Influenza

New vaccine developed yearly

1918 Influenza Pandemic
Influenza A H1N1 virus
Transmitted via aerosols coughing and sneezing
Rapid, lethal
What is an Epidemic?
Given area or among a species group of people over a certain
period of time
What is Pandemic?
Wide area affects large portion
Examples: Cholera, AIDS, Pandemic Influenza
Background
Enveloped virus
- ss RNA virus
Family Orthomyxoviridae
Genome- 8 segments
Haemagglutinin and neuraminidase spikes on envelope
3 types by serotype of ribonucleic protein
o Influenza virus A, B, C
A multiple species
B and C humans
Several serotypes based on antigenic determinates of
hemagglutinin (HA) and neuraminidase (NA).
Type A undergoes antigenic shift and drift.
o Makes it very dangerous
Type B undergoes antigenic drift only and type C is relatively
stable
Influenza A Virus
Undergoes antigenic shifts and antigenic drifts with the
haemagglutinin and neuraminidase proteins.
Shifts in haemagglutinin pandemics.
Drifts in H & N epidemics.
Cause mild febrile illness.
Death may result from complications such as viral/bacterial
pneumonia.

Influenza: disease Pathogenesis

Mild influenza: virus replication is primarily restricted to the
upper respiratory track.
Severe influenza: the virus replicates extensively in the lower
respiratory track inhibiting gas exchange by damaging type 2
pneumocytes and may disseminate further
o Multi organ failure
o Infection and killing of pneumocytes
o Severe hypoxemia
o Secondary infections
o Paralysis of cilia
o Excessive inflammatory response
Influenza Virus Structure
Polymorphic but mainly circular
Influenza viral genome
o (-) ssRNA
o 8 segments (pieces)
o One gene per segment
Nucleoprotein
Matrix proteins
NS (nonstructural proteins, that are not incorporated into viral
particles) gene encodes two different non-structural proteins
Subunits of RNA polymerase
Spikes (about 500)
Two antigen on envelope:
o Hemagglutinin - trimer (HA)
Helps virus enter cell
Type A infects humans, birds and pigs
Type A has ~ 20 different sub types
o Neuraminidase - tetramer (NA)
Helps virus exit cell

9 subtypes
Flu Viruses Currently infecting...

Humans: H1N1, H1N2, and H3N2

Avian Flu Virus: H5N1

HA - hemagglutinin
NA - neuraminidase
helical nucleocapsid (RNA plus
NP protein)
lipid bilayer membrane
polymerase complex

M1 protein

type A, B, C : NP, M1 protein

sub-types: HA or NA protein
Haemagglutinin (HA)
Encoded by RNA segment # 4
Can agglutinate red blood cells - hence the nomenclature
o Used for diagnostic of influenza
Cleavage by host-cell protease is required (resulting in HA1 and
HA2) for infection to occur imp.
Hemagglutinin glycoprotein is the viral attachment protein and
fusion protein, and it elicits neutralizing, protective antibody
responses ***MAIN
Neuraminadase (NA)
Encoded by RNA segment # 6
Removes neuraminic (sialic) acid from cell and permits
dissemination of viruses
Important in releasing mature virus from cells
Contributes production of protective antibodies

Features of viral genera

severity of illness
animal reservoir
human pandemics
human epidemics
antigenic changes
segmented genome
amantadine, rimantidine
zanamivir
surface glycoproteins

TYPE A

TYPE B

TYPE C

++++
yes
yes
yes
shift, drift
yes
sensitive
sensitive
2

++
no
no
yes
drift
yes
no effect
sensitive
2

+
no
no
no (sporadic)
drift
yes
no effect
(1)

Animal Susceptibility and Growth of Virus

Virus human strains can affect different animals; ferrets are
most susceptible. Serial passage in mice increases its
virulence, producing extensive pulmonary consolidation &
death
The developing chick embryo readily supports the growth of
virus, but there are no gross lesions.
Influenza virus reproduction**
Virus (structure shown in a cutaway view) adsorbs to a
respiratory epithelial cell by hemagglutinin spikes and fuses
with the membrane
Virus is endocytosis into a vacuole and uncoated to release its
8 nucelocapside segments into the cytoplasm
The nucleocapsids are transported into the nucleus. There the
(-) sense RNA strand is transcribed into a (+) snese strand
that will be translated into the viral proteins that make up the
capsid and spikes
(+) Sense RNA is used to synthesize glycoprotein spikes
inserted into the host membrane.

