Foundations in Microbiology

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Viruses
Chapter 6
Talaro
1
Viruses of bacteria,
Archaea and eukaryotes
2
Size of Viruses
Smaller than bacteria
Pass through a filter
Once called “filterable agents”
3
Taxonomy of Viruses
• Originally based on disease
• Now based mainly on three criterion
• Nucleic acid type (DNA or RNA)
• Strategy for replication
• Morphology and size
• Naming and classification decided on by International Committee on
Taxonomy of Viruses (ICTV)
•Order, Family, Subfamily, Genus, Species – Subspecies, strain etc.
Naming: -virales Order; -viridae, Family; -virinae Subfamily; -virus
4
Genus, Species Epithet usually in English
Taxonomy of Viruses
(Continued)
• Ex. 1, Poliovirus
Genus & Species: Enterovirus human poliovirus 1
Family: Picornaviridae
Order: Not decided by ICTV
• Ex. 2, Rabies Virus
Genus & Species: Lyssavirus rabies virus
Family: Rhabdoviridae
Order: Mononegavirales
5
Species Concept in Virology
• What is a virus “species”?
After many years of controversy, in 1991, the ICTV accepted
the definition of a virus species proposed by van
Regenmortel (1990), as follows: "A virus species is defined
as a polythetic class of viruses that constitutes a
replicating lineage and occupies a particular ecological
niche." Members of a polythetic class are defined by more
than one property and no single property is essential or
necessary. One major advantage in this definition is that it
can accommodate the inherent variability of viruses and it
does not depend on the existence of a single unique
characteristic.
6
Isolation, Cultivation, and Identification
• Much more problematic than cultivation of organisms
• Requires host cell for replication
• Often times propagation of host cell is challenging
• Mammalian tissue culture expensive and time
consuming
7
Viral Identification
• Usually too small to see without electron microscopy
• Nanometer range
• Antibodies detect viral coat proteins
•Western blot / ELISA technique to identify
(ELISA- Enzyme-Linked ImmunoSorbent Assay)
•May also use tissue culture and observe characteristic
cytopathic effects
• Newer methods use PCR to amplify virus specific DNA or
RNA sequences
8
Composition of Viruses
-Viral genomes
i. double-stranded DNA
ii. single-stranded DNA
iii. double-stranded RNA
iv. single-stranded RNA
- Nucleic acid can be organized as
i. circular molecule
ii. one or more linear molecules
- Viruses range from simple (4 genes) to
complex (100s of genes)
9
dsDNA
(+)
sense, template or transcribed strand
(-)
antisense or non-template stand
(-) & (+) ssDNA
dsRNA
(+) ssRNA (for mRNA)
(-) ssRNA
(+) ssRNA (for DNA)
10
Capsids
- Protein shell enclosing
the genome
- Built of large number
of proteins called
capsomeres
- Capsid can be composed
of 100s of same or
different capsomeres
11
Helical Shaped Viruses
12
Icosahedral Shaped Viruses
• 20-sided with 12 corners
• Vary in the number of capsomers
• One edge is called a facet
13
Viral Envelopes
- Not found in all viruses
- Surrounds capsid
- Derived from host cell (budding)
- May have viral proteins and
glycoproteins embedded
- Assists the virus in infecting
its host
- Example is influenza
14
Viral Life Cycles
Host Ranges
- Viruses can only reproduce within a host cell
- Viruses usually infect hosts by binding
to receptors on the cell surface
- Some viruses can infect many types of cells
or many types of species (rabies for example)
- Most viruses can only infect a few types of
cells = limited host range
- Eukaryotic viruses often attack specific tissues
Cold virus = cells of upper respiratory track
AIDS virus = T cells & macrophages of immune system
Hepatitis = Only liver cells
Polio = Intestinal & nerve cells
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Rabiesvirus = Nerve cells
Viral Mutation Rate
No proof-reading enzymes so
no repair
- Mutation rate  1 in 10,000 nucleotides
(depends upon the virus)
- Low-fidelity RNA polymerase (means not so accurate)
- May recombine with different strains of the same virus
- Reassortment of nucleic acid
- Occasionally (rarely), acquire traits from completely
different virus
- DNA & RNA viruses
16
Viral Replication Cycles
Bacteriophage l dsDNA
Influenza Virus (-) sense ssRNA
Human Immunodeficiency Virus
(+) sense ssRNA for DNA
6 Steps in Phage Replication
1. Adsorption – binding of virus to specific molecule
on host cell
2. Penetration – viral genome enters host cell
3. Replication – viral components produced
