Influenza
Chapter 39
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Seasonal Influenza
Causative Agents
Influenza A virus
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Family Orthomyxoviridae
Negative-stranded RNA genome
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Spiked envelope
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Genome divided into 8 gene segments
H spike – hemagglutinin (subtypes H1-H16)
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Mediates attachment
N spikes – neuraminidase (subtypes N1-N9)
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Cleaves H protein to allow fusion of viral and cellular
membranes (i.e., entry into the cell)
Requires cellular enzyme trypsin to facilitate infection
Influenza B & C viruses only circulate in
humans
Influenza Genes
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Influenza A viruses have 8 gene segments that
encode 10 polypeptides
• Segment 1 (2,341 nt): PB2 (RNA transcriptase)
• Segment 2 (2,341 nt): PB1 (RNA transcriptase)
• Segment 3 (2,233 nt): PA (RNA transcriptase)
• Segment 4 (1,778 nt): HA (hemagglutinin) - 16 known subtypes
• Segment 5 (1,565 nt): NP (nucleoprotein)
• Segment 6 (1,413 nt): NA (neuraminidase) - 9 known subtypes
• Segment 7 (1,027 nt): M1, M2 (matrix proteins)
• Segment 8 (890 nt): NS1, NS2 (inhibits mRNA splicing and IFN response; nuclear
export signal for viral RNPs)
The HA and NA Polypeptides
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HA encodes hemagglutinin spike protein
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Binds to any cellular protein with a α2,6-sialic acid group
Mediates penetration of viral core and RNA into target
cell
It is the major target of neutralizing antibodies for
vaccines and successful immune responses
NA encodes neuraminidase protein
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This enzyme cleaves the HA protein upon cell binding
and facilitates penetration of the viral core
Together they mediate cell infection
Influenza viruses, in part, are named by their HA and NA
subtypes (e.g., H5N1)
Seasonal Influenza
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Symptoms
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Influenza Type A
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Short incubation period
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Averaging 2 days
Headache
Fever
Muscle pain
Dry cough
Acute symptoms abate within a week
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Cough, fatigue and generalized weakness may linger
Influenza A Transmission Cycle
Adaptation/
reassortment
with swine
influenza viruses
Circulates with
limited pathology
Transmission to
domestic fowl
Transmission
to humans
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Seasonal Influenza
Pathogenesis
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Acquired through inhalation of infected respiratory secretions
Virus attaches to host cells via hemagglutinin spikes
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Mature viruses bud from host cell
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Budding allows mature virus to pick up envelope
Infected cells die and slough off
Host immunity quickly controls viral spread
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Once attached viral envelope fuses with host membrane, leading to viral
replication within the cell
Anti-HA neutralizing IgG is protective
Mortality rate is low
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However, hundreds of thousands or millions of people are infected each year in
the U. S.
On average, about 30,000 Americans, mostly elderly and very young children,
die from influenza each year
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Seasonal Influenza
Epidemiology
Outbreaks occur in United States every year
Vaccines are formulated months in advance
using prominent circulating strains
2012-13 vaccine strains
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Type / Geographic origin / Strain/ Year isolated (H & N genes)
A/California/7/2009 (H1N1)-like virus (same strain as 2011-2012 flu
season)
A/Victoria/361/2011 (H3N2)-like virus
B/Wisconsin/1/2010-like virus
Pandemics occur periodically
Most infamous pandemic of 1918 (Spanish flu)
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Spanned the globe in 9 months
Pandemics have higher than normal morbidity
Seasonal Influenza
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Epidemiology
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Spread caused by major
antigenic changes
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Antigenic drift
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Consists of amino acid changes
in spikes (point mutations)
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Particularly hemagglutinin
Changes minimize effectiveness
of immunity to previous strains
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Ensures enough susceptible
people are available for
continued virus survival
Antigenic shiftRepresent more
dramatic changesVirus strains are
drastically antigenically different from
previous strains, importantly
hemagglutininNew virus comes from
genetic re-assortmentOccurs when two
different viruses infect a cell at the same
timeGenetic mixing results in new virus
that is often more virulent
Seasonal Influenza
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Seasonal Influenza
Prevention and Treatment
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Vaccine can be 80% to 90% effective
New vaccine required every year
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Due to antigenic drift
Antiviral medications are 70% to 90%
effective
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Include amantadine, rimantadine, and
Tamiflu
