Effects of High Pressure on the Viability, Morphology, Lysis, and Cell Wall Hydrolase Activity of Lactococcus lactis subsp. cremoris

Abstract

Viability, morphology, lysis, and cell wall hydrolase activity of Lactococcus lactis subsp. cremoris MG1363 and SK11 were determined after exposure to pressure. Both strains were completely inactivated at pressures of 400 to 800 MPa but unaffected at 100 and 200 MPa. At 300 MPa, the MG1363 and SK11 populations decreased by 7.3 and 2.5 log cycles, respectively. Transmission electron microscopy indicated that pressure caused intracellular and cell envelope damage. Pressure-treated MG1363 cell suspensions lysed more rapidly over time than did non-pressuretreated controls. Twenty-four hours after pressure treatment, the percent lysis ranged from 13.0 (0.1 MPa) to 43.3 (300 MPa). Analysis of the MG1363 supernatants by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) confirmed pressure-induced lysis. Pressure did not induce lysis or membrane permeability of SK11. Renaturing SDS-PAGE (zymogram analysis) revealed two hydrolytic bands from MG1363 cell extracts treated at all pressures (0.1 to 800 MPa). Measuring the reducing sugars released during enzymatic cell wall breakdown provided a quantitative, nondenaturing assay of cell wall hydrolase activity. Cells treated at 100 MPa released significantly more reducing sugar than other samples, including the non-pressure-treated control, indicating that pressure can activate cell wall hydrolase activity or increase cell wall accessibility to the enzyme. The cell suspensions treated at 200 and 300 MPa did not differ significantly from the control, whereas cells treated at pressures greater than 400 MPa displayed reduced cell wall hydrolase activity. These data suggest that high pressure can cause inactivation, physical damage, and lysis in L. lactis. Pressure-induced lysis is strain dependent and not solely dependent upon cell wall hydrolase activity.

Reducing Biogenic-Amine-Producing Bacteria, Decarboxylase Activity, and Biogenic Amines in Raw Milk Cheese by High-Pressure Treatments

“…The increase of TDC activity in control cheese can be explained by the production of the enzyme by intact bacterial cells, followed by its release into the surrounding medium, favored by spontaneous cell lysis. In cheeses treated at 400 MPa, counts of bacterial groups after pressurization were lower than in control cheese, but the mild-pressure treatment would most probably enhance the lysis of LAB cells, as shown previously for lactococci (35), without inactivating the enzyme. The minor increase in TDC activity recorded for cheeses treated at 600 MPa might be due to a double effect of high pressure at this level, on the one hand promoting cell lysis, as in the case of cheeses treated at 400 MPa, and on the other hand inactivating the enzyme released into the medium.…”

Correlative Electron and Fluorescence Microscopy of Magnetotactic Bacteria in Liquid: Toward In Vivo Imaging

“…Compressive stresses have been shown to increase membrane permeability and damage efflux systems in E. coli, leading to increased absorption of propidium iodide 55 . High pressures have also been shown to lead to increased rates of cell lysis in Lactobacillus strains 56 . Our systematic fluorescence imaging showed increased propidium iodide absorption into the cells upon sample preparation and over the two hours in the fluid cell; however, cell lysis was not observed.…”

Cloning of the organophosphorus pesticide hydrolase gene clusters of seven degradative bacteria isolated from a methyl parathion contaminated site and evidence of their horizontal gene transfer

“…Zymogram analysis was performed according to the procedures described (Granelli-Piperno & Reich 1978;Vassalli et al 1984;Li et al 2001;Malone et al 2002). The cell pellet from 1 ml cultures was resuspended in 100 ll 1Â SDS-loading buffer without b-mercaptoethanol, heated for 5 min at 100°C, and then subjected to 12% SDS-PAGE.…”