Transmission Electron Microscopy of Unfrozen and Frozen/Thawed Cells of Listeria monocytogenes Treated With Lipase and Lysozyme

Abstract

Unfrozen cells of Listeria monocytogenes typically contained no preplasma space exterior to the plasma membrane (PM) when viewed by transmission electron microscopy. Cells of L. monocytogenes strains Scott A, V7, and California (CA), after freezing and frozen storage, exhibited one or more of the following when viewed with transmission electron microscopy: (a) retraction of cytoplasm and infolding of the PM to form mesosomes, (b) extra-and intracellular rupture of the cell wall (CW), (c) formation of intracellular “bubbles,” and (d) damage to the CW and PM that could have resulted from autolysin activity. Type and degree of effect depended on frozen storage time and strain of L. monocytogenes. Lysozyme treatment of unfrozen or frozen/stored (19 d)/thawed cells of strains Scott A, V7, and CA resulted in protoplast formation and damage to the CW. Three stages of protoplast formation were observed when cells of strain CA were frozen, stored 2 weeks, thawed, and treated with lysozyme. More damage to the CW and PM occurred when frozen storage time was extended for up to 6 weeks before treatment with lysozyme. Lipase and lysozyme treatment of unfrozen or frozen/stored (19 d)/thawed cells of strain Scott A resulted in protoplast formation with some damage to the PM and irregularity in shape of cells. Damage to the PM increased with increasing frozen storage time for up to 6 weeks. Some cells of strain CA resisted freezing, frozen storage for 6 weeks, thawing, and treatment with lipase and lysozyme.