Botulinum toxins

Botulinum toxins are the produce of the Clostridium Botulinum species, a gram positive bacillus (rod), anaerobic, spore forming, which is found everywhere in the environment. The name Botulinum derives from the Latin botulus, which means sausage. Botulism, the disease caused by the toxins, it is originally known in fact from consumption of contaminated, not enough deeply cooked sausages. The contamination of sausages would happen because of the ubiquitous presence of the organism and lack of elementary principle of hygiene. In other words the maker of the sausage would contaminate just by not washing their hands, or working on the minced meat on or over contaminated surfaces. The spore would germinate in anaerobic environment such as within the sausage and in turn the organism would produce the toxins. This has been also a frequent happening at the origin of tinned food industry, when gas would form within a tin this was an indication of the germination of the organism. Toxins do not have any particular flavor or odor, and germination of the bacillus and gas formation is also not smelling, opposite to many anaerobes. Some of the toxins digest food giving to it a spoiled appearance, some do not leaving the food unchanged. Some other Clostridium species (Clostridium butyricum -of the butter- and Clostridium barati) have been known to produce toxins.
Botulinum toxins, there are seven of them, are neurotoxins, they work at the neuromuscular junction and cause generalised paralysis. The clinical picture it is a paralysis beginning in the muscles of the head and then progressing in succession to the neck, upper arms, body trunk and legs. Death supervenes by stop breathing and suffocation.
Toxins are deactivated by home cooking at 100 degrees for ten minutes. However is well know that a variety of food boiled for ten minutes do not reach in the inner part that temperature. The boiling for ten minutes at 100 degrees is therefore a notion valid for fully exposed toxins. It toxins are deeply endowed in a large sausage, as it is possible, the roasting of the sausage, or even its boiling for ten minutes is not to be expected to deactivate the toxins. Spores are killed at 120 degrees for twenty minutes (steam sterilization or pressure cooking) and this procedure is deactivating toxins as well. Toxins are of course resistant to gastric content.
Clinical botulism is traditionally described in four types,
-the first is food borne poisoning, caused by the presence of toxins in food. However the presence of toxin does not mean toxins are introduced in the food, but rather that there were spore and anaerobic condition in the food, spore would germinate because of the anaerobic conditions, and toxin would be produced within the food
-second is wound botulism, the bacillus would contaminate a wound and produce toxin
-infant botulism caused by ingestion of spore and germination in the intestine and toxin production
-undetermined, including some cases of adult with the same mechanism as for infant type.
How a botulinum attack could take place is difficult to say, toxins are relatively heat resistant but no information are available on activity in relation to dry condition. It is not known if they were planned to be used in the alimentary chain or just spread as such in form of aerosol. A possibility would be spreading of botulinum toxins in the water. However these toxins are rapidly inactivated by standard potable water treatment such as chlorination and aeration. It is conceivable though that toxins could remain active for several days in untreated water. Opposite to other agents there are no reports of industrial accidents or terrorist attacks nor of epidemic of paralysis which could be attributed to accidental or not mass spreading of the toxins. All industrial accidents, the last reported caused by toxins present in the thick cream industrially processed, have resulted in one or two death.

The incubation Period
Incubation is the time between assumption of the toxin and the beginning of symptoms. The rapidity of onset and severity of symptoms depend on the rate and amount of toxin absorption. Food borne botulism may give symptoms as early as two hours and as late as 8 (eight) days after ingestion of toxins. Typically cases presents 12 to 72 hours after the implicated meal. In large food borne outbreak, new cases presented during the ensuing 3 days at a fairly even rate before decreasing.
The time to onset of inhalation botulism cannot be stated with certainty. In experimental animals, monkeys showed signs of botulism 12 to 80 (eighty) hours after aerosol exposure four to seven time the monkey median lethal dose. The three known human cases of inhalation botulism had onset of symptoms approximately 72 hours after exposure to an unknown but probably small amount of aerosolized toxin (J.A.M.A.) Like in untreated water, is conceivable that toxin could remain active in aerosol for at least a few days.
One can therefore presume two things

