Production of listeriolysin O by Listeria monocytogenes (Scott A) under heat-shock conditions.
Early stationary phase cells of Listeria monocytogenes (Scott A) were examined to determine the effect of heat-shock on the production of listeriolysin O (LLO) during and after resuscitation at 37 degrees C. Cells were subjected to a heat-shock at 48 degrees C for 1 h. Intracellular and extracellular proteins of the heat-shocked cells were assayed for LLO using a microtiter plate hemolysis assay and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Our results showed that significant amounts of LLO are synthesized under heat-shock conditions that are not detected in the extracellular medium by a functional assay. This situation is evident by the absence of hemolytic activity immediately after heat-shock, and may be due to either a lack of excretion or inactivation of the LLO at 48 degrees C once outside the cell. By studying the intracellular and extracellular proteins using SDS-PAGE and immunoblots of the heat-shocked cells, we substantiated an absence of excretion as an operating mechanism. Heat-shocked cells resumed LLO production within 2-4 h of resuscitation at 37 degrees C, achieving an activity level 2-fold higher compared to the controls and 4-fold higher compared to cells immediately after heat-shock. Most likely, the LLO excreted must have been from LLO accumulated in the cells during heat-shock.
Sampathkumar
B
B
Department of Applied Microbiology and Food Science, University of Saskatchewan, Saskatoon, Canada.
Xavier
I J
IJ
Yu
L S
LS
Khachatourians
G G
GG
D001427:Bacterial Toxins; D004591:Electrophoresis, Polyacrylamide Gel; D006360:Heat-Shock Proteins / Q000096:biosynthesis*; D006460:Hemolysin Proteins; D006358:Hot Temperature; D008089:Listeria monocytogenes / Q000378:metabolism*
D001427:Bacterial Toxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; C038705:hlyA protein, Listeria monocytogenes