CLOSTRIDIUM TETANI – CAUSATIVE AGENT OF TETANUS
The History of Discovery
A causative agent of tetanus was described first by A. Nicolaier in 1884, and isolated the pure culture by S. Kitasato in 1889.
Tetanus causative agent pertains to the family Clostridiaceae, genus Clostridium, species C. tetani.
Structure and Properties of C. tetani
Clostridium tetani is a thin gram-positive rod about 5 μm in length. It is a motile peritrichous bacterium with round terminal spore. Spore- containing cells resemble drumsticks.
The temperature range for optimal propagation of C. tetani is 15-45°C. Usually it grows on the blood or sugar agar, pH 7.0-8.0 at 37°C being cultured in anaerobic conditions within anaerobic jars. At the end of cultivation small smooth colonies surrounded by slight zones of hemolysis are observed. The view of colonies resembles dew drops.
When cultured within the deep agar stabs, fragile cloud-like colonies appear. Cultivation in Kitt-Tarozzi medium results in homogenous turbidity with gas production.
As any other clostridia, C. tetani is obligate anaerobe. It demonstrates generally poor biochemical activity. Tetanus causative agent doesn’t ferment carbohydrates. Nevertheless, these bacteria liquefy gelatin, coagulate milk, and reduce nitrates to nitrites.
Clostridium tetani is divided into 10 serotypes accroding to variations of flagellar H-antigen. Somatic O-antigen is group-specific. Microbial exotoxin has common antigenic properties in all C. tetani.
C. tetani expresses highly poisonous exotoxin composed of two fractions: tetanospasmin that causes the contraction of skeletal muscles, and tetanolysin, which produces hemolysis.
The lethal activity of tetanus exotoxin succumbs only to the action of the most potent botulotoxin – one mouse lethal dose of dry tetanus toxin is about 5 ng. The mortal dose for humans is near 2-2.5 ng per 1 kg of body weight.
Tetanospasmin is Zn-containing metal protease that destroys synaptic vesicle-associated protein and synaptobrevin in the synapses of inhibitory neurons of central nervous system resulting in generalized muscular spasms.
Tetanolysin is the membrane-damaging fraction of exotoxin with hemolytic and cardiotoxic properties. Also it affects medullar nuclei and the neurons of autonomic nervous system.
Heating at 60-70°С inactivates vegetative forms of clostridia within 30 minutes. The bacteria are sensitive to conventional disinfectants.
The spores are extremely resistant and keep viability in soil and dust for many years. They can withstand boiling for more than 1 hour. Standard disinfectants, such us 5% phenol or 1% formaldehyde inactivate clostridia spores only after 5-10 h of exposure.
Pathogenesis and Clinical Findings in Tetanus
C. tetani is the normal inhabitant of human and animal gut. Animals and humans are the major sources of infection. Spores of clostridia appear in the soil with feces and may stay there for years.
Microorganisms enter the body through the injured skin or mucous membranes via soil-contaminated wounds or skin lesions (contact route of the disease transmission). More often the disease affects children or agriculture workers.
C. tetani multiply in the site of primary contact and release exotoxin. Toxin undergoes retroaxonal or perineural lymphatics transport and moves into the spinal cord. It binds to ganglioside receptors of neurons, penetrates into the synapses and blocks the release of inhibitory neuromediators (glycin and γ- aminobutyric acid).
Impairment of inhibitory signalling leads to uncontrolled stimulation of neuromuscular synapses of motor neurons that elicits tonic or myoclonic striated muscle contractions.
At the primary site of pathogen penetration persistent tonic muscular cramps are observed. The symptoms arise in descending order. At first the disease affects jaw muscles with tonic spasms (trismus) and face muscles (risus sardonicus). Then the back muscles and limbs are involved. Generalized tetanus muscular spasm is known as opisthotonus.
The disease prognosis is very serious. In case of delay of treatment the developed tetanus results in lethal outcome in 40-50% of cases. Anti-toxic natural immunity is very weak and can’t prevent next tetanus infection.
Laboratory Diagnosis of Tetanus
In most cases the clinical findings of tetanus are evident enough to make correct diagnosis.
For laboratory confirmation of the diagnosis of tetanus the specimens of wound discharge, biopsy tissue samples, or stitch material can be examined.
Investigation of tetanus toxin in clinical samples is performed by ELISA test, or by indirect hemagglutination test with erythrocyte anti-tetanus diagnosticum, or by neutralization reaction in mice.
For culture isolation the specimens should be previously heated at 80°С for 20 min to inactivate non-sporeforming bacteria. Then they are inoculated into Kitt-Tarozzi medium or upon blood agar that is placed into anaerobic jar. After several days of incubation the grown colonies undergo microscopy. Toxin accumulation is evaluated by experimental mice infection. The diagnosis is confirmed by neutralization reaction with anti-tetanus antibodies.
Treatment and Prophylaxis of Tetanus
Urgent prophylaxis of the disease depends on the level of initial antitoxic immunity of affected person. Prophylaxis covers all patients with traumas, burns, animal bites, etc.
Previously vaccinated individuals are immunized with tetanus toxoid. Non-immune patients obtain tetanus toxoid and human antitoxic anti-tetanus immunoglobulin.
For treatment of developed tetanus the high doses of human antitoxic tetanus immunoglobulin or horse antitoxic serum are used. In addition, anticonvulsive drug therapy is administered (diplacine, aminazine, diazepam, etc.)
Organized active prophylaxis is performed by vaccination with tetanus toxoid. The toxoid is an essential constituent of complex ADPT polyvaccine (adsorbed diphtheria, pertussis, tetanus vaccine with aluminum hydroxide as adjuvant) or of combined toxoid preparation ADT. Tetanus is the completely preventable disease.
The vaccination starts and repeats thrice at the first year of life. Subsequent booster is injected in 9-12 months and then reproduced every 10 years.