Clostridium botulinum is a bacterium that produces dangerous toxins (botulinum toxins) under low-oxygen conditions.
• Botulinum toxins are one of the most lethal substances known.
• Botulinum toxins block nerve functions and can lead to respiratory and muscular paralysis.
• Human botulism may refer to foodborne botulism, infant botulism, wound botulism, and inhalation botulism or other types of intoxication.
• Foodborne botulism, caused by consumption of improperly processed food, is a rare but potentially fatal disease if not diagnosed rapidly and treated with antitoxin.
• Homemade canned, preserved or fermented foodstuffs are a common source of foodborne botulism and their preparation requires extra caution.

Clostridium botulinum is a gram-positive, rod-shaped, anaerobic, spore-forming, motile bacterium with the ability to produce the neurotoxin botulinum.
The botulinum toxin can cause botulism, a severe flaccid paralytic disease in humans and other animals, and is the most potent toxin known to mankind, natural or synthetic, with a lethal dose of 1.3–2.1 ng/kg in humans.
C. botulinum is a diverse group of pathogenic bacteria initially grouped together by their ability to produce botulinum toxin and now known as four distinct groups, C. botulinum groups I–IV, as well as some strains of Clostridium butyricum and Clostridium baratii, are the bacteria responsible for producing botulinum toxin.

Microbiology of Clostridium botulinum

C. botulinum is a gram-positive, rod-shaped, spore-forming bacterium. It is an obligate anaerobe, meaning that oxygen is poisonous to the cells. However, C. botulinum can tolerate traces of oxygen because of the presence of enzyme superoxide disputable, which is an important antioxidant defence in nearly all cells exposed to oxygen.
C. botulinum produce neurotoxin only during sporulation, which can happen only in anaerobic environment.

Isolation of C. botulinum

In the laboratory, C. botulinum is usually isolated in tryptose sulfite cyclosporine (TSC) growth medium in an anaerobic environment with less than 2% oxygen. C. botulinum is a lipase-positive microorganism that grows between pH of 4.8 and 7.0 and cannot use lactose as a primary carbon source.

Isolation of pure culture

Restreak toxic culture in duplicate on egg yolk agar medium. Incubate one plate anaerobically at 35°C. Incubate second plate aerobically at 35°C. If colonies typical of C. botulinum are found only on anaerobic plate (no growth on aerobic plate), the culture may be pure. Failure to isolate C. botulinum from at least one of the selected colonies means that its population in relation to the mixed flora is probably low. Repeated serial transfer through additional enrichment steps may increase the numbers sufficiently to permit isolation. Store pure culture in sporulated state either under refrigeration, on glass beads, or lyophilized.

Pathology

Food borne botulism

Signs and symptoms of food borne botulism typically begin between 18 and 36 hours after the toxin gets into the body. Depending on the amount of toxin ingested, it can range from a few hours to several days.
Following are the symptoms of food borne botulism:
• Double vision
• Blurred vision
• Dropping eyelids
• Nausea, vomiting, and abdominal cramps
• Slurred speech
• Trouble breathing
• Difficulty in swallowing
• Dry mouth
• Muscle weakness
• Constipation
• Reduced or absent deep tendon reactions, such as in the knee

Wound botulism

Most people who develop wound botulism inject drugs several times a day, so it’s difficult to determine how long it takes for signs and symptoms to develop after the toxin enters the body. Most common in people who inject black tar heroin, wound botulism signs and symptoms include:
• Difficulty swallowing or speaking
• Facial weakness on both sides of the face
• Blurred or double vision
• Dropping eyelids
• Trouble breathing
• Paralysis

Infant botulism

Infant botulism occurs mostly in infants under 6 months of age. Different from food borne botulism caused by ingestion of pure-formed toxins in food, it occurs when infants ingest C. botulinum spores, which germinate into bacteria that colonise in the gut and release toxins. In most adults and children older than about 6 months, this would not happen because natural defences in intestines that develop over time prevent germination and growth of the bacterium.
If infant botulism is related to food, such as honey, problems generally begin within 18 to 36 hours after the toxin enters the baby’s body. Signs and symptoms include:
• Constipation (often the first sign)
• Floppy movements due to muscle weakness and trouble controlling the head
• Weak cry
• Irritability
• Drooling
• Dropping eyelids
• Tiredness
• Difficulty sucking or feeding
• Paralysis

Other type of intoxication

Waterborne botulism could theoretically result from the ingestion of the pre-formed toxin. However, as common water treatment processes (such as boiling, disinfection with 0.1% hypochlorite bleach solution) destroy the toxin, the risk is considered low.

Diagnosis and treatment

Diagnosis of botulism is based on clinical history and clinical examination followed by laboratory confirmation including demonstrating the presence of botulinum toxin in serum, stool or food, or a culture of C. botulinum from stool, wound or food. Misdiagnosis of botulism may occur in the confusion of stroke, Guillain-Barré syndrome, or myasthenia graves.
Antitoxin should be administered as soon as possible after a clinical diagnosis. Early administration is effective in reducing mortality rates. Severe botulism require supportive treatment, especially mechanical ventilation, which may be required for weeks or even months.

Prevention

Prevention of foodborne botulism is based on good practice in food preparation particularly during heating/sterilization and hygiene. Foodborne botulism may be prevented by the inactivation of the bacterium and its spores in heat-sterilized (for example, retorted) or canned products or by inhibiting bacterial growth and toxin production in other products. The vegetative forms of bacteria can be destroyed by boiling but the spores can remain viable after boiling even for several hours. However, the spores can be killed by very high temperature treatments such as commercial canning.

Commercial heat pasteurization (including vacuum packed pasteurized products and hot smoked products) may not be sufficient to kill all spores and therefore the safety of these products must be based on preventing bacterial growth and toxin production. Refrigeration temperatures combined with salt content and/or acidic conditions will prevent the growth of the bacteria and formation of toxin.

The WHO Five Keys to Safer Food serve as the basis for educational programmes to train food handlers and educate the consumers. They are especially important in preventing food poisoning.
The Five Keys are:

• keep clean
• separate raw and cooked
• cook thoroughly
• keep food at safe temperatures
• use safe water and raw materials.

Reference:

WHO, sciencedirect, fda

Gaurav Singh

Editor in Chief Medical Microbiology & RDT Labs - RDT Labs Magazine

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