The (+) sense RNA strands are used to synthesize new (-)
sense RNA strands. These are assembled into nucelocapsides
and transported out of the nucleus to the cell membrane
Release of mature virus occurs when viral parts gather at the
cell membrane and are budded off with an envelope
containing spikes
Single-cell reproductive cycle
1. Attachment to the epithalial cells of the host through
hemagglutinin.
2. Endocytosis
3. Uncoating - > This exposes the contents of the virus to the
cytosol.
4.The RNA enter the nucleus of the cell where fresh copies are
made.
5. These copies return to the cytosol where some serve as mRNA
molecules to be translated into the proteins of fresh virus
particles.
6. Progeny virions are formed and released by budding from the
plasma membrane of the cell (aided by the neuraminidase) thus
spreading the infection to new cells.
Epidemiology
Pandemics - influenza A pandemics arise when a virus with a
new haemaglutinin subtype emerges as a result of antigenic
shift. As a result, the population has no immunity against the
new strain. Antigenic shifts had occurred 3 times in the 20th
century.

Epidemics - epidemics of influenza A and B arise through more

minor antigenic drifts as a result of mutation.

Antigenic changes of Influenza A

Viruses can undergo frequent changes due to recombination,
reassortment, insertions and point mutations
Antigenic drift
Antigenic shift occurs every 8-10 yrs
Minor antigenic changes favor persistence of the viruses in the
population and allow recombination that can eventually lead to
severe epidemics and/or pandemics

Orthomyxoviruses. Nomenclature
Human influenza virus
Influenza A/Bangkok/1/79(H3N2)
Influenza A/Singapore/1/57(H2N2)
Influenza B/Ann Arbor/1/86

Influenza type Year of isolation Hemagglutinin subtype

A/Sydney/5/97 (H3N2)
Geographic source Isolate number Neuraminidase subtype

Antigenic Drift
Gradual accumulation of mutations that allow the
hemagglutinin to escape neutralizing antibodies
Epidemic strains thought to have changes in three or more
antigenic sites
Antigenic differences can result from changes in one amino
acid
Can involve any antigenic protein
Can occurs every year
RNA replication is error prone
New HA types are created frequently
Requires new vaccine every season
Antigenic shift
Occurs every 8-10 yrs
Major antigenic change of either H or N antigens or both H and N
Occurs by gene reassortment after simultaneous infection of a
cell with two different viruses
Three different H proteins and 2 major N proteins have evolved

Antigenic shift theories

Reassortment - Reassortment of the H and N genes between
human and avian influenza viruses through a third host. There is
good evidence that this occurred in the 1957 H2N2 and the 1968
H3N2 pandemics. The 2009 pandemic virus was thought to be
novel virus that was a triple re-assortant involving human, bird,
N. American pig and Eurasian pig viruses.
2. Recycling of pre-existing strains this probably occurred in
1977 when H1N1 re-surfaced.
3. Gradual adaptation of avian influenza viruses to human
transmission. There is some evidence that this occurred in the
1918 H1N1 pandemic.
Mechanism of shift
Mallard 16 Has, 9 NAs: non human virus direct to humans
Mallard 16 Has, 9 NAs: non human virus pig reassortant
virus humans
Where does influenza come from?
Type A constantly circulates in natural reservoirs
Birds are the natural reservoir of all subtypes of Influenza A
viruses
o Migratory waterfowl
o Chickens, turkeys, ducks, geese
Humans
Pigs
Horses
Other
Epidemiology
Source of infection: patients and carriers.
AEROSOL
o 100,000 TO 1,000,000 VIRIONS PER DROPLET
Common: large droplets (sneezing, coughing, contact with saliva)
Probably common: contact
o Direct
o Fmite
Rare: airborne over long distance
18-72 HR INCUBATION
Symptoms
FEVER
HEADACHE

MYALGIA
COUGH
RHINITIS
OCULAR SYMPTOMS
CHILLS and/or SWEATS
Infection may be very mild, even asymptomatic, moderate or
very severe
Clinical Responses
Acute Symptoms last one week
Abrupt onset of fever, myalgia, headache and non-productive
cough
Fatigue and weakness can last 2-3 weeks.
Infected individual predisposed to bacterial infections
Staphylococcus, Streptococcus, Hempohilus
Immunity dependent upon localized anti-viral secretory IgA
( strain specific)
Develop long lasting circulating anti-viral IgG
Immunity to influenza
Antibody to HA - >protective

Antibody to NA - > decrease severeity

Serum antibody - > years

Secretory antibody - > months

Laboratory Diagnosis
Respiratory secretions (direct aspirate , gargle , nasal washings)
Virus isolation and growth in embryonated eggs
Cell culture in primary monkey kidney or madindarby canine
kidney cells
Hemagglutination (inhibition)
ELISA
Direct immunofluorescence
Hemaggutination inhibition
Gold standard for diagnosis and used to follow drift and shift
important in vaccine development
Diagnosis Hemagglution inhibition (HI) and near patient or rapid
tests which detects ride in antibodies to influenza or NA activity
Rapid influenza diagnostic test (RIDT)
Detect influenza antigens

