Inhibition of enhanced toxin production by Clostridium difficile in biotin-limited conditions

Abstract

Production of toxins A and B by Clostridium difficile is enhanced in a defined medium with biotin-limited conditions. In the present study compounds inhibitory to enhanced toxin production by a C. dij$jjcile strain were examined. Increases in biotin concentration from 0.05 nM to 50 nM accelerated growth and inhibited enhanced toxin production. Asparagine, glutamic acid and glutamine (10 mM) showed an effect on growth and toxin production similar to that of biotin. Lysine (10 mM) suppressed growth and inhibited toxin production. Addition of these toxin-inhibitory compounds within an incubation period of 2 days inhibited the enhanced toxin production, but later addition showed only slight inhibition of toxin production. Amino acids contained in the defined medium under the biotin-limited conditions were actively utilised in the presence of the three toxin-inhibitory amino acids, but in the presence of lysine, amino-acid utilisation was suppressed. Different mechanisms of action of these toxin-inhibitory molecules, which may be divided into excess biotin, asparagine-glutamic acid-glutamine group, and lysine, are discussed.

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

“…Toxin mRNA levels have been shown to respond to a variety of environmental signals such as temperature, nutrient concentrations and antibiotics (Karlsson et al, 1999(Karlsson et al, , 2003Yamakawa et al, 1998;Nakamura et al, 1982). We checked for the downregulation of toxin mRNAs by nutrient depletion or other signalling molecules using real-time PCR.…”

Induction of toxins in Clostridium difficile is associated with dramatic changes of its metabolism

“…Toxin synthesis is normally upregulated during entry into the stationary growth phase in tryptone/peptone-yeast media (Dupuy & Sonenshein, 1998;Karlsson et al, 1999), and is associated with the induction of many other C. difficile proteins that are repressed together with the toxins by the presence of glucose, cysteine and proline (Karlsson et al, 2000). In addition, arginine, glutamine, biotin and carbonate influence toxin production, further supporting a link between carbon/amino acid metabolism and toxin production (Ikeda et al, 1998;Karasawa et al, 1997;Karlsson et al, 1999;Maegawa et al, 2002;Yamakawa et al, 1996Yamakawa et al, , 1998.…”

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

“…Toxin expression may differ 100,000-fold between toxin-positive isolates of C. difficile in vitro (14) but the underlying reason is not understood. Previous studies have shown that regulation of toxin production in C. difficile may be affected by amino acid levels, as demonstrated in defined media during biotin starvation (25) and in complex media (12,18). High toxin levels are observed in complex media deprived of glucose (4,12).…”

“…The onset of C. difficile growth in the large intestine, followed by C. difficile-associated diarrhea (CDAD), is thought to be caused by the reduction of protective colonic microbiota, especially by antibiotic treatment (14). 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.…”