Sensitivity of various salt forms of Bacillus megaterium spores to the germinating action of hydrostatic pressure

Abstract

Spores of Bacillus megaterium ATCC 19213 were subjected to a complete ion-exchange regimen, which included titration to a pH value of 2, heating at 60 °C for up to 18 h, back titration with various base solutions to a pH value of 8, and heating again at 60 °C. Spore populations maintained high levels of viability throughout this rigorous procedure, and the various salt forms prepared showed a wide range of sensitivities to the germinating effect of hydrostatic pressure. Native spores showed the expected germination response when subjected to pressures of 350–750 atm (1 atm = 101.325 kPa) at 24 °C. There was a threshold pressure for germination of some 300 atm, and the apparent activation volume for the process was calculated to be 188 mL/mol, indicating that these spores had approximately the same pressure sensitivity as those of Bacillus subtilis or Bacillus pumilus. The optimum pH for germination was approximately 8, and the optimum temperature was approximately 45 °C. The hierarchy of resistance of the various salt forms tested to the germinating action of 493 atm pressure was H > K > Ca, Mg, Na > native. The H form was particularly insensitive, even to pressures as high as 1020 atm, but germinated in response to chemical germinants. We concluded that the specific mineralization of bacterial spores has major influence on pressure-induced germination, which can occur even in the absence of added salts. Sensitivities to pressure could not be correlated with previously reported heat sensitivities, electrostasis, or states of dehydration.