Differential protein expression by Pseudomonas fragi submitted to various stresses

We have analyzed the impact of various stressing conditions on the physiological and molecular responses of the main psychrotrophic spoilage bacterium of refrigerated meat and meat products, Pseudomonas fragi. A survival study using conventional plating was first performed to select the stressing agents and parameters. Some of these mimicked cleaning-disinfection processes but with less drastic conditions in order to keep alive enough bacterial cells for the protein expression characterization. Cultures of P. fragi, at the beginning of the stationary phase of growth, were submitted to individual pH (5.4, 10.5), osmotic (8% Na2SO4, pH 7.0), biocide (fatty amine) shocks or combined treatments (8% Na2SO4, pH 10.5; 8% Na2SO4, pH 10.5 + biocide; pH 5.4 + pH 10.5 and pH 10.5 + pH 5.4) and the molecular responses were investigated by comparing autoradiograms of two-dimensional gel electrophoresis (2-DE) patterns of proteins radiolabeled with L-[35S]methionine. The observation of qualitative and relative quantitative variations in protein expression, determined with Melanie II image analysis software (Bio-Rad), revealed the overexpression of a total of 91 proteins for the eight challenges by comparison with the nonshocked controls. Some proteins appeared to be more or less general stress proteins whereas others were specific for one chemical treatment. The appraisal of the type of molecular response according to the type of treatment was analyzed statistically.