Molecular analysis of the virulence determinants of Clostridium perfringens associated with foal diarrhoea

During an epidemiological study of foal diarrhoea, over half of the cases yielded Clostridium perfringens which was significantly associated with disease (Netherwood et al., 1996b). However, the association could not be accounted for by enterotoxigenic isolates which had a low prevalence (Netherwood et al., 1997). Nonetheless, we have hypothesized that the association may be caused by a pathogenic sub-population which would be significantly more common amongst C,. perfringens-positive cases compared with C. perfringens-positive healthy controls if it acted as a pathogen when present. Conversely, if foal diarrhoea caused by C. perfringens was dependent on a predisposing factor, then such an association might not be evident. As a first step to determine if a molecular marker was more frequently to be found in C. perfringens-positive cases than controls, we have genotyped the study isolates (up to five per foal) by polymerase chain reaction (PCR) based on the published gene sequences for the major lethal toxins alpha, beta, epsilon and iota as well as for theta toxin, large and small sialidases, hyaluronidase and virulence regulation. Isolates of major toxin types B, C, D and E, or isolates which were untypeable, were isolated from less than 15% of C. perfringens- positive foals and these were not associated with diarrhoea nor were they more commonly found in C. perfringens- positive cases. Isolates of type A were found in more than 90% of all C perfringens-positive foals. A number of different genotypes were identified by their different patterns of gene possession but types without any of the genes for theta toxin, large and small sialidases, hyaluronidase and virulence regulation were found in only 10% of positive foals. Only type A isolates with all of these genes were associated with diarrhoea overall but they were not more commonly isolated from C. perfringens-positive cases than controls. In conclusion, genotyping by the sequenced virulence genes did not identify a marker for a sub-population of C. perfringens which may be acting more frequently as a pathogen when present.