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15.4D: Botulism - Biology

15.4D: Botulism - Biology


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Learning Objectives

  • Compare and contrast the three major modes of entry for Botulinium toxin (infant botulism or adult intestinal toxemia, foodborne botulism, and wound botulism) and describe its mechanism of action

Overview of Botulism

Botulism is a rare, but sometimes fatal, paralytic illness caused by botulinum toxin. It can affect a wide range of mammals, birds and fish. This toxin is a protein produced under anaerobic conditions by the bacterium Clostridium botulinum. The toxin enters the human body in one of three ways: by colonization of the digestive tract by the bacterium in children (infant botulism) or adults (adult intestinal toxemia), by ingestion of toxin from foods (foodborne botulism), or by contamination of a wound by the bacterium (wound botulism). Person-to-person transmission of botulism does not occur. All forms lead to paralysis that typically starts with the muscles of the face and then spreads towards the limbs. In severe forms, it leads to paralysis of the breathing muscles and causes respiratory failure. In light of this life-threatening complication, all suspected cases of botulism are treated as medical emergencies, and public health officials are usually involved to prevent further cases from the same source. Botulism can be prevented by killing the spores by pressure cooking or autoclaving at 121 °C (250 °F) for 30 minutes or providing conditions that prevent the spores from growing. Additional precautions for infants include not feeding them honey.

C. botulinum is an anaerobic, Gram positive, spore-forming rod. Botulinium toxin is one of the most powerful known toxins: about one microgram is lethal to humans. It acts by blocking nerve function (neuromuscular blockade) through inhibition of the release of the excitatory neurotransmitter acetyl choline from the presynaptic membrane of neuromuscular junctions in the somatic nervous system. This causes paralysis. Advanced botulism can cause respiratory failure by paralyzing the muscles of the chest, which can progress to respiratory arrest. In all cases, illness is caused by the botulinium toxin produced by the bacterium C. botulinum in anaerobic conditions, and not by the bacterium itself. The pattern of damage occurs because the toxin affects nerves that fire (depolarize) at a higher frequency first.

MODES OF ENTRY

Three main modes of entry for the toxin are known. The most common form in Western countries is infant botulism. This occurs in small children who are colonized with the bacterium during the early stages of their lives. The bacterium then releases the toxin into the intestine, which is absorbed into the bloodstream. The consumption of honey during the first year of life has been identified as a risk factor for infant botulism and it is a factor in a fifth of all cases. The adult form of infant botulism is termed adult intestinal toxemia, and is exceedingly rare. Foodborne botulism results from contaminated foodstuffs in which C. botulinum spores have been allowed to germinate in anaerobic conditions. This typically occurs in home-canned food substances and fermented uncooked dishes. Given that multiple people often consume food from the same source, it is common for more than a single person to be affected simultaneously. Symptoms usually appear 12–36 hours after eating, but can also appear within 6 hours to 10 days. Wound botulism results from the contamination of a wound with the bacteria, which then secrete the toxin into the bloodstream. This has become more common in intravenous drug users since the 1990s, especially people using black tar heroin and those injecting heroin into the skin rather than the veins

TREATMENT

The only drug currently available to treat infant botulism is Botulism Immune Globulin Intravenous-Human (BIG-IV or BabyBIG). BabyBIG was developed by the Infant Botulism Treatment and Prevention Program at the California Department of Public Health. There are two primary Botulinum Antitoxins available for treatment of wound and foodborne botulism. Trivalent (A,B,E) Botulinum Antitoxin is derived from equine sources utilizing whole antibodies (Fab & Fc portions). This antitoxin is available from the local health department via the CDC. The second antitoxin is heptavalent (A,B,C,D,E,F,G) Botulinum Antitoxin which is derived from “despeciated” equine IgG antibodies which have had the Fc portion cleaved off leaving the F(ab’)2 portions. This is a less immunogenic antitoxin that is effective against all known strains of botulism where not contraindicated. This is available from the US Army.

Key Points

  • The toxin (s) enters the human body by colonization of the digestive tract by the bacterium, by ingestion of toxin from foods or by contamination of a wound by the bacterium.
  • All forms lead to paralysis that typically starts with the muscles of the face and then spreads towards the limbs.
  • Botulism can be prevented by killing the spores by pressure cooking or autoclaving at 121 °C (250 °F) for 30 minutes or providing conditions that prevent the spores from growing.

Key Terms

  • infant botulism: poisoning caused by the toxin from Clostridium botulinum where the gastro-intestinal tract is colonized by spores prior to the protective intestinal bacterial flora having developed
  • spore: A thick resistant particle produced by a bacterium or protist to survive in harsh or unfavorable conditions.
  • botulism: Poisoning caused by the toxin from Clostridium botulinum, a type of anaerobic bacteria that grows in improperly-prepared food.
  • wound botulism: poisoning caused by the toxin from Clostridium botulinum when spores enter a wound under the skin, and, in the absence of oxygen are activated and release toxin
  • toxin: A toxic or poisonous substance produced by the biological processes of biological organisms.

