In this study, we investigated the mechanisms of spore inactivation by high pressure at moderate temperatures to optimize the sterilization efficiency of high pressure treatments Bacillus subtilis spores were first subjected to different pressure treatments ranging from 90 to 550 MPa at 40 C, with holding times from 10 mm to 4 h These treatments alone caused slight inactivation, which was related to the pressure-induced germination of the spores After these pressures treatments, the sensitivity of these processed spores to heat (80 degrees C/10 mm) or to high pressure (350 MPa/40 degrees C/10 min) was tested to determine the pressure-induced germination rate and the advancement of the spores in the germination process The subsequent heat or pressure treatments were applied immediately after decompression from the first pressure treatment or after a holding time at atmospheric pressure As already known, the spore germination is more efficient at low pressure level than at high pressure level Our results show that this low germination efficiency at high pressure seemed not to be related either to a lower induction or a difference in the induction mechanisms but rather to an inhibition of enzyme activities which are involved in germination process In fact, high pressure was necessary and very efficient in inducing spore germination However, it seemed to slow the enzymatic digestion of the cortex, which is required for germinated spores to be inactivated by pressure Although these results indicate that high-pressure treatments are more efficient when the two treatments are combined, a small spore population still remained dormant and was not inactivated with any holding time or pressure level Biotechnol Bioeng 2010 107 876-883 (C) 2010 Wiley Periodicals, Inc