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The effects of storage conditions on viability of Clostridium difficile vegetative cells and spores and toxin activity in human faeces

Freeman, J. and Wilcox, M.H. (2003) The effects of storage conditions on viability of Clostridium difficile vegetative cells and spores and toxin activity in human faeces. Journal of Clinical Pathology, 56 (2). pp. 126-128. ISSN 0021 - 9746

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Abstract

AIMS: Clostridium difficile is a common nosocomial pathogen and as such diagnostic and research methods may necessitate storage of faecal specimens for long periods, followed by subsequent re-examination. This study investigated the effects of storage conditions upon the viability of this organism and its toxin.

METHODS: Three genotypically distinct strains of C difficile (two clinical isolates including the UK epidemic strain, and an environmental isolate) were grown anaerobically at 37°C for 72 hours in a pool of five faecal emulsions. Aliquots of each emulsion were stored at either -20°C (frozen) or 4°C (refrigerated). Emulsions were assayed for viable cells, spores, and cytotoxin titre before storage and at days 1, 3, 5, 7, 14, 28, and 56. An aliquot of each emulsion was also removed, assayed, and replaced in storage at each time point to investigate the effects of multiple freezing/refrigeration/thawing .

RESULTS: Neither storage temperature nor multiple cycles of refrigeration/freezing and thawing adversely affected the viability of C difficilevegetative cells or spores. Single and multiple exposures of samples to 4°C had little effect upon the C difficile toxin titre. Toxin titres of multiply frozen and thawed faeces became significantly lower than for refrigerated faeces (p < 0.01) by day 5 of the experiment in two of the three strains, and in all strains by day 28. Toxin titres of singly frozen faeces became significantly lower than for refrigerated faeces (p < 0.01) by day 56 of the experiment in two of the three strains.

CONCLUSION: Storage temperature and multiple cycles of freezing (refrigeration)/thawing had minimal effects upon the viability of C difficile or its spores. Storage at 4°C has no discernible effect on C difficile cytotoxin. However, storage at -20°C has a detrimental effect upon C difficile cytotoxin, and multiple cycles of freezing and thawing may further adversely effect toxin titres.

Item Type: Article
Copyright, Publisher and Additional Information: © 2003 BMJ Publishing Group & Association of Clinical Pathologists
Institution: The University of Leeds
Academic Units: The University of Leeds > Faculty of Biological Sciences (Leeds) > Institute of Molecular and Cellular Biology (Leeds)
Depositing User: Sherpa Assistant
Date Deposited: 13 Mar 2006
Last Modified: 06 Jun 2014 07:04
Published Version: http://www.jclinpath.com/
Status: Published
Refereed: Yes
URI: http://eprints.whiterose.ac.uk/id/eprint/253

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