Clostridium: Toxins, Characteristics

Genus Clostridium General Characteristics

• The name "kloster" is derived from a Greek word and means spindle.
• Bacilli are large, and it contains more than 100 species.
• Young cultures are Gram-positive, while older cultures exhibit Gram variability.
• Clostridium is an obligate anaerobic bacteria with unusual features.
• Spore and toxin formation are key features.
• They commonly inhabit the environment or the digestive systems of humans and animals.
• Clostridia spp. are dominant in human intestinal flora, accounting for approximately 50% or 10^13/gm of stool.
• They form pairs or short chains with rounded, pointed ends.
• Most clostridia move via peritrichous flagella.
• All species create endospores and possess a fermentative metabolism.
• Three important qualities include multiplying only in the absence of oxygen, surviving adverse conditions, and releasing potent toxins during multiplication.

Genus Clostridium Culture Characteristics

• Growth on solid medium is slow.
• Some colonies fluoresce.
• Hemolysis occurs on blood agar.
• It is saccharolytic, fermenting carbohydrates into butyric acid, acetic acid, and alcohol.
• It is proteolytic.

Genus Clostridium Culture Media

• Robertson's cooked meat broth contains amino acids.
• They can be cultured on nutrient agar.
• Blood agar targets or produces a double zone of hemolysis (inner-β; outer-α).
• Detection of lecithinase C is possible on lactose egg yolk milk agar.
• Green fluorescent colonies grow on Mac Conkey agar.
• Thioglycolate can be used for culturing.

Clostridium Pathogenesis and Toxins

• Clostridia produce the largest number of toxins of any bacterial class.
• Toxin production is the primary mechanism of pathogenicity.
• Clostridium forms endospores in adverse environmental conditions as a survival mechanism.
• Spores are characterized by their position, size, and shape.
• Most Clostridium species, including C. perfringens and C. botulinum, have ovoid subterminal (OST) spores.
• C. tetani has round terminal (RT) spores.

Clostridium Pathogenesis: Four Broad Types

• Histotoxic group are tissue infections: C. perfringens type A is acquired exogenously more commonly than endogenously, and C. septicum is acquired endogenously.
  • These cause cellulitis, myonecrosis, gas gangrene, and fasciitis.
• Enterotoxigenic group causes gastrointestinal disease.
  • Clostridial foodborne disease occurs 8-24 hours after ingesting large numbers of organisms on contaminated meat products where spores germinate and produce enterotoxins (C. perfringens type A).
  • Necrotizing enteritis is caused by beta toxin-producing C. perfringens type C.
  • C. difficile is acquired endogenously or exogenously from person to person in hospitals, causing antibiotic-associated diarrhea and pseudomembrane colitis.
• Tetanus and other toxins (exogenously acquired) includes the C. tetani neurotoxin.
  • It is generalized (most common), cephalic (primary infection in head, uncommonly ear), localized, or neonatal (contaminated umbilical stump).
• Botulism (exogenously acquired) includes the C. botulinum neurotoxin.
  • It is foodborne (intoxication, 1-2 days incubation), infant (ingestion of spores in honey), or wound (symptoms similar to foodborne, but 4 or more days incubation).

Clostridium tetani

• This spore gives a typical "drumstick" appearance.
• It is motile and grows on blood agar or cooked meat medium with swarming colonies, resulting in β-hemolysis.
• It is the causative agent of tetanus.
• It is extremely sensitive to oxygen toxicity.
• Spores are highly resistant but killed by autoclaving.
• It is found in soil and in the manure of horses, cows, sheep, dogs, chickens, rats, and humans.

Clostridium tetani Toxins and Action

• Tetanospasmin and tetanolysin cause the characteristic "tetanic spasm" and the clinical manifestations of tetanus.
• It releases an A-B toxin when the bacteria lyse (exotoxin).
• The toxin binds gangliosides in synaptic membranes (synapse).
• Tetanospasmin is a heat-labile neurotoxin that blocks the release of inhibitory neurotransmitters, leading to continuous stimulation of excitatory transmitters such as GABA and glycine.
• Tetanolysin is a heat-stable hemolysin of unknown significance.

