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Chapter 4.
Viruses
—— Non-cellular entities
Section A.
Introduction
A-1. Definition
Contain only a single type of nucleic acid, either DNA
or RNA.
Contain a protein coat (sometimes itself enclosed by an
envelope of lipids, protein, and carbohydrates) that
surrounds the nucleic acid.
Multiply inside living cells by using the synthesizing
machinery of the cell.
They are simple, acellular entities, may be regarded as
an exceptionally complex molecular microorganism .
A-2. Development of Virology
•Although the ancients did not understand the nature of
illnesses, such as rabies, smallpox, they were acquainted
with disease that are now known to be viral in origin.
•In 1886,the Dutch chemist Adolf Mayer (German)
showed that tobacco mosaic disease was transmissible. In
1892, the Russian bacteriologist Dimitri Iwanowski
proved that the infectious agent of the disease could pass
through the porcelain filter and contracted tobacco
mosaic disease. In 1898, the Dutch botanist Martinus
Beijeringck observed that the behavior of this agent was
caused by a filterable virus (Latin, for poison) ,which
would multiply only in living plant cell (1890).
tobacco mosaic disease
The Russian microbiologist
Winograsky discovered that soil
bacteria could oxidize iron, sulfur
and ammonia to obtain energy,
and also isolated nitrogen – fixing
bacteria.
Beijerinck made fundamental
contributions to microbial ecology.
He isolated Azotobacter and
Rhizobium.
At the same time, many other diseases were found to be
caused by the filterable virus by other scientists. The
term virus have been commonly used up to now.
In 1935, American chemist Wendell M. Stainley (of the
Rockefeller Institute) isolated the tobacco mosaic virus
and successfully crystallized the virus, making it
possible for the first time to carry out chemical and
structural studies on a purified virus. Stainley was later
awarded the 1946 Nobel Prize in chemistry.
•At about the same time (early 30s), the invention of
electron microscope made it possible to see viruses.
Kausche (German, 1940) observed the rod-like shape
of TMV for the first time.
• From 1950s, molecular biology of virus had made a
great progresses in the feature, components, structure,
even the sequencing of viral genomes.
• Advances in molecular biology techniques have led to
the recognition of new human viruses. Newly
recognized viruses are referred to as emerging virusesviruses that are not necessarily new but that may be
new to Western medicine.
• HIV (human immunodeficiency virus) was recognized
as the agent of AIDS (acquired immunodeficiency
syndrome) in 1983. EHF (Ebola hemorrahgic fever)
have been learned in Zaire in 1995. And the virus
(coronavirus)responsible for the SARS (severe acute
respiratory syndrome) as you know in 2003.
HIV
INFLU
Orf virus：口疮病毒(接触性脓疱皮炎病毒)
（负染技术）
Vaccinia virus 痘苗病毒
（投影技术）
Ebola virus ：埃博拉病毒（正染技术）
SARS病毒
Section B. Virus
B-1. Host Range
The host range of a virus is the spectrum of host
cells the virus can infect. How ever, most viruses are
able to infect specific types of only one host species.
Viruses
bacterial phage
phycophage
Phage
mycophage
Protozoal viruses
Animal viruses invertebrate viruses
vertebrate viruses
Plant virus
B-2. Viral Size
Viral sizes are determined with the aid of electron
microscopy. Different viruses vary considerably in size.
Although most are quite a bit smaller than bacteria,
some of the larger viruses (such as the vaccinia virus or
poxviruses) are about the same size as some very small
bacteria (such as mycoplasmas). Viruse range from 20 to
14,000 nm in length.
Largest: Poxviruses 450 nm
Longest: Filovirus 80-14,000 nm (Ebola virus)
Smallest: Bean distortion dwarf virus (BBDV) 9-11
Thinnest: Coliphage f1 5 × 800
nm
B-3. Viral Structure
3-1. Virion
A virion is a complete, fully developed viral particle
composed of nucleic acid and surrounded by a protein
coat called Capsid that protects it from the
environment and is a vehicle of transmission from one
host cell to another.
Capsid made up of protomers (capsomer)
Virion
Genome (core): DNA or RNA
Envelope
virion(病毒粒子) or virus particle(病毒颗粒)。
3-2. Symmetries and Morphology
Viruses may be classified in to tree basic different
morphological types based on their capsid architecture.
The structure of these capsids has been revealed by
electron microscopy and a technique called X-ray
crystallography.
3-2.1 Helical symmetry
Helical capsids are shaped much like hollow tubes
with protein walls. TMV provides a well-studied
example of helical capsid structure. RNA is wound in a
spiral and positioned toward the inside of the capsid
where it lies within a groove formed by the protomers
(capsomers or protein particles).
