CHAPTER 5

BASIC CONCEPTS OF VIROLOGY

DR SONAL SAXENA

D R A R P I TA S A X E N A
CHARACTERS OF
VIRUSES
Viruses do not have cellular organisation

They range in size from 20–300 nm

They contain only one type of nucleic acid

They are obligate intracellular parasites

They lack the enzymes necessary for protein and nucleic acid
synthesis
They multiply by a complex process and not by binary fission

They are unaffected by antibacterial antibiotics

 MORPHOLOGY
OF VIRUSES
Table 5.1 Important differentiating characteristics of virus and SIZE
bacteria
The extracellular infectious
virus particle: Virion
Range from 300 nm
(Mycoplasma) to 20 nm
(Parvovirus)
Fig. 5.1 Relative
sizes of
representative
viruses,
bacteriophages
(bacterial viruses)
and bacteria,
including
chlamydia
 STRUCTURE
Capsid
protein coat surrounding nucleic acid
capsid and the enclosed nucleic acid
together: Nucleocapsid morphological
Unit of the capsid: Capsomer.
Three kinds of symmetry:
i) Icosahedral (cubical)
ii) Helical
iii) Complex

Fig. 5.2 Illustration of the structure of a

viral particle
ENVELOPE
• Outer covering derived from the host cell membrane when the
progeny virus is released by budding, made up of lipoproteins
• Protein subunits seen as projecting spikes on the surface of the
envelope--peplomers
• Envelopes possess chemical, antigenic, and biological properties
• Enveloped viruses are susceptible to the action of lipid solvents
like ether, chloroform, and bile salts
• Specific neutralisation of virus infectivity depends on antibodies
to the surface antigens
• Biological properties such as attachment to the host cell surface
or hemagglutination depend on the envelope
SHAPE
Roughly spherical
Some are irregular and pleomorphic
Rabies-bullet-shaped
Hepatitis virus-Filamentous
Poxvirus-Brickshaped
TMV-rod-shaped
VIRAL COMPONENTS
Nucleic acid: Either single or double-stranded DNA or RNA
Positive-sense: Viral RNA may itself act as mRNA
Negative-sense: RNA is ‘antisense’, and the complementary strand
synthesised by a viral RNA transcriptase serves as a mRNA
Proteins-capsid, non-structural
Lipids
Other components:

i) Neuraminidase in the influenza virus

ii) RNA-dependent DNA polymerase or ‘transcriptase’ in retroviruses
 Temperature: Very heat-labile; for long-term
storage, they are frozen at –70°C
pH: Viruses vary greatly in their resistance to
acidity
SUSCEPTIBILITY OF Radiations: Viruses are inactivated by sunlight,
VIRUS TO CHEMICAL UV rays and ionising radiation
AND PHYSICAL
AGENTS Disinfectants: In general, viruses are more
resistant than bacteria to chemical disinfectants
Antibiotics: Absolutely ineffective
SUSCEPTIBILITY OF VIRUS TO CHEMICAL AND
PHYSICAL AGENTS
Lipid solvents: Enveloped viruses being sensitive and the naked viruses resistant to them

Disinfectants active against viruses

Weakly virucidal—phenolic disinfectants
Actively virucidal—oxidising agents such as hydrogen peroxide, potassium permanganate and
hypochlorites, organic iodine, formaldehyde, and beta-propiolactone
 REPLICATION OF
VIRUSES
Six sequential phases
Early events in replication: Attachment and
Adsorption → Penetration → Uncoating
→Replication → Assesmbly &
maturation→Release

Fig. 5.3 Stages in the infection of a host cell and replication of a

virus
REPLICATION OF VIRUSES
Attachment and Adsorption
Host cell surface should bear specific receptor sites to which the virus can gain attachment: cell
specificity
Differences in susceptibility to viral infection

Influenza virus: glycoprotein receptor sites on the surface of the respiratory epithelium

