VIRUSES OF BACTERIA,

PLANTS & ANIMALS

Dr. M. Ashraf
PS/Assc. Prof.
Environmental Microbiology,
Soil Science Division,
NIAB, PEAC,
Jhang Road, Faisalabad
mashrafmalik11@hotmail.com
 VIRUSES
Non cellular, infectious entities whose genomes are a
nucleic acid either DNA or RNA
Infectious agents
Obligate intracellular Parasites
20 - 30 nm in size
Passed through the bacterial filters
10-100 times smaller than bacteria
Can be seen only with Electron microscope
Grow only on plant, animal and microbial cells
Reproduce only in living cells
Lack machinery to generate energy or synthesize proteins
Depend on host cells

Genetic material: DNA or RNA

Does not have both unlike host
Possess genetic information for replication, capturing host
cells energy generating and protein synthesis system
Dr. M. Ashraf, PS, NIAB 2
 VIRUSES
Use the host cell biosynthetic machinery to direct synthesis of
specialized particles called VIRION
The particles contain the viral genome and transfer them
efficiently to other cells

Virion
Structurally complete, mature and infectious virus
Cause diseases during multiplication in host cell
Human diseases like Cold, Flu etc.
Also cause Chronic diseases
Insensitive to broad range of antibiotics including Penicillin,
Streptomycin, etc.
Nucleic acid enclosed in highly specialized protein coat of
various designs

Protein coat
Protect viral DNA outside host cell
Serve as vehicle for entry in the host
Dr. M. Ashraf, PS, NIAB 3
 VIRUSES

With thanks from Pearson Education, Inc.

Dr. M. Ashraf, PS, NIAB 4

 BACTERIOPHAGES
Bacterial Viruses, Phages
Discovered independently in 1915 & 1917 by
Frederick W Twort , England
Felix dHerelle , Pasteur Institute Paris
Twort observed Lysis (the dissolution & disappearance
of the bacterial colonies)
Lytic effect can be transmit from colony to colony
Even high dilutions passed through bacterial filters
retained the effect
The lytic agent -called virus

Dr. M. Ashraf, PS, NIAB 5

 BACTERIOPHAGES
Simplest biological entities of self replicating
Considerable importance in viral research
Model for:
Virology (study of viruses)
Molecular Biology (discipline which examines structure,
function and organization of macromolecules in biological
systems)
Help to understood Host-parasite interaction, plant & animal
infections with viral pathogens
Widely used in genetic Research
Phages exists for all bacteria can be isolated easily in Lab
Host can be grown easily in laboratory
Required little time, labour & space compared to plant and
animal cells
Composed of n/a core surrounded by a protein coat occur in
different shapes mostly with tail for inoculation of n/a in the host

Dr. M. Ashraf, PS, NIAB 6

 BACTERIOPHAGES
Bacterial viruses: 2 types
1. Lytic or virulent
2. Temperate or Avirulent
Lytic
Infected cells produce large number of viruses
Lysed or burst the host cells The Lytic Cycle

Temperate
Infection not readily detected
viral n/a incorporated and replicate with host DNA/RNA
from one generation to another generation without cell
lysis
Become virulent at later stages and lysed the host cell
. Filamentous Viruses

simply leaked out of the cells

does not burst the cell

Dr. M. Ashraf, PS, NIAB 7

BACTERIOPHAGES

Rod shaped

Dr. M. Ashraf, PS, NIAB 8

 BACTERIOPHAGES
Structure
n/a core covered by protein coat capsid
Capsid
Protein coat made up of sub-units Capsomeres with no of protein
sub units the Protomers

Six Morphological Types

A. Hexagonal head, rigid tail, contractile sheath, tail fibers
B. Hexagonal head without contractile sheath, flexible tail with or
without tail fibers
C. Hexagonal head with shorter tail, without contractile sheath, flexible
tail with or without tail fibers
D. Hexagonal head with large capsomeres, without tail
E. Hexagonal head with small capsomeres without tail
F. Filamentous
G. Pleomorphic with variable morphology

Dr. M. Ashraf, PS, NIAB 9

 VIRUS MORPHOLOGY
E
D G

A B x174
B F2, MS2
C ssDNA ssRNA MV-L2

T7, (T3) ssDNA

T2 , (T4,
T6)

