Characterization of Mild Thermal Stress in Pseudomonas fluorescens and Its Repair

Abstract

The exposure of exponentially growing Pseudomonas fluorescens P7 cells to heating at 36 C for 2 h in a defined medium, followed by cooling to 25 C and further incubation at this, the optimal growth temperature, resulted in the apparent death of approximately 99% of the cells, as determined by their inability to form colonies on Trypticase soy agar. Continued incubation at 25 C resulted in an extremely rapid increase in the Trypticase soy agar count, demonstrating that the phenomenon observed was not death but rather injury. Presumptive evidence of heat-stimulated ribonucleic acid (RNA) degradation and membrane damage was provided by the observed loss of 260-nm absorbing materials. Confirmation of RNA degradation was obtained by colorimetric analysis. Ribosomal RNA from normal and injured cells, which was electrophoretically separated on polyacrylamide gels, revealed that the 23 S and 16 S species were only partially destroyed. Inhibitor studies demonstrated, however, that RNA synthesis was necessary for recovery. The unusual accumulation of 17 S RNA during recovery pointed to the presence of a heat-induced lesion in the RNA maturation process. A thermally induced membrane lesion is also discussed.