Changes in Listeria monocytogenes membrane fluidity in response to temperature stress

Abstract

Listeria monocytogenes is a food-borne pathogen that has been implicated in many outbreaks associated with ready-to-eat products. Listeria adjusts to various stresses by adjusting its membrane fluidity, increasing the uptake of osmoprotectants and cryoprotectants, and activating the sigma(B) stress factor. The present work examines the regulation of membrane fluidity through direct measurement based on fluorescent anisotropy. The membrane fluidities of L. monocytogenes Scott A, NR30, wt10403S, and cld1 cells cultured at 15 and 30 degrees C were measured at 15 and 30 degrees C. The membrane of the cold-sensitive mutant (cld1) was more rigid than the membranes of the other strains when grown at 30 degrees C, but when grown at 15 degrees C, it was able to adjust its membrane to approach the rigidity of the other strains. The difference in rigidities, as determined at 15 and 30 degrees C, was greater in liposomes than in whole cells. The rates of fluidity adjustment and times required for whole cells to adjust to a different temperature were similar among strains but different from those of liposomes. This suggests that the cells had a mechanism for homeoviscous adaptation that was absent in liposomes.

FIG. 1.

Growth and measurement temperatures (°C) impact membrane rigidity (anisotropy) of Listeria monocytogenes Scott A (hatched bars), NR30 (checkered bars), wt10403S (horizontally striped bars), cld1 (vertically striped bars), and 2-methylbutyrate-treated cld1 (stippled bars) liposomes (a) and whole cells (b). The x axis displays the growth temperature followed by the measurement temperature. Significant differences (P < 0.05) are indicated by solid stars.

FIG. 2.

Real-time whole-cell anisotropy measurement examining the effect of temperature decrease (5°C every 5 min) for Listeria monocytogenes Scott A (a), NR30 (b), wt10403S (c), and cld1 (d) grown at 30°C. Dashed black lines show the linear regression of the data.

FIG. 3.

Real-time whole-cell anisotropy measurements during temperature increases (5°C every 5 min) for Listeria monocytogenes Scott A (a), NR30 (b), wt10403S (c), and cld1 (d) grown at 15°C. Dashed black lines show the linear regression fit performed on the data.

FIG. 4.

Time required for homeoviscous adaptation of Listeria monocytogenes. The graph presents the anisotropy (continuous line) of Scott A (a), NR30 (b), wt10403S (c), and cld1 (d) grown at 15°C and measured at 30°C. The black dotted line represents the average equilibrium value from the static measurement. Replicate experiments yielded consistent results.

FIG. 5.

Real-time dynamic anisotropy of liposomes (squares) and whole cells (triangles) of L. monocytogenes strains wt10403S (a and b) and cld1 grown at 30 and 15°C (c and d) showing the physiological adjustment of whole cells to temperature compared to the solely physical adjustment of liposomes. An average of multiple data is shown.

FIG. 6.

Change of anisotropy of whole cells (triangles) measured at 30°C as a result of equilibration at 15°C. The liposomes (squares) have similar values at 30°C pre- and postequilibration at 15°C. The average of multiple data is shown.