Magic™ Anti-CPB2 monoclonal antibody, clone N29723UB3

The incidence of a new, yet unassigned toxin type of Clostridium perfringens containing the genes for the alpha-toxin and the recently described beta 2-toxin in horses with intestinal disorders is reported. The study included 18 horses suffering from typical typhlocolitis, 7 horses with atypical typhlocolitis, 16 horses with other intestinal disorders, and 58 horses without intestinal disease. In total, 20 samples of ingesta of the small and large intestines, five biopsy specimens of the intestinal wall, and 74 fecal samples were analyzed bacteriologically, C. perfringens isolates were typed for the presenceof the alpha-, beta-, beta 2-, and epsilon-toxin and enterotoxin genes by PCR, including a newly developed PCR for the detection of the beta 2-toxin gene cpb2, beta 2-Toxigenic C. perfringens was detected in samples from 13 of 25 (52%) horses with typical or atypical typhlocolitis, with a particularly high incidence in specimens of ingesta and biopsy specimens (75%), whereas only 6 of 16 specimens from horses with other intestinal diseases yielded PZ-toxigenic C. perfringens. No beta 2-toxigenic C, perfringens was found in the samples from the 58 control horses, of which only one fecal sample contained C, perfringens type A. Among the samples from the 15 horses with fatal cases of typical and atypical typhlocolitis 9 (60%) were positive for beta 2-toxigenic C. perfringens, whereas samples from only 4 of the 10 (40%) animals with nonfatal cases of infection were positive. We found an interesting correlation between the antibiotic-treated horses which were positive for beta 2-toxigenic C. perfringens and lethal progression of the disease. No C, perfringens strains isolated in this study contained genes for the beta- and epsilon-toxins and enterotoxin. The high incidence of beta 2-toxigenic C. perfringens in samples of ingesta, biopsy specimens of the intestinal wall, and feces from horses suffering or dying from typhlocolitis together with the absence of this organism in healthy horses provides strong evidence that beta 2-toxigenic C. perfringens play an important role in the pathogenesis of typhlocolitis.

Clostridium perfringens type A isolates carrying an enterotoxin (cpe) gene are an important cause of human gastrointestinal diseases, including food poisoning, antibiotic-associated diarrhoea (AAD) and sporadic diarrhoea (SD). Using polymerase chain reaction (PCR), the current study determined that the cpb2 gene encoding the recently discovered beta2 toxin is present in < 15% of food poisoning isolates, which typically carry a chromosomal cpe gene. However, > 75% of AAD/SD isolates, which usually carry a plasmid cpe gene, tested cpb2(+) by PCR. Western blot analysis demonstrated that > 97% of those cpb2(+)/cpe(+) AAD/SD isolates can produce CPB2. Additional PCR analyses, sequencing studies and pulsed field gel electrophoresis experiments determined that AAD/SD isolates carry cpb2 and cpe on the same plasmid when IS1151 sequences are present downstream of cpe, but cpb2 and cpe are located on different plasmids in AAD/SD isolates where IS1470-like sequences are present downstream of cpe. Those analyses also demonstrated that two different CPB2 variants (named CPB2h1 or CPB2h2) can be produced by AAD/SD isolates, dependent on whether IS1470-like or IS1151 sequences are present downstream of their cpe gene. CPB2h1 is similar to 10-fold more cytotoxic for CaCo-2 cells than is CPB2h2. Collectively, these results suggest that CPB2 could be an accessory toxin in C. perfringens enterotoxin (CPE)-associated AAD/SD.