Workbook in Microbiology Laboratory Techniques 7e

Clostridium tetani is the agent of tetanus, or "lockjaw."When introduced into deep tissues, this organism produces little or no local tissue damage, but secretes an exotoxin known as a neurotoxin, which adversely affects nerve function. The neurotoxin is absorbed from the infected area and travels along peripheral motor nerves to the spinal cord. Severe muscle spasm and convulsive contraction of the involved muscles result. It is often difficult to make a laboratory diagnosis of this disease because the site of injury may be closed and healed, with no apparent signs of infection, by the time the symptoms of neurotoxicity begin. The organism is difficult to cultivate, but if isolated, is identified by microscopic morphology and patterns of carbohydrate fermentation. The en-dospore of C. tetani is usually at one end of the bacillus (terminal). It is wider in diameter than the vegetative bacillus, giving the cell the appearance of a "drumstick" or tennis racquet. The diagnosis is usually based on clinical signs and symptoms.

Clostridium botulinum produces an exotoxin that causes the deadly form of food poisoning called botulism. This is not an infectious disease, but a toxic disease. If the endospores of this soil organism survive in processed foods that have been canned or vacuum packed, they may multiply in the anaerobic conditions of the container, elaborating their potent exotoxin in the process of growth. If the food is eaten without further cooking (which would destroy the toxin), the toxin is absorbed and botulism results. The disease is difficult to diagnose bacteriologically, but the incriminated food or the patient's blood can be tested to demonstrate the toxin's effect in mice, which confirms the diagnosis.

In infant botulism, when endospores of the bacillus (endospores in honey have been implicated in a few cases) are ingested by children under one year of age, the endospores germinate in the child's intestinal tract in some instances, and the resulting vegetative cells produce toxin. This type of botulism has been implicated in certain cases of sudden infant death syndrome (SIDS).

In this exercise we shall use species of Clostridium to illustrate the general principles of anaerobic culture methods.


To learn basic principles of anaerobic bacteriology


Anaerobe jar

Blood agar plates

Thioglycollate broth

Tubed skim milk

Phenol red broths (glucose, lactose)

Blood agar plate cultures of Clostridium perfringens and Clostridium histolyticum

Nutrient slant cultures of Pseudomonas aeruginosa and Staphylococcus epidermidis

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