Enumeration and Characterization of Clostridium perfringens Spores in the Feces of Food Poisoning Patients and Normal Controls

Abstract

The fecal spore enumeration method for confirming Clostridium perfringens as the cause of food poisoning was evaluated using strains implicated in nine outbreaks in the United States. Confirmed spore counts from 66 stool specimens were made on tryptose-sulfite-cycloserine (TSC) without egg yolk and trypticase soy-sheep blood (TSB) agars. Counts from outbreak stools on TSC agar ranged from 2.0 × 104 to 3.5 × 108 (mean ≥ 1.4 × 106/g) as compared with <103 to 5.0 × 105/g (overall mean 9.5 × 103/g) from normal stools. Similar results were obtained with TSB agar. Isolates from seven of the nine outbreaks were nonhemolytic and produced ≥100 ng enterotoxin/ml in spore broth, as measured by an enzyme-linked immunosorbent assay. Spores in stools from six of the outbreaks were heat-resistant and survived heating for 30 to 60 min at 100°C in cooked meat medium. Strains from the three remaining outbreaks were heat-sensitive and survived heating for only 15 min at 100°C. Enterotoxigenic isolates from all but one of the outbreaks were serotyped. In all instances, the predominant strain in specimens from an outbreak was of the same serotype, indicating that it was the causative strain. Reexamination of five specimens from each of three outbreaks after storage at −20°C for 6 months showed only a minimal reduction in the spore counts.