CLOSTRIDIUM BOTULINUM

PATHOGEN SAFETY DATA SHEET - INFECTIOUS SUBSTANCES

SECTION I - INFECTIOUS AGENT

NAME: Clostridium botulinum.

SYNONYM OR CROSS REFERENCE: Botulism^(1-3), allantiasis⁽²⁾, and botulinum toxin⁽⁴⁾.

CHARACTERISTICS: A gram-positive (at least in early stage of growth), anaerobic, rod- shaped⁽³⁾, spore-forming bacillus^(1-3). Seven types of C. botulinum toxins exist (A-F)^{(1,2, 5)}.Types A, B, E, and rarely F can cause human botulism. Strains consistent with type G were assigned to C. argentinense in 1993. Botulinum neurotoxin is produced when, under anaerobic conditions, C. botulinum spores germinate^(3,5,6). In addition to C. botulinum, C. argentinense (formerly C. botulinum type G), C. butyricum, and C. baratii can also produce botulinum neurotoxin.

C. botulinum (botulinum toxin) is defined as a biothreat level A organism by the Centers for Disease Control and Prevention⁽⁷⁾. Category A organisms are considered to pose the greatest threat to national security.

SECTION II - HAZARD IDENTIFICATION

PATHOGENICITY: Rare but serious paralytic disease^(1,6), caused by a neurotoxin formed during the growth of the spore-forming bacterium C. botulinum (or rarely, C. argentinense, C. butyricum, or C. baratii )^(3,5). This neurotoxin binds to the neuromuscular junction and blocks excitatory synaptic transmission by inhibiting acetylcholine release^(2,8), causing (flaccid) paralysis^(1,5,6), and sometimes fatal respiratory failure^(1,2,5,8). The fatality rate of botulism is 5 to 10%⁽⁸⁾.

Food-borne botulism: The classic form of botulism is caused by the ingestion of preformed toxin in contaminated food^(1,2,4,5). Symptoms include double vision, drooping eyelids (ptosis), slurred speech, difficulty swallowing and muscle weakness that is symmetric and descends through the body (first shoulders are affected, then upper arms, lower arms, thighs, calves, etc.)⁽¹⁾. Death is usually due to respiratory failure^(1,2,6,8) and may occur as soon as 24 hours after onset of symptoms⁽⁸⁾.

Wound botulism: Occurs by contamination of a wound with spores from neurotoxin-producing Clostridium species in the environment and subsequent germination of these spores and production of toxin in the anaerobic milieu of an abscess^(1,4,5). The toxin is released into the bloodstream^(1,4) and symptoms may take up to 2 weeks to appear⁽¹⁾.

Intestinal (infant) botulism: Results almost exclusively from spore ingestion and subsequent growth and toxin production in the intestine^(1,2,5), affecting infants under 1 year old^(1,5,9). The first clinical sign is usually constipation^(1,2), but this disease has a wide spectrum of clinical severity, ranging from mild illness with gradual onset, to sudden infant death due to respiratory failure^(1,2). With appropriate intensive care, almost 100% of infants with botulism make a full recovery⁽⁵⁾. Infants with botulism are lethargic, feed poorly, have a weakened cry, exhibit ptosis, and floppy neck, and may progress to generalised flaccidity and respiratory compromise^(2,4).

Adult infectious botulism: Rare⁽⁵⁾. Caused by the intestinal colonization of C. botulinum / other neurotoxin producing species, followed by in vivo toxin production in a manner similar to infant botulism^(1,2,5). Patients often have a history of immunocompromise, abdominal surgery, bowel disease, or recent antibiotic therapy.

Inhalational botulism: Is not a naturally occurring disease⁽⁵⁾, but has occurred in laboratory workers due to inhalation of aerosolized toxin⁽¹⁾. Inhalational botulism leads to neurological symptoms similar to those of food-borne botulism, but with a longer incubation period^(5,6).

