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Using Comparative Genomics to Explore
the Genetic Code of Influenza
Sangeeta
Venkatachalam
The influenza pandemic of 1918
----“Spanish flu”
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killed 20-50 million people.
considered the most deadly pandemic in recorded human history.
Why??
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The virus antigens were extremely different to those encountered
previously so people carried no immunity to this virus strain and
were highly susceptible to illness and even death.
What was it??
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It was caused by the H1N1 type.
It has been found to be very similar to the bird flu of today mainly
H5N1 and H5N2.
Resurrecting the 1918 influenza virus
Fragments of RNA are retrieved
from fragments of long tissue,
converted into DNA and sequenced.
Flu victim frozen in Alaska
Permafrost since 1918.
The overlapping sequences are pieced
together to give the full gnome
sequence. A DNA version is synthesized in
the lab.
The virus is isolated from the cells and
used to infect mice. They all die within
6 days.
The DNA is injected into canine
kidney cells, which produces tens
of virus particles.
Influenza Virus structure
Hemaglutinin
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The Influenza virus A is a globular particle
about 100 nm in diameter
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covered by in a lipid bilayer .
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Studded in the lipid bilayer are two integral
membrane proteins.
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Neuraminidase
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500 molecules of hemagglutinin ("H")
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100 molecules of neuraminidase ("N").
8 pieces of RNA.
The HA gene.
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3 distinct hemagglutinins, H1, H2, and
H3 are found in human infections
The NA gene.
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2 different neuraminidases N1 and N2
have been found in human viruses
The NP gene
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encodes the nucleoprotein. Influenza A,
B, and C viruses have different
nucleoproteins.
Hemagglutinin & Neuraminidase
Hemagglutinin
Neuraminidase
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the main antigenic
determinant on the virus.
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a glycoprotein expressed on
the viral surface.
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immune system primarily
recognizes and responds to
hemagglutinin by making
antibodies and mounting an
immune defense.
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Its principal biological role is
the cleavage of the terminal
sialic acid residues that are
receptors for the virus'
hemagglutinin (HA) protein.
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Based on how well a person's
immune system can
recognize the hemagglutinin
the severity of the flu can be
decided.
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It is clear that successful virus
replication and spread
depends on tightly coordinated
interactions among the virus’
genes.
Antigenic shift and drift
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Antigenic drift
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The changes in the HA antigen shape, may cause
the antibodies not to match up, and thus allowing
the newly mutated virus to infect the body's cells.
This genetic mutation is called. link
Antigenic shift
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when the genetic change enables a flu strain to
jump from one animal species to another,
including humans. is called antigenic shift. link
Investigation into changes or mutations in
the gene for influenza N antigen.
What did we do ?
 Run a multiple sequence alignment using
versions of H3N2 viral sub-strain.
 Run a multiple sequence alignment using
H3N2, H1N1 (1918) and H5N1 influenza A
viral sub-strains.
 Use the sequence alignment to construct a
phylogenic tree.
Multiple Sequence alignment of
H3N2 sequences
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The NA gene sequences used here
come from influenza viruses isolated
from infected humans. The virus strain is
H3N2 (referring to the H and N antigens
on the virus).
Each sequence has a name
e.g. A_HongKong_68_H3N2 This
indicates it is an influenza A virus from
Hong Kong isolated in 1968.
Findings
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Some nucleotides not conserved (shown in black)
most of the time these led to the same amino acid.
most of the changes take place in the third nucleotide position.
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Because
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the third nucleotide is where the binding between the tRNA and the mRNA is the weakest and
mistakes in translation are most likely to take place here.
Also the there often are several codons for a single amino acid and that the first two letters in
a codon usually are the important ones , but that the third letter is occasionally significant.
The NA gene is highly conserved between viral strains, especially the active
site.
If the amino acid sequence is altered too significantly,
then the protein shape changes and
can no longer function to cut the virus particle from the host cell membrane.
As a result, the virus can no longer spread to infect new host cells.
Multiple Sequence alignment of H3N2 and
H1N1 sequences
We did alignment of this particular H1N1 virus
from 1918 and the others found recently.
The results show that the H5N1 virus seen in
Thailand in 2004 are very close to each other in
the tree.
This may mean that the H5N1 might be able to
create a pandemic like the H1N1 of 1918
Conclusion
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Knowledge of the genome of the 1918 virus gained so far may provide
clues to help us avoid or prepare for another pandemic.
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resemble bird flu genomes more closely than those of human strains.
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Using the techniques available through bioinformatics we can analyze
the virus strain which emerge newly and find out how deadly it can be
by finding the closeness to viruses which have caused pandemics
before.
References
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R. J. Webby, R. G. Webster, Science 302, “Are We Ready for Pandemic Influenza?”, 15191522 (2003).
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Tumpey, T.N. et al. Science 310, “Characterization of the Reconstructed 1918 Spanish
Influenza Pandemic Virus”, 77-80 (2005).
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Palese, P. & Compans, R. W. Inhibition of influenza virus replication in tissue culture by 2deoxy- 2,3-dehydro-N-trifluoroacetylneuraminic acid (FANA): mechanism of action (1976)
J. Gen. Virol. 33, 159-163
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A. H. Reid et al., Initial Genetic Characterization of the 1918 "Spanish" Influenza Virus
Proc. Natl. Acad. Sci. U.S.A. 97, 6785-6790 (2000)
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Reid, A. H., Fanning, T. G., Hultin, J. V. & Taubenberger, J. K. Origin and evolution of the
1918 "Spanish" influenza virus hemagglutinin gene (1999) Proc. Natl. Acad. Sci. USA 96,
1651-1656
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Using bioinformatics to explore the genetic code http://www.gtac.edu.au/site/bioinformatics/