| Co-Investigator(Kenkyū-buntansha) |
KOZAKI Shunji University of Osaka Prefecture, College of Agriculture, 農学部, 助手 (10109895)
ASAO Tsutomu Osaka Prefectural Institute of Public Health, 主任研究員 (00250318)
SHINAGAWA Kunihiro Iwate University, Faculty of Agriculture, 農学部, 助教授 (60133906)
UEMURA Takashi University of Osaka Prefecture, College of Agriculture, 農学部, 助教授 (40081591)
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| Research Abstract |
We studied on pathogenic roles of bacterial toxins in food poisoning and improved immunological methods for its diagnosis and prevention. Clostridium botulinum neurotoxin, Clostridium perfringens enterotoxin, Aeromonas hydrophila hemolysin, Bacillus cereus enterotoxin, and Staphylococcus aureus enterotoxin were examined for their physicochemical, immunological, and molecular structures and also the toxin-cell interaction on molecular level. Type G botulinum toxin was purified. C. botulinum types A through G produce a complex toxin, consisting of a neurotoxic and nontoxic components. The neurotoxin consists of at least three fragments named L, H-1, and H-2. They possess distinct but cooperative functions; H-1 fragment mediates the internalization of the toxin molecule into the hydrophobic area of the membranes and H-2 binding to a certain receptor(s) of neural cell membranes. The antigenicity of botulinum toxin was more stable in animal bodies than its biological activity. Immunological
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detection of toxins in patients' specimens, foodstuffs and other materials should help increase the rate of diagnosis of food-borne intoxications. A neutralizing monoclonal antibody did not inhibit the binding of C. perfringens enterotoxin to the receptor of the target cell. A nontoxic fragment binding to the receptor of the target cell was purified by chemical cleavage of the toxin molecule. C. perfringens enterotoxin may consist of at least two functional sites, one binding to the receptor and the other causing diarrhea. Two immunologically distinct A. hydrophila hemolysins were purified and both demonstrated to possess enterotoxic activity. A monoclonal antibody neutralized both the activities, indicating that the two activities are elicited by the same site on the toxin molecule. A method for purifying Bacillus cereus enterotoxin was established.Monoclonal antibodies against the toxin were produced and used for analysis of the antigenic structure. Monoclonal antibodies against Staphylococcus aureus enterotoxin were prepared. The enterotoxin was purified by only one step of immunoaffinity chromatography of culture supernatant on a column coupled with the monoclonal antibody. Less
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