Iatrogenic botulism: Side effects resulting from the therapeutic intramuscular injection of Botox (purified, diluted A neurotoxin)⁽⁵⁾. Characterized by clinical weakness and electrophysiological abnormalities^(10,11).

EPIDEMIOLOGY: Sporadic. Family and general outbreaks occur worldwide in association with food products prepared or preserved by means that do not destroy spores and permit the formation of toxin^(1,2,5).

HOST RANGE: Humans, and various animals, e.g., fowl, fish, cows, dogs, and minks^(1,2).

INFECTIOUS DOSE: Cells/spores are not normally toxic for healthy adults⁽²⁾. Botulinum toxin is the most potent toxin known, with an estimated oral or injected toxic dose (serotype A) of 0.001 μg/kg body weight, and an estimated lethal dose by inhalation exposure in humans of approximately 0.07 μg/kg body weight^(5,6). Type A toxin is more potent than types B and E and causes the longest lasting disease⁽⁹⁾.

MODE OF TRANSMISSION: Food borne botulism: Ingestion of contaminated food containing toxin^(1,2,5). Infection is commonly associated with commercially processed foods that had undergone poor processing, storage, and improper preservation⁽¹²⁾.

Wound botulism: Contamination of wounds with spores of neurotoxin producing Clostridium species^(1,2,5) and is seen almost exclusively in injection drug users, particularly those who partake in injection of black-tar heroin into skin tissue^(5,13).

Intestinal (infant) botulism: Ingestion of spores. Sources include honey and infant milk powder^{(1, 5,9)}.

Adult infectious botulism: Ingestion of clostridial spores, rather than toxin, which then colonize the gut to produce their neurotoxin directly in the gut^(1,5,9).

Iatrogenic botulism: Side effect of injection of purified toxin⁽⁵⁾.

Inhalational botulism: Occurs due to absorption of botulinum toxin by the mucous membrane of the nose^(5,6).

INCUBATION PERIOD: The shorter the incubation period, the more severe the disease and the higher the case fatality rate⁽¹⁾.

Food-borne botulism: Usually 12 to 72 hrs after ingestion of toxin, depending on the dose⁽⁹⁾.

Wound botulism: The median period is 7 days⁽⁹⁾.

Adult infectious botulism: Unknown⁽¹⁾.

Intestinal botulism: Unknown⁽¹⁾.

Inhalational botulism: Not well defined, but it is longer than for food borne botulism^(1,6), and is estimated at 12-80 hours⁽¹⁴⁾.

COMMUNICABILITY: No evidence of person-to-person transmission^(1,5).

SECTION III - DISSEMINATION

RESERVOIR: Spores are found in soil, aquatic sediments, the intestinal tract of birds, animals and fish, and agricultural products, including honey and vegetables^(1,2,13).

ZOONOSIS: No epidemiological relationship between human and animal botulism has been established⁽²⁾.

VECTORS: None.

SECTION IV - STABILITY AND VIABILITY

DRUG SUSCEPTIBILITY: Susceptible to penicillin, metronidazole, clindamycin, cephalothin, cefoxitin, cefotaxime, chloramphenicol, tetracycline, erythromycin, rifampin, and vancomycin (with some strain variation)^(15,16).

DRUG RESISTANCE: Usually resistant to the aminoglycosides⁽³⁾, and may be resistant to tetracyclines and cephalosporins (with some strain variation)^(16,17). Also resistant to nalidixic acid and sulphamethoxazole-trimethoprim (SMX-TMP)⁽¹⁸⁾.

SUSCEPTIBILITY TO DISINFECTANTS: The vegetative state is susceptible to disinfectants such as 70% ethanol, 0.1% sodium hypochlorite, and 0.1N NaOH⁽¹⁹⁾. Spores may be resistant to disinfectants. Toxins are inactivated (more than 99.7%) by 20 minutes exposure to 3 mg/L free available chlorine (FAC; similar to the military disinfection procedure), and 84% inactivated by a treatment of 20 minutes at 0.4 mg/L FAC (similar to municipal water treatment procedures)⁽⁶⁾.

