General Properties of Viruses including Morphology, Multiplication, Cultivation, Classification and Nomenclature

1. Kalpesh Zunjarrao

2.  Viruses do not fall under any category of unicellular
organisms because:
o Do not possess cellular organisation
o Contain only 1 type of N. A. (DNA / RNA)
o Obligate intracellular parasites
Lack enz necessary for protein & n. a. synthesis
For replication depend on synthetic machinery of host cell
o Multiply by complex method

3. MORPHOLOGY

4. Size:
 Extracellular infectious viral particle is called ‘Virion’
 Viruses are much smaller than bacteria
For a time, they were known as ‘filterable agents’
 Can not be seen under light microscope
 Size range: 20-300 nm
 Parvovirus: 20 nm
 Pox virus: 300 nm (can be seen under light microscope)

5. Estimation of Size:
 Earliest method:
o Passing through membrane filter of graded pore size
o Average pore size of finest filter that allows passage of
virion gave an estimate of size
 Next method:
o Ultracentrifuge: depending on rate of sedimentation,
particle size was calculated
 Latest & direct method:
o Electron microscope

7. Structure:
 Virion consists of nucleic acid core surrounded by protein
coat called ‘capsid’
 Capsid: made up of subunits called ‘Capsomers’
 Genome + capsid: nucleocapsid
 Functions of capsid:
o Protection of n.a. core from inactivation by nucleases
o Introduction of viral genome into host by adsorbing on the
host cell surface
o Antigenic in nature

8. Types of symmetries:
1. Icosahedral:
 Icosahedron: polygon with 12 vertices or
corners & 20 facetes or sides
 Each facete has shape of equilateral triangle
 It’s a rigid structure
2. Helical:
 N.A. & capsomers are wound together to form
helical tube
 Tube can be rigid or pliable
 Some viruses show complex symmetry

9.  Viruses can be enveloped or non-enveloped
 Envelope is lipoprotein in nature
o Lipid derived from host cell
o Protein: virus coded
o Protein subunits are seen as projecting spikes on surface of
envelope: called ‘Peplomers’

10.  Overall shape of virus varies with different groups of
viruses:
o Most animal viruses: spherical, some: irregular
o Rabies virus: bullet shaped
o Ebola virus: filamentous
o Pox virus: brick shaped
o TMV: rod shaped
o Bacteriophage: complex morphology

11. Chemical properties:
 Nucleic acids:
o Viruses contain only 1 type of n. a.
o Single or double stranded RNA or DNA
o N. A. can be extracted by treatment with certain chemicals
 Proteins:
o Capsid & envelope
o Protects n.a. & determines antigenic properties
 Some viruses contain small amount of carbohydrates
 Most viruses don’t possess enzymes but some of them
may possess (neuraminidase, reverse transcriptase)

12. Viral hemmaglutination:
 Large number of viruses agglutinate erythrocytes of many
species
 Hemmaglutination by influenza virus is due to presence of
protein spikes ‘Hemagglutinin’
 Hemagglutinin has ability to bind glycoprotein receptor
sites on erythrocytes
 Convenient method of detection of viruses
 Procedure:
RBCs are added to serial dilutions of viral suspension →
highest dilution producing hemmaglutination is ‘titre’

13.  Non agglutinated RBCs settle down at bottom in the form
of ‘button’
 Agglutinated RBCs spread into shield like pattern
ButtonTitre

14.  Hemagglutination is inhibited by Antibodies to virus.
This principle can be used in ‘Hemagglutination inhibition
test’.
This test is used for detecting antiviral antibodies
 Some viruses carry surface enzymes (neuraminidase)
which act on receptors on erythrocytes
- They are called ‘Receptor destroying enzymes’ (RDE)
- Destruction of receptor leads to reversal of
hemagglutination. Called as ‘Elution’

15. MULTIPLICATION

16.  Genetic information required for viral replication is
present in viral NA but they lack enzymes
 Viruses depend on synthetic machinery of host cell
 Viral multiplication cycle is divided into 6 sequential
phases:
o Adsorption
o Penetration
o Uncoating
o Biosynthesis
o Maturation
o Release

