GW is the Principal Investigator of the funded

projects. Antifection chemical She coordinated the study and helped to draft the manuscript. All authors read and approved the final manuscript.”
“Background Clostridium difficile is a Gram-positive, spore-forming, obligately anaerobic bacterium. It is the leading cause of nosocomial diarrhoea among patients undergoing antibiotic treatment [1, 2]. The severity of C. difficile-associated disease (CDAD) ranges from mild diarrhoea to pseudomembranous colitis, toxic megacolon, and intestinal perforation [3–6]. Mortality rates of CDAD reportedly range from 6 to 30% [5, 7, 8]. During the last decade, the incidence of CDAD has increased significantly in North America [9–12] and Europe [4, 8, 13, 14]. In the USA and Canada, this increase has been associated with the emergence of a novel, hypervirulent strain designated NAP1/027 [11, 15]. Strains with the same genotype and associated outbreaks have also been reported from several European countries [14, 16–18]. For infection control investigations and epidemiological studies, it is mandatory to track the emergence and spread of epidemic strains. For this purpose, appropriate genotyping methods are needed. The utility of a typing method will depend on its inter-laboratory reproducibility and data portability, its discriminatory power and concordance

of identified groupings with epidemiology, the temporal stability of the genetic markers investigated, JAK inhibitor and the universal typeability of isolates [19]. Multilocus variable number of tandem repeats Interleukin-2 receptor analysis (MLVA) is the most discriminatory method presently available for typing C. difficile [20, 21]. Recently reported results suggested that the level of resolution achieved through MLVA may be highly useful for detecting epidemiological clusters of CDAD within and between hospitals [21, 22]. The genetic loci currently exploited for MLVA-typing of C. difficile accumulate variation so rapidly, however, that

longer-term relationships between isolates get obscured [23]. It is therefore advisable – and has been a common practice – to combine MLVA with the analysis of more conserved genetic markers [20–23]. Most commonly applied approaches to genotyping C. difficile at present are DNA macrorestriction analysis (based on pulsed-field gel electrophoresis, mostly used in Canada and the USA [12, 15, 24]) and PCR ribotyping (in Europe [25–27]). These two methods yield largely concordant results [23, 27]. While DNA macrorestriction has slightly higher discriminatory power than PCR ribotyping, it is also more labour-intensive and time consuming [23, 27–29]. A major disadvantage of PCR ribotyping, DNA macrorestriction, and other band-based typing techniques (including restriction endonuclease analysis (REA) [30]) is the poor portability and interlaboratory comparability of the generated data.