Clostridium botulinum - Botulism Food Poisoning

Table of Contents

• What is Botulism Food Poisoning?
• Source of Clostridium botulinum contamination
• Clostridium botulinum neurotoxin production
• Clinical manifestation of Botulism Food Poisoning
• Epidemiology of Botulism Food Poisoning
• Botulinum toxin action
• Detection methods of Botulism Food Poisoning

1. Bioassay
2. Immunoassay
3. Endopeptidase assay
4. PCR-based assays

• Treatments of Botulism Food Poisoning
• Control measures of Botulism Food Poisoning

What is Botulism Food Poisoning?

Botulism is a potentially fatal food-borne illness that causes neuroparalysis when botulinum neurotoxins are consumed. (BoNTs). Botulism is a rare disease with a high mortality rate; even a tiny quantity of poison may kill millions of people.

• Clostridium botulinum produces intoxication, which happens as a result of either eating preformed BoNTs in foods or producing BoNTs in the gut.
• Botulinum toxins (Types A–G) are responsible for sickness in humans and animals.
• Food-borne botulism, newborn botulism, and wound botulism are the clinical-epidemiological manifestations of botulism.
• Botulinum toxins A, B, and E have been linked to food-borne botulism in humans.
• BoNTs have a fatal dosage of 0.2g to 2g per kg body weight for humans.
• Botulinum toxin is a potentially harmful biological substance in the environment.

Source of Clostridium botulinum contamination

• C. botulinum is an anaerobic, spore-forming, gramme-positive bacteria whose spores are widely disseminated throughout the environment, including soil, sewage, muck, lakes, sea and ocean sediments, and the intestines of land and aquatic animals.
• Because of the bacterium's ability to spread far, its spores can infect foods where it colonises and creates toxins.
• C. botulinum spores in honey and syrup, the primary cause of newborn botulism.
• Because the spores can resist high heating temperatures during processing, poorly treated canned goods are also a major source of infection.
• Other food sources include smoked, salt-dried, and fermented meat items, as well as numerous varieties of lake and sea fish.
• Type A and B BoTNs are also commonly present in soil and biofertilizers, therefore, they are likely to contaminate vegetables and fruits.
• Beans, baked potatoes, corn, celery, mushrooms, onions and olives that have been improperly cooked.

Clostridium botulinum neurotoxin production

• BoNTs are extracellular proteins weighing roughly 150kDa generated by C. botulinum.
• BoNTs have seven unique serotypes based on bacterial physiology and phylogeny, ranging from Type A to H.
• Based on their proteolytic and non-proteolytic action, the seven toxins are divided into four groups: Groups I, II, III, and IV. 
• Group I proteolytic Types A, B, and F can proliferate at temperatures ranging from 10 to 20°C, whereas Group II non-proteolytic Types B, E, and F may proliferate at 2.5 to 3°C.
• Types C and D are non-proteolytic in Group III, whereas Type G is in Group IV.
• C. botulinum is a stringent anaerobe that can withstand high temperatures. It produces toxins in the gut and causes intoxication when swallowed.

Figure: (a) Molecular processes of natural C2 toxin poisoning. To bind to C2I, heavy chain C2II must be activated by a protease and oligomerized. After receptor-mediated endocytosis, acidification of early endosomes leads C2I to be delivered into the target cell's cytosol via the pore produced by C2IIa oligomers to begin its enzymatic activity. (b) Neural delivery model based on the C2II-C1 and C2It transport systems. C2It is changed separately to contain a conjugated molecule as a payload for delivery through activated C2II-C1. This modular system has the potential to reach targets in the cytosol of the peripheral nerve. The requirements of conjugation and subsequent translocation by C2II-C1 restrict payloads. 

