of host cells
Dr. Gene Mayer
Medical Microbiology, MBIM 650/720 Lecture 31
- James A. Sullivan
describe the general composition and structure of
discuss the infectious process and the lytic multiplication
explain the lysogenic cycle and its regulation.
- Bacteriophage (phage) are obligate intracellular parasites that
multiply inside bacteria by making use of some or all of the host
biosynthetic machinery (i.e., viruses that infect bacteria.).
There are many
similarities between bacteriophages and animal cell viruses. Thus,
bacteriophage can be viewed as model systems for animal cell
viruses. In addition a knowledge of the life cycle of bacteriophage
is necessary to understand one of the mechanisms by which bacterial
genes can be transferred from one bacterium to another.
At one time it
was thought that the use of bacteriophage might be an effective way
to treat bacterial infections, but it soon became apparent that
phage are quickly removed from the body and thus, were of little
clinical value. However, bacteriophage are used in the diagnostic
laboratory for the identification of pathogenic bacteria (phage
typing). Although phage typing is not used in the routine clinical
laboratory, it is used in reference laboratories for epidemiological
purposes. Recently, new interest has developed in the possible use
of bacteriophage for treatment of bacterial infections and in
prophylaxis. Whether bacteriophage will be used in clinical medicine
remains to be determined.
AND STRUCTURE OF BACTERIOPHAGE
T4 Bacteriophage (TEM x390,000) ©
Dennis Kunkel, University of Hawaii. Used with permission
T4 bacteriophage Negative stain electron micrograph
- Although different bacteriophages may contain different materials
they all contain nucleic acid and protein.
the phage, the nucleic acid can be either DNA or RNA but not both
and it can exist in various forms. The nucleic acids of phages often
contain unusual or modified bases. These modified bases protect
phage nucleic acid from nucleases that break down host nucleic acids
during phage infection. The size of the nucleic acid varies
depending upon the phage. The simplest phages only have enough
nucleic acid to code for 3-5 average size gene products while the
more complex phages may code for over 100 gene products.
The number of
different kinds of protein and the amount of each kind of protein in
the phage particle will vary depending upon the phage. The simplest
phage have many copies of only one or two different proteins while
more complex phages may have many different kinds. The proteins
function in infection and to protect the nucleic acid from nucleases
in the environment .
Fig 1. Structure of T4 bacteriophage
- Bacteriophage come in many different sizes and shapes. The basic
structural features of bacteriophages are illustrated in Figure 1,
which depicts the phage called T4.
- T4 is among the largest phages; it is approximately 200 nm
long and 80-100 nm wide. Other phages are smaller. Most phages
range in size from 24-200 nm in length.
or Capsid - All phages contain a head structure which can
vary in size and shape. Some are icosahedral (20 sides) others
are filamentous. The head or capsid is composed of many copies
of one or more different proteins. Inside the head is found the
nucleic acid. The head acts as the protective covering for the
- Many but not all phages have tails attached to the phage head.
The tail is a hollow tube through which the nucleic acid passes
during infection. The size of the tail can vary and some phages
do not even have a tail structure. In the more complex phages
like T4 the tail is surrounded by a contractile sheath which
contracts during infection of the bacterium. At the end of the
tail the more complex phages like T4 have a base plate and one
or more tail fibers attached to it. The base plate and tail
fibers are involved in the binding of the phage to the bacterial
cell. Not all phages have base plates and tail fibers. In these
instances other structures are involved in binding of the phage
particle to the bacterium.
III. INFECTION OF
- The first step in the infection process is the adsorption of the
phage to the bacterial cell. This step is mediated by the tail
fibers or by some analogous structure on those phages that lack tail
fibers and it is reversible. The tail fibers attach to specific
receptors on the bacterial cell and the host specificity of the
phage (i.e. the bacteria that it is able to infect) is
usually determined by the type of tail fibers that a phage has. The
nature of the bacterial receptor varies for different bacteria.
Examples include proteins on the outer surface of the bacterium,
LPS, pili, and lipoprotein. These receptors are on the bacteria for
other purposes and phage have evolved to use these receptors for
attachment - The attachment of the phage to the bacterium via
the tail fibers is a weak one and is reversible. Irreversible
binding of phage to a bacterium is mediated by one or more of the
components of the base plate. Phages lacking base plates have other
ways of becoming tightly bound to the bacterial cell.
Contraction - The irreversible binding of the phage to the
bacterium results in the contraction of the sheath (for those phages
which have a sheath) and the hollow tail fiber is pushed through the
bacterial envelope (Figure 2). Phages that don't have contractile
sheaths use other mechanisms to get the phage particle through the
bacterial envelope. Some phages have enzymes that digest various
components of the bacterial envelope.
Fig. 2 Contraction of the tail sheath of T4
Acid Injection - When the phage has gotten through the bacterial
envelope the nucleic acid from the head passes through the hollow
tail and enters the bacterial cell. Usually, the only phage
component that actually enters the cell is the nucleic acid. The
remainder of the phage remains on the outside of the bacterium.
