Comparison of culture based methods for the isolation of Clostridium difficile from stool samples in a research setting

Research output: Contribution to journalArticlepeer-review


  • Michelle Lister
  • Emma Stevenson
  • Daniela Heeg
  • Nigel P Minton
  • Sarah Kuehne

Colleges, School and Institutes

External organisations

  • Clostridia Research Group, Centre for Biomolecular Sciences, School of Life Sciences, Nottingham Digestive Diseases Centre, NIHR Biomedical Research Unit, The University of Nottingham, Nottingham, United Kingdom.
  • University of Nottingham


Effective isolation of Clostridium difficile from stool samples is important in the research setting, especially where low numbers of spores/vegetative cells may be present within a sample. In this study, three protocols for stool culture were investigated to find a sensitive, cost effective and timely method of C. difficile isolation. For the initial enrichment step, the effectiveness of two different rich media, cycloserine-cefoxitin fructose broth (CCFB) and cycloserine-cefoxitin mannitol broth with taurocholate and lysozyme (CCMB-TAL) were compared. For the comparison of four different, selective solid media; Cycloserine-cefoxitin fructose agar (CCFA), Cycloserine-cefoxitin egg yolk agar (CCEY), ChromID C. difficile and tryptone soy agar (TSA) with 5% sheep's blood with and without preceding broth enrichment were used. As a means to enable differentiation between C. difficile and other fecal flora, the effectiveness of the inclusion of a pH indictor (1% Neutral Red), was also evaluated. The data derived indicated that CCFB is more sensitive than CCMB-TAL, however, the latter had an improved recovery rate. A broth enrichment step had a reduced sensitivity over direct plating. ChromID C. difficile showed the best recovery rate whereas CCEY egg yolk agar was the most sensitive of the four. The addition of 1% Neutral Red did not show sufficient colour change when added to CCEY egg yolk agar to be used as a differential medium. For a low cost, timely and sensitive method of isolating C. difficile from stool samples we recommend direct plating onto CCEY egg yolk agar after heat shock.


Original languageEnglish
Pages (from-to)226-229
Number of pages4
Early online date16 Jul 2014
Publication statusPublished - Aug 2014


  • Animals, Bacteriological Techniques, Clostridium difficile, Cost-Benefit Analysis, Culture Media, Feces, Mesocricetus, Sensitivity and Specificity, Time Factors, Comparative Study, Evaluation Studies, Journal Article, Research Support, Non-U.S. Gov't