clostridium botulinum clostridium botulinum clostridium botulinum is a gram-positive, rod-shaped, anaerobic, spore-forming, motile bacterium with the ability to produce the neurotoxin botulinum. clostridium botulinum clostridium botulinum stained with gentian violet. scientific classification domain: bacteria phylum: bacillota class: clostridia order: eubacteriales family: clostridiaceae genus: clostridium species: c. botulinum binomial name clostridium botulinumvan ermengem, 1896 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. c. botulinum is responsible for foodborne botulism (ingestion of preformed toxin), infant botulism (intestinal infection with toxin-forming c. botulinum), and wound botulism (infection of a wound with c. botulinum). c. botulinum produces heat-resistant endospores that are commonly found in soil and are able to survive under adverse conditions. c. botulinum is commonly associated with bulging canned food; bulging, misshapen cans can be due to an internal increase in pressure caused by gas produced by bacteria. microbiology 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. 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. groups physiological differences and genome sequencing at 16s rrna level support the subdivision of the c. botulinum species into groups i-iv. groups of c. botulinum groupserotypes 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 iiitype c and d ivtype 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. however, group ii can ferment various carbohydrates like sucrose and mannose, and both of them can degrade the derived protein, gelatin. human botulism is predominantly caused by group i or ii c. botulinum. group iii organisms mainly cause diseases in non-human animals. group iv c. botulinum has not been shown to cause human or animal disease. botulinum toxin main article: botulinum toxin 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 toxins to be absorbed from the intestines into the bloodstream. 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). 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. 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. 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. botulinum toxin types 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, while type e is 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. 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. as no antitoxin to type h is yet available, discovered in 2013 and by far the deadliest, details are kept under shroud. a few strains from organisms genetically identified as other clostridium species have caused human botulism: c. butyricum has produced type e toxin and c. baratii had produced type f toxin. 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. 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. sporogenesc. putrificum c. butyricumc. beijerinickii c. haemolyticumc. novyi type a c. subterminalec. haemolyticum laboratory isolation 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. taxonomy history genomic informationncbi genome id726ploidyhaploidgenome size3.91 mbnumber of chromosomes1year of completion2007 c. botulinum was first recognized and isolated in 1895 by emile van ermengem from home-cured ham implicated in a botulism outbreak. 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.) 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. 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. group i c. botulinum strains that do not produce a botulin toxin are referred to as c. sporogenes. the complete genome of c. botulinum has been sequenced at wellcome trust sanger institute in 2007. pathology foodborne botulism "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." 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 is 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 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 beneficial effects of botulinum toxin 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 adult intestinal toxemia 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 dysarthria imbalance weakness in arms and hand area c. botulinum in different geographical locations 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. north america type a c. botulinum predominates the soil samples from the western regions, while type b is the major type found in eastern areas. 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. 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). europe c. botulinum type e is prevalent in aquatic sediments in norway and sweden, denmark, the netherlands, the baltic coast of poland, and russia. 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. 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. australia c. botulinum type a was found to be present in soil samples from mountain areas of victoria. type-b organisms were detected in marine mud from tasmania. 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. use and detection 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, 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, and quantitative pcr can detect the toxin genes in the organism. growth conditions and prevention