HIV: the CD4 receptor on host cells

Polioviruses—the lipoprotein present on the surface of primate cells
 2. Penetration
Virus particles may be engulfed: Pinocytosis
Viral envelope may fuse with the plasma membrane of the
host cell and release the nucleocapsid into the cytoplasm
REPLICATION 3. Uncoating
OF VIRUSES Stripping the virus of its outer layers and capsid

Nucleic acid is released into the cell

The next phase of viral replication is gene expression and

genome replication
 REPLICATION
OF VIRUSES
4. (a) Synthesis of DNA
Viruses
synthesis of viral nucleic acid
and capsid protein, enzymes
DNA viruses synthesise their
nucleic acid in the host cell
nucleus
- exceptions to this are the
poxviruses,

Fig. 5.4 Replication of a DNA

(herpes) virus
 REPLICATION
OF VIRUSES
4. (b) Synthesis of RNA viruses

 They synthesise all their components in the

cytoplasm
 Exceptions are orthomyxoviruses, some
paramyxoviruses and retroviruses

Fig. 5.5 Replication of RNA viruses

STEPS IN THE REPLICATION OF VIRUSES
Types of mRNA replication in viruses
 von Magnus phenomenon: some daughter virions
produced are not infective due to defective assembly.
Defective virus: Viruses that are genetically deficient;
incapable of producing infectious virions without the helper
ABNORMAL activity of another virus.

REPLICATION Abortive infection: Infection by a virus does not lead to the

production of infectious progeny.
CYCLES
Co-infection: Some viruses are genetically defective in that
when they infect cells, they are unable to produce fully
formed progeny. hepatitis D virus and HBV
VIRAL GENETICS
Two main mechanisms for genetic modification in viruses
 Mutation

 Recombination

 Variations due to gene product interactions

MUTATION

Spontaneously or may be induced by mutagens, physical

agents such as irradiation or chemical agents such as 5-
fluorouracil.
MUTATION
Types of mutants
Conditional lethal mutant :
Can grow under certain conditions (called permissive
conditions)
But not in certain other specified conditions, which are
lethal (called non-permissive or restrictive conditions)
Ts or temperature-sensitive mutant
Host-dependent mutants
 GENETIC INTERACTIONS
i) Recombination
Two different but related viruses infect a cell simultaneously

They exchange segments of their nucleic acids so that a hybrid

results

ii) Reassortment
Two different strains of the same virus are grown together,
recombinants or reassortant viruses may be produced,

iii) Reactivation
 PATHOGENESIS OF
VIRAL INFECTIONS

Table 5.2 Common viruses causing human infections

 PATHOGENESIS OF
VIRAL INFECTIONS
ROUTES OF ENTRY
• Inhalation (droplet infection—influenza, SARS-
CoV)
• Ingestion (hepatitis A, rotavirus)
• Direct transfer from in infected host
- Sexual contact (HIV, hepatitis C)
- Transplacental (rubella, HIV)

Fig. 4.7 Organ-specific infections by some common viral agents

 Direct inoculation (bite of dogs—rabies, through sexual
contact—HIV)
ROUTES OF Bites of vectors (arbovirus)
ENTRY Blood transfusion (hepatitis B and C, HIV)
 Virus settles down in the target cells to replicate, causing cell
death and tissue damage

Host Interactions are influenced by various defense mechanisms of

the body and can result in:
Virus Cell death: Cytocidal effect or lysis cytolysis, as seen with
poliovirus
Interactio Cellular proliferation: As seen with molluscum contagiosum
Malignant transformation: As seen with oncogenic viruses
n Steady-state infection: The virus and host cells co-exist
peacefully, both replicating independently; in such cases,
there
OUTCOMES OF VIRAL INFECTIONS
Types of infections
1. Full-blown acute infection
2. Persistent (chronic) infection
3. Latent infection
Virus–cell interactions
1) cytocidal effect or lysis (cytolysis)
2) cellular proliferation
3) steady-state infection.

