GENERAL VIROLOGY

Viruses
The smallest infectious agents
(20-300 nm)
only seen under

Electron Microscope
(except Poxviruses)

Electron Microscope
- The source of illumination is a beam of electrons
- Electromagnetic lenses

- Magnification is 100.000 or more

Electron Microscopy

Bacterial
fimbriae

Structure of viruses

Virus particle (virion)

Structure of Viruses

Viral Symmetry
Ico

1. Icosahedral symmetry:

e.g., all DNA viruses, except

poxviruses & some RNA viruses

2. Helical symmetry:

e.g., many RNA viruses (e.g. rabies

virus)

3. Complex symmetry:

e.g., poxviruses (brick-shaped)

Icosahedral Symmetry

Icosahedral Symmetry

Rotavirus

Calicivirus

Astroviru
s

Helical
Symmetry

Complex symmetry

Differences between viruses

& bacteria
1. They are obligatory intracellular
parasites
They can not be cultivated on artificial
culture
media.

can only replicate inside living cells.

2. Viruses contain only one type of

nucleic acid
(DNA or RNA), never both.

3. They can not be cultivated on

artificial culture media.

4. They are not susceptible to

antibacterial agents.

Diagnosis of viral
infections

B- Indirect methods

A- Direct
methods
I. Direct
detection of
viruses and /
or their
components

II. Isolation
of viruses

I.
Serolog
y

II. Skin
tests

A- Direct
methods
I. Direct detection of viruses & / or
their components:
1. Light microscopy:
Examination of large viruses
as Poxviruses
Detection of giant cells
in Herpes infection
Detection of inclusion bodies
e.g. Negri bodies in nerve
cells in rabies

2. Electron microscopy (EM):

Large number of viruses
sample.
Size and shape of viruses.

in

the

3.

Immunoelectron
(IEM):

Sample (unknown
specific antibody

virus)

microscopy
+

known

aggregation of unknown virus

particles
e.g. hepatitis A virus in stools

4. Fluorescent microscopy:
Direct immunofluorescent antibody technique (IF)
e.g. diagnosis of rabies in brain smears.

5. Immunoassays :
For detection of the virus antigens by ELISA /
RIA
e.g. hepatitis B antigens in blood

ELISA

6. Nucleic acid hybridization:

A highly sensitive and specific method.
Viral nucleic acid in sample
labeled probe

+ Specific

hybridization
fluorescence

7. Polymerase chain reaction (PCR)

Amplification of a specific sequence of nucleic acid
Detection e.g. by gel electrophoresis.

A- Direct
II. Isolation of methods
viruses:
Obligatory intracellular parasites
Replicate only in living
susceptible cells

a. Laboratory animals
b. Embryonated eggs
c.
Cell
culture)

culture

(tissue

Cultivation of viruses
a. Laboratory
animals:
Used in:
Isolation of coxsackie viruses by inoculation in
white suckling mice.
Rabies virus inoculation in mice.
For research.

Cultivation of viruses
b. Embryonated eggs

Cultivation of viruses
c. Cell culture (tissue culture)
The
most
method

widely

used

Tissues + trypsin
separate cells

Cells + growth media in

flat-sided bottles

Virus inoculation incubation at 37C (CO2 incubators)

Types of cell lines

Primary cell
lines

Diploid cell
lines

Continuous cell
lines

Prepared
from

Organ
fragments

Human fibroblasts
derived from
embryonic tissues

Tumor cells

Examples

Monkey kidney

Number of
Passages
(subcultures)

5-10

Human embryo lung

HeLa cells from
tissue
carcinoma of cervix

50-100

Unlimited

Detection of Virus Replication in

Tissue Culture:
1- Cytopathic Effect (CPE):
i- Cell death or lysis

ii- Syncytial formation (multinucleated

giant cells ):

2- Transformation:
Viral nucleic acid + cellular DNA cell
transformation foci of malignant cells.

