Characterization of conserved properties of hemagglutinin of H5N1 and human influenza viruses: possible consequences for therapy and infection control (Egypt)

[HenryN]

Characterization of conserved properties of hemagglutinin of H5N1 and human influenza viruses: possible consequences for therapy and infection control

Veljko Veljkovic , Nevena Veljkovic , Claude P Muller , Sybille Muller , Sanja Glisic , Vladimir Perovic and Heinz Kohler
BMC Structural Biology 2009, 9:21doi:10.1186/1472-6807-9-21

Published:	7 April 2009

Abstract (provisional)

Background

Epidemics caused by highly pathogenic avian influenza virus (HPAIV) are a continuing threat to human health and to the world's economy. The development of approaches, which help to understand the significance of structural changes resulting from the alarming mutational propensity for human-to-human transmission of HPAIV, is of particularly interest. Here we compare informational and structural properties of the hemagglutinin (HA) of H5N1 virus and human influenza virus subtypes, which are important for the receptor/virus interaction.
Results

Presented results revealed that HA proteins encode highly conserved information that differ between influenza virus subtypes H5N1, H1N1, H3N2, H7N7 and defined an HA domain which may modulate interaction with receptor. We also found that about one third of H5N1 viruses which are isolated during the 2006/07 influenza outbreak in Egypt possibly evolve towards receptor usage similar to that of seasonal H1N1.
Conclusions

The presented results may help to better understand the interaction of influenza virus with its receptor(s) and to identify new therapeutic targets for drug development.

[HenryN]

Full paper at

http://www.biomedcentral.com/1472-6807/9/21

http://www.biomedcentral.com/content...-6807-9-21.pdf

[HenryN]

The isolates in the paper with the H1N1 characteristics have a 3 BP deletion, as did many of the mild cases in 2007.

[Commonground]

Dr. Niman, thanks for this document. These are some excerpts that I found interesting (and could comprehend!) Is this the correct listing for the Egypt-2 sequences they used for this study?

H5N1 (Egypt-2) (Figure 2b): A/Egypt/0636-NAMRU3/2007 [EF382359];
A/Egypt/14725-NAMRU3/2006 [EF200513]; A/duck/Egypt/2253-3/2006 [DQ862002];
A/duck/Egypt/1301-NAMRU3/2007 [EF441281]; A/chicken/Egypt/1081NAMRU3/
2006 [EF441279]; A/chicken/Egypt/1080-NAMRU3/2006 [EF441278];
A/chicken/Egypt/1079-NAMRU3/2007 [EF441277]; A/Egypt/14724-NAMRU3/2006


[EF200512]; A/chicken/Egypt/1889N3-SM26/2007 [EF469653];
A/chicken/Egypt/1892N3-HK49/2007 [EF469660]; A/duck/Egypt/1888N3-SM25/2007
[EF469657]; A/Egypt/1394-NAMRU3/2007 [EF535817]; A/Egypt/1604-NAMRU3/2007
[EF535818]; A/Egypt/1731-NAMRU3/2007 [EF535819]; A/Egypt/2621-NAMRU3/2007
[EF535826]; A/ goose/Egypt/R4/2007 [EU183330]; A/chicken/Egypt/F6/2007
[EU183326]; A/chicken/Egypt/R6/2007 [EU183332]; A/duck/Egypt/R5/2007
[EU183331]; A/chicken/Egypt/R3/2007 [EU183329] ; A/chicken/Egypt/R2/2007
[EU183328]; A/Egypt/2629-NAMRU3/2007 [EU095025]; A/Egypt/2631NAMRU3/
2007 [EU095027]; A/Egypt/2947-NAMRU3/2006 [EF042617];
A/chicken/Egypt/9385NAMRU3-CLEVB125/2007 [EU371910]; A/chicken/Egypt/
9390NAMRU3-CLEVB157/2007 [EU371915]; A/chicken/Egypt/9392NAMRU3CLEVB167/
2007 [EU371917]; A/chicken/Egypt/ 9387NAMRU3-CLEVB148/2007
[EU371912]; A/chicken/Egypt/3051NAMRU3-CLEVB78/2007 [EU371905];
A/chicken/Egypt/9386NAMRU3-CLEVB/136/2007 [EU371911]; A/chicken/Egypt/
9391NAMRU3-CLEVB158/2007 [EU371916];

Excerpts:

Despite its low infectivity for humans, there has been evidence in Egypt of several
clusters of human-to-human transmission with very high mortality rate
-snip-

Figure 4 shows the IS spectra of peptide VIN1 and of the domains identified by
consensus IS of H1N1, H3N2, H5N1 and H7N7 viruses (Table 2) and the position of
these domains in the molecule. As can be seen, the receptor targeting site of H5N1 virus
from the group Egypt-1 (A/Egypt/0636-NAMRU3/2007) is closer to the receptor binding
site than in the other viruses of Figure 4. It may be speculated that this may affect the
efficacy of the virus/receptor interaction.

