106 ◾ Fundamental Food Microbiology
that some cell components are damaged by almost all types of stresses studied. In addition, specific components can be damaged by specific stresses. The structural and functional components
known to be damaged by sublethal stresses are the cell wall (or outer membrane, OM), cytoplasmic membrane (or inner membrane, IM), ribosomal RNA (rRNA) and DNA, and some enzymes
(see Figure 2.2). Damages in the cell wall (or OM) and cytoplasmic membrane (or IM) are more
evident in injury caused by freezing and drying, whereas damage to rRNA is more extensive in
sublethal heating and DNA damage following radiation of cells.
In Gram-positive and -negative bacteria, freezing and drying cause changes in cell surface
hydrophobicity and the inability to form compact pellets and to adsorb some phages. In Grampositive bacteria, surface layer proteins are also lost. In sublethally stressed Gram-negative bacteria, the lipopolysaccharide (LPS) layer undergoes conformational alteration and loses its barrier
property to many chemicals (such as SDS, bile salts, antibiotics, lysozyme, and RNase), which can
easily enter the injured cells and kill them. Very little LPS is lost from the cells into the environment. The alteration in conformation of LPS is a result of loss of divalent cations, which are necessary for the normal stability of LPS. In both Gram-positive and -negative cells, the cytoplasmic
membrane (or IM) remains intact in injured cells, but it loses its permeability barrier function. The
cells become sensitive to NaCl and also lose different cellular materials. It is suggested that protein
molecules in this structure probably undergo conformational changes in the injured cells. rRNA is
extensively degraded by the activated RNase. DNA can undergo single- and double-strand breaks.
In some strains, autolytic enzymes can be activated by stress, causing lysis of the cells.
In bacterial spores, depending on the type of sublethal stress, different structural and functional components can be injured. High heat causes damage to the lytic enzymes, necessary for
the lysis of the cortex before spore germination and to the spore membrane structures, causing
loss of permeability barrier functions. Damages by irradiation (UV and γ) are mainly confined to
DNA in the form of single-strand breaks and, by some chemicals (H 2 O 2 , antibiotics, or chlorine),
to the lytic enzymes of the germination system. Hydrostatic pressure, in combination with heat,
damages the cortex, whereas γ irradiation damages both the cortex and DNA. Mild to strong acid
treatments can cause the spores to become dormant by removing Ca 2
+
from the spores and making
them sensitive to heat.
Repair of Reversible Injury
One of the most important characteristics of injured bacterial cells is the ability to repair the injury
in a suitable environment and become similar to normal cells. The repair process and the rate of
repair can be measured by specific methods. One of them is to measure regain in resistance of
injured cells to the surface-active agents by repair in the cell wall or the OM. Suspending a sublethally stressed population in a repair medium and simultaneously enumerating the colony-forming
units (CFUs) during incubation in nonselective and selective plating media help determine the
rate of repair (Figure 10.4). Initially, the injured survivors fail to form colonies in the selective, but not
in the nonselective, media. However, as they repair and regain resistance to the selective agent,
they form colonies on both media, as indicated by the increase in counts in the selective media
only. Injured cells differ in the levels of injury (from low to high) as manifested from the differences in recovery time in a suitable medium.
The injured cells can repair in a medium devoid of selective compounds but containing the
necessary nutrients during incubation at optimum pH and temperature. In general, the cells
repair well in a medium rich in metabolizable carbon and nitrogen sources and several vitamins.
that some cell components are damaged by almost all types of stresses studied. In addition, specific components can be damaged by specific stresses. The structural and functional components
known to be damaged by sublethal stresses are the cell wall (or outer membrane, OM), cytoplasmic membrane (or inner membrane, IM), ribosomal RNA (rRNA) and DNA, and some enzymes
(see Figure 2.2). Damages in the cell wall (or OM) and cytoplasmic membrane (or IM) are more
evident in injury caused by freezing and drying, whereas damage to rRNA is more extensive in
sublethal heating and DNA damage following radiation of cells.
In Gram-positive and -negative bacteria, freezing and drying cause changes in cell surface
hydrophobicity and the inability to form compact pellets and to adsorb some phages. In Grampositive bacteria, surface layer proteins are also lost. In sublethally stressed Gram-negative bacteria, the lipopolysaccharide (LPS) layer undergoes conformational alteration and loses its barrier
property to many chemicals (such as SDS, bile salts, antibiotics, lysozyme, and RNase), which can
easily enter the injured cells and kill them. Very little LPS is lost from the cells into the environment. The alteration in conformation of LPS is a result of loss of divalent cations, which are necessary for the normal stability of LPS. In both Gram-positive and -negative cells, the cytoplasmic
membrane (or IM) remains intact in injured cells, but it loses its permeability barrier function. The
cells become sensitive to NaCl and also lose different cellular materials. It is suggested that protein
molecules in this structure probably undergo conformational changes in the injured cells. rRNA is
extensively degraded by the activated RNase. DNA can undergo single- and double-strand breaks.
In some strains, autolytic enzymes can be activated by stress, causing lysis of the cells.
In bacterial spores, depending on the type of sublethal stress, different structural and functional components can be injured. High heat causes damage to the lytic enzymes, necessary for
the lysis of the cortex before spore germination and to the spore membrane structures, causing
loss of permeability barrier functions. Damages by irradiation (UV and γ) are mainly confined to
DNA in the form of single-strand breaks and, by some chemicals (H 2 O 2 , antibiotics, or chlorine),
to the lytic enzymes of the germination system. Hydrostatic pressure, in combination with heat,
damages the cortex, whereas γ irradiation damages both the cortex and DNA. Mild to strong acid
treatments can cause the spores to become dormant by removing Ca 2
+
from the spores and making
them sensitive to heat.
Repair of Reversible Injury
One of the most important characteristics of injured bacterial cells is the ability to repair the injury
in a suitable environment and become similar to normal cells. The repair process and the rate of
repair can be measured by specific methods. One of them is to measure regain in resistance of
injured cells to the surface-active agents by repair in the cell wall or the OM. Suspending a sublethally stressed population in a repair medium and simultaneously enumerating the colony-forming
units (CFUs) during incubation in nonselective and selective plating media help determine the
rate of repair (Figure 10.4). Initially, the injured survivors fail to form colonies in the selective, but not
in the nonselective, media. However, as they repair and regain resistance to the selective agent,
they form colonies on both media, as indicated by the increase in counts in the selective media
only. Injured cells differ in the levels of injury (from low to high) as manifested from the differences in recovery time in a suitable medium.
The injured cells can repair in a medium devoid of selective compounds but containing the
necessary nutrients during incubation at optimum pH and temperature. In general, the cells
repair well in a medium rich in metabolizable carbon and nitrogen sources and several vitamins.