Cloning of a DNA sequence unique to Clostridium botulinum group I by selective hybridization

Nucleic acid sequences were isolated from a strain of Clostridium botulinum type A by a selective hybridization method known as deletion enrichment. Nontoxigenic C. sporogenes was used to produce a C. botulinum type A sequence-enriched library. A probe, pCBM44, which showed specific hybridization to a 4.0-kb HindIII fragment present in all of the C. botulinum type A strains tested was isolated, and there was no hybridization to any strains of C. sporogenes. Upon further investigation, pCBM44 was found to hybridize to all of the group I proteolytic C. botulinum strains tested (toxin types A, B, and F) but not to hybridize to groups II, III, and IV (toxin types B, C, D, or E). The probe did not cross-react with nine other Clostridium spp. Such a probe, which differentiates between nontoxigenic C. sporogenes and neurotoxigenic C. botulinum group I strains, should prove extremely useful.