Finally, we compared informational properties of H1N1 pandemic strains from 1918
from GenBank and seasonal H1N1 strains. The consensus IS of these pandemic isolates
(Figure 3) is characterized by a dominant peak of the frequency F(0.258) which is different from the frequency F(0.236) characteristic of other seasonal flu H1N1 isolates
(Figure 1d). Table 2 shows the domain corresponding to the frequency F(0.258). In the
model of A/South Carolina/1/18 (Figure 4i) the position of this domain does not overlap
with the corresponding domain of other seasonal H1N1 strains, but overlaps with the
corresponding domain of Egypt-2 H5N1 viruses.
-snip-

H5N1 already replicates efficiently in humans, and cause case fatality rates that are ten
times higher than those seen in the 1918 pandemic. Thus, an infectivity of H5N1 similar
to seasonal flu would cause a catastrophic pandemic. The main obstacle for this worst
case scenario is poor human-to-human transmission of H5N1 viruses, which is attributed
to the paucity of sialic acid a2,3 receptor in the epithelium of the human upper respiratory
tract, and the inability of the virus to replicate efficiently at this site. Interestingly, the
ISM approach identifies important differences between H5N1 viruses from Egypt. Some
have the characteristics of most H5N1 strains whereas about one third of the viruses
display characteristics that are also found in human H1N1 seasonal virus. Interestingly
the proportion of the latter viruses has increased from 25 to about 50: between 2006 and
2007.
Similarly the results of H5N1 strains from Egypt (Figure 2) may be indicative of a
possible viral evolution towards receptor usage similar to that of H1N1 viruses, which
efficiently replicate in the upper respiratory tract. The protein domain, which seems to be involved in this subtle change, corresponds to amino acid domain 99-132 (Figure 4g).
However, the role of this domain for enhanced infectivity in humans remains elusive.
Interestingly the corresponding domain of Spanish flu viruses and Egypt-2 H5N1 viruses
are much closer to the receptor binding site of HA1 than in all other H1N1 and H5N1
viruses (Figures 4e-4i and Table 2). This closer proximity may indicate more efficient
virus/receptor interactions in these influenza viruses.*

Finally, we will discuss some of recently reported experimental results which point out
functional and immunological role of H5 HA domain encompassing peptide VIN1. In
order to identify mutations which increase the recognition of H5 HA by SA•2,6Gal
human type receptor, Su and co-workers compared HA from
A/chicken/Ffujian/1042/2005 as wild type with isolates identified in both poultry and
humans in mainland China, Hong Kong, Thailand, and Vietnam during outbreaks
between 1996 and 2005 [29]. Unexpectedly, this analysis revealed six amino acid
substitutions (K35R, D45N, D94N, K35R/D45N, K35R/45N/D94N, A247T) outside the
receptor-binding domain of HA, which could enhance interaction between H5 HA and
human-type SA•2,6Gal receptor. As can be seen, three of these mutations encompass
mutation D45N which is located within peptide VIN1 and two other mutations (K35R
and D94N) are located in its vicinity. It is the first report that naturally occurring
mutations in region of H5 HA which encompasses peptide VIN1 play an important role
in virus transmission from avian to human. It is of note that Egyptian strains contain all
of these mutations, except mutation in position K35. These results point out need for
future testing of evolution of Egyptian strains using hemiadsorption assays for HA receptor-binding activity in order to identify possible new mutations in this domain of
HA which could increase affinity of H5N1 viruses to human-type receptor.
-snip-

It is of note that recent Egypt group-2 strains are characterized by N43, in
contrast to Egypt group-1 strains which contain D43. It means that the 4G6 MAb can not
be used for detection and neutralization of H5N1 viruses belonging to the Egypt group-2.
-snip-

(iv) at least in Egypt H5N1 viruses
have acquired features that may adapt them for H1N1-like receptor usage possibly
allowing more efficient human-to-human transmission.
-snip-

Figures:

Figure 1 -ISM analysis of HA1 proteins of H5 influenza viruses.

(a) Consensus IS of HA1 of all H5N1 sequences in GenBank (n=1407); (b) IS of H5N3
(A/swan/Hokkaido/51/96), the progenitor of H5N1, and (c) of the first isolated H5N1
virus (A/Goose/ Guangdong/1/96); (d) consensus IS of H1N1 (n=30), (e) of H3N2 (n=30)
and (f) of H7N7 (n=30).

Figure 2 -The ISM analysis of HA1 proteins of H5N1 viruses isolated in Egypt in
2006/2007.

(a) consensus IS of Egypt-1 and (b) of Egypt-2 strains, (c) IS of a representative Egypt-1
(A/teal/Egypt/9885-NAMRU3/2005) and (d) Egypt-2 virus (A/chicken/Egypt/R6/2007).
Figure 3 -Consensus IS of HA1 from three Spanish flu H1N1 viruses.

Figure 4 -Overview of H5 HA trimer (PDB: 2ibx) and details of the VIN1 region.