17. Adsorption (Attachment):
 Contact between virion & host cell: by random collision
 Adsorption takes place only if there is affinity between
them
 Cell surface contains some receptors to which viruses can
attach
 In case of influenza viruses: hemagglutinin on virus
surface attaches to glycoprotein receptors sites on
respiratory epithelium
 Destruction of receptors by RDE prevents viral adsorption
 In HIV virus: attachment between CD4 receptors on host
cell & viral surface glycoprotein ‘gp120’
 Susceptibility to viral infection depends on presence or
absence of receptors on cells

18. Penetration:
 Bacterial cells possess rigid cell wall. Thus, viruses can
not penetrate into the cell. Only nucleic acid is introduced
 Animal cells → no cell wall → whole virus can enter into
the cell
 Virus particle may be engulfed by process resembling
phagocytosis, called ‘Viropexis’
 In case of enveloped viruses: viral envelop fuse with
plasma membrane of host cell → nucleocapsid released
into the cytoplasm

19. Uncoating:
 Stripping the virus of its outer layer & capsid
 In most cases, uncoating is effected by action of lysosomal
enzymes
 In pox virus: Uncoating is 2 step process.
1st step in phagosome: outer coat removed by lysosomal
enz
2nd step in cytoplasm: viral uncoating enz removes protein
covering

20. Biosynthesis:
 Synthesis of viral nucleic acid, protein capsid & various
enzymes required for synthesis, assembly & release
 Certain ‘regulator proteins’ are also synthesized
 Regulator proteins: shut down normal cellular metabolism &
stimulates production of viral components
 Site of viral synthesis depends on type of virus
 Most DNA viruses: synthesize n. a. in host cell nucleus
(exception: poxvirus which synthesizes all components in
host cytoplasm)
 Most RNA viruses: synthesize all components in cytoplasm
(Exceptions: Orthomyxoviruses, some paramyxoviruses
synthesized partly in nucleus)
 Proteins: always synthesized in cytoplasm

21.  Biosynthesis consists of following steps:
1. Transcription of mRNA from viral nucleic acid
2. Translation of mRNA into ‘early proteins’
Early/non-structural proteins are enzymes which initiate
& maintain synthesis of virus components
They may also shut down production of host proteins
3. Replication of viral nucleic acid
4. Synthesis of ‘late / structural proteins’ required for viral
capsid
 Critical step in biosynthesis: transcription of mRNA from
viral nucleic acid
 Once this is achieved, host cell resources can be used for
translating mRNA into viral components

22. Maturation (Assembly):
 Assembly of daughter virions follows the synthesis of
viral nucleic acid & proteins
 Assembly may take place in cytoplasm or nucleus
 Herpes & Adenoviruses are assembled in nucleus
 Picorna & Poxviruses are assembled in cytoplasm
 At this stage, non-enveloped viruses are present
intracellularly as fully developed virions but in case of
enveloped viruses, only nucleocapsid is complete
 Envelops are derived from host cell membrane during
process of budding
 Host cell membrane that becomes envelope is modified by
addition of virus-specific antigens

23. Release:
 In case of bacteriophages, release takes place by lysis of
bacterium
 In animal viruses, release usually occurs without cell lysis
 Certain viruses are released by process of budding from the
cell membrane over period of time. Host cell is unaffected &
may even divide, daughter cells continuing to release virions
 Progeny virions released into surrounding medium may
infect other cells
 In case of some viruses, transmission occurs directly from
cell to cell, very little free viruses being demonstrable
extracellularly in the medium
 Poliovirus causes profound damage to host cell & may be
released by cell lysis

25.  From the stage of penetration till appearance of mature
daughter virions, virus can not be demonstrated inside host
cell
 This period during which virus seems to disappear or go
‘underground’ is called as ‘eclipse phase’
 Single life cycle of replication takes 15-30 mins in
bacteriophages & about 15-30 hours for animal viruses
 Single infected cell releases large number of progeny
viruses. This can be demonstrated in bacteriophages but
difficult in case of animal viruses which are released over
a prolonged period

26. CULTIVATION

27.  Viruses: Obligate intracellular parasites
 Can not be grown on inanimate culture medium
 3 methods employed for cultivation of viruses:
o Inoculation into animals
o Embryonated eggs
o Tissue culture

28. Animal Inoculation:
 Earliest method for cultivation: human volunteers → high
risk involved → used only when virus is relatively
harmless
 Monkeys were used for isolation of poliovirus → limited
application due to cost
 Use of white mice: most widely employed in virology
 Guinea pigs, rabbits: used in some situations
 Growth of virus in animal can be indicated by death,
disease or visible lesion
 Disadvantage: immunity may interfere with viral growth