There are some exceptions to this rule. This is different from
animal cell viruses in which most of the virus particle usually gets
into the cell. This difference is probably due to the inability of
bacteria to engulf materials.
or Virulent Phages
- Lytic or virulent phages are phages which can only multiply on
bacteria and kill the cell by lysis at the end of the life cycle.
cycle - The life cycle of a lytic phage is illustrated in
Figure 3 .
Fig. 3. Life cycle of a lytic phage
period - During the eclipse phase, no infectious phage
particles can be found either inside or outside the bacterial
cell. The phage nucleic acid takes over the host biosynthetic
machinery and phage specified m-RNA's and proteins are made.
There is an orderly expression of phage directed macromolecular
synthesis, just as one sees in animal virus infections. Early
m-RNA's code for early proteins which are needed for phage DNA
synthesis and for shutting off host DNA, RNA and protein
biosynthesis. In some cases the early proteins actually degrade
the host chromosome. After phage DNA is made late m-RNA's and
late proteins are made. The late proteins are the structural
proteins that comprise the phage as well as the proteins needed
for lysis of the bacterial cell.
Accumulation Phase - In this phase the nucleic acid and
structural proteins that have been made are assembled and
infectious phage particles accumulate within the cell.
and Release Phase - After a while the bacteria begin to
lyse due to the accumulation of the phage lysis protein and
intracellular phage are released into the medium. The number of
particles released per infected bacteria may be as high as 1000.
for Lytic Phage
assay - Lytic phage are enumerated by a plaque assay. A
plaque is a clear area which results from the lysis of bacteria
(Figure 4). Each plaque arises from a single infectious
phage. The infectious particle that gives rise to a plaque is
called a pfu (plaque forming unit).
Figure 4. Assay for lytic phage
or Temperate Phage
- Lysogenic or temperate phages are those that can either multiply
via the lytic cycle or enter a quiescent state in the cell. In
this quiescent state most of the phage genes are not transcribed;
the phage genome exists in a repressed state. The phage DNA in
this repressed state is called a prophage because it
is not a phage but it has the potential to produce phage. In most
cases the phage DNA actually integrates into the host chromosome
and is replicated along with the host chromosome and passed on to
the daughter cells. The cell harboring a prophage is not adversely
affected by the presence of the prophage and the lysogenic state
may persist indefinitely. The cell harboring a prophage is termed
Leading to Lysogeny - The Prototype Phage: Lambda
Fig. 5. Circularization of phage chromosome: cohesive ends
of the phage chromosome - Lambda DNA is a double
stranded linear molecule with small single stranded regions at
the 5' ends. These single stranded ends are complementary (cohesive
ends) so that they can base pair and produce a circular
molecule. In the cell the free ends of the circle can be ligated
to form a covalently closed circle as illustrated in Figure 5.
recombination - A recombination event, catalyzed by a
phage coded enzyme, occurs between a particular site on the
circularized phage DNA and a particular site on the host
chromosome. The result is the integration of the phage DNA into
the host chromosome as illustrated in Figure 6.
Fig. 6. Site-specific recombination
of the phage genome - A phage coded protein, called a repressor,
is made which binds to a particular site on the phage DNA,
called the operator, and shuts off transcription
of most phage genes EXCEPT the repressor gene. The result is a
stable repressed phage genome which is integrated into the host
chromosome. Each temperate phage will only repress its own DNA
and not that from other phage, so that repression is very
specific (immunity to superinfection with the same phage).
Leading to Termination of Lysogeny
lysogenic bacterium is exposed to adverse conditions, the
lysogenic state can be terminated. This process is called induction.
Conditions which favor the termination of the lysogenic state
include: desiccation, exposure to UV or ionizing radiation,
exposure to mutagenic chemicals, etc. Adverse conditions lead to
the production of proteases (rec A protein) which destroy the
repressor protein. This in turn leads to the expression of the
phage genes, reversal of the integration process and lytic
Fig. 7. Termination of lysogeny
vs Lysogenic Cycle
for lambda to enter the lytic or lysogenic cycle when it first
enters a cell is determined by the concentration of the repressor
and another phage protein called cro in the cell.
The cro protein turns off the synthesis of the repressor and thus
prevents the establishment of lysogeny. Environmental conditions
that favor the production of cro will lead to the lytic cycle
while those that favor the production of the repressor will favor
for animal virus transformation - Lysogeny is a model
system for virus transformation of animal cells
conversion - When a cell becomes lysogenized,
occasionally extra genes carried by the phage get expressed in
the cell. These genes can change the properties of the bacterial
cell. This process is called lysogenic or phage conversion. This
can be of significance clinically. e.g. Lysogenic phages
have been shown to carry genes that can modify the Salmonella O
antigen, which is one of the major antigens to which the immune
response is directed. Toxin production by Corynebacterium
diphtheriae is mediated by a gene carried by a phage. Only
those strain that have been converted by lysogeny are pathogenic