Fig. 4.8 Outcomes of viral infection

ACTION OF VIRUSES ON
CELLS
Shutdown of host protein and DNA synthesis

Distort the cellular architecture and leads to autolysis

Polykaryocytosis or syncytium formation
Appearance of virus-coded antigens on the surface of the
infected cells that may confer new properties on the cells
Damage to the chromosomes of host cells
Formation of inclusion bodies in infected cells
INCLUSION BODIES
 Histological feature of virus-infected cells
 Have distinct size, shape, location and
staining properties
 Demonstrated in virus-infected cells with
the help of light microscopy.
 Crystalline aggregates of virions or made up
of virus antigens present at the site of virus
synthesis
 Confer altered staining properties on the
cell.
Fig. 5.8 Negri bodies (intracytoplasmic
 Situated in the cytoplasm(poxviruses), inclusion bodies) in a case of rabies virus
nucleus (herpesviruses) or both (measles infection seen in the pyramidal cells of Ammon’s
horn (Source: CDC, PHIL, Image ID 3372)
virus)
INCLUSION BODIES
TA B L E 5 . 4 S O M E V I RA L I N C LU S I O N B O D I E S , T H E I R S TA I N I N G C H A RA C T E R I S T I C S A N D
SITES IN THE CELL
 Apparent absence of an extracellular dormant phase
(virion)
Very small genome

VIROID Protein-free, low-molecular-weight RNA, which is

resistant to heat and organic solvents but sensitive to
nucleases
Cause diseases in plants
Disease-causing potential in humans is doubtful
 Discovered by Stanley B. Prusiner in 1982
Small proteinaceous infectious particles that have a MW
50,000 and diameter of approximately 4–6 nm, lack any
detectable nucleic acid
Resistant to heat (90°C for three minutes), UV rays, and
PRIONS nucleases but are sensitive to proteases, hypochlorites, and high
temperature autoclaving
May be associated with some chronic neurological
degenerative diseases of humans, resulting in fatal dementia
Also cause slow degenerative disorders such as scrapie, Kuru,
and Creutzfeldt–Jakob disease
Table 5.5 : A brief outline of laboratory diagnosis of viral
infections
LABORATORY DIAGNOSIS OF VIRAL DISEASES

Table 5.5 Types of

specimens to be sent
to the laboratory for
the diagnosis of viral
infections
LABORATORY DIAGNOSIS OF VIRAL DISEASES
Microscopy
1. Inclusion bodies can be demonstrated by light microscopy

2. Electron microscopy

Immune-electron microscopy, fluorescent antibody technique

Immunological Tests

1. Demonstration of Virus Antigens

- Immunofluorescence

- Enzyme immune assays

LABORATORY DIAGNOSIS OF VIRAL DISEASES
2. Antibody Detection (Serological Tests)
Enzyme-linked immunosorbent assay (ELISA) and other
immuoassays

Neutralisation test (to demonstrate protective

neutralising antibodies)

Complement fixation test

Hemagglutination inhibition tests

Fig. 4.13 Viral
hemagglutination
LABORATORY DIAGNOSIS OF VIRAL DISEASES
Molecular Diagnosis
- In situ hybridisation using nucleic acid probes
- Viral nucleic acid amplification techniques like PCR, reverse transcriptase PCR
(RT-PCR) and real-time PCR

- Quantitative RT-PCR (qRT-PCR)

- Sequencing of viral genes: to detect mutations or to genetically engineer
viruses and other microbial agents.
 CULTIVATION
1. Animal inoculation OF VIRUSES
Infant (suckling) mice
guinea pigs, rabbits and ferrets
growth of the virus indicated by Death, Disease, Visible
lesions

2. Embryonated egg inoculation

Embryonated hen’s egg
Inoculation on the chorioallantoic membrane (CAM)
produces visible lesions (pocks).
Fig. 5.10 10-day-old embryonated
hen’s egg
 EMBRYONATED EGG INOCULATION
Inoculation into the allantoic cavity provides a rich yield of influenza and some paramyxoviruses.