F

T1, T5 fd type (fd1)

ssDNA
dsDNA

Dr. M. Ashraf, PS, NIAB 10

 BACTERIOPHAGES
Polyhedron found in Icosahedron
a regular polyhedron with 20 triangular Facets
12 Vertices (corners) e.g., T2, T4
Icosahedron
Three dimensional 20 sided polygon with 12 evenly
spaced corners
Most icosahedron Capsid are built up of two types of
Capsomeres
1. Triangular hexagon that forms flat faces
2. Round Pentons that forms the corners

Dr. M. Ashraf, PS, NIAB 11

 CAPSID
Vertex

Capsomere

Hexon Capsomere

Protein sub-
units Penton
Capsomere

Nucleic acid 12
Dr. M. Ashraf, PS, NIAB
 BACTERIOPHAGES
A - C: Bacterio-phages
D - E: Plant, animal, insect
F: Plant viruses
G: Pleomorphic discovered recently
with a lipid containing envelope
no detectable capsid
with double stranded DNA (ds-DNA) - Phage MV-L2
Cubical or helical symmetry -Most phages
Cubical -Polyhedron
Helical Rod shaped
Capsomeres of rod shaped arranged in helical symmetry
e.g., M13
Complex structure with head, tail + Binal Symmetry
Virion: Icosahedron
Head: Hollow helical tail
e.g., T-even phages

Dr. M. Ashraf, PS, NIAB 13

 BACTERIOPHAGES
Coliphages
Infect non-motile strain of E. coli
Designated as T1 to T7
All with DNA and protein in equivalent amounts
T1, T2, T4, T5, T6
Tadepole shape with polyhedral heads and long tails
T3 & T7
very short tails
Size 65-200 x 50-80 nm L x W
DNA continuous or circular
Double stranded (ds)
50 um long, 1000 times as high as phage
Tightly packed in protein head

Dr. M. Ashraf, PS, NIAB 14

 BACTERIOPHAGES
Coliphages
f2 phages
Smaller than T phages, single stranded linear RNA, (ssRNA)
without tail
x174
icosahedral with ssDNA
Fd & f1
filamentous, circular ssDNA
M13,
ssDNA circular
Mu
dsDNA
R17,
ssRNA
B
ssRNA

Dr. M. Ashraf, PS, NIAB 15

 BACTERIOPHAGES
Coliphages
Nucleic Acids
All tailed phages with double stranded DNA (dsDNA)
With large capsomeres (type D) and filamentous (group F)
With single stranded DNA (ssDNA)
with small capsomeres and short tail (E) single stranded RNA
(ssRNA) DNA either circular closed loop loosely folded coil
packed inside
Capsid
x174 circular both in virion & host cell
Phage (Lambda), linear virion in host cell circular by cohesive
ends

Replication:
Adsorption, Penetration, Transcription, Assembly and Release
(Replication, Lysogeny, Prophage)
Lysogeny, Lysis, Temperate and virulent, Plaques

Dr. M. Ashraf, PS, NIAB 16

 REPLICATION
LYSOGENY
Host chromosome carrying bacterio-phage DNA is lysogeny
Incorporation of virus genome into host chromosome
Multiplication with host DNA replication
Inactive viral DNA -Prophage

Steps involved in Lysogeny

Adsorption
Penetration
Replication
Maturation
Release

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 REPLICATION
1. Adsorption
Recognition process b/w a virus & host cell
Cell membrane OmpC Protein and Lipopolysaccharides
Attachment of virus to external surface of the cell
Long tail fiber absorption to cell membrane surface (I)
Tail Pin adsorption to cell surface (II)

2. Penetration
Entrance of Virion (either a whole virus or its n/a) into host cell
Contraction of tail sheath and insertion of needle (III)
Cell Peptidoglycan layer
Approach of cytoplasmic membrane to outer membrane (IV)
Membrane potential
Interaction of cor tip with cytoplasmic membrane (V)
Phosphotidylglycerol or Cordipin
Viral DNA injection into to cytoplasm (VI)
Heating

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 REPLICATION
3. Replication
Copying and expression of viral genome at expanses of the host
synthetic equipment
Production of various viral components

