Influenza virus characterisation - Summary Europe, August and September 2011 (ECDC, October 7 2011, extract, edited)
Influenza virus characterisation - Summary Europe, August and September 2011 (ECDC, October 7 2011, extract, edited)
[Source: European Centre for Disease Prevention and Control (ECDC), full PDF document: (
LINK). Extract, edited.]
TECHNICAL DOCUMENT
Community Network of Reference Laboratories (CNRL) for Human Influenza in Europe
Influenza virus characterisation - Summary Europe, August and September 2011
Influenza virus characterisation
Summary
- Influenza A(H1N1)pdm09, influenza A(H3N2), and influenza B/Victoria- and B/Yamagata-lineage viruses have been characterised genetically and antigenically.
- A(H1N1)pdm09 viruses collected since 1 February 2011 fall into several genetic groups but all groups show antigenic similarity to the currently recommended vaccine virus A/California/7/2009.
- A(H3N2) viruses collected since 1 February 2011 fall into distinct genetic groups. There is no consistent correlation of altered antigenicity compared to the vaccine virus, A/Perth/16/2009, with any genetic group.
- Influenza B viruses of the B/Victoria/2/87 lineage have predominated over those of the B/Yamagata/16/88 lineage. Most of the B/Victoria/2/87 lineage viruses remain genetically similar to the currently recommended vaccine virus B/Brisbane/60/2008 and antigenically similar to cell-propagated viruses that are genetically closely related to the vaccine virus B/Brisbane/60/2008.
- The majority of influenza B viruses of the B/Yamagata-lineage are in the B/Bangladesh/3333/2007 genetic clade.
Table 1 provides an updated summary since the last influenza virus characterisation report on the influenza viruses collected between February and August 2011 in ECDC-affiliated countries and received at the WHO CC in London.
As noted previously, viruses were predominantly of type A(H1N1)pdm09 and type B of the B/Victoria/2/87 lineage, although type A(H3N2) viruses and type B viruses of the B/Yamagata/16/88 lineage have continued to be received. In Table 1, batches of specimens for which analysis has yet to be completed are shown as in progress.
Influenza A(H1N1)pdm09 virus analysis
Table 2 shows HI results for viruses recovered and analysed since the previous report; these viruses were from Slovenia and Portugal. The majority of viruses continued to react well with the panel of post-infection ferret antisera, including antiserum raised against the vaccine virus A/California/7/2009.
Two viruses, A/Slovenia/808/2011 and A/Slovenia/861/2011, showed more than a four-fold reduction in HI titre with the antiserum raised against the vaccine virus, A/California/7/2009, compared to the titre with the vaccine virus itself.
An updated representative phylogenetic tree of the HA gene is shown in Figure 1. The phylogenetic tree shows residues that define genetic groups predominant over the last six months; residues in black are located in the HA1 polypeptide chain, those in brown indicate residues in HA2.
Certain sporadic amino acid substitutions or polymorphisms are also shown: K154X, G155X/E and N156D are frequently associated with low reactivity in HI assays and commonly result from propagation of viruses in certain tissue culture cells as they are observed rarely in clinical samples.
The sequence change of D222G, or the polymorphism D222X, is postulated to be detected more often in viruses recovered from patients suffering with severe disease.
The amino acid substitution Q223R is associated with the isolation and passage of virus in hens? eggs.
We have described at least six genetic groups for the circulating A(H1N1)pdm09 viruses. These groups can be defined by amino acid substitutions and are numbered groups 2 to 7:
- Group 2:
- N31D, S162N (adding a glycosylation site) and A186T in HA1, e.g. A/Czech Republic/32/2011;
- Group 3:
- A134T and S183P, e.g. A/Toulon/1173/2011 with some viruses also carrying the substitutions A141S, I295V in HA1 and V152A in HA2, e.g. A/Umea/1/2011
- Group 4:
- N125D in HA1, observed originally as an emerging genetic group in the Southern Hemisphere e.g. A/Christchurch/16/2010;
- Group 5:
- D97N, R205K, I216V and V249L in HA1, e.g. A/Astrakhan/1/2011;
- Group 6:
- D97N and S185T in HA1 and S124N in HA2, e.g. A/St Petersburg/27/2011;
- Group 7:
- S143G, S185T and A197T in HA1 and S124N in HA2, e.g. A/St Petersburg/100/2011.
The sequence of the HA gene of A/Slovenia/861/2011, one of the pair of viruses with reduced reactivity against the panel of sera, was determined.
The virus fell into genetic group 7 but carried four additional amino acid substitutions in the HA1 coding region that might account for its unusual reactivity in the HI test.
Viruses collected since February 2011 in EU and EU-affiliated countries fall into each of the genetic groups. None of the groups shows a consistently marked change in antigenicity compared with the vaccine virus.