Domains of H1N1, H3N2, H5N1, H7N1 and Spanish flu identified by consensus IS
(Table 2) and their position in the 3D structure of HA1 and the IS of the peptide
sequence. (e) A/New York/383/2004 (H3N2); (f) A/equine/Prague/56 (H7N7); (g)
A/Egypt/0636-NAMRU3/2007(H5N1); (h) A/New Caledonia/20/99 (H1N1); (i) A/South
Carolina/1/18 (H1N1).

[HenryN]

Yes, the Egypt-2 sequences, which have characteristics that match H1N1 include the sequences with the 3 BP deletion as well as the Gharbiya cluster.

[HenryN]

Originally Posted by niman

The isolates in the paper with the H1N1 characteristics have a 3 BP deletion, as did many of the mild cases in 2007.

It should be noted that the 3 BP deletion removes S129, and S129 is involved in receptor binding.

[HenryN]

Originally Posted by niman

The isolates in the paper with the H1N1 characteristics have a 3 BP deletion, as did many of the mild cases in 2007.

The mild cases in 2007 included the siblings from Qena who were hospitalized 4 days apart, signaling H2H. Both siblings (6F and 4M) had identical sequences, which included the 3 BP deletion.

[HenryN]

Commentary

http://www.recombinomics.com/News/04...t_S129del.html

[AlaskaDenise]

Originally Posted by niman

Commentary

http://www.recombinomics.com/News/04...t_S129del.html

Commentary

H5N1 Receptor Binding Domain Changes In Egypt

Recombinomics Commentary 04:27
April 15, 2009

Similarly the results of H5N1 strains from Egypt (Figure 2) may be indicative of a possible viral evolution towards receptor usage similar to that of H1N1 viruses, which efficiently replicate in the upper respiratory tract. The protein domain, which seems to be involved in this subtle change, corresponds to amino acid domain 99-132.

The above comments, from “Characterization of conserved properties of hemagglutinin of H5N1 and human influenza viruses: possible consequences for therapy and infection control” describe a subset of H5N1 sequences from Egypt that have chacteristics that match H1N1 seasonal flu. These matches include sequences from the Ghabiya cluster, as well as two Qena siblings with mild H5N1 in the spring of 2007. The sequences from the siblings had a 3 BP deletion that produce S129del, which is within the region listed above, Although the disease onset date of the index case was withheld from the WHO update, the hospital admission dates were four days apart, signaling the infection of the brother (4M) by his sister (6F). The sequences from the Qena siblings were identical, further supporting H2H transmission. Moreover, additional family members had symptoms, but tested negative.

This year there has been another cycle of mild cases in Egypt, but instead of affecting children between the ages of 3 and 10, like the siblings above, 10/12 cases are toddlers. However, like the cases in the spring of the 2007, the case fatality rate is low (in 2007 only 1/17 died, and this year 0/12 have died) and most patients did not develop pneumonia. These mild cases, coupled with the younger age group, raise concerns that H5N1 spread in the past two year was far more extensive than indicated by the confirmed cases, and were missed by current testing, which requires a poultry contact for H5N1 PCR testing.

Although NAMRU-3 has not released any human or poultry sequences this year, the first two human isolates were represented in a recent HA tree of vaccine targets. Although public sequences are not on the branch with the 2009 isolates, the location of the branch suggests these isolates also have the 3 BP deletion and are closely related to the sequences in the siblings from 2007.

The four day gap in admission dates for the Qena siblings, matches the four day gap in the disease onset dates for two cousins (both 2M) from Beheira. The cousins are next door neighbors, and the four day gap also indicated human to human transmission, further supporting the spread of mild H5N1.

The 3 BP deletion produces S129del and position 129 is in the receptor binding domain. This deletion maps to the region in H5N1 with similarities with H1N1, suggesting more efficient transmission, consistent with the clusters in Qena and Beheira.

Release of the sequences from the H5N1 cases this year would be useful. If these sequences have the same deletion as seen in the Qena cluster, an extensive screening of toddlers without a poultry connect is critical.

The failure to recognize the significance of the mild cases in 2007 is cause for concern. The failure to aggressively act at this time is beyond scandalous, and is hazardous to the world’s health.

.

[gsgs]

> We also found that about one third of H5N1 viruses which
> are isolated during the 2006/07 influenza outbreak in Egypt
> possibly evolve towards receptor usage similar to that of seasonal H1N1.

"1/3 possibly evolve as H1N1"

so they exclude this for the other 2/3 and don't really know (can't exclude,
"possibly") for the other 1/3


"evolves" seems to imply long chains of H2H2H--2H , possibly
with birds (swine?..) inbetween

[HenryN]

Originally Posted by gsgs

> We also found that about one third of H5N1 viruses which
> are isolated during the 2006/07 influenza outbreak in Egypt
> possibly evolve towards receptor usage similar to that of seasonal H1N1.

"1/3 possibly evolve as H1N1"

so they exclude this for the other 2/3 and don't really know (can't exclude,
"possibly") for the other 1/3


"evolves" seems to imply long chains of H2H2H--2H , possibly
with birds (swine?..) inbetween

The sequence profile matches H1N1, as described in the paper (which is open access).

[HenryN]

Commentary

http://www.recombinomics.com/News/04..._Pandemic.html