USE OF THE MTT TEST FOR THE DETECTION OF BACILLUS-CEREUS ENTEROTOXIN

Use of Caco-2 cell line and a modified, simultaneous test performance enabled the preparation of a sensitive and reproducible MTT cytotoxici ty assay for the detection of B. cereus diarrhoeal toxin. Characteriza tion of some properties of this enterotoxin, particularly heat labilit y, susceptibility to trysin and stability during storage at various te mperatures, showed that 10-20% (corresponding to a cytotoxicity titre of approx. 1:10 in the MTT-Test) of the cytotoxic activity of B. cereu s culture supernatants is not caused by the toxicity of the diarrhoeal toxin. In detail, studies on the in vitro stability of the toxin reve aled the following results: after heating of non-treated culture super natants up to 56 degrees C, the cytotoxic activity was reduced to appr ox. 14 % of the initial value; storage of culture supernatants at 21 a nd 4 degrees C caused a considerable loss of cytotoxicity after 8 and 28 days, respectively. The toxic activity, however, could be completel y conserved at storage temperatures of -20 and -80 degrees C for a per iod of 439 days. Treatment with trypsin resulted in a decrease of toxi c activity by 69 and 91 % after 10 and 120 min respectively. The analy sis of 152 B. cereus isolates from foods revealed that only 4 strains showed similary high cytotoxic titres as the enterotoxic reference str ain DSM 4384 (1 : 35 to 1 : 52). The majority of the isolates showed a titre in the range of 1 : 6 to 1 : 15 (66 %), 5.9 % were atoxic. Ther e was no correlation between the toxicity titres and the results obtai ned with a commercial available enzyme immunoassay for B. cereus enter otoxin (TECRA-BDE visual immunoassay).