Microbiology laboratories where MALDI-TOF MS is available can benefit from its capacity to identify most clinically interesting non-tuberculous mycobacteria in a rapid, reliable, and inexpensive manner.

first aligned the genome sequences of the 250 isolates against the NRCS-A reference genome CR01, resulting in a total of 22,621 single nucleotide polymorphisms (SNPs). To quantify recombination, we used ClonalFrameML 12 , which is specifically aimed at analysing whole-genome sequence data (see Supplementary Information). The results indicated that the impact of recombination (r) on the genome-wide substitution rate in S. capitis overall is almost equal to the impact of mutation (m), with r/m = 0.85. ClonalFrameML identified 190 recombination events in the global genealogy (Extended Data Fig. 1). The largest detected events (up to 26 kb) are probably products of horizontal gene transfer, some of which correspond to the insertion of pathogenicity islands. Clonal specialization and geographical dispersion of NRCS-A.The reconstructed maximum-likelihood tree (Fig. 1a) enabled us to draw a clear distinction between NRCS-A isolates that harbour the previously described specific NRCS-A pulsed-field gel electrophoresis pattern 8 (n = 197) and all the other strains found in basal positions (n = 53; hereafter 'basal'). These reconstructions revealed that this NRCS-A population is composed of at least three sublineages, which we named in chronological order of divergence on the basis of the observed branching order in the tree: 'proto-outbreak 1' (n = 18), 'proto-outbreak 2' (n = 17) and 'outbreak' (n = 162) (Fig. 1a,b). These three clades are supported both by bootstrap values greater than 95% and by the trimodal distribution of the

Clostridium difficile infection (CDI) remains poorly controlled in many European countries, of which several have not yet implemented national CDI surveillance. In 2013, experts from the European CDI Surveillance Network project and from the European Centre for Disease Prevention and Control developed a protocol with three options of CDI surveillance for acute care hospitals: a 'minimal' option (aggregated hospital data), a 'light' option (including patient data for CDI cases) and an 'enhanced' option (including microbiological data on the first 10 CDI episodes per hospital). A total of 37 hospitals in 14 European countries tested these options for a three-month period (between 13 May and 1 November 2013). All 37 hospitals successfully completed the minimal surveillance option (for 1,152 patients). Clinical data were submitted for 94% (1,078/1,152) of the patients in the light option; information on CDI origin and outcome was complete for 94% (1,016/1,078) and 98% (294/300) of the patients in the light and enhanced options, respectively. The workload of the options was 1.1, 2.0 and 3.0 person-days per 10,000 hospital discharges, respectively. Enhanced surveillance was tested and was successful in 32 of the hospitals, showing that C. difficile PCR ribotype 027 was predominant (30% (79/267)). This study showed that standardised multicountry surveillance, with the option of integrating clinical and molecular data, is a feasible strategy for monitoring CDI in Europe.

Outbreaks of Clostridium difficile infections (CDI) with increased severity, high relapse rate and significant mortality have been related to the emergence of a new, hypervirulent C. difficile strain in North America and Europe. This emerging strain is referred to as PCR ribotype 027 (Type 027). Since 2005, individual countries have developed surveillance studies about the spread of type 027. C. difficile Type 027 has been reported in 16 European countries. It has been responsible for outbreaks in Belgium, Germany, Finland, France, Ireland, Luxembourg, The Netherlands, Switzerland and the United Kingdom (England, Wales, Northern Ireland and Scotland). It has also been detected in Austria, Denmark, Sweden, Norway, Hungary, Poland and Spain. Three countries experienced imported patients with CDI due to Type 027 who acquired the infection abroad. The antimicrobial resistance pattern is changing, and outbreaks due to clindamycin-resistant ermB positive Type 027 strains have occurred in three European countries. Ongoing epidemiological surveillance of cases of CDI, with periodic characterisation of the strains involved, is required to detect clustering of cases in time and space and to monitor the emergence of new, highly virulent clones.

Objectives: Clostridium difficile is a major global human pathogen divided into five clades, of which clade 3 is the least characterized and consists predominantly of PCR ribotype (RT) 023 strains. Our aim was to analyse and characterize this clade. Methods: In this cohort study the clinical presentation of C. difficile RT023 infections was analysed in comparison with known 'hypervirulent' and non-hypervirulent strains, using data from the Netherlands national C. difficile surveillance programme. European RT023 strains of diverse origin were collected and whole-genome sequenced to determine the genetic similarity between isolates. Distinctive features were investigated and characterized. Results: Clinical presentation of C. difficile RT023 infections show severe infections akin to those seen with 'hypervirulent' strains from clades 2 (RT027) and 5 (RT078) (35%, 29% and 27% severe CDI, respectively), particularly with significantly more bloody diarrhoea than RT078 and non-hypervirulent strains (RT023 8%, other RTs 4%, p 0.036). The full genome sequence of strain CD305 is presented as a robust reference. Phylogenetic comparison of CD305 and a further 79 previously uncharacterized European RT023 strains of diverse origin revealed minor genetic divergence with >99.8% pairwise identity between strains. Analyses revealed distinctive features among clade 3 strains, including conserved pathogenicity locus, binary toxin and phage insertion toxin genotypes, glycosylation of S-layer proteins, presence of the RT078 four-gene trehalose cluster and an esculinase-negative genotype. Conclusions: Given their recent emergence, virulence and genomic characteristics, the surveillance of clade 3 strains should be more highly prioritized.