4. Maturation
Assembly of virus parts into whole intact virion

5. Release
Escape from the host cell of active infectious viral particles by
Bursting or Lysis of the cell (Lytic cycle)
Leak out of the intact host cells (filamentous virion)

Dr. M. Ashraf, PS, NIAB 19

REPLICATION

Dr. M. Ashraf, PS, NIAB 20

REPLICATION

Dr. M. Ashraf, PS, NIAB 21

 REPLICATION
Viral proteins & nucleic acids multiplication in the host
1. Attachment of virion with host at specific site (Adhesion)
2. Whole virus or n/a penetrate the host cell (Penetration)
3. Uncoating (if whole virion penetrate) of the virus to release n/a
(uncoating)
4. Reproduction in cytoplasm, nucleus or both (Transcription and
Replication)
5. Assembling of viral proteins & n/a into viral particles (Assembling)
6. Release of virus from cell (Release)

Adsorption Penetration Uncoating Component replication & Biosynthesis

Assembling Release

Dr. M. Ashraf, PS, NIAB 22

 REPLICATION
Adsorption/Attachment
1. Ionic bonds or charges involved -Reversed by pH or salts
2. Firm attachment irreversible
Receptors on host cell bound with capsid or envelope proteins
Plant viruses -no receptors required
Penetration
1. Animals -Two mechanisms
a. Viropoxis: Engulfment of virus by phagocytes
Uncoating or removal of capsid /envelope in
phagocyte vaccuoles by enzyme lysozyme
protease
b. Enveloped virus lipoproteins fuses with host cell surface
membrane
Release of nucleo-capsid in cytoplasm of the host
cell , uncoating in the host cell
2. Plants
a. Virus penetrates through transient pores (Ectodesmata)
protrude out of the cell wall at intervals
-Link interior of the cell with outside
-for water, nutrient uptake or release of waxes
b. Whole virion engulfed
i. Insect inoculate plant virus into cell
Insect feeding the most important plant virus
transfer mechanism 23
Dr. M. Ashraf, PS, NIAB
 REPLICATION
Reproduction/multiplication/replication
Latent Period: Time from infection to lysis (bursting) of the cell
Intracellular accumulation of mature viruses
uncoating of the virion
n/a get free from capsid
accessible to enzymes for translation, transcription, replication
Transcription of viral RNA into mRNA
Positive (+) strand viruses If viral RNA act as mRNA
Negative (-) strand viruses -if host mRNA is used

Eclipse Period: No phage can be recovered by disruption of bacterial

cell after 10 min of infection -Called Eclipse Period

Rise Period: Increase in phages to a constant number after latent period

1. Increase in extracellular titre (number assayed) -constt.
2. No more increase in extracellular accumulation of phages

Burst Size: Phage yield per bacterium

Nos. of phages required to burst the cell

Spontaneous Induction:
Removal of prophage from host DNA & entering into Lytic Cycle
24
during Lysogeny
Dr. M. Ashraf, PS, NIAB
 REPLICATION
Reproduction/multiplication/replication
Biosynthesis of viral components
1. Early or Immediate early phage genes
2. Delayed early phage genes
3. Late phage genes

1. Early
a. Process in cytoplasm or nucleus
-dsDNA viruses use host RNA polymerase
-RNA viruses use viral-coded RNA polymerase
b. Part of n/a transcribed into mRNA
c. Codes for early enzymes involved in n/a replication
d. Proteins to stop cellular macromolecules
e. Breakdown of polyribosomes -make for viral transcription
f. n/a replication follows early proteins synthesis
-demarcation line b/w early and late viral replication
Since late mRNA is not synthesized until viral DNA
replication

