tcdC Genotypes Associated with Severe TcdC Truncation in an Epidemic Clone and Other Strains of Clostridium difficile

Abstract

Severe Clostridium difficile associated disease is associated with outbreaks of the recently described BI/NAP1 epidemic clone. This clone is characterized by an 18-bp deletion in the tcdC gene and increased production of toxins A and B in vitro. TcdC is a putative negative regulator of toxin A&B production. We characterized tcdC genotypes from a collection of C. difficile isolates from a hospital that experienced an outbreak caused by the BI/NAP1 epidemic clone. Sequence analysis of tcdC was performed on DNA samples isolated from 199 toxigenic C. difficile isolates (31% BI/NAP1) from 2001 and 2005. Sequences obtained from 36 (18.6%) isolates predicted wild-type TcdC (232 amino acid residues), whereas 12 (6.1%) isolates had tcdC genotypes with previously described 18-or 39-bp deletions. The remaining isolates comprised 15 unique genotypes. Of these, 5 genotypes contain 18-or 36-bp deletions. Of these five genotypes, one is characterized by a single nucleotide deletion at position 117 resulting in a frameshift that introduces a stop codon at position 196, truncating the predicted TcdC to 65 amino acid residues. All 62 of the isolates in this collection comprising the epidemic clone are characterized by this genotype. This result suggests that severe truncation of TcdC is responsible for the increased toxin production observed in strains belonging to the BI/NAP1 clone and that the 18-bp deletion is probably irrelevant to TcdC function. Further investigations are required to determine the effect of this and other tcdC genotypes on toxin production and clinical disease.

Fourteen-Genome Comparison Identifies DNA Markers for Severe-Disease-Associated Strains of Clostridium difficile

“…Future work may reveal insertions or deletions in SDA strains in addition to those previously identified in tcdC (5,32,33,36). Indeed, whereas deletions in tcdC have previously been hypothesized to lead to increased toxin production (33,57,62), findings from a recent study (39) indicated that deletions in tcdC do not predict the biological activity of the PaLoc toxin genes, providing further motivation to identify all sources of genetic variation within the C. difficile genomes that may correlate with disease severity.…”

Toxinotype VClostridium difficilein Humans and Food Animals

“…Current literature suggests that this considerable truncation of TcdC may impair its negative regulatory function and contribute to the increased toxin production observed in BI/NAP1/027 strains (27,36). Molecular analysis of toxinotype V C. diffi cile has demonstrated a similarly truncated TcdC (61 aa compared with 65 aa in BI/NAP1/027 strains and 232 aa in wild-type TcdC) (15), which may imply hypervirulence for this strain as well.…”

Multiplex PCR Method for Detection of Clostridium difficile tcdA, tcdB , cdtA , and cdtB and Internal In-Frame Deletion of tcdC

“…In this respect, PCR ribotype 027 (North America pulsed-field type 1), which has been responsible for the most severe outbreaks around the world, is often reported to have an 18-bp in-frame deletion and ⌬117. Evidently, such genetic changes in the negative regulator have been proposed to be responsible for increased toxin production and hence hypervirulence (9,15), but a clear association with clinical manifestations has not been determined (8,14,16,20). No matter what role tcdC deletions and mutations may have in relation to clinical outcome, the internal in-frame deletion of 18 bp and ⌬117 have been targets for real-time PCR screening for PCR ribotype 027 (4,11,18,21).…”