Whole genome sequences of three Clade 3 Clostridium difficile strains carrying binary toxin genes in China

Abstract

Clostridium difficile consists of six clades but studies on Clade 3 are limited. Here, we report genome sequences of three Clade 3 C. difficile strains carrying genes encoding toxin A and B and the binary toxin. Isolates 103 and 133 (both of ST5) and isolate 106 (ST285) were recovered from three ICU patients. Whole genome sequencing using HiSeq 2500 revealed 4.1-Mb genomes with 28–29% GC content. There were ≥1,104 SNP between the isolates, suggesting they were not of a single clone. The toxin A and B gene-carrying pathogenicity locus (PaLoc) of the three isolates were identical and had the insertion of the transposon Tn6218. The genetic components of PaLoc among Clade 3 strains were the same with only a few nucleotide mutations and deletions/insertions, suggesting that the Tn6218 insertion might have occurred before the divergence within Clade 3. The binary toxin-genes carrying CDT locus (CdtLoc) of the three isolates were identical and were highly similar to those of other Clade 3 strains, but were more divergent from those of other clades. In conclusion, Clade 3 has an unusual clade-specific PaLoc characteristic of a Tn6218 insertion which appears to be the main feature to distinguish Clade 3 from other C. difficile.

Genome Characterization of a Novel Binary Toxin-Positive Strain of <em>Clostridium difficile</em> and Comparison with the Epidemic 027 and 078 Strains

“…Like the PaLoc reported in other clade 3 strains before [16,33,42], the PaLoc carried by the three ST201 strains discussed in this study as well as another clade 3 strain ZJCDC-S82 contained a mobile genetic element designated Tn6218 inserted between tcdE and tcdA ( Figure 2). It is suggested that the insertion of Tn6218 in PaLoc is clade-specific [16]. Consistence with this, this insertion element was not found in the PaLoc of R20291 in clade 2 and M120 in clade 5.…”

“…What is more, the clade 3 branch also included another China-sourced toxigenic C. difficile strain ZJCDC-S82 which is also reported as a severe diarrhea-associated strain [42]. In addition, the other three recently-reported binary toxin-positive C. difficile (strains 103, 133, and 106) recovered from three ICU patients in China are also clade 3 strains [16]. These findings suggest that C. difficile clade 3 strains might contribute to CDI outbreak in China.…”

“…The 027 has not been detected in China before 2013, and cases of C. difficile 078 have not been reported yet [14]. Instead, a number of severe diarrhea-associated C. difficile toxigenic strains belonging to clades distinct from the 027/ST1 and 078/ST11 strains have been reported in China [16,42]. This might indicate that the dominant genotypes of C. difficile spreading in China are different from those circulating in North America and Europe.…”

“…In addition, the Tn6218 in the three ST201 genomes were found to be flanked by two AT rich sequences. Previous studies suggested that those two AT rich sequences might have inserted into clade 3 PaLoc prior to the insertion of Tn6218 and provide the insertion site of Tn6218 [16]. For the other components of the PaLoc, it is worth to mention that although phylogenetic analysis using either MLST or whole genome comparison demonstrated a closer evolutionary relationship between the ST201 strains and the 027 strain R20291 (Figure 1), the ST201 strains and the 078 strain M120 shared a more homologous tcdA and/or tcdB (Table 5).…”

Characteristics of Clostridium difficile isolates and the burden of hospital-acquired Clostridium difficile infection in a tertiary teaching hospital in Chongqing, Southwest China

“…Data on C. difficile strains in clade 3 with binary toxins are not well documented in mainland China. WGS of three clade 3 C. difficile strains carrying binary toxin genes in a university hospital found that clade 3 has unusual clade-specific PaLoc characteristic of Tn6218 insertion, which may be the main feature to distinguish clade 3 from other C. difficile [41]. The identification of seven novel RTs indicated the diversity of C. difficile strains in this hospital.…”