29. Embryonated eggs:
 Embryonated hen’s egg was 1st used for cultivation by
Goodpasture (1931) & method was further developed by
Burnet
 Embryonated eggs offer several sites for cultivation of
viruses
 Inoculation on chorioallantoic membrane (CAM)
o produces visible lesions (pocks)
o Different viruses: different pock morphology
o Each infectious viral particle can form 1 pock. Thus, pock
counting can be used for assay of pock-forming viruses
like variola & vaccinia

30.  Inoculation into allantoic cavity:
o provides rich yield of influenza & paramyxoviruses
o Used for growing influenza virus for vaccine production
 Inoculation into amniotic sac: used for primary isolation of
influenza virus
 Yolk sac inoculation:
for cultivation of some
viruses, Chlamydiae &
Rickettsiae

31. Tissue culture:
 Tissue & organ culture: used for study of morphogenesis
& wound healing
 1st application of tissue culture in virology: for
maintaining vaccinia virus in fragments of rabbit cornea
 Major obstacle in using tissue culture: bacterial
contamination
 Antibiotics: prevention of contamination
 Every human virus can be grown in tissue culture

32. Types of tissue culture:
 Organ culture:
o Small bits of organs can be maintained in vitro preserving
their architecture & function
o Useful for isolation of viruses which appear to be
specialised parasites of certain organs
o Tracheal ring organ culture: for coronavirus isolation
 Explant culture:
o Fragments of tissues can be grown as explants embedded
in plasma clots or in suspension
o Originally known as tissue culture
o Adenoid tissue explant culture: for Adenovirus isolation

33.  Cell culture:
o Routinely employed for growing viruses
o Tissues dissociated into cells by proteolytic enzymes →
cells washed → counted → suspended in growth medium
o Cell culture medium components:
Amino acids, vitamins, salt, glucose, buffer (HCO3
-), fetal
calf serum, antibiotics & phenol red
o Cell suspension is dispensed in bottles/ petri plates →
incubated → cells adhere to glass surface → form
monolayer of cells
o Bottles incubated at stationary condition or in roller drums
for aeration

34.  Cell cultures are classified into 3 types:
1. Primary cell culture:
o Normal cells freshly taken from body
o Capable of only limited growth in culture
o Eg: monkey kidney, human embryonic kidney, Human
amnion
2. Diploid cell culture:
o Cells of single cell type that retain original diploid
chromosome number & karyotype
o Can be subcultured for limited number of times (due to
senescence)
o Eg: Human fibroblasts

35. 3. Continuous cell culture:
o Cells of single cell type derived from cancer cells
o capable of continuous serial cultivation
o Eg: HeLa, HEp-2, Vero cell lines

36. Detection of virus growth in cell cultures:
 Cytopathic effects (CPE):
o Morphological changes in cultured cells → can be
observed by microscopic examination
o Viruses causing CPE are called ‘cytopathogenic viruses’
o Help in presumptive identification of viruses
o Eg:
- Enteroviruses produce rapid CPE by crenation of cells
- Measles virus produces syncytium
- Adenovirus produces large granular clumps

37.  Metabolic inhibition:
o In normal cell cultures, medium becomes acidic due to
metabolism
o When viruses grow → metabolism inhibited → no acid
 Hemadsorption:
o Hemagglutinating viruses can be identified by addition of
guinea pig erythrocytes
o If viruses are multiplying, erythrocytes adsorb onto cell
surface

38.  Interference:
o Growth of non-cytopathogenic virus in cell culture can be
tested by subsequent challenge with known
cytopathogenic virus
o Growth of first inhibits infection by second virus
 Transformation:
o Tumor forming viruses induce cell transformation → loss
of contact inhibition → piled-up growth ‘Microtumors’
 Immunofluorescence:
o Cells from virus infected cultures → stained by
fluorescent conjugated antiserum → examined under UV
microscope

39. CLASSIFICATION
&
NOMENCLATURE

40.  Till 1950, little was known about viruses
 They were named haphazardly, based on the disease they
caused or site of isolation
 They were grouped according to tropism or affinity to
different organs
 Were classified as Dermotropic, Neurotropic,
pneumotropic & viscerotropic
 Bawden suggested that nomenclature & classification
should be based on properties of viruses & not the
responses