Helical symmetry
Icosahedral symmetry
Complex symmetry
3-2.2 Icosahedral symmetry or Polyhedral symmetry
The icosahedron is one of nature ’ s favorite
shape.( the helix is probably most popular) Viruses
employ the icosahedral shape because it’s the most
efficient way to enclose a space. A few genes, sometimes
only one, can code for proteins that self-assemble to
form the capsid. In this way, a small number of linear
genes can specify a large three-dimensional structure.
The capsids are constructed from ring-or knob-shaped
units called capsomers or protomers, they usually form
a unit of five or six protomers named pentons and
hexons which form the 12 corners (apexex) and 20
triangular faces. N=10 ( n-1 )2 +2
Icosahedral symmetry
3-2.3 Complex symmetry
head
Some viruses do not fit into
either the category. The
Core or
poxviruses and T-even
tube
coliphages are two important
examples. They are complex
symmetries with both heads Fiber
(icosahedral) and tails
(helical) combined together
(such as T2 , T4 , T6)
collar
Helical
sheath
Base
plate
pins
T-2 coliphage
3-2.4 Enveloped viruses
Many animal viruses, some plant viruses are bounded
by an outer membranous layer called an envelope.
Animal viral envelopes usually arise from host cell
nuclear or plasma membranes; their lipids and
carbohydrates are normal host constituents. In contrast,
envelope proteins are coded by virus genes and may even
project from the envelope surface as spikes or
peplomers . Influenza virus is a well-studied example of
an enveloped virus.
3-3 Chemical Compositions
Nucleic acid
ssDNA
DNA dsDNA
RNA ssRNA
dsRNA
proteins
+RNA
-RNA
Structural
(protomer)
functional
(enzymes)
Functional proteins are of non-structural proteins mostly
exist freely from the structures and functioned as
enzymes, or the infectious factors. Such as attachment or
penetrating associate glycoprotein spikes on the surface
of HIV, influenza viruses, or the tail fibers of bacterial
phages. Certain viruses contain enzymes of
neuraminidase responsible to the damage of host plasma
membrane or cell wall. Most functional proteins found in
viruses are duplication associate enzymes, such as
polymerases of DNA or RNA, reverse transcriptase, also
the protein synthesis associated enzymes.
Other components
Other components refer mainly to the substances
found in envelopes of complexity virions. Lipids and
carbohydrates are often found within the envelopes
that usually arise from host cell nuclear or plasma
membranes. That is why they resemble the membrane
components and structures. Carbohydrates exist
generally in forms of glycoprotein and responsible for
their antigenicities.
B-4. Replication or Multiplication
Although the means by which a virus enters and exits a
host cell may vary, the basic mechanism of viral
multiplication is similar for all viruses. The best
understood viral life cycles are those of the bacteriophage.
So that we will take the T-even coliphages as an example
of the life (lytic) cycle.
The multiplication (life) cycle of phages, like that of all
viruses, occurs in five distinct stages:
attachment (adsorption)
penetration
biosynthesis
assembly (maturation)
release or burst
4-1. Attachment
After a chance collision between phage particles and
bacteria, attachment or adsorption, occurs. During
this process, an attachment site on the virus attaches
to a complementary receptor site on the bacterial cell.
This attachment is a chemical interaction in which
weak bonds are formed between the attachment and
receptor sites. T-even coliphages use fibers at the end
of the tail as attachment sites. The complementary
receptor sites are on the bacterial cell wall. This gives
the specific feature of adsorption. The receptors and
the attachment sites are the chemical fundamentals of
the specific adsorption. All the viruses and hosts have
a similar mechanism.
Attachment
4-2. Penetration
After attachment, the phage fixed onto the surface of
the host cell, and a stimulation was given. Then the tail
sheath would contract to be half of its length, and the
tail core (tube) insert into the host cell wall and the
membrane. This process needs the help of ATPase and
phage lysozyme. They are carried within the tube and
helpful to the ATP release and the peptidoglycan
digestion (degradation).
After that, the DNA from the phage head,pass
through the tail core (tube), and enters (be injected
into) the cell. The capsid of the phage remains outside
of the cell wall.
4-3. Biosynthesis (Replication)
Once the phage DNA has reached the cytoplasm of
the host cell, the biosynthesis of viral nucleic acid and
protein occurs. Host protein synthesis is stopped by
virus-induced degradation of the host DNA, viral
proteins that interfere with transcription or the
repression of translation.