Genomes, neurotoxins and biology of Clostridium botulinum Group I and Group II

Recent developments in whole genome sequencing have made a substantial contribution to understanding the genomes, neurotoxins and biology of Clostridium botulinum Group I (proteolytic C. botulinum) and C. botulinum Group II (non-proteolytic C. botulinum). Two different approaches are used to study genomics in these bacteria comparative whole genome microarrays and direct comparison of complete genome DNA sequences. The properties of the different types of neurotoxin formed, and different neurotoxin gene clusters found in C. botulinum Groups I and II are explored. Specific examples of botulinum neurotoxin genes are chosen for an in-depth discussion of neurotoxin gene evolution. The most recent cases of foodborne botulism are summarised.

Keywords: Botulinum neurotoxin Botulism Clostridium botulinum Genomics Neurotoxin gene cluster.

Copyright © 2014 The Authors. Published by Elsevier Masson SAS.. All rights reserved.


Diagnosis and Treatment

Infant botulism is the most common kind of botulism in the United States. Learn about the diagnosis and treatment of this disease.

Diagnosis

If you or someone you know has symptoms of botulism, immediately see your doctor or go to the emergency room.

Your doctor will ask you questions and examine you to find out the cause of your symptoms. However, these clues are usually not enough for your doctor to diagnose you because some botulism symptoms also occur with other diseases &ndash such as Guillain-Barré syndrome, meningitis, myasthenia gravis, and stroke &ndash and even opioid overdose.

Your doctor may need to order tests to make a diagnosis. Some of these tests are:

  • Brain scan
  • Spinal fluid examination
  • Nerve and muscle function tests (nerve conduction study and electromyography)
  • Tensilon test for myasthenia gravis

If these tests don&rsquot show what is making you sick, your doctor might order laboratory tests to look for the toxin or bacteria that cause botulism. These laboratory tests are the only way to know for certain whether you have botulism. It may take several days to get the results of your tests from the laboratory. If your doctor suspects you have botulism, you may start treatment right away.

Treatment

Botulism is caused by a toxin that attacks the body&rsquos nerves and causes difficulty breathing, muscle paralysis, and even death.

Doctors treat botulism with a drug called an antitoxin, which prevents the toxin from causing any more harm. Antitoxin does not heal the damage the toxin has already done. Depending on how severe your symptoms are, you may need to stay in the hospital for weeks or even months before you are well enough to go home.

If your disease is severe, you may have breathing problems. You may even have respiratory (breathing) failure if the toxin paralyzes the muscles involved in breathing. If that happens, your doctor may put you on a breathing machine (ventilator) until you can breathe on your own. The paralysis caused by the toxin usually improves slowly. The medical and nursing care you receive in the hospital is to help you recover.

People with wound botulism sometimes need surgery to remove the source of the bacteria and may need to take antibiotics.

Survival and Complications

The development of antitoxin and modern medical care means that people with botulism have a much lower chance of dying than in the past, when about 50 in every 100 people with botulism died. Today, fewer than 5 of every 100 people with botulism die.

Even with antitoxin and intensive medical and nursing care, some people with botulism die from respiratory failure. Others die from infections or other problems caused by being paralyzed for weeks or months.

Patients who survive botulism may have fatigue and shortness of breath for years afterward and may need long-term therapy to help them recover.


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Pathogenesis

Transmission:

It is ubiquitous in nature, widely distributed as a saprophyte in soil, animal manure, vegetables, and sea mud. Homemade canned foods, condiments, and fish products are the most common sources of infection with C. botulinum. Ingestion of contaminated honey is the major cause of infant botulism.

Insufficient cooking temperature followed by packaging in anaerobic conditions facilitates the germination of spores and synthesis neurotoxins.

Mechanism of action of Botulinum toxin (BoNT)

Clostridium botulinum is non-invasive. Its pathogenesis is due to the production of powerful neurotoxin ‘botulinum toxin’ (BoNT), probably the most toxic substance known to be lethal to mankind. It produces flaccid paralysis. There are 7 serological types of botulinum neurotoxin labeled as types A, B, C [C1 C2], D, E, F, and G. Human botulism is caused mainly by types A, B, E and F (rarely).

C. botulinum toxin is categorized as a potential bioterrorism agent but botox is in use to smooth facial wrinkles.

After entry (either ingested, inhaled, or produced in a wound), botulinum toxin is transported via the blood to peripheral cholinergic nerve terminals. The most common nerve terminal sites are neuromuscular junctions, postganglionic parasympathetic nerve endings, and peripheral ganglia. It does not affect the CNS.