Clinical Forms of Tetanus

• Generalized tetanus involves bulbar and paraspinal muscles, causing trismus or lockjaw, risus sardonicus, difficulty swallowing, irritability, and opisthotonos and also involves the autonomic nervous system, leading to sweating, hyperthermia, cardiac arrhythmias, and blood pressure fluctuations.
• Cephalic tetanus is a primary infection in the head, particularly the ear, with isolated or combined involvement of cranial nerves and a poor prognosis.
• Localized tetanus involves muscles in the area of primary injury, which may precede generalized disease and has a favorable prognosis.
• Neonatal tetanus is a generalized disease in neonates, typically originating from the umbilical stump, and has a very poor prognosis in infants whose mothers are nonimmune.
• Tetanus results in spastic paralysis
• Germination of the spore and production of toxin are aided by conditions that lead to low oxidation-reduction potential like necrotic tissue, calcium salts, and associated pyogenic infections.

Tetanus Diagnosis, Prevention, and Treatment

• Diagnosis depends on the clinical picture and a history of injury.
• Prevention and control involve active immunization with toxoid and a 'booster shot' for previously immunized individuals, which may be accompanied by antitoxin injected into a different area of the body, as birds are naturally immune.
• Treatment includes TAT at 100,000 to 200,000 units as a single dose, penicillin, surgical debridement to remove necrotic tissue, and muscle relaxants, sedation, and assisted ventilation.

Clostridium botulinum

• It has fastidious growth requirements.
• It is motile and produces heat-resistant spores (ovoid, subterminal).
• Produces 7 distinct botulinum toxins that all cause flaccid paralysis.
• The toxins bind neuromuscular junctions and alter the mechanism for acetylcholine release.
• Strains associated with human disease produce lipase, digest milk proteins, hydrolyze gelatin, and ferment glucose.
• It causes disease mostly through intoxication.
• It causes lamziekte and limberneck and results in flaccid paralysis due to the nerve cells' inability to release acetylcholine.
• Swine and carnivorous animals are resistant.
• Forms include foodborne, infant, and wound botulism.

Clostridium botulinum Diagnosis, Treatment, and Prevention

• Diagnosis is via ELISA.
• Treatment includes trivalent antitoxin (A, B, E), ventilatory support, and gastric lavage, penicillin, and metronidazole.
• Prevention includes toxoid and heating of home-canned food.

Clostridium toxin Comparison

• Clostridium tetani has wounds as a site of toxin production and acts centrally by blocking synaptic inhibition, causing spastic paralysis with tetanospasmin as one antigenic type.
• Clostridium botulinum acts the site of toxin production is carcasses, decaying vegetation, and occasionally wounds and intestines, it acts peripherally by blocking neuromuscular transmission, causing flaccid paralysis. There are eight different toxins produced by types A-G.

Clostridium perfringens (C. welchii)

• It is characterized by large, rectangular bacilli.
• Spores are rarely seen in vitro (ovoid, subterminal).
• It is non-motile but rapidly spreads on blood agar.
• It is aerotolerant on specially supplemented media with blood.
• There are five types of strains (A - E) and four lethal toxins: Alpha, Beta, Epsilon, and Iota.
• Epsilon-toxin increases intestinal permeability, causing vascular damage and edema in major organs, liver damage, and higher blood pressure.
• Iota-toxin causes food-borne illness.
• Alpha-toxin and Beta-toxins lead to gas gangrene by necrotizing cell membranes, primarily in humans.
• It can cause food-borne illness.

Clostridium perfringens Diagnosis

• Direct smear and gram stain can be used.
• Rapid growth culture occurs in chopped meat media at 45°C, then isolated in blood agar for four to six hours.
• Double zone of hemolysis, Nagler reaction (precipitation in serum or egg yolk media), and stormy fermentation (coagulation) of milk are all diagnostic tools.
• Treatment involves removal of necrotic tissue (debridement) and Penicillin G.