Reconsidering the Sporulation Characteristics of Hypervirulent Clostridium difficile BI/NAP1/027
Minton NP (2011) Reconsidering the Sporulation Characteristics of Hypervirulent Clostridium difficile BI/NAP1/027. PLoS
ONE 6(9): e24894. doi:10.1371/journal.pone.0024894
Reconsidering the Sporulation Characteristics of Hypervirulent Clostridium difficile BI/NAP1/027
David A. Burns. 0
Daniela Heeg. 0
Stephen T. Cartman 0
Nigel P. Minton 0
Peter Setlow, University of Connecticut, United States of America
0 Clostridia Research Group, School of Molecular Medical Sciences, Centre for Biomolecular Sciences, University of Nottingham , Nottingham , United Kingdom
Clostridium difficile is the leading cause of antibiotic-associated diarrhoea and a major burden to healthcare services worldwide. In recent years, C. difficile strains belonging to the BI/NAP1/027 type have become highly represented among clinical isolates. These so-called 'hypervirulent' strains are associated with outbreaks of increased disease severity, higher relapse rates and an expanded repertoire of antibiotic resistance. Spores, formed during sporulation, play a pivotal role in disease transmission and it has been suggested that BI/NAP1/027 strains are more prolific in terms of sporulation in vitro than 'non-epidemic' C. difficile types. Work in our laboratory has since provided credible evidence to the contrary suggesting that the strain-to-strain variation in C. difficile sporulation characteristics is not type-associated. However, the BI/NAP1/027 type is still widely stated to have an increased rate of sporulation. In this study, we analysed the sporulation rates of 53 C. difficile strains, the largest sample size used to-date in such a study, including 28 BI/NAP1/027 isolates. Our data confirm that significant variation exists in the rate at which different C. difficile strains form spores. However, we clearly show that the sporulation rate of the BI/NAP1/027 type was no higher than that of non-BI/NAP1/027 strains. In addition, we observed substantial variation in sporulation characteristics within the BI/NAP1/027 type. This work highlights the danger of assuming that all strains of one type behave similarly without studying adequate sample sizes. Furthermore, we stress the need for more rigorous experimental procedures in order to quantify C. difficile sporulation more accurately in the future.
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Funding: The research leading to these results has received funding from the UK Medical Research Council (grant G0601176) and the European Communitys
Seventh Framework Programmes CLOSTNET (PEOPLE-ITN-2008-237942) and HYPERDIFF (HEALTH-F3-2008-223585). The funders had no role in study design,
data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
. These authors contributed equally to this work.
Clostridium difficile, a Gram-positive, spore-forming bacterium, is
the major underlying cause of antibiotic-associated diarrhoea.
Outbreaks of C. difficile infection (CDI) have led to patient
isolation, ward closures and, sometimes, hospital closure. In the
United States of America alone, CDI is estimated to affect over
500,000 people each year and cost the healthcare system over $3
billion per year [1,2]. Endospores, formed during sporulation, are
able to resist a variety of industrial cleaning agents and can persist
on surfaces in healthcare settings for prolonged periods of time
[3,4]. Following ingestion by susceptible individuals, spores return
to vegetative cell growth through germination which allows for
colonisation and production of the characteristic toxins [5].
Consequently, CDI can cause intestinal perforation, toxic
megacolon and a potentially fatal pseudo-membranous colitis
[6]. The spore form of C. difficile is, therefore, crucial for disease
transmission.
The challenge of CDI has increased with the emergence of
socalled hypervirulent strains belonging to restriction endonuclease
type BI, North American pulsed-field type 1 and PCR-ribotype
027 (BI/NAP1/027). Strains of the BI/NAP1/027 type have
become highly represented among clinical isolates from recent
outbreaks and are associated with an expanded repertoire of
antibiotic resistance, more severe disease and higher relapse rates
[7,8]. Unsurprisingly, there is widespread interest in
understanding the underlying factors that have led to the emergence of strains
such as those of the BI/NAP1/027 type.
Some BI/NAP1/027 strains are believed to produce higher
levels of toxin in the laboratory than strains belonging to other
types [9] and a number of recent studies have concluded that
strains of the BI/NAP1/027 type are also more prolific in terms of
sporulation in vitro [10,11,12,13,14,15]. However, work in our
laboratory has since provided credible evidence to the contrary
suggesting that the strain-to-strain variation in C. difficile
sporulation characteristics is not type-associated [16,17]. In spite
of this evidence, the BI/NAP1/027 type is still widely stated to
have an i (...truncated)