The MLST of seven housekeeping

The MLST of seven housekeeping genes of 53 ribotypes revealed 40 different STs clustering to 5 clades. Although the MLST was performed only in one isolate of each identified CE-ribotyping profile, we found the correlation with STs identified in ribotypes represented in the Leeds-Leiden C. difficile reference strain collection published by Knetsch et al. (Table 1, marked with b) (Knetsch et al., 2012). The most heterogeneous was MLST clade 1, which included 44 CE-ribotyping profiles of 53 CE-ribotyping profiles. MLST clade 1 heterogeneity was also observed in the study by Stabler et al., who found that this clade contained 106 STs of the 141 studied STs (Stabler et al., 2012). Knetsch et al. typed 35 STs out of 56 as belonging to clade 1 (Knetsch et al., 2012), whereas Griffiths et al. concluded that 31 STs out of 40 belonged to clade 1 (Griffiths et al., 2010). Similarly, Dingle et al. found 60 STs out of 69 belonging to clade 1 (Dingle et al., 2011). Several isolates belonging to a different RT or WTR revealed the same ST (clade) and the specific deletion in tcdC gene that suggests their phylogenetic relationship. RTs 027 and 176 revealed ST1 (clade 2), as was published by Knetsch et al. (Knetsch et al., 2012), as well as the presence of one GSK1324726A pair deletion at nucleotide position 117, which is a target site for commercial molecular systems (Krutova et al., 2014a, Mentula et al., 2015), and 18bp deletions in the tcdC gene. RT 023 and WRT 438 revealed ST5 (clade 3) and had 54bp deletions in the tcdC gene. RTs 078, 126 and WRT 413 showed ST11 and 39bp deletions in the tcdC gene. Isolates harbouring 54bp and 39bp deletions (except WRT 413) as previously described above revealed a nonsense mutation C184T (Spigaglia and Mastrantonio, 2002, Curry et al., 2007). All these isolates (RT 023, 027, 126, 176 and WRTs 413 and 438) revealed the presence of binary toxin genes, another important C. difficile virulence factor (Gerding et al., 2014). The Czech Republic is a country with increasing CDI incidence (1.1 cases per 10,000 patient bed-days in 2008–4.4 cases in 2011–2012 and 6.2 cases per 10,000 patient bed-days in 2012–2013) (Bauer et al., 2011, Davies et al., 2014) and relatively high rates of antibiotic resistant C. difficile strains (Freeman et al., 2015). Implementation of CDI surveillance based on the recently released CDI surveillance protocol Control (ECDC, 2015) in the Czech Republic would fill the gap in Czech CDI epidemiology with national CDI incidence data, including clinical case information and C. difficile isolate antibiotic susceptibility results.
Conclusion The molecular characterisation of 2201 Czech clinical C. difficile isolates revealed 53 different CE-ribotyping profiles and 40 multi-locus sequence types. Of 2201C. difficile isolates, 2024 were toxigenic (tcdA and tcdB), and of these, 677 isolates carried genes for binary toxin production (cdtA, cdtB). The results of molecular characterisation showed a high diversity of C. difficile strains circulating in the Czech Republic with prevailing representation of RTs 001 and 176 (027-like).