Genotypic and phenotypic characterization of Clostridium perfringens isolates from Darmbrand cases

Clostridium perfringens type C strains are the only non-type-A isolates that cause human disease. They are responsible for enter-itis necroticans, which was termed Darmbrand when occurring in post-WorldWar II Germany. Darmbrand strains were ini-tially classified as type F because of their exceptional heat resistance but later identified as type C strains. Since only limited in-formation exists regarding Darmbrand strains, this study genetically and phenotypically characterized seven 1940s era Darmbrand-associated strains. Results obtained indicated the following. (i) Five of these Darmbrand isolates belong to type C, carry beta-toxin (cpb) and enterotoxin (cpe) genes on large plasmids, and express both beta-toxin and enterotoxin. The other two isolates are cpe-negative type A. (ii) All seven isolates produce highly heat-resistant spores withD100 values (the time that a culture must be kept at 100°C to reduce its viability by 90%) of 7 to 40 min. (iii) All of the isolates surveyed produce the same variant small acid-soluble protein 4 (Ssp4) made by type A food poisoning isolates with a chromosomal cpe gene that also pro-duce extremely heat-resistant spores. (iv) The Darmbrand isolates share a genetic background with type A chromosomal-cpe-bearing isolates. Finally, it was shown that both the cpe and cpb genes can be mobilized in Darmbrand isolates. These results sug-gest that C. perfringens type A and C strains that cause human food-borne illness share a spore heat resistance mechanism that likely favors their survival in temperature-abused food. They also suggest possible evolutionary relationships between Darm-brand strains and type A strains carrying a chromosomal cpe gene. Clostridium perfringens, an anaerobic, Gram-positive bacte-rium, exists ubiquitously in soils, sewage, food, and the nor