virus

1. Medical virology
For Medical laboratory students
Level III
Dr/ Samira Hameed Hanash
Associated professor in medical microbiology and
immunology
2022
1

3. Introduction
What``s the virus?
Viruses are the smallest infectious agents. They can infect man, animal, insects, plants and
bacteria. They are able to replicate in live susceptible cells.
General properties of viruses
Characters of viruses that can distinguish them from other microorganism.
1. They are very small in size.
2. They contain one kind of nucleic acid DNA or RNA.
3. They are obligate intracellular parasites i.e virus replicates only inside living cells.
4. They cannot be grown on artificial media (culture) they grow only in tissue culture or
embryonated egg and in living animal.
5. They depend on the ribosome, enzymes of the infected cells for protein synthesis.

4. Discovery of viruses
In 1892 a Russian Pathologist named Dmitri I Winooski
described the causative agent of a disease. That effect the tobacco
leafs as filterable virus because it can pass the bacterial filter, this
virus called tobacco mosaic virus (TMV) and causes a disease is
called tobacco mosiac disease.
Virus Size & Structure
Size of viruses is variable from 10nm to 500nm, as a result these
are small in size:
1- They can pass through bacterial filters.
2- They require high speeds for their sedimentation (called ultra
centrifugation 10.000-30.000rpm.
3- They are only seen by electron microscope.

5. Structure of viruses
Each virus particle is composed of a protein coat or capside and a nucleic acid
core. The genetic material as nucleic acid with capisd is called nucleocapsid.
1. viral genome (genetic material):
- Viruses contain either DNA or RNA but not both.
- Most DNA viruses are double stranded while most RNA viruses are single
stranded.
- The nucleic acid may be liner or circular.
- Some RNA viruses have segmented genome such as Rotavirus and influenza
virus.
- All viruses have one copy of their genome except retroviruses have two copies.

6. Function of Nucleic acid
a. It is the infectious part of the virus.
b. It carries the genetic information for replication, virulence, Antigenic specificity of the protein coat.
1. Viral capsid:-
Protein coat is made up of many protein subunits called capsomeres.
Function of capsid
1. Protect the viral genome against inactivation by nucleases enzyme.
2. It is responsible for the structural symmetry of virion (virion = complete virus particle)
3. Help for attachment of virion to susceptible cells.
4. Capsid proteins are important antigens that induce antibodies that neutralize virus infectivity.
5. Viral envelope:-
Many viruses are surrounded by envelope which consists of proteins, glycoprotein, and host lipids-derived from
host membrane, also there are a glycoprotein spike like projections, which attach to host cell receptors during the
entry of the virus in to the cell.

7. For example Haemagglutinine (HA). Neuraminidase (NA) in influenza virus helps
the virus attachment to the host cell.
The envelope is derived or acquired from the host membrane during release from
the cell.

9. 1. Virus Enzymes
Present in some viruses, Require for virus live cycle and Not available in host.
e.g: Integras, RNA dependant – RNA polymerase, RNA dependent – DNA polymerase (R-T).
Virus symmetry (shapes)
a. Icosahedral symmetry shape.The virus capsid has 20 triangular faces, 12 corners e.g. Polio virus
and herpes simplex virus.
b. Helical shape in which the in which the capsomers are arranged in a hollow coil that appears
rod-shaped. The helix can be either rigid or flexible. All human viruses that have a helical
nucleocapsid are enclosed by an outer membrane called an envelope.
Such as Rabies virus and tobacco mosiac virus.
complex shape. The viruses are complicated in structure (in this shape). They are a combination of both a
helical and icosahedral.such as Pox virus and bacteriophage

10. Virus nomenclature
a. Name after disease causes , for example hepatitis virus.
b. Name of virus related to the scientist who first discovered the virus.
For example .Epstein Barr virus (E.B.V).
a. Name related to the city-state-country where the virus was first discovered. For example . Hong
Kong influenza virus.
b. Complex name such as HIVand CMV.
Classification of viruses:
Viruses are classified according to:
1. Virion morphology including size, presence or absence of envelope, type of symmetry.
2. Mode of transmission the vectors and pathogenicity
3. Host infected (bacteria, parasite, plant, animal ….. and soon.
4. Genetic material (RNA or DNA.) single or double, circular or liner, segmented or non- segmented.
5. Type of target cell- type of cells that virus can infect (dermotopic, neurotopic, pneumotopic, lymphotopic,
viscerotopic, spleen and liver).

