Protective effect of calcium on inactivation of Escherichia coli by high hydrostatic pressure

The effect of divalent cations on the inactivation of Escherichia coli by high hydrostatic pressure was investigated. The presence of 0.5 mmol l-1 of CaCl2, MgCl2, MnCl2 and FeCl2 reduced pressure inactivation of E. coli MG1655, while 0.5 mmol l-1 of ZnCl2, NiCl2, CuCl2 and CoCl2 increased inactivation. Baroprotection by Ca2+ was found to be dose-dependent up to at least 80 mmol l-1 and was studied in more detail in terms of inactivation kinetics. Logarithmic survivor plots against time deviated from first order kinetics, suggesting that MG1655 cultures were heterogeneous with regard to pressure resistance. All cultures were shown to contain a small proportion of cells that were only slowly inactivated. Addition of Ca2+ increased the proportion of these tolerant cells in the cultures up to 1000-fold at 80 mmol l-1, but did not affect their inactivation rate. The addition of EDTA resulted in the opposite effect, lowering the proportion of pressure-tolerant cells in the cultures. Three pressure-resistant mutants of E. coli MG1655 were found to be more resistant to EDTA under pressure compared with MG1655, and were unaffected by Ca2+ under pressure. In addition, these mutants had a 30-40% lower Ca2+ content than MG1655. Based on these results, it is postulated that pressure killing of E. coli MG1655 is mediated primarily by the destabilization of Ca(2+)-binding components, and that the mutations underlying pressure resistance have resulted in pressure-stable targets with reduced Ca(2+)-binding affinity.