Transfer to neurotoxigenicity from Clostridium butyricum to a nontoxigenic Clostridium botulinum type E-like strain

Two Clostridium butyricum strains from infant botulism cases produce a toxic molecule very similar to C. botulinum type E neurotoxin. Chromosomal, plasmid, and bacteriophage DNAs of toxigenic and nontoxigenic strains of C. butyricum and C. botulinum type E were probed with (i) a synthesized 30-mer oligonucleotide encoding part of the L chain of type E botulinum toxin and (ii) the DNA of phages lysogenizing these cultures. The toxin gene probe hybridized to the chromosomal DNA of toxigenic strains but not to their plasmid DNA. All toxigenic and most nontoxigenic strains tested were lysogenized by a prophage on the chromosome. Prophages of toxigenic strains, irrespective of species, had related or identical DNAs which differed from the DNAs of prophages in nontoxigenic strains. The prophage of toxigenic strains was adjacent or close to the toxin gene on the chromosome. Phage DNAs purified from toxigenic strains did not hybridize with the toxin gene probe but could act as the template of the polymerase chain reaction to amplify the toxin gene. The toxin gene was not transferred between C. botulinum and C. butyricum (either direction) when different pairs of a possible gene donor and a recipient strain were grown as mixed cultures. Nontoxigenic C. butyricum or C. botulinum type E-like strains did not become toxigenic when grown in both containing the phage induced from a toxigenic strain of the other species. Interspecies transfer of the toxin gene did occur when a recipient strain (nontoxigenic C. botulinum type E-like with chloramphenicol resistance) was incubated in filter-sterilized culture fluid obtained by growing a 'helper' strain (nontoxigenic C. butyricum) in broth containing phage induced from the toxin gene donor (toxigenic C. butyricum). The toxin gene was detected in 1 of about 10(4) colonies obtained by plating out the recipient strain.