11. Methods of virus cultivation
Viruses are obligatory intracellular parasites that grow in living cells but they can survive in certain
conditions as non-replicating particles.
Requirement for viral growth:
- Heat: there is great variability in the heat stability of different viruses. Most viruses are pathogenic viruses
and are inactivated at 55-60co because their capsid protein is destroyed.
- Cold: most viruses can be preserved at sub-freezing temperature .Some can with stand lyophlisation and
can be stored in the dry state at 4co or at R.T which envelope Viruses lose their infectivity after prolonged
storage at -70co.
or lower in liquid nitrogen
Concentration of hydrogen ion (PH)
Most viruses are stable in the PH range 5 to 7 enteroviruses that have to pass through the
stomach resist the low PH. All viruses are destroyed by alkaline conditions.

12. * Radiation
Radiation such as U.V causing damaging of DNA that can cause inactivation of the virus, x-ray, gamma rays and
beta particles inactivate viruses.
* Stabilization by salts
Magnesium chloride stabilises polioviruses, magnesium sulphate stabilises influenza viruses, sodium sulphate
stabilizes herpes virus. They are Important in the preparation of vaccines.
Ether susceptibility and lipid solubility:-
- Enveloped viruses are inactivated by ether.
Non ionic detergents solubilise lipid constituents but do not denature the proteins of the capsid.
Anionic detergents solubilise the lipid constituents and disrupt the capsid in to separated polypeptides.
 50% glycerol
Many viruses remain alive in 50% glycerol for many years while bacteria are killed.
• Formaldehyde
Destroys viral infectivity used in the production of inactivated viral vaccines.
Hydrogen peroxide and hypochlorite are disinfectants for most viruses.

13. - Non ionic detergents solubilise lipid constituents but do not
denature the proteins of the capsid.
- Anionic detergents solubilise the lipid constituents and
disrupt the capsid in to separated polypeptides.
50% glycerol
Many viruses remain alive in 50% glycerol for many years
while bacteria are killed.
Formaldehyde
Destroys viral infectivity used in the production of inactivated
viral vaccines.
Hydrogen peroxide and hypochlorite are disinfectants for most
viruses.

14. Growth of viruses in the laboratory
There are 3 systems for cultivation of viruses in the laboratory:-
1. Cell culture.
2. Embryonated eggs.
3. Living laboratory animals.
Cell culture
The recent method for cultivation growing of virus.
- Pieces of animal or human tissues.
- It is a homogenous collection of cells.
- Prepared by treating animal tissue with enzyme to get separate cells.
- Cell grow and adhere to glass or plastic container to from a monolayer or sheet of
cell within few days. (these growth media containing serum)
- This media kept from microbial contamination by adding antibiotics for preventing
the contamination.

15. Type of tissue culture
1. Primary cell lines
These are prepared from organ fragments, for example, monkey kidney.
1. Secondry cell lines
Derived from primary cell lines by subculture.
1. Human diploid cell lines
These are usually fibroblasts derived from human embryo tissue such as human embryo lung tissue.
1) Continuous cell lines
These are derived from tumor cells. They can be divided indefinitely such as Hela cell line derived from
carcinoma of the cervix.
Detection of virus replication in cell culture
Many viruses kill the cells in which they replicate;
Cytopathogenic effects (CPE)
Altered shapes (rounding and group like cluster formation.
Cell lyses, cell death, detachment from glass surface.
Giant cell formation (cell fusion).
Cell transformation.