3- Inclusion bodies
Site of virus assembly or degenerative
changes
Their
location
and
appearance
are
diagnostic for a particular virus.

a- Intracytoplasmic:
e.g. Rabies (Negri

b- Intranuclear:
. e.g
bodies)
Herpes
viruses
c- Both: e.g. Measles virus
and
CMV

4- Plaque formation:
Infected monolayer + vital dye unstained
areas (plaques)

5- Haemadsorption:
Monolayer + hemagglutinating virus + RBCs
hemadsorption (RBCs clumping) to infected cells

6. Interference phenomenon:
Monolayer + rubella virus no change for

weeks

Add CPE-producing virus NO CPE

interference)
7- Detection of viral antigens

(due to

8- Direct fluorescent antibody staining of

infected cells (DFA):

:Neutralization test- 9
Monolayer + unknown virus + known
specific Ab
NO CPE (due to neutralization)

Diagnosis of viral
infections
A- Direct
methods
I. Direct
detection of
viruses and /
or their
components

B- Indirect methods

II. Isolation
of viruses

I.
Serolog
y

II. Skin
tests

B- Indirect methods
I. Serological diagnosis:
Detection of antiviral antibodies

2 serum samples
acute phase & 2-3 weeks later,
to demonstrate a rising titer
(4 fold increase or more is diagnostic).
Only one sample may be used in the
acute stage to detect IgM

I. Serological diagnosis:
Serological methods include:

Neutralization test
Complement fixation test
Haemagglutination inhibition test
Indirect IF
ELISA
RIA

II. Skin tests

Used as an indication of cell-mediated
immunity (CMI) in some viral infections, e.g.
mumps.

MCQs

1) Viruses differ from bacteria in

all of the following EXCEPT:
a) Viruses are very small in size.
b) Viruses are obligatory intracellular
parasites.
c) Viruses contain both DNA and RNA.
d) Viruses cannot be cultivated on
artificial culture media.
e) Viruses are not susceptible to
antibiotics.

2) Sites of viral assembly in tissue

culture are called:
a)Plaques
b)Inclusion bodies
c)Viral capsid
d)CPE
e)Areas of transformation

3) Regarding continuous cell lines, all

of the following are true EXCEPT:
a)They allow unlimited number of
passages.
b)They are prepared from tumor cells.
c)After viral inoculation, they should be
sterilized by autoclaving.
d)They are used for isolation of viruses.
e)HeLa cell line is an example of
continuous cell lines.

4) PCR may be used to diagnose

viral infections by detecting:
a)Antiviral IgM antibodies
b)Viral antigens
c)Rising titer of antiviral antibodies
d)Viral nucleic acid
e)CPE produced by the virus in tissue
culture

5) Direct detection of viruses

and/or their components can be
done by all of the following
EXCEPT:
a)Fluorescent microscope
b)Electron microscope
c)Light microscope
d)PCR
e)Biochemical reactions

6) Cultivation of viruses can be

done on:
a)Blood agar
b)Tissue culture
c)MacConkeys medium
d)Nutrient broth
e)Anaerobic media

THANK
YOU

1) Viruses differ from bacteria in all

the following EXCEPT:
a) Viruses are very small in size.
b) Viruses are obligatory intracellular
parasites.
c) Viruses contain both DNA and RNA.
d) Viruses cannot be cultivated on
artificial culture media.
e) Viruses are not susceptible to
antibiotics.

2) Sites of viral assembly in tissue

culture are called:
a)Plaques
b)Inclusion bodies
c)Viral capsid
d)CPE
e)Areas of transformation

3) Regarding continuous cell lines,

all of the following are true
EXCEPT:
a)They allow unlimited number of
passages.
b)They are prepared from tumor cells.
c)After viral inoculation, they should be
sterilized by autoclaving.
d)They are used for isolation of viruses.
e)HeLa cell line is an example of
continuous cell lines.

4) PCR may be used to diagnose

viral infections by detecting:
a)Antiviral IgM antibodies
b)Viral antigens
c)Rising titer of antiviral antibodies
d)Viral nucleic acid
e)CPE produced by the virus in tissue
culture

5) Direct detection of viruses

and/or their components can be
done by all of the following
EXCEPT:
a)Fluorescent microscope
b)Electron microscope
c)Light microscope
d)PCR
e)Biochemical reactions

6) Cultivation of viruses can be

done on:
a)Blood agar
b)Tissue culture
c)MacConkeys medium
d)Nutrient broth
e)Anaerobic media