In normal condition: Upon stimulation of peripheral and cranial nerves, acetylcholine is normally released from vesicles at the neural side of the motor endplate. Acetylcholine then binds to specific receptors on the muscle, inducing contraction.

Mechanism of Botulinum toxin
(Image source: lumenlearning.com)

Botulinum toxin acts by binding to synaptic vesicles of cholinergic nerves, thereby preventing the release of acetylcholine (Ach) at the peripheral nerve endings, including neuromuscular junctions. This results in a lack of stimulus to the muscle fibers, irreversible relaxation of the muscles, and flaccid paralysis.

As botulinum toxin produces flaccid paralysis it can be used therapeutically for the treatment of spasmodic conditions such as strabismus (misaligned eyes), blepharospasm (uncontrollable blinking), and myoclonus.

Clinical manifestations

  1. Diplopia (double vision) or blurring of vision
  2. Dysphagia (difficulty swallowing)
  3. Dysarthria (difficulty in speech) or slurring of speech
  4. Descending symmetric flaccid paralysis of voluntary muscles.
  5. Decreased deep tendon reflexes
  6. Fatigue
  7. Dizziness
  8. Nausea
  9. Constipation
  10. Respiratory muscle paralysis may lead to death.

There is no sensory or cognitive deficits

Types of Botulism

  1. Foodborne botulism: It results from the consumption of foods contaminated with preformed botulinum toxin such as homemade canned food.
  2. Wound botulism: It is a systemic intoxication resulting from the growth of C. botulinum and toxin production in the wounds. It presents like foodborne botulism except for the absence of gastrointestinal features.
  3. Infant botulism: Infant botulism is much milder than the adult version. It results from the ingestion of food (usually honey) contaminated with spores of C. botulinum by children ≤1 year of age. Spores germinate in the intestine, and the vegetative cells secrete botulinum toxin. Clinical manifestations include the inability to suck and swallow, weakened voice, ptosis, floppy neck, and extreme weakness hence called floppy child syndrome. It is a self-limiting disease prognosis is excellent if managed by supportive care and assisted feeding.

Spores do not normally germinate in adult intestine, however may germinate in the intestine of infants.


Contents

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 tolerates traces of oxygen due to the enzyme superoxide dismutase, which is an important antioxidant defense in nearly all cells exposed to oxygen. [5] C. botulinum is able to produce the neurotoxin only during sporulation, which can happen only in an anaerobic environment. C. botulinum is divided into four distinct phenotypic groups (I-IV) and is also classified into seven serotypes (A–G) based on the antigenicity of the botulinum toxin produced. [6] [7]

Groups Edit

Physiological differences and genome sequencing at 16S rRNA level support the subdivision of the C. botulinum species into groups I-IV. [8]

Groups of C. botulinum [8] [9]
Group Serotypes
I (Proteolytic) All type A and proteolytic strains of types B and F
II (Non proteolytic) All type E and nonproteolytic strains of types B and F
III Type C and D
IV Type G

One of the fundamental differences between group I and group II is that C. botulinum group I can lyse native proteins like coagulated egg white, cooked meat particles, whereas group II cannot. [9] However, group II can ferment various carbohydrates like sucrose, mannose. And both of them can degrade the derived protein, gelatin. [9] Human botulism is predominantly caused by group I or II C. botulinum. [9] Group III organisms mainly cause diseases in animals. [9] Group IV C. botulinum has not been shown to cause human or animal disease. [9]

Botulinum toxin Edit

Neurotoxin production is the unifying feature of the species. Eight types of toxins have been identified that are allocated a letter (A–H), several of which can cause disease in humans. They are resistant to degradation by enzymes found in the gastrointestinal tract. This allows for ingested toxin to be absorbed from the intestines into the bloodstream. [3] However, all types of botulinum toxin are rapidly destroyed by heating to 100 °C for 15 minutes (900 seconds). Botulinum toxin, one of the most poisonous biological substances known, is a neurotoxin produced by the bacterium Clostridium botulinum. C. botulinum elaborates eight antigenically distinguishable exotoxins (A, B, C1, C2, D, E, F and G). [ citation needed ]

Most strains produce one type of neurotoxin, but strains producing multiple toxins have been described. C. botulinum producing B and F toxin types have been isolated from human botulism cases in New Mexico and California. [10] The toxin type has been designated Bf as the type B toxin was found in excess to the type F. Similarly, strains producing Ab and Af toxins have been reported. [ citation needed ]

Evidence indicates the neurotoxin genes have been the subject of horizontal gene transfer, possibly from a viral (bacteriophage) source. This theory is supported by the presence of integration sites flanking the toxin in some strains of C. botulinum. However, these integrations sites are degraded (except for the C and D types), indicating that the C. botulinum acquired the toxin genes quite far in the evolutionary past. Nevertheless, further transfers still happen via the plasmids and other mobile elements the genes are located on. [11]