Genetic controls regulate when different regions of
phage DNA are transcribed into mRNA during the
multiplication cycle.
Since the viruses contain different genetic material
(DNA or RNA) as their genetic information depend on
the species, the modes of mRNA transcription are
different from each other. There are six patterns
found in viral mRNA transcription:
ssDNA (±DNA)
+ DNA
-DNA
mRNA
dsDNA (± DNA)
-DNA
mRNA
dsRNA ( ± RNA)
-RNA
mRNA
± RNA
+RNA ± DNA
ssRNA
- RNA mRNA
-RNA
+DNA
mRNA
- DNA mRNA
4-4. Assembly (Maturation)
In the next sequence of events, maturation (assembly)
occurs. In this process, the phage DNA and capsids are
assembled into complete virions. The viral component
essentially assembled into a viral particle spontaneously,
eliminating the need for many non-structural genes and
gene products. The phage heads and tails are separately
assembled from protein subunits, and the head is filled
with phage DNA and attached to the tail.
4-5. Release
The final stage of multiplication is the release of virions
from the host cell. The term lysis is generally used for
this stage in T-even phages because of the cell membrane
actually broken up and releasing virions.
B-5. One-Step Growth Curve
One-Step growth curve is a classical experiment
designed for demonstrating the multiplication of phages.
In this procedure, a phage suspension is diluted until a
sample containing only a few phage particles is obtained.
These particles are then introduced into a culture of host
cells. Periodically, samples of phage particles are
removed from the culture and inoculated onto a plate
culture of susceptible host cells; the plaque method is
used to determine the number of infective phage
particles on this culture. The curve drawn out contains
three important stages: Latent phase; Burst (rise phase);
and plateau. The number of phages newly released from
a single cell is referred as to Burst Size (50-200)
One-step growth curve was designed by Max Delbrük
and Emory Ellis in 1939. It is significant to the
understanding of bacteriophages with three important
aspects:
The Latent phase; Burst (rise phase) and Burst size.
This experiment is a remarkable event which starts
the beginning of modern virology. And it is also an
evidence of becteriophage reproduction mode of
mutiplication, other than fission.
Latent phase: is the period from the infection to the completely
formation of the first phage. There is no release of virions. At the
very beginning of this phase, host cell contains no complete,
infectious viral particles. This phenomenon can be observed by
lysing the cells with chloroform. The initial period is called
eclipse period.
Burst (or rise) phase: This is the period that closely following the
latent phase. In this rise phase, the bacterial cells lyse rapidly
and release mature, infectious virions. Since the host cells are not
synchronized individually, the burst of cells last for a period,
other than a theoretically sudden burst.
Plateau phase: This period is the final stage that the plaque is
formed and no more viruses are released. The total number of
virions released per cell is called burst size.
B-6. Lysogeny
Some phages( viruses) do not cause lysis and death of
the host cell when they infect and multiply.( such as
coliphage λ ) They can either proceed through alytic
cycle (in some certain conditions) or incorporate
(integrate) their DNA into the host DNA. In the latter
state, called Lysogeny, the phage remains latent
(inactive).
The phages that certainly cause lysis and death of the
host cells are called Virulent Phage.
The phages that can integrate their DNA into the host
DNA and usually do not cause lysis of the cells are
named Temperate Phage.
Lysogenic cycle of bacteriophage lambda in E.coli
• The temperate phage ’ s DNA that integrated in the
host ’ s DNA is called Prophage.
• The temperate phage can remain in the host cell in
three forms: Prophage; Free form; Vegetative form.
• The bacteria that contain a Prophage are called
Lysogen or lysogenic bacteria. They are different from
the normal ones by the following characters:
1. Spontaneous lysis
2. Inductive lysis
3. Re-conversion (recovery)
4. Super infection immunity
5. Lysogenic conversion (ß-phage, diphtheria toxin)
6. Hereditary stability
Viroids and Prions:
Viroids are small, circular, single-stranded RNA
molecules that are the smallest known pathogens. The
extracellular form of the viroid is naked RNA-there is
no capsid of any kind.
Prions have a distinct extracellular form, but the
extracellular form seems to be entirely protein. It
apparently does not contain any nucleic acid, or if it
does, the molecule is not long enough to encode the
single kind of protein of which the prion is composed
Prusiner (1982) discoverd proteinaceous infectious
particle and named it Prion or Virion--朊病毒
1997年，Stanley B. Prusiner owned Nobelpris
In 1982, American neurobiologist Stanley Prusiner
proposed that infectious proteins caused a neurological
disease in sheep called scrapie. Experiments suggested
that the infectious agent is pure protein. Prusiner
coined the name PRION for proteinaceous infectious
particle.
