PURIFICATION AND CHARACTERIZATION OF ENTEROTOXIN PRODUCED BY AEROMONAS-SOBRIA

We purified the toxin of Aeromonas sobria capable of inducing a positi ve response in the mouse intestinal loop assay. The purified toxin sho wed a positive response not only in the loop assay but also in a hemol ytic assay. Subsequently, we cloned the toxin gene and demonstrated th at the product of this gene possessed both hemolytic and enterotoxic a ctivities. These results showed that the enterotoxin of A. sobria poss esses hemolytic activity. Nucleotide sequence determination of the tox in gene and amino acid sequence analysis of the purified toxin reveale d that it is synthesized as a precursor composed of 488 amino acid res idues, and that the 24 amino-terminal amino acid residues of the precu rsor is removed in the mature toxin, As antiserum against the purified toxin neutralized the fluid accumulation induced by living cells not only of A. sobria but also of A. hydrophila, this and antigenically re lated toxin(s) are thought to play an essential role in the induction of diarrhea by these organisms. The toxin-injured Chinese hamster ovar y (CHO) cells induced the release of intracellular lactose dehydrogena se (LDH), The release of LDH from CHO cells and the lysis of erythrocy tes by the toxin were repressed by the addition of dextran to the reac tion solution, indicating that the toxin forms pores in the membranes and that the cells were injured by the osmotic gradient developed due to pore formation. However, the histopathological examination of intes tinal cells exposed to the toxin showed that it caused fluid accumulat ion in the mouse intestinal loop without causing cellular damage.