This study has investigated the isolation and characterisation of organisms resembling C. botulinum type C from the gastrointestinal tract of horse with grass sickness and of control horses. Five of the organisms isolated from the GI tract of horses with equine grass sickness were found to produce detectable levels of BoNT/C by ELISA. The BoNT/C gene was detected by PCR and Southern blotting in four of the seven horses with equine grass sickness and none of the control horses investigated. The BoNT/C gene was detected in several of the sites sampled along the GI tract and was detected in the duodenum of all four horses. These results support the hypothesis that a toxicoinfection with C. botulinum type C is associated with equine grass sickness. Forty four strains identified as group III clortridia by phenotype were investigated fro the distribution of toxin genes by PCR. Several combinations of the species-defining genes and the C2 genes were found. The reference C. botulinum types C and D strains and one non-neurotoxic strain of C. botulinum type C carried the genes for both components of the C2 toxin. Of the 40 C. novyi type A strains, the gene for the alpha toxin was found in 22 with 19 of these strains carrying the gene for one component of the C2 toxin. .In the 22 alpha toxin-negative strains, 2 carried the genes for both components of the C2 toxin and a further 11 strains carried the gene for the component I only. The detection of the C2 toxin genes in C. novyi type A is novel and may suggest a role for one or both components of the C2 toxin in disease.