Inoculation into the amniotic sac is employed for the primary isolation of the influenza virus.

Yolk sac inoculation is used for the cultivation of some viruses such as chlamydiae and rickettsiae

Allantoic inoculation is employed for growing the influenza virus for vaccine production
 CULTIVATION
3. Tissue culture OF VIRUSES
Types of tissue cultures
1. Organ culture
2. Explant culture: Fragments of minced tissue
3. Cell culture
i) Primary cell cultures: These are normal cells freshly
taken from the body and cultured
Fig. 5.11 Vero cell
ii) Diploid cell strains: developed from human fibroblasts
monolayer

iii) Continuous cell lines: cell lines derived from human

cancers
Features of types of cell cultures
 DETECTION OF VIRUS GROWTH IN CELL CULTURES
1. Cytopathic effect: morphological changes in the cultured cells in
which they grow, which can be readily observed microscopically

2. Metabolic inhibition

3. Hemadsorption

4. Interference

5. Transformation

6. Immunofluorescence

Fig. 5.12 Vervet monkey kidney

cells (Vero cell line) infected with
measles virus stained with crystal
and viewed under × 100
magnification (note syncytium
 BACETRIOPHAGE
virus that infects bacteria.
Used in treating antibiotic resistant organisms

functions of phages
- genetic material transfer through transduction.
- phage conversion
- cloning vectors in genetic manipulations.

- Phage therapy
BACETRIOPHAGE
Morphology

- Head of a phage: tightly packed core of nucleic

acid (double-stranded DNA) surrounded by a
protein coat or capsid

- Tail is composed of a hollow core, a contractile

sheath surrounding the core and a terminal base
plate which has prongs, tail fibres or both attached
to it

Fig. 5.15 (a) Morphology of

bacteriophage and (b) electron
micrograph of a bacteriophage
BACETRIOPHAGE
Life Cycle

- Phages exhibit two types of life cycles.

1. Lytic or virulent cycle:

intracellular multiplication of the phage culminates in the lysis of the host bacterium and the release of
progeny virions.

2. Lysogenic or the temperate cycle:

phage DNA becomes integrated with the bacterial genome, replicating synchronously with it, causing no
harm to the host cell

The integrated phage nucleic acid is known as the prophage

BACETRIOPHAGE

Fig. 5.16 Lytic and lysogenic cycles of

bacteriophages
 BACETRIOPHAGE
Transmission of Genetic Information

1. Restricted transduction:

- Only bacterial genes contiguous to the prophage are transmitted

2. Generalised transduction:

- Any bacterial gene may be transferred to the prophage.

Significance of Phages in Medical Microbiology and Clinical Practice

- Phage assay

- Phage typing
 BACETRIOPHAGE
Phages may lyse:
- Members of a bacterial genus (e.g., genus-specific
bacteriophage for Salmonella)

- Species (e.g., specific bacteriophage for B. anthracis)

- A biotype or subspecies (e.g., Mukherjee’s phage IV which

lyses all strains of classical V. cholerae but not V. cholerae
biotype El Tor

Fig. 5.17 Mukerjee’s phage IV

classical lysis of V. cholera culture
(Source: CDC, PHIL, Image ID
3915)
SIGNIFICANCE
Phage therapy
- to treat bacterial infections

- conventional : naturally occurring bacteriophages that infect and lyse bacteria at the site of
infection.

- bioengineered phages

- hope of using bacteriophages as an alternative or a supplement to antibiotic treatment

Phage assay
When phages are applied on the lawn culture of a susceptible bacterium, areas of clearing
appear after incubation. These zones of lysis are called plaques.
Phage typing
The specificity of the phage–bacterium interaction is applied to the identification and typing of
bacteria. Phages exhibit different degrees of host specificity.