Prophylaxis
Hand washing
o Generally perceived to be useful
o No studies specifically performed for influenza
o Easy to recommend
Masks
o Effectiveness not shown for influenza
o However, could reduce transmission associated with large
droplets
Types of Vaccine
o Killed Whole Virus
o Inactivated virus cacinne grown in embyonated eggs
o 70-90% effective in health persons <65 years of age
o 30-70% in persons > 65 years
o Live virus
o Attenuated strains were widely used in Russia but not
elsewhere
o Virus subunit
o HA extracted from recombinant virus forms the basis of
todays vaccines
o Synthetic
o Much research is being done to try and find a neutralizing
epitope that is more stable, and can therefore be used for
a universal vaccine
Trivalent Influenza virus vaccines
1999-2000

A/Sydney/05/97 (H3N2)

A/Beijing/262/95 (H1N1)

2000-2001

B/Yamanashi/166/98

A/Moscow/10/99(H3N2)-like
A/New Caledonia/20/99 (H1N1)-like

B/Beijing/184/93-like
2006-2008
A/Brisben/59/2007 (H1N1)
A/Brisben/10/2007 (H3N2)
B/Florida/4/2006

Prevention and Treatment

o 70-90% effective in preventing illness
o RIMANTADINE (blocks the M2 ion channel) (M2)
o type A only, needs to be given early
o AMANTADINE (blocks the M2 ion channel) (M2)
o type A only, needs to be given early
o ZANAMIVIR (neuraminidase inhibitors) (NA)
o types A and B, needs to be given early
o OSELTAMIVIR (neuraminidase inhibitors) (NA)
o types A and B, needs to be given early
Antivirals: Adamantanes and Neuraminidase Inhibitors
Prophylaxis and antivirals targeting influenza
1.Seasonal Vaccination #1 strategy for prevention
o Risk groups first pregnant women, medical personnel,
young children people with medical insufficiencies, older
people, general population
2. Reducing risk: quarantine and suspected contact vaccination
o vaccination of older adults with latest polyvalent
pneumococcal vaccine
3.Therapy
o Anti virals targeting influenza proteins, M2 protein,
amantadine and neuraminidase , zanamivir and oseltamir
Secondary Bacterial Infections
o Cleavage of the HA0and HA1 and HA2 ** imp. virulence and high
mortality with influenza A infections
o Spread of influenza virus to lungs and other tissue
o In addition NA action on respiratory
o Exposes bacterial adhesion
o Molecules resulting in Viral Pneumonia ,
o Secondary Bacterial infections high morbidity and
mortality
o Bacterial proteases from secondary
o Infections can contribute to pathogenicity
Synergy between influenza and bacteria resulting in
pneumonia: proposed mechanism
o Vicious Cycle develops
1) Initial Influenza infection
1. Lung cilia damaged
2 Macrophages killed
3 bacterial receptors exposed by NA
4 pneumocytes killed

2) Increased susceptibility to Bacterial infection inflammation,

proteases; inflammatory and bacterial
3) Enhanced Virus replication: viral pneumonia
4) Enhanced Bacterial replication: bacterial pneumonia
o Archival evidence suggests bacterial infections contributed
o To the high mortality during 1918 1919 high rate of carriage
o In the developing world also contributed 29% deaths from novel
o H1N1 2009 pandemic evidence of bacterial infection
Emergence of New Influenza Subtypes: H5N1
o Antigenic shift due to genome reassortment within intermediate
hosts drives flu epidemics and pandemics
1. Nonpathogenic H5 influenza virus: Wild fowl domestic ducks
and geese, domestic chickens.
2. H5 virus became highly pathogenic in chickens domestic ducks
and geese.
3. Highly Pathogenic H5 virus reassorted its genome with those of
other influenza viruses in aquatic birds, spread to poultry
farms, humans, and occasionally to pigs.
Avian influenza
Wild birds are the reservoir.
Circulation of low pathogenic avian flu in domestic poultry leads
to mutations to highly pathogenic forms over time.
Co-infection with swine or humans infected with human influenza
can result in genetic reassortment and highly pathogenic strains.
Why do new strains of influenza and bird flu arise in Asia?
o In 2003, an outbreak of chicken flu necessitated killing tens of
millions of birds
o H5N1 in wild birds, poultry & humans 5/19/06