4. Assembly - viral components assembled
5. Maturation – completion of viral formation
6. Release – viruses leave cell to infect other cells
Lytic Cycle
Virulent
Lysogenic Cycle
Temperate
Bacteria eater
19
l
Fig 6.11
20
-Viral genome has a promoter
-Bacterial RNA polymerase transcribes viral genes
-EARLY STAGE PROTEINS
-Viral genes adjacent to the promoter are transcribed
- Proteins that stop host transcription
- Proteins that stimulate viral replication
- Viral nucleases digest host genome
- Stimulate late gene transcription
-LATE
STAGE PROTEINS
- Viral genes that code for viral capsid proteins
- Proteins that lyse the host cell
21
Virus infects host
Host’s RNA polymerase
transcribes EARLY genes
Lytic Stage
operon
Early proteins
stop transcription
of host’s genes
Early protein
stimulates viral genome
reproduction, digest host
genome & stimulate late
gene transcription
Viral capsid
proteins
Early protein
stimulates transcription
of LATE genes.
Lyse
host
22
Lysogenic Phage Cycle
- Phage able to reproduce without destroying host
- Temperate phages are able to use both the lytic
and lysogenic cycles
- When phage DNA enters cell, it uses site-specific
recombination to insert its DNA into the host’s
chromosome = prophage
- The phage DNA is replicated and passed to
daughter cells each time the bacteria divides
- Occasionally, the prophage exits the bacterial
DNA and engages in the lytic cycle
- Exit from the lysogenic stage is often triggered
23
by environmental cues/stresses
24
Temperate phage
Bacterium growing well  lysogenic cycle
Bacterial host stressed or damaged  lytic cycle
Regulatory proteins
cI
Cro
Compete for promoter sites
Constant Growth
STRESS or UV Damage
Cro 
cI 
Lysogenic
cI 
Cro 
Lytic
25
STRESS or DAMAGE
Cro protein activates promoters
for phage DNA replication &
cell lysis
GOOD GROWTH
cI accumulates & activates
promoters for integration
Lysogenic Cycle
26
27
Animal VirusAdsorption
Penetration
Duplication/Synthesis
Assembly
Release
28
Adsorption
- Glycoproteins on the envelope bind to
specific receptors on the host membrane
- Cell receptor is also a glycoprotein
- Envelopes fuse, viral capsid &
genome are transported inside cell
- The receptors are involved in normal
cell functions
- Rabies virus attaches to acetylcholine
receptor on nerve cells
-HIV attaches to the CD4 receptor on T
and some other cells of the immune system
29
Penetration
Endocytosis
Fusion with cell membrane
30
Release by Budding or Exocytosis
31
Cytopathic Effects
1.
2.
3.
4.
5.
6.
7.
Changes in size & shape
Cytoplasmic inclusion bodies
Nuclear inclusion bodies
Cells fuse to form multinucleated cells
Cell lysis
Alter DNA
Transform cells into cancerous cells (called
Transformation but do not get confused with
bacterial transformation)
•
Can see these effects microscopically in tissue
culture cells infected with virus
32
Influenza
(-) sense RNA virus
Spanish Flu Epidemic
20 million deaths
Negative Sense RNA Virus
(+) RNA = mRNA
- An RNA virus whose genome is complementary to
mRNA
- Contains an RNA-dependent RNA polymerases
required for the synthesis of mRNA
Influenza virus
- Genome consists of eight different negative sense
ss RNA strands
- Strands 1 through 6 each encode one protein
- Strands 7 & 8 encode two proteins
34
Lipoprotein envelope
Glycoprotein spikes
Hemagglutinin
Binds to a receptor found
on eukaryotic
respiratory mucosa cells
RNA + nucleoprotein
Matrix
Neuraminidase
Hydrolyzes the protective
mucous coating, assists in viral
budding, prevents virions from
sticking together & helps in penetration
Nonspecific
Polymerase
35
Viral RNA is
released
(-)
Viral glycoproteins bind to
host cell receptors
(-)
Virus enters by endocytosis
Viral RNA dependent
RNA polymerase makes (+) RNA
Viral RNA is translated
- Enveloped viruses do
Virion
not always kill host cell
assembled
- Virus can be continuously
shed from infected cell
(factory)
36
- Influenza viruses can infect humans, pigs, sea mammals
& birds
- Typically, one virus can only infect one type of host
- Occasionally, some viral strains can infect another
animal host
- Genome segments can be exchanged when two different
viral strains infect the same host
- Some of the exchanged RNA segments encode for
glycoproteins
- The virus can have glycoproteins from several hosts:
swine, human & bird
- The same virus will appear as a “new” virus to
the human immune system because of different
glycoproteins
37
Each year, new strains of influenza
emerge from southern China.
