title: Comparison of culture based methods for the isolation of Clostridium difficile from stool samples in a research setting.
abstract: 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.
authors.affiliation: 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.
authors.forename: Michelle
authors.initials: M
authors.lastname: Lister
authors.affiliation: 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.
authors.forename: Emma
authors.initials: E
authors.lastname: Stevenson
authors.affiliation: 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.
authors.forename: Daniela
authors.initials: D
authors.lastname: Heeg
authors.affiliation: 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.
authors.forename: Nigel P
authors.initials: NP
authors.lastname: Minton
authors.affiliation: 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. Electronic address: sarah.kuehne@nottingham.ac.uk.
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Kuehne
mesh_terms: D000818:Animals; D001431:Bacteriological Techniques / Q000191:economics / Q000379:methods*; D016360:Clostridioides difficile / Q000254:growth & development / Q000302:isolation & purification*; D003362:Cost-Benefit Analysis; D003470:Culture Media / Q000737:chemistry; D005243:Feces / Q000382:microbiology*; D008647:Mesocricetus; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D003470:Culture Media
keywords: Clostridium difficile; Clostridium difficile infection; Culture media; Culture methods; Stool culture