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Must be taken early
Not a substitute for vaccine
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Avian Influenza
There are hundreds,
if not thousands, of
influenza A viruses circulating in nature
Seasonal influenza occurs from mammalian
viruses
Pigs in SE Asia are frequently a source of
these viruses
New reassortants arise every year, but most
are not pathogenic to humans
HA attaches to α2,6-sialic acid receptor
Avian influenza viruses routinely circulate
among wild birds
Some species can be infected without
conspicuous pathology
These species often carry the viruses along
migratory routes, exposing other birds
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Avian Influenza
Most avian influenza viruses are highly
inefficient at infecting humans
However, some cultures have domestic birds
and pigs in close periodomestic proximity
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This practice increases the chance of
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Reassortment with mammalian influenza
viruses
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1957, 1967 pandemic strains were reassortant
mammalian viruses with avian segments (antigenic shift)
Emergence of mutant avian strains that can
infect humans
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1918 pandemic strain was an avian virus that adapted to
efficient human to human transmission (antigenic drift)
Recovery of the Spanish Flu Virus
Using Reverse Genetics
• Spanish Flu pandemic: 1918-1919
• Estimated 20 to 50 million dead
• High death rate among young, health
people
• Played a role in ending World War 1
• No viable virus was saved
Spanish Influenza
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Rescue of the 1918 pandemic strain
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Virology did not exist in 1918
The virus could not be isolated, thus went
extinct when the pandemic ended
In 2005 a group resurrected the 1918 strain
from bodies buried in Alaskan permafrost
Viral genome sequencing indicated it was
an avian influenza A virus
Molecular Recovery of Gene Segments
• Collect infected tissues from victims of the
virus buried in permafrost or formalin-fixed
tissues
• Use reverse transcription-PCR to amplify
gene segments
• Clone into plasmids
• Transfect MDCK (canine kidney cell line)
• Recover infectious viruses
Characteristics of the Spanish Flu Virus
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It is an avian influenza virus (i.e., directly jumped from birds to
humans)
In infects cells in culture without trypsin (an indicator of
pathogenicity in mammals)
It was an H1N1 virus
Kills chicken eggs in 4.5 days
Pathology was largely confined to the lungs of experimentallyinfected mice (similar to humans)
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Mice died in as little as 3 days post infection
Substantial damage occurred in the lungs (similar to humans)
Very high titers of virus were obtained from the lungs
Avian Influenza
Tumpey et al.,
310:77. 2005
Lungs from Mice Infected with Rescued 1918 H1N1 Pandemic Virus
1918 Strain
1918 Strain
1918 Strain with
Texas 1991 H
segment
1918 Strain
1918
Strain(HN)/Texas
1991 Strain hybrid
Texas 1991
strain (control)
Is Another Avian Influenza
Pandemic Upon Us?
• The current avian influenza (H5N1) virus
that has been circulating since 1997 has
reached Europe and Africa
• All human cases thus far are confined to
SE Asian countries
• Sporadic person to person transmission
has occurred
• Will this virus become efficient at person
to person transmission?
Avian Influenza
Feature
SI1
H5N1 AI2
1918 H1N13
Transmission efficiency
High
Very low/none
High
Replication site
Upper and lower
respiratory tract
Lower RT only
Likely upper &
lower RT
Viral CPE4
Limited
Substantial
Substantial
Immunopathology
Limited
Substantial
Substantial
Kills embyronated chicken eggs?
No
Yes
Yes
Requires trypsin for infection of
cell cultures?
Yes
No
No
Vaccine
Yes
No
N/A
Fatality rate
0.03% (U.S.)
57% (global)
About 1-2% (U.S)
Demographic
Young children,
elderly
Young adults
Young adults
1 Seasonal
influenza
2 Currently circulating H5N1 avian influenza virus
3 Rescued 1918 pandemic H1N1 avian influenza virus
4 Cytopathic effect (damage directly caused by the virus)
Red - more pathogenic feature
Yellow - less pathogenic feature
• For seasonal influenza, viruses are grown in
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Will A Vaccine
Be Easy
to Produce?
chicken
eggs for 7 days
to produce
high-enough
titers for vaccines
Avian influenza viruses kills eggs in 4.5 days
An insufficient titer is generated for vaccines
The world’s annual vaccine capacity is 300 million
doses
If the vaccine can be produced by cell culture,
then adequate supplies should be able to be
produced
A reverse genetics vaccine may be the only way to
prevent an avian influenza pandemic
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A DNA vaccine has recently been described that protects mice from lethal
challenge to the 1918 strain
DNA vaccines are plasmids that express viral antigens under the control of
• For seasonal influenza, viruses are grown in
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•
•
Will A Vaccine
Be Easy
to Produce?
chicken
eggs for 7 days
to produce
high-enough
titers for vaccines
Avian influenza viruses kills eggs in 4.5 days
An insufficient titer is generated for vaccines
The world’s annual vaccine capacity is 300 million
doses
If the vaccine can be produced by cell culture,
then adequate supplies should be able to be
produced
A reverse genetics vaccine may be the only way to
prevent an avian influenza pandemic
•
•
A DNA vaccine has recently been described that protects mice from lethal
challenge to the 1918 strain
DNA vaccines are plasmids that express viral antigens under the control of