Botulinum toxin types Edit

Only botulinum toxin types A, B, E, F and H cause disease in humans. Types A, B, and E are associated with food-borne illness, with type E specifically associated with fish products. Type C produces limber-neck in birds and type D causes botulism in other mammals. No disease is associated with type G. [12] The "gold standard" for determining toxin type is a mouse bioassay, but the genes for types A, B, E, and F can now be readily differentiated using quantitative PCR. [13] As no antitoxin to type H is yet available, discovered in 2013 and by far the deadliest, details are kept under shroud. [14]

A few strains from organisms genetically identified as other Clostridium species have caused human botulism: C. butyricum has produced type E toxin [15] and C. baratii had produced type F toxin. [16] [17] The ability of C. botulinum to naturally transfer neurotoxin genes to other clostridia is concerning, especially in the food industry, where preservation systems are designed to destroy or inhibit only C. botulinum but not other Clostridium species. [ citation needed ]

Phenotypic groups of Clostridium botulinum
Properties Group I Group II Group III Group IV
Toxin Types A, B, F B, E, F C, D G
Proteolysis + weak
Saccharolysis +
Disease host human human animal
Toxin gene chromosome/plasmid chromosome/plasmid bacteriophage plasmid
Close relatives C. sporogenes
C. putrificum
C. butyricum
C. beijerinickii
C. haemolyticum
C. novyi type A
C. subterminale
C. haemolyticum

Laboratory isolation Edit

In the laboratory, C. botulinum is usually isolated in tryptose sulfite cycloserine (TSC) growth medium in an anaerobic environment with less than 2% oxygen. This can be achieved by several commercial kits that use a chemical reaction to replace O2 with CO2. 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, characteristics important for biochemical identification. [18]

C. botulinum was first recognized and isolated in 1895 by Emile van Ermengem from home-cured ham implicated in a botulism outbreak. [19] The isolate was originally named Bacillus botulinus, after the Latin word for sausage, botulus. ("Sausage poisoning" was a common problem in 18th- and 19th-century Germany, and was most likely caused by botulism.) [20] However, isolates from subsequent outbreaks were always found to be anaerobic spore formers, so Ida A. Bengtson proposed that the organism be placed into the genus Clostridium, as the genus Bacillus was restricted to aerobic spore-forming rods. [21]

Since 1959, all species producing the botulinum neurotoxins (types A–G) have been designated C. botulinum. Substantial phenotypic and genotypic evidence exists to demonstrate heterogeneity within the species. This has led to the reclassification of C. botulinum type G strains as a new species, C. argentinense. [22]

Group I C. botulinum strains that do not produce a botulin toxin are referred to as C. sporogenes. [23]

The complete genome of C. botulinum has been sequenced at Wellcome Trust Sanger Institute in 2007. [24]

Foodborne botulism Edit

"Signs and symptoms of foodborne botulism typically begin between 18 and 36 hours after the toxin gets into your body, but can range from a few hours to several days, depending on the amount of toxin ingested." [25]

  • 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 Edit

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: [25]

  • Difficulty swallowing or speaking
  • Facial weakness on both sides of the face
  • Blurred or double vision
  • Dropping eyelids
  • Trouble breathing
  • Paralysis

Infant botulism Edit

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 [25]

Beneficial effects of botulinum toxin Edit

Purified botulinum toxin is diluted by a physician for treatment:

  • Congenital pelvic tilt
  • Spasmodic dysphasia (the inability of the muscles of the larynx)
  • Achalasia (esophageal stricture)
  • Strabismus (crossed eyes)
  • Paralysis of the facial muscles
  • Failure of the cervix
  • Blinking frequently
  • Anti-cancer drug delivery [26]

Adult intestinal toxemia Edit

A very rare form of botulism that occurs by the same route as infant botulism but is among adults. Occurs rarely and sporadically. Signs and symptoms include:

  • Abdominal pain
  • Blurred vision
  • Diarrhea
  • Imbalance
  • Weakness in arms and hand area [27]

A number of quantitative surveys for C. botulinum spores in the environment have suggested a prevalence of specific toxin types in given geographic areas, which remain unexplained. [ citation needed ]

North America Edit

Type A C. botulinum predominates the soil samples from the western regions, while type B is the major type found in eastern areas. [28] The type-B organisms were of the proteolytic type I. Sediments from the Great Lakes region were surveyed after outbreaks of botulism among commercially reared fish, and only type E spores were detected. [29] [30] [31] In a survey, type-A strains were isolated from soils that were neutral to alkaline (average pH 7.5), while type-B strains were isolated from slightly acidic soils (average pH 6.23). [ citation needed ]

Europe Edit

C. botulinum type E is prevalent in aquatic sediments in Norway and Sweden, [32] Denmark, [33] the Netherlands, the Baltic coast of Poland, and Russia. [28] The type-E C. botulinum was suggested to be a true aquatic organism, which was indicated by the correlation between the level of type-E contamination and flooding of the land with seawater. As the land dried, the level of type E decreased and type B became dominant. [ citation needed ]

