CLOSTRIDIUM

146 plus species including:

SPECIES

Clostridium amniovalercium
Clostridium argentinense
Clostridium barati
Clostridium bifermentans
Clostridium botulinum
Clostridium butyricum
Clostridium cadaveris
Clostridium carnis
Clostridium chauvoei
Clostridium cocleatum
Clostridium difficile
Clostridium fallax
Clostridium histolyticum
Clostridium nigrificans
Clostridium novyi
Clostridium perfringens
Clostridium ramosum
Clostridium septicum
Clostridium sordellii
Clostridium sporogenes
Clostridium tertium
Clostridium tetani

MICROSCOPIC APPEARANCE

Gram Stain: They usually stain gram-positive, at least in very early stages of growth.
Morphology: Cells of most strains occur as straight or slightly curved rods.
Size: 0.3-2.0 micrometers by 1.5-20.0 micrometers in length.
Motility: Motility occurs by peritrichous flagella.
Capsules: None.
Spores: Clostridium spp. form oval or spherical endospores that usually distend the cell.
Other: Most have round, tapering ends; long filaments are formed by some species.

MACROSCOPIC APPEARANCE

Clostridia possess no one typical colony morphology. They are generally a large colony (>2mm) with irregular edges or swarming growth. Some Clostridia form small, convex, non-hemolytic colonies with a smooth edge. Other Clostridia produce several different-looking colony types, so the culture appears mixed. A few Clostridium spp. have distinctive colony characteristics. Clostridium perfringens usually produces a double zone of beta-hemolysis. The inner zone shows complete hemolysis, whereas the outer zone may display partial hemolysis. Clostridium difficile produces a yellow ground-glass colony on CCFA. On blood agar, Clostridium difficile are usually 2mm or more in diameter after 24 hours of incubation, fluoresce yellow-green, and emit a horse stable odor. (Fluorescence should not be observed on CCFA because it itself is fluorescent).

METABOLIC PROPERTIES

Obligately anaerobic, though some strains (e.g. Clostridium perfringens ) can carry out nitrate respiration; growth often poor in/on basal media found in soils and in the intestines of man and other animals; some species pathogenic.

KEY BIOCHEMICAL REACTIONS

Usually catalase-negative.

HABITAT

Clostridium are ubiquitous in soil and street dust throughout the world and are able to survive in this environment or years. They are commonly found in soil, sewage, marine sediments, animal and plant products, the intestinal tract and in wounds of man and animals. They enter the human body through a wide variety of wounds, which are in turn contaminated by feces or soil.

PATHOGENICITY

Clostridium tetani produces two primary toxins; the highly toxic spasmogenic neurotoxin, tetanospasmin, and the hemolytic toxin, or tetanolysin. Although the first of these is the only one largely responsible for the characteristic signs of tetanus, there is some evidence that the tetanolysin may also be involved. Early signs and symptoms include tension or cramps and twitching in muscles around the wound, increased reflexes in the wounded extremity, slight dysphagia, stiffness of the neck and jaws muscles, headache, backache, general irritability, and anxious facial expressions. These are followed by lockjaw, spasms of the jaw and mouth, and finally spasticity of the neck trunk, and limbs.

Most infections are acquired endogenously and are part of a mixed flora with other anaerobes and aerobes. Clostridial disease syndromes of exogenous origins occur less frequently and can include gas gangrene. Clostridial syndromes involving toxigenicity botulism ( Clostridium botulinum ) and food poisoning ( Clostridium perfringens ) have also been noted. Some species are involved in several types of disease. Clostridium perfringens and Clostridium ramosum are the most common clinical isolates. The primary etiological agent responsible for antibiotic-associated diarrhea is Clostridium difficile .

RECOMMENDED MEDIA

For culture: Anaerobic Blood Agar or Brucella with H & K.
For selective isolation: Anaerobic PEA, Cycloserine-Cefoxitin Fructose Agar (CCFA; for Clostridium difficile ).
For identification: Egg Yolk Agar.
For maintenance: Cooked Meat Media or Thioglycollate with Supplements.

INCUBATION

Temperature: 35 degrees C.
Time: 24-48 hours.
Atmosphere: Anaerobic.
Optimum pH: 7.0 +/- 0.2.

REFERENCES

1. Holt, J.G., et al. 1994. Bergey's Manual of Determinative Bacteriology , 9th ed. Williams & Wilkins, Baltimore, MD.

2. Holt, J.G., et al. 1986. Bergey's Manual of Systemic Bacteriology , Vol. I & II. Williams & Wilkins, Baltimore, MD.

3. The Oxoid Vade-Mecum of Microbiology . 1993. Unipath Ltd., Basingstoke, UK.

4. Internet: www.hardlink.com /Bacterial Database Search, February, 1998.

5. Murray, P.R., et al. 1995. Manual of Clinical Microbiology , 6th ed. American Society for Microbiology, Washington, D.C.


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