Prevalence and Characterization of a Binary Toxin (Actin-Specific ADP-Ribosyltransferase) from Clostridium difficile

Abstract

In addition to the two large clostridial cytotoxins (TcdA and TcdB), some strains of Clostridium difficile also produce an actin-specific ADP-ribosyltransferase, called binary toxin CDT. We used a PCR method and Southern blotting for the detection of genes encoding the enzymatic (CDTa) and binding (CDTb) components of the binary toxin in 369 strains isolated from patients with suspected C. difficile-associated diarrhea or colitis. Twenty-two strains (a prevalence of 6%) harbored both genes. When binary toxin production was assessed by Western blotting, 19 of the 22 strains reacted with antisera against the iota toxin of C. perfringens (anti-Ia and anti-Ib). Additionally, binary toxin activity, detected by the ADP-ribosyltransferase assay, was present in only 17 of the 22 strains. Subsequently, all 22 binary toxin-positive strains were tested for the production of toxins TcdA and TcdB, toxinotyped, and characterized by serogrouping, PCR ribotyping, arbitrarily primed PCR, and pulsed-field gel electrophoresis. All binary toxin-positive strains also produced TcdB and/or TcdA. However, they had significant changes in the tcdA and tcdB genes and belonged to variant toxinotypes III, IV, V, VII, IX, and XIII. We could differentiate 16 profiles by using typing methods, indicating that most of the binary toxin-positive strains were unrelated.

Toxigenic status of Clostridium difficile in a large Spanish teaching hospital

“…0 to 12 . 0 % (Goncalves et al, 2004;Stubbs et al, 2000). All our strains had both components of the binary toxin, which was not always the case in the reports of other authors (Perelle et al, 1997).…”

“…The genetic diversity of strains producing both LCTs varies in different series, depending on the sporadic or epidemic character of each one (Kato et al, 2001;Spigaglia et al, 2001). Several authors report a high homogeneity of variant strains (Alfa et al, 2000;Pituch et al, 2001) and one study reported that strains producing binary toxin are usually epidemiologically unrelated and not derived from a common ancestor (Goncalves et al, 2004), although in this report the isolates were obtained from 17 different institutions. LCTs and binary toxin prevalence in C. difficile…”

Variant forms of the binary toxin CDT locus and tcdC gene in Clostridium difficile strains

“…It has been recognized previously that the presence of functional binary toxin genes encoded by the 4.3 kb CDT locus correlates well with variant toxinotypes including A + B + , A 2 B + and A 2 B 2 strains (Stubbs et al, 2000;Rupnik et al, 2001Rupnik et al, , 2003aGeric et al, 2004;Spigaglia & Mastrantonio, 2002;Goncalves et al, 2004). This study presents an overview of all of the known variant toxinotypes and has confirmed that the full-length CDT locus is found in variant toxinotypes with significantly changed LCT toxin genes (shown in bold in Table 1), whereas the truncated version of the CDT locus is predominant in variant toxinotypes with minor changes in the LCT toxin genes (Table 1).…”

“…Interestingly, binary toxin-producing strains often seem to be associated with community-acquired C. difficile infections (Terhes et al, 2004;Barbut et al, 2005). The prevalence of binary toxin-producing strains has been estimated to be between 1.6 and 6 % (Stubbs et al, 2000;Rupnik et al, 2001Rupnik et al, , 2003bGeric et al, 2003;Goncalves et al, 2004;Terhes et al, 2004; Alonso et al, 2005) et al, 2006), regions with significant similarity to binary toxin have been found to be present, representing a truncated CDT locus (Fig. 1).…”

Detection of binary-toxin genes (cdtA and cdtB) among Clostridium difficile strains isolated from patients with C. difficile-associated diarrhoea (CDAD) in Poland

“…Five A þ B þ isolates (8 . 6 %) harboured binary-toxin genes, a prevalence similar to that reported by Goncalves et al (2004). Binary-toxin-positive strains were isolated from adult patients hospitalized at different times (between 1999 and 2001) and in different units, and were considered epidemiologically unrelated.…”