Differential kinetic patterns for Shiga toxin production by Escherichia coli

We studied the differential kinetic patterns for Shiga toxin (Stx) production (i.e. Stx1, Stx2 and Stx2c) in different reference Escherichia coli strains and in those isolated from hemolytic uremic syndrome (HUS) patients. These results were correlated with those obtained by specific cytotoxic activity assays on Vero cells and hybridization tests with DNA probes for Stx1 and Stx2. Strains cultured in Penassay broth were sampled at 1.5; 3; 5; 9 and 24 hours to determine bacterial growth and its association with cell-bound and free cytotoxicity. Stx1 showed an intracellular/extracellular concentration ratio (ic/ec) between 32 and 200 times after 3 h-growth. At 24 h both Stx1 concentrations were equal or, in some strains, the ec resulted 2-fold higher that the ic. The ic-Stx1 was equal or just 2-fold higher that ec after 3 h-growth. However, at 24 h the released toxin level was 16 to 32 times higher that cell-bound toxin. The ec-Stx2c increased logarithmically, with maximal yields at 5 h, remaining constant up to 24 h. At that time ic-toxin was 2-fold higher than the released one. When the same experiments were performed on strains isolated from HUS patients they showed that the kinetic patterns obtained corresponded to Stx2. These results were confirmed by hybridization assays. In this study we have shown that Stx1 production decreases dramatically during stationary phase while Stx2 is detected at high level at that time. This could explain the higher frequency of association of Stx2-producing E. coli strains and HUS in some countries, including Argentina.