A study of the influence of proteolytic enzymes on various types of Clostridium botulinum toxin

Abstract

Botulism is a food poisoning which is caused by taking in nerve poison produced by Cl. botulinum which multiplies in foodstuffs.In Japan, since 1951 occurrence has been mainly in Hokkaido and Tohoku (northern part of Honshu) districts. Up until the present time 63 outbreaks have been reported and 93 people out of 359 patients died.Toxin produced by Cl. botulinum is classified into 6 different types: A, B, C, D, E and F according to their immunological nature. Of these 6 types of toxin, types A, B, E and F are the cause of poisoning in human beings whereas types C and D affect cattle and fowl.Botulism in Japan is mainly attributable to fermented preserved foods such as “IZUSHI” and “KIRIKOMI” which utilize raw fish as their component. But it should be mentioned that type B poisoning, which occurred in Miyazaki Prefecture in 1969, is thought to have been caused by imported caviare, 21 people were affected, 3 died.Therefore, while only type B episodes have been previously reported in Japan, the discovery of types A and B strains in the soil of Tohoku district should make it clear that outbreaks due to other types are now possible. The possibility seems to be higher since a poisoning from imported food has occurred.The study of botulism in Japan has been conducted mainly on type E which has frequently broken out. The geographical distribution of type E organisms in the soil, the multiplication in “IZUSHI”, the conditions of toxin production, the chemical nature of the toxin, and its remedy using antitoxic serum have all been studied. But the author has taken up the following problems broadly not only on type E toxin but also other types of toxin due to the above mentioned reasons.Why is botulinus toxin not affected by the proteolytic enzymes in the digestive tract? Though the effect of proteolytic enzymes on the toxin in the digestive tract is a very urgent problem, no unanimous conclusion has been reached.In other words, no report on the study of the effect of proteolytic enzymes of all the six toxins, already known to us, under the same conditions has yet been published. Bearing this in mind, the author made a study on the effect of various proteolytic enzymes using crude botulinus toxins (cultural super-mates) under simulated natural conditions.The results obtained are as follows:(1) With type A, strain 190, type B, strain Lamanna, and type F, strain Denmark which belong to the Proteolytic Group, a remarkable attenuation of their toxicity was observed on both acid and alkaline sides. But inactivation of the toxins due to enzymes was not particularly notable.(2) With toxin type C, strain Stockholm and type D, strain 1873 which belong to the Non-Proteolytic Group, the effects of fluctuation of PH and the action of various types of enzymes were comparatively slight.(3) With type B, strain QC, type E, strain Iwanai and type F, strain 70 F which belong to the Non-Proteolytic Group, activation of the toxins was observed by treating them with trypsin at PH 6.0. Generally speaking, except for type F, effects of fluctuation of PH and of various enzymes were slight.(4) When botulinus toxin is taken orally the toxin changes its toxicity in varying degrees by fluctuation of PH in the stomach and the intestinal tract or by action of digestive enzymes, and it is presumed that a considerable amount of active toxin remains in the digestive tract.