Influence of peptone source on sporulation of Clostridium perfringens type A

Clostridium perfringens is a foodborne disease agent that produces a sporulation-specific enterotoxin. To produce enterotoxin for experimental purposes or spores for challenge or physiological studies, the use of a convenient sporulation medium is required. The most commonly used is Duncan-Strong medium. Few isolates sporulate at high levels in this medium. We investigated the effectiveness of peptones from a variety of sources on the sporulation of this organism compared with the peptone in the original formulation, proteose peptone (control). Seven strains were used to screen 32 peptones, with starch or raffinose as the carbohydrate source. In most cases, raffinose was more effective than starch in stimulating sporulation, confirming our previous study. Two promising peptones, potato peptone, and Proteose Peptone no. 3, were selected and tested against 49 additional enterotoxin-positive and -negative strains, with raffinose as the carbohydrate. For 49 strains, 5 sporulated best (>10%) in the control peptone, 6 sporulated best in Peptone no. 3, and 23 sporulated best in the potato peptone. Of the 23 strains, 16 sporulated at levels 25% more than the control peptone. The increase in sporulation rates was reflected in the enterotoxin and heat-resistant spore levels. The methylxanthines caffeine and theobromine were effective in increasing the sporulation of less than half of 19 enterotoxin-positive strains. Our results suggest that the replacement of proteose peptone with potato peptone be considered if difficulty in obtaining spores of specific strains of C. perfringens is encountered.