Clostridium perfringens type A: in vitro system for sporulation and enterotoxin synthesis

Abstract

Polysomes were isolated from an enterotoxigenic strain of Clostridium perfringens during vegetative growth and at 1-h intervals after transfer into Duncan-Strong sporulation medium. During vegetative growth, about 67% of the ribosomes were in polysomal complexes. This proportion decreased to about 20% during the first 2 h in sporulation medium and then gradually increased to a maximum of 45% at 6 h. Ribosomes isolated from cells in vegetative or in sporulation phase could equally translate vegetative, sporulation, and natural viral R17 messenger ribonucleic acid with either vegetative or sporulation initiation factors. When polysomes were allowed to complete their nascent chains with labeled amino acids in vitro, most of the polypeptides synthesized by the vegetative phase and by the sporulation phase polysomes appeared to be identical. There were, however, notable differences upon further investigation. Specifically, when antiserum against the enterotoxin was reacted with the completed polypeptides, no counts were precipitated from the vegetative products. On the other hand, up to 12% of the total labeled protein was precipitated from the products obtained with the sporulation phase polysomes. Upon electrophoresis on sodium dodecyl sulfate, the putative enterotoxin synthesized in vitro ran as a major band with a molecular weight of 35,000, and as two minor bands with molecular weights of 17,000 and 52,000, respectively.