INFLUENZA AND PRODUCTION OF VACCINES AGAINST INFLUENZA VIRUSES
RIMJHIM ROY CHOUDHURY
• Influenza virus belongs to the orthomyxoviridae family.
• Influenza virus have segmented negative-sense single
stranded RNA molecules, which are eight in number.
• It’s an enveloped virus which is made up of proteins,
glycoproteins, and lipid bilayer.
• Helical in symmetry.
HA - hemagglutinin
NA - neuraminidase
helical nucleocapsid (RNA plus
lipid bilayer membrane
type A, B, C : NP, M1 protein
sub-types: HA or NA protein
Types of Influenza virus
Severity of illness
HA attaches to the membrane of epithelial
cells via sialic acid-galactose
H+ ion passes through the viral M2
matrix ion protein channels
After attachment endocytosis occurs
Fusion of endosome membrane and viral
membrane; release of RNA and NP
Acidification of the core leads to
conformation changes in HA making it a
Inside the endosome pH gets lowered
Mode of action of H1 N1
Virus infects upper respiratory system
Hydrolyzing the mucus membrane by neuraminidase
Virus adheres to epithelial cells
(receptor mediated endocytosis)
Action of lysosome on endosome
When endosome pH decreases, haemagglutine molecule undergoes
conformational change. The hydrophobic end of haemagglutine spring outward
& extends towards the membrane of endosome.
Mode of action of H1 N1
Nucleocapsid is released into the cytoplasm
Virus replication in epithelium cell nucleus
Assembly of viron particles
Released through budding
Undigested virus particles engulf by
Ag presentation by class 1 MHC molecule
Tc cells get activated
As a result, there is reduced clearance of infectious agents from
the respiratory tract. Gaps in the protective epithelium provide
other pathogens with access to other cells.
• Inactivated vaccine (TIV)
flu shot (injection)
Purified virus chemically inactivated by formalin or propiolactone
trivalent (three strains; usually A/H1N1, A/H3N2, and B)
contain much more antigen than live vaccines
works by putting into the bloodstream those parts of three
strains of flu virus that the body uses to create antibodies
• Live attenuated vaccine (LAIV)
nasal spray (FluMist®)
Live virus, completely devoid of pathogenicity
provide continuous antigenic stimulation
works by inoculating against those same three strains that have
been genetically modified to minimize symptoms of illness.
• Each of the three strains is a reassortant of internal proteins of a
master donor virus (MDV) that contains the CA and TS
• surface proteins (HA, NA) are from wild-type influenza virus.
Flu vaccine production (Overview)
Preparation of seed virus
Seed passaging and selection
Large scale production
Purification and testing
Packaging and shipping
Strain selection and Preparation of seed virus
2 gene segments
representing the HA
and NA antigens are
selected from the
6 gene segments
come from a lab
virus strain that
a hybrid is formed which
contains the inner
components of the
laboratory strain, and the
outer components of the
Seed passaging and Selection
Accredited laboratories distribute seed viruses to
manufacturers to begin the production process.
Once the seed virus has been received, the working
seed virus can be prepared by passaging the seed
virus in eggs.
These passages are necessary to determine the
optimum growth conditions to improve virus yield
in the industrial environment.
Large – scale production
Millions of specially - prepared chicken eggs are
used to produce the vaccine. Throughout the year,
fertilized eggs are delivered to the manufacturer.
Each egg is injected with the working seed.
The eggs are incubated for several days to allow the
virus to multiply. After incubation, the virus –
loaded fluid is harvested.
Purification and Testing
Manufacturers test the vaccine concentrate using
specially prepared reagents provided by WHO
Collaboratoring Centers to measure the quantity of
virus produced and guarantee the optimal dosage of
ready to use vaccines.
• A limitation of current vaccines is that the antigenic regions of
HA are highly susceptible to continuous mutation in circulating
epidemic virus strains.
•Thus, the currently available influenza vaccines need to be
updated every year to match the antigenicity of the virus strains
that are predicted to circulate in the next season.
•However, current vaccines would not be effective in preventing
the spread of a new pandemic strain containing a substantially
different HA protein. Therefore, new approaches are being
investigated to develop broadly crossprotective vaccines, focused
primarily on type A influenza viruses
Universal influenza vaccine
1. The extracellular domain of M2 (M2e) is highly conserved
among multiple influenza A viruses, indicating that M2 is an
attractive antigenic target for developing a universal influenza
vaccine. Also, an inactivated influenza vaccine supplemented
with M2 VLP(virus like particles) prevents disease symptoms.
2. HA is a homotrimeric molecule consisting of a disulfide-linked
globular head of HA1 and a stem domain composed of part of
HA1 and all of HA2. It possesses conserved structural features
in the HA2 segment involved in anchoring to the viral
membrane. It has recently been recognized that this segment,
termed the stalk, is a potential target for inducing broadly
• Non – egg – based production
•Targeting generation of virus specific CTLs
•Increasing immunogenicity and adjuvants
• Novel vaccines against influenza viruses: Virus Research,
Volume 162, Issues 1–2, December 2011, Pages 31-38. S.M.
Kang, J.M. Song, R.W. Compans
• The Influenza Virus Enigma: Cell, Volume 136, Issue 3, 6
Rachelle Salomon, Robert G. Webster
• The 2009 A (H1N1) influenza virus pandemic: A review:
Vaccine, Volume 28, Issue 31, 12 July 2010, Pages 4895-4902
Marc P. Girard, John S. Tam, Olga M. Assossou, Marie Paule