The domestic duck is the main
reservoir of influenza viruses in
southern China. Geese are
infrequently infected and chicken
rarely.
Reassortment
The domestic pig serves as the
host for both avian and
human influenza viruses. These viral
strains recombine their genes to give
rise to a novel influenza virus capable
of human-to-human transmission.
Functional virions that
contain combinations of
of genomic segments from
intracellular “pools” of viral strains
38
Influenza A viruses
Infect humans, horses, pigs, ferrets and birds.
A major human pathogen, that incites epidemics and pandemics.
15 known haemagglutinin (HA) serotypes and 9 known neuraminidase
(NA) serotypes have been identified. Pigs and ducks are important
reservoirs, that produce genetically/antigenically diverse viruses. The
viruses are transferred back to humans via close contact between humans
and animals.
Influenza B viruses
Only infects mammal and incite a mild form of influenza.
Does not have distinguishable serotypes like influenza A.
Influenza C viruses
Only infects mammals, but it is unclear if they cause disease.
Genetically and morphologically distinct from A and B types.
39
Novel Influenza
A Strains Incite Pandemics
1918 Spanish flu
1957 Asian flu
1968 Hong Kong flu
1977 Russian flu
Human Flu Transfers May Exceed Reports
By DONALD G. McNEIL Jr.
Published: June 4, 2006
The New York Times
Avian flu (H5N1) 1997
- Type A virus
- Easily transferred & highly virulent in chickens & wild birds
- Not easily transferred to humans to date
- 105 human deaths in 186 confirmed cases (WHO Mar 24, 2006)
- Human deaths are associated with close exposure to infected chickens
- H5N1 is not transmissible via aerosol droplets
- Concern & debate regarding the no. of mutations required to
make this possible
40
How will H5N1 reach North America?
Figure 2. Map of known routes for natural interhemispheric bird movement: route 1, migrants breeding in Alaska and
wintering in East Asia; route 2, migrants breeding in East Asia and wintering along the Pacific Coast of North America;
route 3, migrants breeding in Iceland or northwestern Europe and wintering along the Atlantic Coast of North America;
route 4, vagrants from West Africa carried by tropical storm systems across the Atlantic to eastern North America.
Birds and Influenza H5N1 Virus Movement to and within North America
John H. Rappole* and Zdenek Hubálek†
*Smithsonian Institution, Washington, DC, USA; and †Academy of Sciences, Valtice, Czech Republic
Emerging Infectious Diseases
Vol. 12, No. 10 • October 2006
41
“The bird flu virus, known as A(H5N1), belongs to a group of influenza viruses
known as Type A, which are the only ones that have caused pandemics. It has been
steadily advancing around the world, first appearing in Asia, then Europe and
Africa. The apparent lethality of A(H5N1), combined with its
inexorable spread, are what have made scientists take it seriously. The
virus lacks just one trait that could turn it into a pandemic: transmissibility,
the ability to spread easily from person to person. If the virus acquires that ability, a
worldwide epidemic could erupt.
The A(H5N1) strains circulating now are quite different from the A(H5N1) strain
detected in Hong Kong in 1997, which killed 6 of 18 human victims. Over time,
A(H5N1) seems to have developed the ability to infect more and more
species of birds, and has found its way into mammals -- specifically,
cats that have eaten infected birds.”
The New York Times Sunday, June 4, 2006
42
Aerosols – suspensions of fine dust
and moisture in the air that contain
pathogens
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