Cultural characteristics
Biochemical characters
Phylum Firmicutes, Class Clostridia, Order Clostridiales, Family Clostridiaceae, Genus Clostridium, Cluster XI (non-Clostridium
sensu stricto),
Clostridium difficile (Hall and O’Toole 1935) Prevot 1938.

Historical synonym:
Bacillus difficile  Hall and O’Toole 1935.
Gram-positive, straight rods, 0.5-0.9 x 3.0-16.9 µm. Motility is variable. Peritrichous
flagella. Spores are oval, subterminal / terminal, swelling  the cell. Sporulation of
most strains occurs on Brucella blood agar incubated for 2 days; sporulation may be
enhanced on solid media containing 0.1% sodium taurocholate.
Surface colonies on blood agar plates are 2–5 mm in diameter, circular, occasionally
rhizoid, flat or low convex, opaque, grayish or whitish, and have a matt to glossy surface.
All strains produce an evanescent pale green fluorescence under long wavelength
ultraviolet light after 48 hours incubation on Brucella blood agar supplemented with
hemin and vitamin K1. Blood is not hemolysed. Cultures in PYG broth are turbid with a
smooth sediment and have a pH of 5.0–5.5 after incubation for 5 days. Moderate growth
in nutrient broth with granular sediment. Grow at temperature: 25 and 45 ºC, optimum 30-37 ºC.
Abundant  gas is produced in PYG deep agar cultures. Major products of metabolism in PYG broth: H
2 , acetic, isobutyric, butyric,
isovaleric, valeric, isocaproic, formic, and lactic acids.
Isolated from marine sediment, soil, sand, the hospital environment; camel, horse, and donkey dung; feces of dogs, cats, and
domestic birds; the human genital tract; feces of humans without diarrhea; and rarely from blood and pyogenic infections in humans
and animals.
Susceptible to penicillin G,  ampicillin, vancomycin, rifampin and metronidazole. Resistant to aminoglycosides.
Pathogenic for laboratory animals. Produces two large protein toxins (toxins A and B). Toxin A is lethal when given orally to hamsters
but toxin B is not; either toxin is lethal when injected intraperitoneally into these rodents. Toxins A and B are lethal for mice. Toxin A
has been referred to as the enterotoxin. Toxin B  is extremely cytopathic for all tissue-cultured cells tested.
Pseudomembranous colitis in humans is caused by overgrowth of the organism in the colon, usually after the flora has been
disturbed by antimicrobial therapy. A  similar disease can be produced in hamsters and several other rodents by administration of
antibiotics. Has been reported to cause cecitis in rabbits and hares.
  1. N.A. Logan and P. De Vos, 2009. Genus I. Clostridium Prazmowski 1880. In: (Eds.) P.D. Vos, G. Garrity, D. Jones, N.R. Krieg, W.
    Ludwig, F.A. Rainey, K.-H. Schleifer, W.B. Whitman. Bergey’s Manual of Systematic Bacteriology, Volume 3: The Firmicutes,
    Springer, 738-828.
  2. Smith L.D.S. and Hobbs G., 1975. Genus III. Clostridium Prazmowski 1880. In: (Eds.) Buchanan R.E. and Gibbons N.E., Bergey’s
    Manual of Determinative Bacteriology, Eighth Edition , The Williams & Wilkins Company, Baltimore, 551-572.
  3. Macovei A., 2009. Identificarea bacteriilor anaerobe. In: Tratat de Microbiologie Clinica (Ed. Buiuc D. si Negut M.), editia a IIIa,
    Editura Medicala, Bucuresti, 900-927.
  4. Secasiu V., 2001. Boli produse de germeni din genul Clostridium. In: Boli infectioase ale animalelor, Moga Manzat R., Ed. Brumar,
    Timisoara, 481-612.
H2 is produced in large amounts. Milk reaction and meat digestion are negative.

Positive results for H
2 production, esculin hydrolysis, gelatin hydrolysis (very slowly), substrate utilized and/or acid produced from:
cellobiose (weak), fructose & glucose.

Negative results for casein hydrolysis, indole production, lecithinase, lipase, nitrate reduction, starch hydrolysis,
urease,Voges-Proskauer, substrate utilized and/or acid produced from: amygdalin, arabinose, galactose, glycogen, glycerol, inositol,
inulin, lactose, maltose, melibiose, raffinose, rhamnose, ribose, sorbose, starch & sucrose.

Variable results for H
2S production, substrate utilized and/or acid produced from: dulcitol, mannitol, mannose, melezitose, salicin
(weak), sorbitol (weak), trehalose (weak) & xylose.
Clostridium difficile
(c) Costin Stoica
Culture media
Biochemical tests
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