In soil and sediment from the United Kingdom, C. botulinum type B predominates. In general, the incidence is usually lower in soil than in sediment. In Italy, a survey conducted in the vicinity of Rome found a low level of contamination all strains were proteolytic C. botulinum types A or B. [34]

Australia Edit

C. botulinum type A was found to be present in soil samples from mountain areas of Victoria. [35] Type-B organisms were detected in marine mud from Tasmania. [36] [ verification needed ] Type-A C. botulinum has been found in Sydney suburbs and types A and B were isolated from urban areas. In a well-defined area of the Darling-Downs region of Queensland, a study showed the prevalence and persistence of C. botulinum type B after many cases of botulism in horses. [ citation needed ]

C. botulinum is used to prepare the medicaments Botox, Dysport, Xeomin, and Neurobloc used to selectively paralyze muscles to temporarily relieve muscle function. It has other "off-label" medical purposes, such as treating severe facial pain, such as that caused by trigeminal neuralgia.

Botulinum toxin produced by C. botulinum is often believed to be a potential bioweapon as it is so potent that it takes about 75 nanograms to kill a person (LD50 of 1 ng/kg, [37] assuming an average person weighs

75 kg) 1 kilogram of it would be enough to kill the entire human population. For comparative purposes, a quarter of a typical grain of sand's weight (350 ng) of botulinum toxin would constitute a lethal dose for humans.

A "mouse protection" or "mouse bioassay" test determines the type of C. botulinum toxin present using monoclonal antibodies. An enzyme-linked immunosorbent assay (ELISA) with digoxigenin-labeled antibodies can also be used to detect the toxin, [38] and quantitative PCR can detect the toxin genes in the organism. [13]

C. botulinum is a soil bacterium. The spores can survive in most environments and are very hard to kill. They can survive the temperature of boiling water at sea level, thus many foods are canned with a pressurized boil that achieves even higher temperatures, sufficient to kill the spores. [ citation needed ] This bacteria is widely distributed in nature, and can be assumed to be present on all food surfaces. Its optimum growth temperature is within the mesophilic range. In spore form, it is a heat resistant pathogen that can survive in low acid foods and grow to produce toxin. The toxin attacks the nervous system and will kill an adult at a dose of around 75 ng. [37] This toxin is detoxified by holding food at 100 °C for 10 minutes. [ citation needed ]

Botulism poisoning can occur due to preserved or home-canned, low-acid food that was not processed using correct preservation times and/or pressure. [39] Growth of the bacterium can be prevented by high acidity, high ratio of dissolved sugar, high levels of oxygen, very low levels of moisture, or storage at temperatures below 3 °C (38 °F) for type A. For example, in a low-acid, canned vegetable such as green beans that are not heated enough to kill the spores (i.e., a pressurized environment) may provide an oxygen-free medium for the spores to grow and produce the toxin. However, pickles are sufficiently acidic to prevent growth even if the spores are present, they pose no danger to the consumer. Honey, corn syrup, and other sweeteners may contain spores, but the spores cannot grow in a highly concentrated sugar solution however, when a sweetener is diluted in the low-oxygen, low-acid digestive system of an infant, the spores can grow and produce toxin. As soon as infants begin eating solid food, the digestive juices become too acidic for the bacterium to grow. [ citation needed ]

The control of food-borne botulism caused by C. botulinum is based almost entirely on thermal destruction (heating) of the spores or inhibiting spore germination into bacteria and allowing cells to grow and produce toxins in foods. Conditions conducive of growth are dependent on various environmental factors. Growth of C. botulinum is a risk in low acid foods as defined by having a pH above 4.6 [40] although growth is significantly retarded for pH below 4.9. There have been some cases and specific conditions reported to sustain growth with pH below 4.6. [41] [42]

Physicians may consider the diagnosis of botulism based on a patient's clinical presentation, which classically includes an acute onset of bilateral cranial neuropathies and symmetric descending weakness. [43] [44] Other key features of botulism include an absence of fever, symmetric neurologic deficits, normal or slow heart rate and normal blood pressure, and no sensory deficits except for blurred vision. [45] [46] A careful history and physical examination is paramount in order to diagnose the type of botulism, as well as to rule out other conditions with similar findings, such as Guillain-Barre syndrome, stroke, and myasthenia gravis. Depending on the type of botulism considered, different tests for diagnosis may be indicated.

Foodborne botulism: serum analysis for toxins by bioassay in mice should be done, as the demonstration of the toxins is diagnostic. [47]

Wound botulism: isolation of C. botulinum from the wound site should be attempted, as growth of the bacteria is diagnostic. [48]

Adult enteric and infant botulism: isolation and growth of C. botulinum from stool samples is diagnostic. [49] Infant botulism is a diagnosis which is often missed in the emergency room.

