Clostridium Difficile Agar is a formula recommended by George et al. for isolation of C. difficile. The medium contains fructose which enhances the recovery and growth of C. difficile. The addition of the antibiotics, cefoxitin and cycloserine, inhibit the growth of Enterobacteriaceae, Enterococcus faecalis, staphylococci, Gram negative non-sporulating anaerobic bacilli, and Clostridia sp. but not C. difficile. Supplementing the medium with horse blood and egg yolk further improves the recovery of C. difficile.

Directions per Liter:

Dissolve 78.1grams per liter of distilled/ deionized (DDI) water, heating with stirring until completely solubilized.

Dispense into appropriate containers, loosen caps and autoclave for 15 minutes at 121ºC (15psi).

Cool to 45-50°C and add 7% of horse blood and Cycloserine (0.5 g/L) and Cefoxitin (0.016 g/L). Alternately, neutral red (0.03 g/L) may be added prior to autoclaving and Cycloserine and Cefoxitin added after autoclaving.

Storage and Stability:

Store powdered media at 4°C. Opened bottles should be capped tightly and kept in a dark, low humidity environment. Prepared media should be kept refrigerated and used within a short period of time.