Clinical manifestation of Botulism Food Poisoning

• Within 18 to 38 hours of consumption, symptoms that impact the gastrointestinal tract become noticeable.
• Abdominal discomfort, nausea, vomiting, diarrhea, and constipation are common symptoms.
• Neurological symptoms such as flaccid paralysis, symmetric descending weakness, slurred speech, trouble swallowing, dizziness, impaired vision, dry mouth, and extraocular muscular weakness emerge after intoxication.
• Babies under one year of age are especially affected by baby botulism due to a poorly established gut microbiota where the toxins are generated in the intestine.
• Infants may exhibit signs such as a weak harsh scream, ptosis, poor suck, inability to hold the head erect, and weak muscles.
• In the case of wound botulism, the organism colonises the infected site, such as a wound, abscess, or pus, where Clostridium spores germinate and create toxins.
• Wound botulism symptoms include hypotension, breathing issues, eye muscle paralysis, ataxia, and dilated fixed pupils.
• In the event of a fatality, death generally occurs between 3 to 6 days following intoxication.

Epidemiology of Botulism Food Poisoning

• Botulinum toxin was utilised as a biological agent by the British, American, and Japanese militaries during WWII.
• The largest epidemic to date happened in Egypt in 1991, caused by the eating of Type E toxin found in a typical salted fish dish known as fesaikh, which resulted in 91 instances of an outbreak and 18 deaths.
• The fish was salted and preserved in barrels, which provided an ideal habitat for botulinum toxin development.
• In 2004, an osteopathic physician sought to use botulinum toxin on himself and his girlfriend as a botox. Both were later hospitalised for respiratory failure and required ventilation for several months.
• Approximately 90% of botulinum infections occur in the United States, with 9 to 10 outbreaks reported each year.
• Home-preserved food items, specifically canning, bottling, and preserving food in oil, were linked to those outbreaks.

Botulinum toxin action

• Orally administered botulinum toxin or toxin manufactured in-situ must withstand stomach acid in order to transit into the gut, where it is absorbed and colonises in the duodenum and jejunum.
• The poison then enters the circulation and is taken up by endocytosis before reaching the peripheral nervous system.
• Once within the neuron, its chain is broken, allowing it to function as a zinc-dependent protease and assault SNARE proteins. (soluble N-ethylmaleimide sensitive fusion protein attachment receptor).
• It binds at the nerve-muscle junction and blocks the release of acetylcholine, causing the transmission of impulses to halt and resulting in flaccid paralysis.

Detection methods of Botulism Food Poisoning

1. Bioassay

• The most sensitive and extensively used approach for assessing toxin toxicity is the bioassay using a mouse.
• Symptoms appear in mice 4 hours after treatment.
• The botulinum toxin is characterised by abdominal vibration, wasp-shaped belly, limb paralysis, and trouble breathing.

2. Immunoassay

• The most sensitive immunoassays used to determine the botulinum toxin is ELISA.
• ELISA binds the antigen (toxin) to the antibody (antitoxin) present in the solid surface.
• Then another enzyme-labeled antibody binds to the antigen and the amount of toxin can be determined by an enzymatic reaction.
• The specificity and sensitivity of ELISA are determined by the quality of the anti-serum used.

3. Endopeptidase assay

• Endopeptidase test is an in-vitro assay used to assess the toxin's therapeutic potential. (Eg: Botox, Dysport, Xeomin).
• A bioluminescent signal, fluorescence resonance energy transfer (FRET), and mass spectrometry are used to detect it.
• Endopeptidase tests are more reliable than immunoassays because they only detect active botulinum toxins.

4. PCR-based assays

• On agarose gel electrophoresis, a standard multiplex PCR identifies the particular toxin gene using a high annealing temperature primer. It is capable of detecting toxins of types A, B, E, and F.
• BoNT A, B, and E are detected using real-time PCR.
• Within 1 to 2 hours, fluorescent-based PCR amplifies gene fragments and detects the toxin with excellent sensitivity.

Treatments of Botulism Food Poisoning

• An individual with identical botulism symptoms should be admitted to critical care and given parenteral nourishment.
• If a toxin is detected, antitoxin should be administered within 24 hours for maximum efficiency since it neutralises all free toxins in the bloodstream before they reach the neuromuscular junction.
• Bivalent (AB) or trivalent (ABE) equine immunoglobulins are currently accessible as antitoxins.
• In the instance of wound botulism, medicines such as penicillins should be used to treat the wound.

Control measures of Botulism Food Poisoning

• Foods should be properly heated (above 121°C) and refrigerated (below 4°C).
• Reduced spore development is a benefit of acidifying chemicals like citric acid in canned goods.
• Avoid food that you believe has been improperly reheated or undercooked.