Practical Virology

Dr. Katan Sabir Ali

Hawler Medical University
College of Health sciences
Department: Medical Microbiology

Lecture:1
 Diagnostic Virology objectives
Describe the techniques, and interpretation of different methods
used in diagnosis of viral infections

Obtain a general understanding of the principles of viral

diagnosis by laboratory

 Become familiar with diagnostic methods typically used to

diagnose viral diseases

Determining the stage of viral infection (acute, chronic and

convalescent)

 Be familiar with the typical amount of time needed to obtain
results from each of the different methods of virus detection
Methods of diagnosis for viral infections
• Direct detection

• Serology

• Viral growth in cell culture

• Viral nucleic acid detection

 Indication for the diagnosis of
viral infection
1. For proper management of diseases caused by
viruses such as:
a. Rubella (abortion in first trimester),
b. Post exposure immunization in the establishment of
rabies
c. Caesarian section is required if the patient has
primary genital herpes.
d. The baby borne in an HBs Ag positive mother
(HBV) should be immunized
2. Prevention of spread of viruses such as HIV, HBV and
HCV from carriers to other is possible by screening of
blood donors.

3. Dangerous epidemics like poliomyelitis is importance

for planning to start the control measures and
immunization

4. Reporting of new viruses.

1. Direct detection of the virus

A. Detection of viral cytopathic effect by light

microscopy

B. Detection of viral particles by electron

microscope.
A. Light Microscopy
•Light Microscopy has been traditionally used in
directly demonstrating viral infections by detecting
the viral cytopathic effect in smear & tissue.
•Inclusion bodies: are discrete area of dense of
stainable substances, usually viral proteins or viral
particles
•They can be either intra nuclear (present inside the
nucleus) of infected cell or intracytoplasmic (present
inside the cytoplasm of the infected cell).
• Viruses that are assembled in the nucleus
(usually DNA viruses) such as
• Parvoviruses
• Papova viruses .
• Viruses that are assembled in the cytoplasm
(usually RNA viruses) such as
• Respiratory syncytial virus
• Rabies virus ( Negri bodies )
Resolving Power and Magnification
The resolution power: is the ability to distinguish
between two objects that are very close to each other

The resolving power of a microscope depends mainly on

the quality of its objective lens.

The magnification power: increase in the size of the

optical image over the size object being viewed
 Optical Instrument Resolving Power
Human eye 0.2 mm
(200 µm )
Light microscope 0.0002 mm
Magnified 1000–1500 times (0.20 µm)

Scanning Electron Microscope (SEM ) 0.01µm

Magnified 100 and 100,000 times 10 nm

Transmission Electron Microscope 0.0002 µm

(TEM) Magnified 500 to 500,000 times 0.2 nm
 B.Direct demonstration of virus:
 Viruses are very small and cannot be visualized by light microscopy.
Electron Microscope
It is a scientific instrument that use a beam of highly energetic
electrons to examine objects on a very fine scale

Electron microscopes use electrons instead of (light) to illuminate a

specimen and detector instead of eyes

Wavelength of electron beam is much shorter than light, resulting in

much higher resolution.
 Electron Microscopy

Light microscoe

Electron Microscopy
 • Types of Electron Microscope :
A. Transmission Electron Microscope (TEM):
 TEM is used to study the inner structure of objects (tissues,
cells, viruses)
 The electrons are released by heating a very thin metallic
(tungsten) filament (the cathode) in a vacuum.
 The electrons are released and then submitted to a high voltage
between the cathode and the anode (a metallic plate with a hole in
its center )
 Electrons are thus attracted to the anode and accelerated to high
speeds.
 They pass through the central opening in the anode, forming a
beam of electrons that penetrates the tube of the microscope.
 The first lens is a condenser that focuses the beam of electrons
on the section.
Some electrons interact with atoms of the section, whereas others
simply cross the specimen without interacting.
Most electrons reach the objective lens, which forms a magnified
flat image that is then projected through other magnifying lenses.
Because the human eye is not sensitive to electrons, the image is
finally projected on a fluorescent screen for detection
A. Transmission Electron Microscope (TEM):
Transmission Electron Microscopy
(TEM)
A transmission Electron Microscope
is anologous to a slide projector
A transmission Electron Microscope
image
 B- Scanning Electron Microscope (SEM)
 SEM is applied to detect the three-dimensional views of the
surfaces
 This electron microscope produces an electron beam that is
moved sequentially (scanned) across the specimen.
 The electron beam interacts with a very thin heavy metal
coating previously applied to the specimen and produces
reflected or emitted electrons.
 heavy metals such as lead, gold, platinum, or uranium are used
to introduce electron density into a biological specimen by
staining, to make conductive coatings
 The reflected electrons are captured by a detector that transmits
them to amplifiers and the signal is projected into (a monitor),
resulting in a image with three dimension
Scanning Electron Microscope
Scanning Electron Microscopy
A three-dimensional image of a poxvirus
Limitation of Using Electron Microscope :
However, it is not a tool that is routinely used to
identify viruses in a diagnostic setting.

This is because the test lack sensitivity: a concentration

of approximately (1,000,000) virus particles per
milliliter of fluid is required in order to see the virus of
interest.
The viruses are usually present in very small numbers in
clinical specimens and other contaminating material
tends to obscure their presence.
Electron microscopy (EM) of
herpes virus
Creeping appearance
Electron microscopy ( EM) of adenoviruses
Capsid possess projecting fibers
 EM of Hepatitis B virus (HBV )
(Spherical and filamentous shape )
EM of influenza viruses (spherical shape).
EM of Rabies virus (Bullet shape) .
 EM of Rotaviruses
The term "rota", meaning wheel, is derived from the
appearance of the complete double-capsid particle
EM of Ebola virus ( filament shape)
Questions?