25
Dr. M. Ashraf, PS, NIAB
 REPLICATION
1. Early
a. Immediate early
a. Codes for nuclease to breakdown host DNA
b. Synthesize the enzymes to alter bacterial RNA polymerase
b. Delayed early
a. Codes for phage enzymes to produce
i. 5-hydroxy methylcytosine instead of bacterial DNA cytosine
ii. Glycosylate nucleotides
iii. Destroy precursors of cytosine deoxynucleotides
Bacterial restriction enzymes are unable to degrade
phage DNA with glycosylated methylcytosine
b. Synthesis of polymerase & ligase
2. Late genes
a. Late protein synthesis -after n/a replication
b. Late mRNA for structural proteins
For virion promotor formation
c. Translation in cytoplasm
Use tRNA & enzymes in cytoplasm
d. Codes for
a. structural components of virus (e.g., Head, tail, fibers, etc.)
b. Phage lysozymes (endolysin) for lysis of host cell & release of virion
e. Rise period after latent period Nos. of phages rises to constant number
26
Dr. M. Ashraf, PS, NIAB
 REPLICATION
Assembly & Release
a. Synthesis of structural proteins and the n/a
b. Phage components assemble into Mature phages
i. Process occurs in cytoplasm or in the nucleus (animals and
plants)
ii. DNA viruses with exception of Poxvirus assembled in the
nucleus
iii. RNA viruses assembled in the cytoplasm
c. Released of the viruses from the host through:
1. Bursting/Lysis of the host cell (Bacteria, Animals)
2. Extruded by reverse phagocytosis (Animals)
3. Enveloped viruses released through budding from special areas
(Animals, Plants)
i. Host - not destroyed
ii. Virions -acquire portion of host membrane
iii. Tubules or channels -established in the host cell
iv. Virus yield several
-Thousand to a million/cell (animal, plants) , few
hundred to thousand (bacteria)

27
Dr. M. Ashraf, PS, NIAB
Mechanism of Lysogeny -Studied in temperate phage
i. Lytic cycle with immediate multiplication
ii. Lysogeny -multiplication delayed or repressed
Switching of genes required for viral multiplication & host cell
Lysis
Phage genes -codes for repressor protein
Make cell resistant to Lysis (induced by Prophage or
Chemicals or radiation use to release Phages)
Repressor proteins or Immunity Repressor
iii.An acidic protein of 26000 MW
iv. React at two operator sites of genome to prevent expression of genes
required for
a. Lytic cycle
b. Formation of mature phage particles
The regulatory region (Immunity Operon) in phage governed Lysogeny by:
i. Immunity to external phage infection
ii. Integration of phage genome into host DNA
Phage infection (spontaneous induction)
i.transcription of Cro genes
Produce repressor protein for inhibition of Immunity Operon
repressor proteins
ii. Immunity repressor and the Cro repressor
Antagonistic production and maintenance of Lysogenic state
Phage Lytic cycle -induced by UV light
iii. UV at molecular level -induce synthesis of protein encoded by rec A
genes of E. Coli (the Phage Host)
iv. Protein with proteolytic activity
v. cleaves the Immunity repressor prevents its biding 28
with prophage
Dr. M. Ashraf, PS, NIAB
 PLANT & ANIMAL VIRUSES
Composed of central core of n/a surrounded by Capsid made up of
Capsomeres
Characteristic Icosahedron in Spherical, Helical in rod shaped viruses
Complex in miscellaneous groups symmetry basic criterion for
classification
Animal Viruses Nucleocapsid (n/a + Capsid) covered by
outer membrane the Envelope - made up of lipoproteins
Virions with envelop are sensitive to lipid solvents e.g., Choloropharm,
Ether, etc. the capacity to infect is inactivated by solvents
Naked virions not effected by solvents
Icosahedron Viruses Poliovirus, Adenovirus, Poliomyelitis and respiratory
disease viruses
Helical symmetry TMV (Plant), Measles, Mumps, Influenza and Rabies
(Animals)

Dr. M. Ashraf, PS, NIAB 29

 PLANT: VIRUS CULTIVATION
1. Virus inoculation -through rubbing of leaves
Carborundum abrassive used to enhance
penetration
Hair cell of Tobacco carries 107 TMV constitute 10%
of DW of leaves
2. Transfer of virus in leaves
-through Plasmodesmata (intercellular bridges)
3. Plant Cell Protoplasts
e.g., mespphyll cells of tobacco
4. Monolayer culture of cells
From insects carriers of viruses
e.g., rhabdovirus cultivated in Leafhoffer cell
culture produce 10,000 virions/cell

Dr. M. Ashraf, PS, NIAB 30

THANX

Dr. M. Ashraf, PS, NIAB 31