A plasmid screening was performed on 150 strains out of 75 clostridial species using a modification of the alkaline-lysis procedure. In 26 strains representing 21 species one or more plasmid bands were detected ranging in size from 3 to more than 100 kilobase pairs. Clostridium aceticum proved to contain a single small plasmid (pCA1) of 5.4 kbp as revealed by restriction analysis and electron microscopy. A physical map of pCA1 has been constructed. Spontaneous mutants of C. aceticum defective in autotrophic growth have been isolated. No direct correlation between plasmid content and autotrophy could be found.