PHYSICAL INACTIVATION: Toxin is destroyed after heating for 5 minutes at greater than 85°C^(1,8,9). Toxins are detoxified in air within 12 hours and following exposure to sunlight within 1 to 3 hours⁽⁹⁾. Spores are highly resistant to heat⁽²⁾ and desiccation⁽²⁾; therefore, it is recommended to sterilize with dry heat (2 hours at 160°C) by autoclaving (20 minutes at 121°C, 1 atm pressure) and/or by irradiation⁽⁹⁾.

SURVIVAL OUTSIDE HOST: Survives well in soil, water and agricultural products^(2,5).

SECTION V - FIRST AID / MEDICAL

SURVEILLANCE: Since botulism is a life threatening condition, a rapid diagnosis is essential⁽⁹⁾and may require testing to differentiate botulism from other neurological diseases⁽⁶⁾.

Food borne botulism: Can be diagnosed by demonstration of toxin in serum^(1,2,5,8,15), stool^{(1,2, 6,8)}, gastric aspirate^(1,2,5) or implicated food^(1,2,5), or by culture of C. botulinum from a patient's gastric aspirate or stool in a clinical case^(1,5). The mouse bioassay is the most reliable method for detection of botulinum^(5,6,8).

Wound botulism: Can be diagnosed by demonstration of toxin in serum, or by positive wound culture^(1,2).

Adult infectious botulism: Can be diagnosed by demonstration of C. botulinum (or other neurotoxin producing species) and/or toxins in a patient's faeces or in autopsy specimens⁽¹⁾.

Intestinal (infant) botulism: Since the toxin is rarely found in the sera of infants^(1,2), faeces should be examined⁽¹⁾. An ELISA has been developed for the detection of A and B toxins in children's faecal samples⁽²⁾.

Inhalational botulism: Aerosolized toxin can not usually be identified in serum or faeces, but may be detected by ELISA from nasal swabs⁽⁸⁾.

Iatrogenic botulism: Should be suspected if patient has recently received Botox^(10,11). An immuno-PCR assay capable of detecting neurotoxin type A in the femtogram range has been developed⁽²⁰⁾.

Note: All diagnostic methods are not necessarily available in all countries.

FIRST AID/TREATMENT:

Foodborne botulism: Within 1 hour of ingestion of suspected food, the recommended course of action is a gastric lavage^(1,2), or enemas⁽¹⁾, and the administration of a cathartic (sorbitol)⁽¹⁾. In some cases intravenous administration of AB or ABE^(1,2,6,8) botulinum antitoxin^(1,2,5,6,8) is required, and assisted ventilation if respiratory failure occurs. Treatment may be required for weeks or months⁽⁵⁾.

Wound botulism: Antitoxin^(1,5), wound debridement, drainage and irrigation^(1,5,13), and antibiotic treatment⁽¹⁾.

Infant botulism: Requires meticulous supportive care⁽¹⁾. Instead of antitoxin which can cause sensitization and anaphylaxis^(1,6,8), an investigational human-derived botulinal immunoglobulin (BIG) is available for the treatment⁽¹⁾ and assisted respiration is given if required^(1,5).

Intestinal botulism: Repeated administration of ABE antitoxin⁽⁶⁾ and assisted respiration is given, if required^(1,5).

IMMUNISATION: None⁽⁵⁾.

PROPHYLAXIS: Individuals known to have eaten contaminated food should be purged with cathartics⁽¹⁾, given a gastric lavage and high enemas⁽¹⁾, and may be given equine botulinum AB or ABE antitoxin^(1,6).

SECTION VI - LABORATORY HAZARDS

LABORATORY-ACQUIRED INFECTIONS: Rarely reported⁽⁵⁾.