41.  Classification & nomenclature are now official
responsibility of International committee on Taxonomy of
Viruses (ICTV)
 Viruses are classified into 2 main divisions:
Riboviruses & Deoxyriboviruses
 Further classification is based on properties such as
strandedness of n.a., symmetry of nucleocapsid, presence
of envelop, size & shape of virion & number of capsomers

42. DNA viruses:
 Poxviridae family:
o Large, brick-shaped or ovoid (300 X 240 X 100 nm)
o Complex structure, having lipid containing outer coat & core
carrying single linear ds-DNA
o Multiplication & maturation: in cytoplasm
o Several genera
 Herpesviridae family:
o Medium sized containing linear ds-DNA
o Icosahedral nucleocapsid has 162 capsomers surrounded by
lipid containing envelope
o Multiplication: in nucleus
o Maturation: by budding through nuclear membrane
o Only 1 genus: Herpesvirus

43.  Adenoviridae family:
o Medium sized (70-90 nm) non-enveloped, icosahedral
viruses with 252 capsomers
o 2 genera: Mastadenovirus & Aviadenovirus
 Papovaviridae family:
o Small (40-55 nm), non-enveloped, ds-DNA viruses with
72 capsomers
o 2 genera: Papillomavirus & Polyomavirus

44.  Parvoviridae family:
o Very small (18-26 nm) non-enveloped, ss-DNA viruses
with 31 capsomers
o 3 genera: Parvovirus, Adenosatellovirus, Densovirus
 Hepadnaviridae family:
o Spherical (42 nm) virus with core surrounded by envelope
having specific antigens
o Human Hepatitis type B virus & related viruses of animals

45. RNA viruses:
 Picornaviridae family:
o Small (20-30 nm), non-enveloped, icosahedral, ss-RNA
viruses
o 3 genera: Enterovirus, Rhinovirus, Hepatovirus (HAV)
 Orthomyxoviridae family:
o Medium sized (80-120 nm), spherical or elongated,
enveloped viruses with hemagglutinin & neuraminidase
peplomers
o Genome consists of ss-RNA in several pieces
o 1 genus: Influenzavirus

46.  Paramyxoviridae family:
o Pleomorphic virions with lipid envelope having surface
projections
o Genome: un-segmented, linear ss-RNA
o 3 genera: Paramyxovirus, Morbillivirus, Pneumovirus
 Togaviridae family:
o Spherical viruses (40-70 nm) with lipoprotein envelope &
ss-RNA
o Multiply in arthropods & vertebrates
o 3 genera: Alphavirus (Group A arboviruses), Rubivirus,
Pestivirus

47.  Flaviviridae family:
o Formerly grouped as group B arboviruses under togaviridae
 Bunyaviridae family:
o Spherical, enveloped virions (90-100 nm)
o Arthropod-borne viruses
o 5 genera: Bunyavirus, Hantavirus, Nairovirus, Phlebovirus,
Ukuvirus
 Arenaviridae family:
o Spherical or pleomorphic viruses (50-300 nm) with number of
electron dense particles giving sandy appearance
o Rodent parasites but can infect humans rarely
o 1 genus: Arenavirus

48.  Rhabdoviridae family:
o Bullet shaped viruses (130-300 nm long & 70 nm wide) with
lipoprotein envelope carrying peplomers
o 2 genera: Vesiculovirus, Lyssavirus
 Reoviridae family:
o Icosahedral, non-enveloped viruses (60-80 nm) with double
layered capsid
o Genome: ds-RNA in 10-12 pieces
o 3 genera: Reovirus, Orbivirus, Rotavirus
 Coronaviridae family
o Pleomorphic, enveloped viruses (100 nm) with club-shaped
peplomers. Only 1 genus: Coronavirus

49.  Retroviridae family:
o Icosahedral viruses (100 nm) with lipoprotein envelope
o They have RNA dependent DNA polymerase (Reverse
transcriptase)
o 3 subfamilies: Oncovirinae, Splumivirinae, Lentivirinae
 Calciviridae family:
o Naked spherical particles (35-39 nm) with 32 cup shaped
depressions arranged in symmetry
 Filoviridae family:
o Long, filamentous, enveloped viruses (80 nm in diameter &
14,000 nm long) with helical nucleocapsid & ss-RNA