16. Embryonated eggs:-
This method is used for virus cultivation. The virus suspension is injected in to the fluid
of the egg. Viral growth is detected by death of the embryo or lesion on the membrane of
the egg.
It is the most common method for virus growth and isolation as well as used for
preparation of virus vaccines.
Living animals
Mice, rabbit, quineapig
Animal inoculation was mainly used in the past before tissue culture method were not
known. This method still used for study of
- Immunoresponse.
- Viral pathogenesis.
- Viral oncogenesis.
- Viral disease.
- Primary isolation of viruses.

17. 1. Plaque formation
Plaque are virally infected area in cell culture.
1. Inclusion bodies
Result from accumulation of viruses are eosinophilic or basophilic bodies that appear
within the cell as a result of infection.
They are intracytoplasmic or intranuclear or both.
For example, Rabies-negribodies in cytoplasm of nerve cells present in the brain tissue.
Diagnostic Test for rabies.
1.
Neutralization test
This is done by mixing the virus with specific antisera then the mixture is added to cell
line. absence of CPE indicates virus neutralization.
1. Heamadsorption
When RBCs are added to infected cells they will appear as rosettes shape on the area
where the virus is growing.

18. Vir us r eplication
Viruses are strict intracellular parasites, replicating only within
the host cells.
Steps of virus replication
1- Attachment or adsorption. 2- Penetration.
3- Uncoating. 4- Macromolecules synthesis.
5- Assembly. 6- Release.
Infection to occur depends on
1) Suitable site.
2) Number of viruses (Infective dose).
3) Immune response of the host.
4) Virulence of the microbes.

20. Attachment or adsorption
This step depends on the presence of specific receptors and selection of the cells by viruses and depends on the –
nature of receptors which differ for different viruses (all viruses have receptors -binding protein) on their outside
surface. The virus receptors on the cells are often glycoprotein or glycolipids.
Examples
- Hamagglutinine in influenza virus, sialic acid which is found on most epithelial cells.
- Glycoprotein 120 in (HIV) → with CD4 receptor on T-helper lymphocyte.
Concentration of receptors play role in attachment each susceptible cell contain about 104 -105 receptors on the
cell surface.
.G. protein in rabies virus with acetylecholine
Penetration or entry:
Viruses enter the cell by different mechanisms
Membrane fusion mechanism
These viruses have fusion protein – which mediate the fusion between the lipids of the viruses and the lipids of
the cell membrane and viral nucleic acid enters the cell directly. (e.g. enveloped viruses).
Receptors-mediated endocytosis:-
The virus is endocytosed by the host cell in acidification states.phagolysosme is formed. The viral envelope is
digested by hydrolyzing enzyme of the phagolysosme lead to relase of the capsid and uncoating occurs. Both (1)
and (2) occur in enveloped viruses.

21. 1. Endosome lysis
Non enveloped viruses lack the lipid membranes found in enveloped viruses as a result these
viruses cannot enter cells via a simple process of membrane fusion between the virus
envelope and the host cell membrane. Endocytosis occurs to form endosome which
dissolved by virus enzyme, the virus is released in to the cytoplasm of the host cells and
uncoating occurs.
Uncoating
This process main the hydrolysis of virus capisd protein to release the viral nucleic acid in to the
host cytoplasm under the effect of hosts enzyme or viral enzymes (proteases enzymes).
Macromolecules synthesis:- (synthesis of proteins) N.A.
Most DNA viruses replicate in nucleus (because these viruses do not have RNA dependent RNA
polymerase enzyme except pox virus replicate in cytoplasm and (have RNA dep. RNA. Poly).
Most RNA viruses replicate in the cytoplasm because they have:
(RNA dependent-RNA polymerase) except influenza virus replicate in nucleus due to these
viruses do not have (RNA dep-RNA poly) enzyme.