SOURCES/SPECIMENS: Food products, and clinical materials such as serum/blood, stool, vomit, and gastric aspirates⁽⁵⁾. For wound botulism: wound exudates, debrided tissue, or swab sample. Other sources include, nasal swabs from inhalational botulism patients, and environmental samples (soil, surface water)^(1,2,6,9).

PRIMARY HAZARDS: Exposure to the toxin. The toxin may be absorbed after ingestion, or following contact with the non-intact skin, the eyes, or mucous membranes, including the respiratory tract⁽⁵⁾. Inhalation of the toxin has occurred under laboratory conditions^(5,6).

SPECIAL HAZARDS: None.

SECTION VII - EXPOSURE CONTROLS / PERSONAL PROTECTION

RISK GROUP CLASSIFICATION: Risk Group 2⁽²¹⁾.

CONTAINMENT REQUIREMENTS: Containment Level 2 facilities, equipment, and operational practices for work involving infectious or potentially infectious materials, animals, or cultures.

PROTECTIVE CLOTHING: Lab coat. Gloves when direct skin contact with infected materials or animals is unavoidable. Eye protection must be used where there is a known or potential risk of exposure to splashes⁽¹⁹⁾.

OTHER PRECAUTIONS: All procedures that may produce aerosols, or involve high concentrations or large volumes should be conducted in a biological safety cabinet (BSC). The use of needles, syringes, and other sharp objects should be strictly limited. Additional precautions should be considered with work involving animals or large scale activities⁽¹⁹⁾.

SECTION VIII - HANDLING AND STORAGE

SPILLS: Allow aerosols to settle, and, while wearing protective clothing, gently cover the spill with paper towels and apply appropriate disinfectant starting at the perimeter, working inwards towards the centre. Allow sufficient contact time before clean up⁽¹⁹⁾.

DISPOSAL: Decontaminate before disposal, using steam sterilisation, incineration, or chemical disinfection⁽¹⁹⁾.

STORAGE: In locked, leak-proof containers that are appropriately labelled and secured⁽¹⁹⁾.

SECTION IX - REGULATORY AND OTHER INFORMATION

REGULATORY INFORMATION: The import, transport, and use of pathogens in Canada is regulated under many regulatory bodies, including the Public Health Agency of Canada, Health Canada, Canadian Food Inspection Agency, Environment Canada, and Transport Canada. Users are responsible for ensuring they are compliant with all relevant acts, regulations, guidelines, and standards.

UPDATED: September 2010.

PREPARED BY: Pathogen Regulation Directorate, Public Health Agency of Canada.

Although the information, opinions and recommendations contained in this Pathogen Safety Data Sheet are compiled from sources believed to be reliable, we accept no responsibility for the accuracy, sufficiency, or reliability or for any loss or injury resulting from the use of the information. Newly discovered hazards are frequent and this information may not be completely up to date.

Copyright ©
Public Health Agency of Canada, 2010
Canada

REFERENCES:

1. An Official Report of the American Public Health Association. (2004). In D. L. Heymann (Ed.), Control of Communicable Diseases Manual. (pp. 69-74). Washington, DC, USA: American Public Health Association.
2. Acha, P. N., & Szyfres, B. (2001). Bacterial Infection and Mycoses. Zoonoses and Communicable Diseases Common to Man and Animals. (pp. 3-283, 28-40). Washington, D.C, USA: Pan American Health Organization.
3. Johnson, E. A., Summanen, P., & Finegold, S. M. (2007). Clostridium. In P. R. Murray, E. J. Baron, J. H. Jorgensen, M. L. Landry & M. A. Pfaller (Eds.), Manual of Clinical microbiology (9th ed., pp. 889-910). Washington, DC: ASM press.
4. Davis, L. E. (1993). Botulinum toxin: From poison to medicine. Western Journal of Medicine, 158 (1), 25-29.
5. Sobel, J. (2005). Botulism. Clinical Infectious Diseases, 41 (8), 1167-1173.
6. US Army Medical Research Institute of Infectious Diseases. (2004). Medical Management of Biological Casualties Handbook. Medical Management of Biological Casualties Handbook. (5th ed., ). USA: US Army Medical Research Institute of Infectious Diseases.
7. Humes, R., & Snyder, J. W. (2007). Laboratory Detection of Potential Agents of Bioterrorism. In P. R. Murray, E. J. baron, J. H. Jorgensen, M. L. Landry & M. A. Pfaller (Eds.), Manual of Clinical Microbiology (9th ed., pp. 107-117). Washington, D.C.: ASM Press.
8. Franz, D. R., Jahrling, P. B., McClain, D. J., Hoover, D. L., Byrne, W. R., Pavlin, J. A., Christopher, G. W., Cieslak, T. J., Friedlander, A. M., & Eitzen E.M., J. (2001). Clinical recognition and management of patients exposed to biological warfare agents. Clinics in Laboratory Medicine, 21 (3), 435-473.
9. Lindstr*m, M., & Korkeala, H. (2006). Laboratory diagnostics of botulism. Clinical Microbiology Reviews, 19 (2), 298-314.
10. Bakheit, A. M., Ward, C. D., & McLellan, D. L. (1997). Generalized botulism-like syndrome after intramuscular injections of botulinum toxin type A: a report of two cases J. Neurol. Neurosurg. Psychiatry, 62 (2), 198.
11. Cheng, C. M., Chen, J. S., & Patel, R. P. (2006). Unlabeled uses of botulinum toxins: A review, part 2. American Journal of Health-System Pharmacy, 63 (3), 225-232.
12. McLauchlin, J., Grant, K. A., & Little, C. L. (2006). Food-borne botulism in the United Kingdom. Journal of Public Health (Oxford, England), 28 (4), 337-342. doi:10.1093/pubmed/fdl053
13. Reller, M. E., Douce, R. W., Maslanka, S. E., Torres, D. S., Manock, S. R., & Sobel, J. (2006). Wound botulism acquired in the Amazonian rain forest of Ecuador. American Journal of Tropical Medicine and Hygiene, 74 (4), 628-631.
14. Project Tripwire. (2007). Botulism. Retrieved 11/24, 2010, from www.wildlifecenter.org/bioweapons/analysis.php?dis=botulism&fact=transmission
15. Swenson, J. M., Thornsberry, C., & McCroskey, L. M. (1980). Susceptibility of Clostridium botulinium to thirteen antimicrobial agents. Antimicrobial Agents and Chemotherapy, 18 (1), 13-19.
16. Dezfulian, M., & Dowell Jr., V. R. (1980). Cultural and physiological characteristics and antimicrobial suspectibility of Clostridium botulinum isolates from foodborne and infant botulism cases. Journal of Clinical Microbiology, 11 (6), 604-609.
17. Boyanova, L., Kolarov, R., & Mitov, I. (2007). Antimicrobial resistance and the management of anaerobic infections. Expert Review of Anti-Infective Therapy, 5 (4), 685-701.
18. Fenicia, L., Ferrini, A. M., Anniballi, F., Mannoni, V., & Aureli, P. (2003). Considering the antimicrobial sensitivity of the intestinal botulism agent Clostridium butyricum when treating concomitant infections. European Journal of Epidemiology, 18 (12), 1153-1154.
19. Public Health Agency of Canada. (2004). In Best M., Graham M. L., Leitner R., Ouellette M. and Ugwu K. (Eds.), Laboratory Biosafety Guidelines (3rd ed.). Canada: Public Health Agency of Canada.
20. Chao, H. -., Wang, Y. -., Tang, S. -., & Liu, H. -. (2004). A highly sensitive immuno- polymerase chain reaction assay for Clostridium botulinum neurotoxin type A. Toxicon, 43 (1), 27-34.
21. Human pathogens and toxins act. S.C. 2009, c. 24, Second Session, Fortieth Parliament, 57- 58 Elizabeth II, 2009. (2009).