Growth inhibition of Clostridium difficile by intestinal flora of infant faeces in continuous flow culture

Abstract

Summary. Growth of Clostridium dzficile was inhibited more strongly in continuous flow (CF) culture with C. dzficile-negative faeces of infants than with C. dificile-positive faeces. Culture of faecal flora of infants yielded a greater variety of bacterial species in C. dzficilenegative than in C. dzficile-positive faeces. In the mixed CF culture of C. dzficile with Enterococcus avium, Bacteroides distasonis, Eubacterium lentum, C. ramosum, C. perfringens and either Escherichia coli or Klebsiella pneumoniae isolated from C. dzficile-negative faeces, inhibition of growth of C. dzficile was demonstrated when the pH of the culture medium was decreased. Amino-acid analysis of CF cultures showed considerable utilisation of aspartic acid, serine, threonine, arginine and asparagine. A marked increase in concentrations of citrulline and ornithine was found in the culture that inhibited growth of C. dzficile. The addition of citrulline and ornithine into a Gifu anaerobic medium (GAM) broth produced no inhibition of growth of C. dzficile. The addition of the mixture of the depleted amino acids (aspartic acid, serine, threonine, arginine and asparagine) to the culture filtrate or adjustment of the pH of the culture filtrate induced considerable growth of C. dzficile. These results suggest that the inhibition of growth of C. dzficile may be due to consumption of amino acids by intestinal flora, and not to the presence of inhibitors produced by the intestinal flora.

Toxins, Butyric Acid, and Other Short-Chain Fatty Acids Are Coordinately Expressed and Down-Regulated by Cysteine in Clostridium difficile

“…The large intestine is a milieu essentially lacking free glucose (2), and the availability of amino acids is likely to be crucial to the growth of C. difficile and other bacteria in vivo. In continuous cultures containing human feces, significant competition for amino acids between C. difficile and other microorganisms of the biota was shown (26). The apparent association between amino acid limitation and toxin production may be of clinical relevance in that protein malnutrition may be a risk factor for CDAD.…”

“…Toxin production by C. difficile has been demonstrated to be dependent on the nutrient level of the growth medium (7,10,12,18,24,25). The type and amount of nutrients present have been shown to affect growth of C. difficile in a continuous culture system containing a complex microflora (23,26), suggesting that competition for nutrients in the colon plays a role in colonization by the pathogen and in the development of CDAD. Toxin expression may differ 100,000-fold between toxin-positive isolates of C. difficile in vitro (14) but the underlying reason is not understood.…”

Inhibition of the cytotoxic effect of Clostridium difficile in vitro by Clostridium butyricum MIYAIRI 588 strain

“…For example, faecal emulsions from well patients inhibit C. difficile growth and toxicity in vitro (Borriello & Barclay, 1986;Yamamoto-Osaki et al, 1994). Previous studies on other probiotics have also reported growth and toxicity inhibition in co-culture with various species of Bifidobacterium and Lactobacillus (Banerjee et al, 2009;Trejo et al, 2010).…”

Clostridium difficileinfection

“…The inhibition correlated with a decrease in pH and the depletion of specific amino acids; restoration of the pH and addition of the depleted amino acids relieved the growth inhibition on C. difficile, suggesting that colonization resistance may be mediated by a direct competition for nutrients. 124 In the intestine, commensal bacteria may additionally compete for attachment sites favored by C. difficile, since a non-toxigenic strain of C. difficile can effectively interfere with colonization by toxigenic C. difficile strains. 125 While the mechanism for this interference has not been established, our in vitro experiments suggest that the non-toxigenic strain directly interferes with subsequent attachment by other C. difficile strains.…”