22. Virus assembly
Virus assembly occurs in the nucleus or in the cytoplasm which depends on the viruses.
during assembly Virus nucleic acid is surrounded by the capsid proteins and all the accessory parts as the genomic enzyme and
other capsid enzyme.
Virus Release
Release of virus depends on the type of virus some release by budding through the cell membrane. These are called enveloped
viruses which acquired lipoproteins envelope (enveloped virus). Others release by a process called cytolysis, rupturing the cell
membrane and Release (Non enveloped viruses).
Virion , complete virus particle.
prions , Infectious particle, are protein, without nucleic acid. Prions Highly resistance to heat,formaldehyde, U.V. These prions
cause a disease called spongy form encephalopathies.
Examples,
Scrapie in sheep.
Mad cow disease in cattle called brain spongi form encephalopathies.
Kuru, creutzfeldt Jakob disease ( CJD) in human.
Transmitted through contaminated surgical instrument and consumption of contaminated meat.
Viroids. Small infection agents cause disease of plants. They are nucleic acid molecules without a protein coat (ss RNA).

23. Pathogenesis:
The ability of viruses to cause disease can be viewed on two distinct levels: (1) the changes that occur
within individual cells (discussed previously) and (2) the process that takes place in the infected patient.
The Infected Patient
Viral infection in the person typically has four stages: incubation period, prodromal period,
specific-illness period, and recovery period.
Types of virus infection
1. Local virus infection:
This type of infection occurs at the site of entry with no
viraemia characterized by:
- short incubation period.
- Short lasting immunity.
- IgA, interferon are present
e.g. human papilloma virus, rotavirus and influenza virus.
2--Systemic infection:
Systemic infection occurs after primary replication at the site of entry then virus travels to the different
organs via blood stream or lymphatic ducts characterized by:
- Long incubation period.
- Long immunity.
- IgM, IgG present.

24. 3-Persistent infection:-
- Carrier state e.g. hepatitis c virus
- Latent infection herpes simplex virus
- Slow virus infection . Creutzfeldt-Jakob disease, are caused by prions
Host Defenses
Host defenses against viruses fall into two major categories:
(1) nonspecific, of which the most important are interferons and natural killer cells and
(2) specific, including both humoral and cell-mediated immunity.and Interferons which are an
early, first-line defense, whereas humoral immunity and cell-mediated immunity are effective only
later because it takes several days to induce an immune response.
General Action of interferon's:
Interferon's are small proteins released by macrophages, Lymphocytes, and tissue cells infected
with a virus, when atissue cell is infected by a virus, it releases interferon.Interferon will diffuse to
the surrounding cells, when it binds to receptors on the surface of those adjacent cells, It is inhibit
virus replication by blocking the production of viral proteins, primarily by degrading viral mRNA.
They induce the synthesis of a ribonuclease that specifically cleaves viral mRNA but not cell mRNA.
Three types of interferon's: alpha, beta and gamma. Alpha and beta interferon’s have a stronger
antiviral action than gamma interferon. The latter acts primarily as an interleukin that activates
macrophages

25. Other Nonspecific Defenses
 Natural killer (NK) cells are lymphocytes that destroy cells
infected by many different viruses, i.e., they are
nonspecific. NK cells do not have an antigen receptor on
their surface, unlike T and B lymphocytes. Rather, NK cells
recognize and destroy cells that do not display class I MHC
proteins on the surface. They kill cells by the same
mechanisms as do cytotoxic T cells, i.e., by secreting
perforins and granzymes.
 Phagocytosis by macrophages and the clearance of mucus
by the cilia of the respiratory tract are also important
defenses. Damage to these defenses predisposes to viral
infection.

26. Specific Defenses
Active immunity to viral infection is effected by both
antibodies and cytotoxic T cells. It can be elicited either by
exposure to the virus or by immunization with a viral
vaccine.
Passive immunity consists of antibodies preformed in
another person or animal. Two specialized examples of
passive immunity include the transfer of IgG from mother to
fetus across the placenta and the transfer of IgA from
mother to newborn in colostrum.
The duration of active immunity is much longer than that of
passive immunity. Active immunity is measured in years,
whereas passive immunity lasts a few weeks to a few
months.

27. : General action of interferon's