Other tests that may be helpful in ruling out other conditions are:

    (EMG) or antibody studies may help with the exclusion of myasthenia gravis and Lambert-Eaton myasthenic syndrome (LEMS). [50]
  • Collection of cerebrospinal fluid (CSF) protein and blood assist with the exclusion of Guillan-Barre syndrome and stroke. [51]
  • Detailed physical examination of the patient for any rash or tick presence helps with the exclusion of any tick transmitted tick paralysis. [52]

In the case of a diagnosis or suspicion of botulism, patients should be hospitalized immediately, even if the diagnosis and/or tests are pending. If botulism is suspected, patients should be treated immediately with antitoxin therapy in order to reduce mortality. Immediate intubation is also highly recommended, as respiratory failure is the primary cause of death from botulism. [53] [54] [55]

In Canada, there are currently only 3 antitoxin therapies available, which are accessible through Health Canada Special Access Program (SAP). [56] The 3 types of antitoxin therapies are: 1) GlaxoSmithKline trivalent Types ABE, 2) NP-018 (heptavalent) Types A to G, and 3) BabyBIG®, Botulism Immune Globulin Intravenous (Human) (BIG-IV) for pediatric patients under the age of one year. [57]

Outcomes vary between one and three months, but with prompt interventions, mortality from botulism ranges from less than 5 percent to 8 percent. [58]

There used to be a formalin-treated toxoid vaccine against botulism (serotypes A-E), but it was discontinued in 2011 due to declining potency in the toxoid stock. It was originally intended for people at risk of exposure. A few new vaccines are under development. [59]


Streaming

The transmission of botulism occurs mainly through the consumption of contaminated food and untreated water. In addition, the causative bacteria can reach the body through injuries.

Note that botulism is not a contagious disease, so it is not transmitted between people.

Types

There are several types of botulism, namely:

  • Infant botulism: also called lactating botulism or intestinal botulism, this type of disease affects babies less than a year old. The bacteria reaches the gastrointestinal system, causing abdominal pain and constipation.
  • Food botulism : transmitted by contaminated food, especially canned (canned) that are expired, for example, meats, hearts of palm, pickles, honey, etc. This is because this rod-shaped bacteria can survive in environments with little oxygen (anaerobic bacillus).

Can of expired and rusty canning

  • Wound botulism : skin lesions can be favorable for the contamination of the bacteria that causes botulism. Through the released toxins, this type can cause serious skin infections. Injecting drug users are highly prone to contamination.

Canine Botulism

Canine botulism can manifest in domestic animals such as dogs. These animals contract the disease through the consumption of canned food, garbage, contaminated water and carcasses of dead animals.

The symptoms are very similar to those of humans, for example, facial paralysis, weakness and difficulty in swallowing. In most cases, dogs die.

Bovine Botulism

Botulism in ruminants, also called “fallen cow disease”, affects the animal’s central nervous system causing paralysis.

This usually occurs when these animals ingest the spores of the disease-causing bacteria in pastures.

Bovine botulism can also occur with ingestion of contaminated water or due to the conditions of the environment in which they live. In Brazil, it has been a major cause of cattle deaths.


Definition of Botulism

Botulism refers to a clinical condition during which the neurotoxin of Clostridium botulinum paralyzes the involuntary muscles by inhibiting the motor nerve endings from synthesizing acetylcholine. Clostridium botulinum commonly releases neurotoxin (Botulinum neurotoxin), which accumulates in the small intestine. A botulinum neurotoxin mainly affects the nervous system and paralyzes the involuntary muscles of the body. Botulism can occur in both humans and animals.

Causes of Botulism

Botulism is caused by a neurotoxin produced by Clostridium botulinum. Neurotoxin is an exotoxin, which affects the nervous system. Botulism is a clinical condition, paralyzes the involuntary muscles and further spreads to the respiratory system and the heart. Based on serological specificity, neurotoxins are categorized into the following seven serological types:

  1. Type A: It is a type of human botulism, which is very common in the western regions of the United States.
  2. Type B: It is also a type of human botulism that occurs frequently, and it is less toxic than type-A.
  3. Type-C: It is not a type of human botulism and causes botulism in cattle, fowls etc.
  4. Type-D: It mostly causes forage poisoning of cattle in South African countries.
  5. Type-E: This type of neurotoxin is obtained chiefly from fish and fish products. Type-E is very toxic to humans.
  6. Type-F: It was first isolated in Denmark and is a type of human botulism.
  7. Type-G: This has been recently isolated from the soil in Argentina, and it is not a type of human botulism.