  1. in case of use in the alimentary chain there could be an epidemic of afebrile symmetric descending paralysis without sensory deficit, with some death within 24h and other cases, presenting according to the timing described, beginning with diplopia, then dysartria and or dysphagia, then progressive weakness to involve neck muscle then arms then thorax then legs. Same for, if at all possible, air contamination on large number of people.
  2. Sporadic cases of the above described afebrile symmetric descending paralysis without sensory deficit, here and there caused by sporadic use here and there of the toxins in the food or, if at all possible, in the air
Though the afebrile symmetric descending paralysis is the predominant symptoms, associated to these there can be dry mouth and throat, nausea, vomiting and abdominal pain, as well as dizziness, blurred vision if not diplopia as said above, and paralytic ileus. Constipation and urinary retention are common.
Because of the mechanism described there is not place for antibiotic use in case of a botulinum toxins terrorist attack. Nor there is any antidote. Botulinum toxins are proteins acting in the presynaptic nerve ending expansion of the acetylcholine mediated (cholinergic) junction to prevent release of acetylcholine and block neurotransmission. The blockade is most evident clinically in the cholinergic autonomous nervous system (causing nausea vomit constipation paralytic ileus and urinary retention) and in the neuromuscular junction (causing the descending flaccid muscular paralysis).
Precisely the toxins attack the membrane of the small vesicle containing the neurotransmitter acetylcholine stored in the nerve ending and somehow inhibit its release-crossing in to the synaptic space to reach the receptor . No acetylcholine is consequently available for the normal muscle function and activity and the normal tone and function maintenance of secretion, intestinal motion, urinary motion etc. It is like if all those cholinergic nerves were cut.
The attack to the presynaptic membrane causes a damage which was thought to be irreversible. In fact pharmacological preparation of Botulinum toxins are used therapeutically to release the muscle contracture in strabismus, blepharospasm, and torticollus, all conditions caused by pathological muscle chronic contraction. In these case a defined amount of toxin is injected in to the muscle to intentionally damage the mechanism of acetylcholine release so that the contracted muscle becomes flaccid (de-contracted). Though the damage to the nerve ending (presynaptic) mechanism is probably irreversible, new nerve ending can sprout anew (axon regeneration) so that neurotransmission can be restored after sprouting takes place. This is assumed because clinically some patients have been kept for months in ventilator and then their condition have cleared and muscular function restored. Also from the therapeutic experience is known that often injection in to the muscle have to be repeated after few months.
The only defense are serum antitoxin and vaccination. Serum antitoxin is obtained from horses sensitized. Equine antitoxin should be given to suspected cases, whether from ingested toxin or aerosol inhaled toxins. It is apparently of some efficacy even after symptoms have developed. After testing for horse serum hypersensitivity, as for all equine type antitoxin serum, one dose of trivalent antitoxin is given IM and one IV. Trivalent refers to three of the toxins, the prevalent ones (called A, B and E). Equine antitoxin is useless in infant type botulism, probably because of the continuous toxin production. An heptavalent equine antitoxin has recently been made available (versus type A,B,C, D, E F and G toxins) but not extensively tested. An experimental human serum antitoxin is under investigation. Needles to say neither are presently available in North Iraqi Kurdistan. Vaccine is still on experimental phase, is a is a pentavalent toxoid , that is similar to tetanus vaccine, should be given at 0 week, then 2 then 12 weeks thereafter, then one year after. This is presently indicated for selected groups at high risk of exposure. Is not presently available.
Affected cases could be placed and kept on ventilator after tracheostomy and they should spontaneously recover after several weeks or months.

Note on the Sodium Hypochlorite (bleach) solutions:

While concentration of 0,025 of bleach in water (5 cc of bleach per liter of water) are the one used for infected wound irrigation because they have bactericidal property and no tissue toxicity, (J Burn Care Rehabil 1991 Sep-Oct, 12(5) 420-4), this concentration is irritating eyes and mucosae. A more diluted one should be used , 1 to 2 cc per liter (0,005 to 0,01), when the purpose is purely decontamination of eyes from both mustard and nerve agents. Conversely a more concentrated solution (10 cc of bleach per liter of water ) can be used in recently contaminated intact or superficially burned skin, followed by water rinsing. These dilution are referred to normal commercial preparation of bleach, which have a sodium hypochlorite concentration of around 5%. More diluted preparation should be used accordingly. If the commercial preparation is said to be 2,5% the given amount of bleach should be doubled. More concentrated commercial preparation should conversely diluted more. When in doubt the prepared concentration should be tested in the mouth, the 0,05 concentration is definitely felt strongly in the oral mucosa, and is revolting to drink, while the 0,005 should still give a flavor-taste of diluted chlorine as for instance in a swimming pool water.
CDC gives these approximate relation between sodium hypochlorite concentration and ppm (part per millions): 500 ppm correspond to a dilution 1:100 of household bleach, equal to 10 cc of 5% bleach in one liter of water. 5000 ppm is 100 cc of 5% bleach per liter of water. These concentrations are relevant for sterilization or disinfection purposes (Clinical Microbiology Reviews Oct 1997, p.597-610).
By contrast drinkable water has a chlorine concentration of 0,5 ppm (dilution of household bleach of 0,01 cc per liter of water). Depending on the bacterial or viral contamination of water to be chlorinated, concentration of 0,1 ppm to 0,5 ppm, or up to 2,0 ppm of free chlorine are reached. Some time this is expressed in weight by volume since water chlorination is done with solid chlorine releasing substances. So the Sphere project indications for drinkable water at the tap are of 0,2 mg to 0,5 mg of residual free chlorine per liter of water. One should remember that there are substantially three methods of water potabilization (Chlorination). One is the addition of Calcium hypochlorite solution, which will then be given as a final concentration of weight by volume. A slight confusion may arise from the fact that Calcium hypochlorite, a powder, is said to give around 70% of free chlorine when dissolved in water. It is then better to refer to the actual residual free chlorine (residual refer to the chlorine that has not interacted with organic material) when talking of final free chlorine concentration. At any rate talking abstractly about units of measurement, 1 (one) mg per liter (1 mg/L) is the same than 1 (one) part per million (ppm). This is because one milligram is = one thousandth of a gram, while a liter (of distilled water) is = to thousand grams. So 1000 x 1000 = 1,000,000. Calcium hypochlorite is the most commonly used. A second one is chlorine gas, which is becoming more and more obsolete. A third one is Sodium hypochlorite, which comes in liquid form (bleach) because the method to produce it was originally from salted water electrolysis (H2O + NaCl------ NaOCl). For practical purpose only Calcium hypochlorite (powder or tablets), or Sodium hypochlorite solution (bleach) are used in emergency situation.

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