text: how to culture c. difficile??.? want to know the composition of the medium required. would like to know if any special conditions to be taken care off. clostridium shows optimimum growth when plated on blood agar at human body temperatures. when the environment becomes stressed, however, the bacteria produce spores that tolerate the extreme conditions that the active bacteria cannot.  clostrium species also form spores which survive boiling water, but the closterium species can not form spores in the presence of oxygen. most clostridia are aerotolerant (they survive exposure to air), but most species of clostridium do not resume growth if oxygen is present.  minimal requirements of amino acids and vitamins were determined in chemically defined medium for five strains of clostridium difficile. cysteine, isoleucine, leucine, proline, tryptophan and valine were essential amino acids for growth of c. difficile. arginine, glycine, histidine, methionine and threonine enhanced growth. biotin, pantothenate and pyridoxine were essential vitamins. a defined medium containing the minimal requirements of amino acids and vitamins produced a rapid and heavy growth which was comparable to that in modified brain heart infusion, a complex medium. adenine was able to substitute for glycine and threonine, suggesting that the two amino acids may be utilized as precursors of purine nucleotides. the defined medium developed here will assist physiological and biochemical studies on c. difficile.