Here is one hypothesis for how an infectious agent can
lack nucleic acid . The major prion protein (PrP) and
its gene are found in normal host, and the PrP gene is
located on chromosome 20 in humans. An abnormal
PrP designated PrPSc is found in brains of animals
with scrapie. PrPSc may cause a change in PrP to
refold into PrPSc . Anyway, its genetic mechanism is in
debate.
Viruses Taxonomy
Viruses have been detected in almost every kind of living
organisms, Usually they are separated into several large groups
based on their hosts, such as animal viruses, plant viruses,
bacterial viruses, and so on. But at present, the classification of
viruses is still in a unsatisfactory state. However, a uniform
classification system has been developed since 1971, beginning
with the report on the International Committee on Taxonomy of
Viruses(). The report recorded a universal taxonomy scheme
consisting of three orders, 56 families, 9 subfamilies, 233 genera
with over 4000 members. The system still contains hundreds of
unassigned viruses, and of cause, new viruses are still being
discovered.
Viruses, nowadays, are divided into certain taxonomic groups
according to their characteristics including the kind of hosts
infected, structures of virions and nucleic acids, mode of
reproduction.
1.
Type of the host
2.
Type of the virus nucleic acids (DNA or RNA)
3.
Molecular weight of nucleic acids
4.
Capsid symmetry of virions (cubic, helical, complex)
5.
Envelope presence
6.
Number of capsomers of cubic viral capcid
7.
Type of the virus release
8.
The presence of reverse transcriptase
9.
Intracellular location of viral replication
Mode of transmission and disease caused
1. Type of the virus nucleic acids / single or double strand
2. Molecular weight of nucleic acids / content of nucleic acids
3 Shape of virion / shape of capsid
4 Host / vector of infection
Coliphage T4,
Poxviridae,
D/2 : 137/40 : X/X : B/0
D/2 : 130-240/5-7.5 : X/* : I. V/ 0. R. Ve/Ac. Di. Si
Coronaviridae, R/1 : 9/* : Se/E : V/I. R
In the first group, R = RNA, D = DNA, 1 = single stranded, 2 =
double stranded. In the second group, the unit of molecular
weight is 106 Da, and content of nucleic acids is expressed by
their percentage. The next group is more complex, S represent
spherical in shape, E or U code for rodlike, and X represent
complex or binal. The last group is the most complex that
expressed usually with more codes. A = actinomycetes, B =
bacteria, F = fungi, I = invertebrate, V = vertebrate, P =
pteridophyte, S = seed plant. Vectors of infection are often
coded with the first two letters, such as Fu means fungi and Ne
represents nematodes. 0 means no vectors needed. * is used to
represent undefined characteristics and ( ) represent suspected
data. Here are some examples of taxonomic codes used in viral
classification.
Microbial viruses, here, refer to as the viruses that infect
microbes including bacteria, fungi and unicellular algae. Viruses
that infect bacteria are called bacteriophage, or simply, the
phage. The viruses infectious to fungi are termed mycovirus.
Those infectious to unicellular algae resembles many
bacteriophages in having a tail.
Mycovirus was found firstly in mushroom Agaricus bisporus.
Most mycoviruses are obtained from higher fungi other than the
lowers, such as Lentinulas edodes, Penicillium, and Aspergillus.
Much less is known about the viruses of lower fungi, although
both of dsRNA and dsDNA viruses have been discovered.
Plant Viruses are often difficult to cultivate and purify than that
of animal viruses or bacteriophages. Therefore, they are not as
well studied as the other viruses. Their information are mainly
come from the research on tobacco mosaic virus (TMV), because
it can grow in isolated protoplasm resembling animal viruses
grow in cell suspensions.
Vertebrate virions are usually spherical shaped with
icosahedral symmetry. Some are helical symmetry. Only a few
of them have a complex symmetry in structure, such as bullet
shaped rabies virions and brick liked Poxviruses. Enveloped
virions are more common among this group than that of plant
or bacteria. Nearly almost types of nucleic acid are detected in
these viruses.
Many insect virus infections are often accompanied by the
formation of inclusion bodies within the infected cells. The
inclusion bodies of either granular or polyhedral form are
protein in nature and enclose one or more virions. There are at
least three types of inclusion bodies according to their shape
and location intracellular. Anyway, insect viruses are interested
in their use as a possible means of biological pest control and
advantages over chemical toxins have been made. At least, we
believe that they are safety and stability to the environment
since they are invertebrate-specific and have a long shelf life.