The emetic toxin of Bacillus cereus, found to cause immobilization of spermatozoa and swelling of their mitochondria, was purified and its structure found to be identical to the earlier known toxin cereulide. It increased the conductance in black-lipid membranes in KCl solutions in an ionophore-like manner. It formed adducts with K + , Na + , and NH 4 + but the conductance was highly selective for K + in relation to Na + and H + (three orders of magnitude). The increase in the kinetics of conductance indicated a stoichiometric ratio between the cereulide and K + . Its ionophoretic properties are thus similar to those of valinomycin. In addition, its effects on rat liver mitochondria were similar: it stimulated swelling and respiration in respiring mitochondria in the presence but not in the absence of K + , it reduced the transmembrane potential under these conditions. In nonrespiring mitochondria, swelling was seen in KNO 3 -but not in NaNO 3 -containing media, less in acetate. In NaNO 3 media addition of the cereulide caused a transient diffusion potential which was reduced by adding K + . It is concluded that the toxic effects of cereulide are due to it being a K + ionophore.Keywords: cereulide; ionophore; mitochondria; spermatozoa; toxin.The emetic toxin of strains of Bacillus cereus, that may cause serious food poisoning, has been isolated and found to be structurally related to valinomycin [1]. It was reported to cause vacuole formation and the vacuoles were then found to be swollen mitochondria in mouse liver and HeLa cells [2]. The emetic toxin, also called cereulide [1], stimulated respiration like the uncoupling agent 2,4-dinitrophenol. Similar observations were made in liver failure following a fatal food poisoning due to the toxin which caused lowering of the respiratory control ratio in isolated rat liver mitochondria [3]. These findings are analogous to the early findings on the mechanism of action of valinomycin, found to cause a mitochondrial energy-dependent uptake of K + , ejection of H + , stimulation of respiration in the presence of K + and swelling [4,5]. Several other bacterial toxins have ionophoretic or channel-forming properties: examples are diphtheria toxin, the enterohemorrhagic haemolysin of Escherichia coli [6] and an insecticidal toxin from B. thuringiensis [7].We have earlier reported swelling of mitochondria in boar spermatozoa induced by extracts of water-damaged indoor building materials heavily contaminated by microbes [8]. The cessation of spermatozoic motility was used as a sensitive test for cereulide and related toxins [9]. In this study we report the purification from B. cereus of the active agent, its structural features and ionophoretic properties in black-lipid membranes (BLM) and its effect on mitochondrial swelling and function. It is shown that the agent is structurally identical to cereulide and that it acts like valinomycin, i.e. it is a specific K + ionophore.
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Electron microscopyElectron microscopy was performed as described...
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