Food Sources

The spores of Clostridium botulinum sporulate to produce a neurotoxin, which can be present in the following food sources:

  • Canned meat
  • Canned fish
  • Honey syrup
  • Sweet corn
  • Fermented beans
  • Other low acid foods

Spores of Clostridium botulinum generally grows in favourable conditions like:

  1. Low acid
  2. Anaerobic environment
  3. Low sugar
  4. Low salt
  5. And low temperature.

Types of Botulism

There are three forms of botulism caused by Clostridium botulinum:

Foodborne botulism: It spreads by eating food contaminated with the spores produced by the Clostridium botulinum. Foodborne botulism develops after the consumption of unprocessed, undercooked, canned and unrefrigerated food.

  • Symptoms include:
    • Blurred vision
    • Difficulty in swallowing
    • Muscle weakness
    • Dry mouth
    • Nausea
    • Vomiting
    • Abdominal cramps
    • Paralysis etc.

    Wound botulism: It spreads when the spores of the bacteria (Clostridium botulinum) enters the body through a cut or any mechanical injuries. The spores of Clostridium botulinum sporulate to produce neurotoxin within the wound. The risk factors include people who are addicted to drugs like heroin.

    • Symptoms include:
      • Slurred speaking
      • Double vision
      • Drooping eyelids
      • Paralysis etc.

      Infant botulism: It spreads by the ingestion of the Clostridium botulinum spores that grow in the infant’s intestinal tract of age limit between 2-8 months. Infant botulism develops after the consumption of honey syrup.

      • Symptoms include:
        • Constipation
        • Floppy movements
        • Irritation
        • Drooping eyelids
        • Paralysis etc.

        In addition to this, there are some other types also that occur very rarely. Adult botulism is rarely seen, in which the spores of the Clostridium botulinum colonize in the digestive tract of adults. Iatrogenic botulism is very lethal, which occurs by an overdose of botulinum neurotoxin or botox.

        Types of Neurotoxin Effects

        Botulinum neurotoxin can produce the following type of effects:

        1. Local effect: The local effects of Clostridium botulinum includes the following symptoms:
          • Blurred vision
          • Muscle weakness
          • Dry mouth
          • Reduced gag reflex etc.
        2. Immunological effect: It occurs by the administration of equine antitoxin.
        3. Metabolic effect: It is characterized by a symptom (respiratory acidosis).

        Symptoms of Botulism disease

        A neurotoxin produced by Clostridium botulinum can affect the cardiovascular, respiratory and central nervous system etc.

        1. Effect on the cardiovascular system: Botulinum neurotoxin blocks the automatic nervous system and leads to cause tachycardia and hypertension.
        2. On respiratory system: A neurotoxin paralyzes the respiratory muscle and results in ventilatory failure, and ultimately leads to death.
        3. Effect on the nervous system:
          • Central nervous system: Botulinum neurotoxin paralyzes the cranial nerves and causes symptoms like:
            • Blurred vision
            • Diplopia
            • Dysphonia
            • Dysarthria
            • Dysphagia etc.
          • Peripheral nervous system: After paralyzing cranial nerves, botulinum neurotoxin causes symmetrical descending paralysis, affecting the respiratory muscles.
          • Automatic nervous system: Botulinum neurotoxin blocks the autonomic cholinergic junctions and causes the following symptoms:
            • Blurred vision
            • Orthostatic hypotension
            • Constipation
            • Urinary retention etc.
        4. Skeletal and smooth muscles: Botulinum neurotoxin affects the skeletal and smooth muscles by causing gallbladder dysfunction and necrotic fasciitis.
        5. Effect on Gastrointestinal system: A neurotoxin shows the gastrointestinal symptoms after 18-36 hours of incubation, and the symptoms include:
          • Nausea
          • Vomiting
          • Abdominal cramps
          • Diarrhoea
          • It sometimes leads to cause constipation, gastric dilatation and paralytic ileus.
        6. Neurotoxin does not directly affect the hepatic system (liver) and the urinary bladder.

        Pathogenesis

        A neurotoxin produced by the Clostridium botulinum causes botulism by following a given pathogenic cycle:

        1. First, neurotoxin enters the bloodstream through the ingestion of the bacterial spores or from the mucosal surface.
        2. Botulinum neurotoxin blocks the nerve terminal ends.
        3. Then, it binds to the neuronal membrane.
        4. After that, a neurotoxin enters the cytoplasm of the axon terminal.
        5. Then, botulinum neurotoxin blocks the excitatory synaptic transmission and results in interference with the synthesis of neurotransmitter “Acetylcholine”.
        6. Finally, it causes flaccid paralysis, as the nervous system cannot function without the action of acetylcholine. Acetylcholine plays an essential role in producing nerve impulse and the process of muscle contraction.

        Prognosis

        There are many cases of botulism, and if it remains untreated, then the mortality rate reaches about 50-60%. Treated patients may also concur with botulism, with a mortality rate of about 3-5%. The diagnosis or treatment of botulism should be made early otherwise, it can cause major outbreaks.

