Peroral administration of purified enterotoxin to human volunteers provoked diarrhoea and abdominal pain, symptoms identical with those encountered in outbreaks of Clostridium perfringens food poisoning. Eight milligrams of enterotoxin caused diarrhoea in one of two volunteers. All of five subjects given 10 and 12 mg of purified enterotoxin or crude enterotoxin developed the classical symptoms of this food poisoning. Passive haemagglutination anti‐enterotoxin titres in serum increased in only 5 of 9 volunteers after exposure to enterotoxin. As such levels of anti‐enterotoxin can be detected in normal serum samples, titration of anti‐enterotoxin may be of little use in diagnosing Cl. perfringens food poisoning. Enterotoxin was detected in all diarrhoeal faecal specimens, and the enterotoxin level varied from 0·2–16 μg/g. Detection of enterotoxin in diarrhoeal faeces may be the most reliable procedure in diagnosing outbreaks of Cl. perfringens food poisoning.
Sixty-nine strains of Clostridium perfringens of different toxigenic types were investigated for enterotoxin production. Enterotoxin was definitively detected only in strains of types A and C. This is the first report where enterotoxin production has been demonstrated in a toxigenic type other than type A. The enterotoxin-positive type C strains were isolated from cases of enteritis necroticans ("pig bel") in New Guinea. The major enterotoxin from type C showed a reaction of complete identity with enterotoxin from type A in immunodiffusion using anti-enterotoxin serum prepared against the latter; it induced erythema when injected intradermally into depilated guinea pigs and caused fluid accumulation in the rabbit ileal loop. The results indicate that the major enterotoxin from type C was serologically and biologically similar to enterotoxin from type A. In some type C strains, an enterotoxin was detected that showed a reaction of partial serological identity. Spore coat proteins were extracted from 14 strains by alkaline dithiothreitol, and the extracts were assayed for enterotoxin-like spore protein. Enterotoxin could be extracted from type A and type C spores, and all positive strains showed a reaction of complete identity in immunodiffusion with enterotoxin obtained from cell extracts of type A. Disc immunoelectrophoresis demonstrated that two distinct components that reacted serologically with anti-enterotoxin serum were present in both the cell extract and in extracted spore protein from one type C strain. These distinct components differed in molecular weight. Clostridium perfringens produces a multiplicity of toxins and, according to the kinds and amount of toxin formed, the species is subdivided into five toxigenic types, A to E. This classification is based on the ability to produce the four major toxins, alpha-, beta-, epsilon-, and iota-toxin as assayed by lethality test in mice or skin test in guinea pigs. Only certain strains of types A and C have been implicated in food-borne diseases in humans (14). C. perfringens type A is one of the most common causes of human food poisoning (3). The course of the intoxication is usually mild, and fatal cases are relatively rare.
Strains of Clostridium perfringens from a variety of sources were examined for their ability to produce enterotoxin in vitro. Fifty‐six of 65 (86%) strains isolated from separate outbreaks of food poisoning were found to be enterotoxigenic, only two of 174 strains from other sources produced enterotoxin. The ability to produce this toxin was not confined to particular serotypes: types frequently encountered as the cause of outbreaks were also isolated as enterotoxin‐negative strains from faeces, minced beef and meat carcasses. Loss of toxigenicity was also observed in different serotypes. Five strains of lecithinase‐negative Cl. perfringens produced high levels of enterotoxin. Four strains of Clostridium plagarum failed to produce enterotoxin although they were serologically typable with the Cl. perfringens antisera.
The effect of natural spices and oleoresins on the fermentation properties of three commercially available starter cultures of Lactobacillus plontanrm has been investigated. In liquid medium it was shown that natural spices stimulated growth of the three bacteria and thus enhanced the cultures to ferment glucose and form more lactic acid. When similar oleoresin spices were supplied to the growing bacteria no effect was seen. It was also found that natural spices accelerated the fermentation process in the production of salami dry sausage while the oleoresins showed no effect.
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