        Prevention

        There are a few first aid measures that can control the transmission of the disease to some extent.

        Foodborne botulism: To prevent its transmission

        • Avoid eating canned food
        • Properly wash and boil the vegetables before eating.
        • Trash the canned food that is bulging or leaking.
        • Avoid eating foods that are added with preservatives.
        • Avoid eating undercooked food.

        Wound botulism: To prevent its transmission

        • Properly sanitize the wounded area.
        • Maintain proper body hygiene.
        • Do not let the wound open to the air, as the spores of Clostridium botulinum can penetrate inside the cells through any cuts or injuries.

        Infant botulism: To prevent infant botulism, keep the body hygienic and don’t give corn or honey syrup to the infants.

        Treatment

        The early symptoms of botulism can be treated by the oral administration of polyvalent antitoxin. Enemas are found effective in the removal of the unabsorbed toxin.

        Diagnosis

        Diagnosis of botulism involves the following tests:

        Mouse inoculation test: It is the most reliable and popular method to detect the presence of C.botulinum. Mouse inoculation test involves two series of tests. In one experiment, a patient’s stool is injected into the peritoneal cavity of mice. And, in the second test, an equal amount of the patient’s stool plus multivalent antitoxin is injected into the other mice.

        If the mice live, by the injection of antitoxin and stool sample and die by the infusion of an untreated serum sample, then it gives a positive result for botulism. Mouse inoculation test detects the presence of particularly A, B and E serotypes of C. botulinum.

        Physical tests: It includes tests like brain scan, spinal fluid examination, nerve conduction test (electromyography).


        Mode of VAMP substrate recognition and inhibition of Clostridium botulinum neurotoxin F

        Clostridium botulinum neurotoxins (BoNTs) cleave neuronal proteins responsible for neurotransmitter release, causing the neuroparalytic disease botulism. BoNT serotypes B, D, F and G cleave and inactivate vesicle-associated membrane protein (VAMP), each at a unique peptide bond. The specificity of BoNTs depends on the mode of substrate recognition. We have investigated the mechanism of substrate recognition of BoNT F by determining the crystal structures of its complex with two substrate-based inhibitors, VAMP 22-58/Gln58 D -cysteine and 27-58/Gln58 D -cysteine. The inhibitors bind to BoNT F in the canonical direction (as seen for BoNTs A and E substrates) but are positioned specifically via three major exosites away from the active site. The cysteine sulfur of the inhibitors interacts with the zinc and exists as sulfinic acid in the inhibitor VAMP 27-58/Gln58 D -cysteine. Arg133 and Arg171, which form part of two separate exosites, are crucial for substrate binding and catalysis.


        Future directions

        Although some progress has been made in recent years, identification and characterization of the protein receptors for the BoNTs and determination of the mechanism of specificity of CNT binding domains for their receptors is an outstanding problem. Further, understanding the mechanism of LC translocation and activation within the motorneuron, including the effects of pH on the tertiary structures of BoNTs, will be crucial for rational design of engineered BoNT therapeutics. Further structural studies on the endopeptidase domains of BoNTs, including the structural basis behind BoNT substrate specificity, might lead to the development of serotype-specific inhibitors.

        It has been proposed that the extreme neurospecificity of BoNT heavy chains could be applied to deliver engineered molecule in to nerve cells. This can be achieved by the replacement of light chain with desired therapeutic agent that could be reached in the nerve endings without iatrogenic complications which might otherwise occur 182 . Use of fragments of BoNT for the therapeutics of the future is also exciting. For example, harnessing the properties of the BoNT LC endopeptidase fragments for the creation of a range of �signer’ therapeutics is a real possibility following the successful retargeting of the LC/A domain to cells of neuronal and non-neuronal origin 183 . Additionally, the ability of BoNTs to transport large polypeptides across the membranes could be harnessed for the delivery of biopharmaceuticals to cytosolic targets 184 . Derivatives of BoNT/A and BoNT/B can target compounds specifically to human neuroblastoma cells. The therapeutic potential of clostridial toxins is not limited to the neurotoxin for the inhibition of neurotransmitter release, but also has potential as an anticancer drug 62 . The technology termed 𠆌lostridia directed enzyme pro-drug therapy’ (CDEPT) in which intravenously injected clostridial spores are used to target hypoxic regions of solid tumours. Spores get localized to solid tumours exclusively for germination, as they cannot grow in healthy tissues. Genetic modification of the clostridial host to express anti cancer compounds or pro-drug converting enzymes (as in CEDPT), has the potential to lead the localized destruction of solid tumour tissue.

        Botulinum neurotoxins are of great interest to the medical and scientific communities. Despite causing disease, they have become valuable research tools and have wide-ranging applications as pharmaceuticals. As the structure and mechanism of action of the toxins are further dissected, the development of vaccines, serotype-specific inhibitors and novel therapeutics will undoubtedly follow.