Influenza A(H3N2) virus analysis
For specimens collected since February 2011, influenza A(H3N2) viruses have been successfully isolated and propagated from seven ECDC-affiliated countries. The difficulties with antigenic characterisation of recent H3N2 viruses have been described previously. Shown in Table 3 are the results of HI assays using guinea pig red blood cells in the presence of oseltamivir to reduce any effect of the virus neuraminidase on the agglutination of the red blood cells (Lin et al. 2010) for three recently received viruses from Norway.
The results showed that one of the viruses tested yielded an eight-fold reduction in titre with the post-infection ferret antiserum raised against the vaccine virus A/Perth/16/2009 compared with the homologous reaction between the antiserum and the vaccine virus, and one of the three viruses showed a four-fold reduction.
All three test viruses showed good reactivity with antisera raised against A/Alabama/5/2010, A/Stockholm/18/2011 and A/South Australia/3/2011, three reference viruses propagated in tissue culture cell lines.
A phylogenetic tree of the HA1 coding region of the HA gene of representative A(H3N2) viruses is shown in Figure 2. Ten reference viruses and the vaccine virus A/Perth/16/2009 used in the HI test are highlighted.
The majority of viruses from ECDC-affiliated countries continue to fall within the A/Victoria/208/2009 genetic clade and a minority in the A/Perth/16/2009 clade. Distinct genetic groups within the HA gene are seen within both genetic clades.
Seven genetic groups can be described in recently circulating A(H3N2) viruses.
Within the A/Perth/16/2009 genetic clade there are two genetic groups (1 and 2) defined by amino acid substitutions:
- Group 1:
- I260M, R261Q, e.g. A/Victoria/210/2009 with some viruses also carrying P162S and substitutions of either E50K, e.g. A/Norway/1186/2011, or N81D, e.g. A/Niigata/510/2011;
- Group 2:
- N133D (resulting in the loss of a glycosylation site), R142G, T212A and V213A, e.g. A/Norway/1330/2010. This group has not been shown in Figure 2 since very few of these viruses have been circulating in recent months.
There are at least four genetic groups in the A/Victoria/208/2009 genetic clade of A(H3N2) viruses. These are defined by additional amino acid substitutions to those that define viruses of the A/Victoria/208/2009 clade compared with the A/Perth/16/2009 clade (K62E, K144N resulting in the gain of a glycosylation site and T212A):
- Group 3:
- V223I, with some viruses having the substitution N144D that results in the loss of a glycosylation site, and N145S e.g. A/Stockholm/18/2011; and another group with the substitution A198S with either N145S, e.g. A/Norway/1762/2011, or the substitutions S45N (resulting in the gain of a glycosylation site), T48I and N312S, e.g. A/Slovenia/537/2011;
- Group 4:
- N312S, with many viruses also carrying T48A and K92R, e.g. A/Iraq/7/2011;
- Group 5:
- D53N, Y94H, I230V and E280A, e.g. the reference viruses A/Alabama/05/2010 and A/Perth/10/2010;
- Group 6:
- D53N, Y94H, S199A, I230V and E280A e.g. A/Iowa/19/2010 and A/Ireland/M27357/2011; with some encoding an additional glycosylation site as a result of S45N substitution e.g. A/Hong Kong/3951/2011.
A further genetic group can be considered but is not presently numbered:
- S45N, which results in an additional glycosylation site, e.g. A/Norway/685/2011.
The viruses shown in Table 3, all from Norway and collected in July or August, fall into genetic group 3.
Viruses from genetic groups 1, 3, 5 and 6, as well as the unnumbered group, have been collected in ECDC-affiliated countries since February 2011. Analysis of viruses from each of the emerging genetic groups has indicated that no group shows consistent significant change in antigenicity compared with the vaccine virus, A/Perth/16/2009.
Influenza B virus analyses
Influenza B viruses of the B/Victoria/2/87 lineage (83%) have continued to predominate over those of the B/Yamagata/16/88 lineage (17%).
B/Victoria-lineage viruses
Antigenic analysis of influenza B/Victoria-lineage viruses recently received and propagated is shown in Table 4 ? all the viruses were from Slovenia.
In the HI assay all showed low reactivity with antiserum raised against B/Brisbane/60/2008, the egg-propagated vaccine virus. As described previously, HI assays for influenza B/Victoria lineage viruses propagated only in cells frequently show reduced HI titres when tested with antisera raised against egg-propagated reference viruses, including vaccine viruses (Schild et al. 1983 and see also previous ECDC/CNRL Influenza Virus Characterisation Technical Reports). As a consequence, the antigenic properties of cell-propagated viruses are assessed with antisera raised against viruses which are genetically closely related to the vaccine virus but propagated in cells.
In Table 4 the cell-propagated reference viruses B/Paris/1762/2008, B/Hong Kong/514/2009 and B/Odessa/3886/2010 are genetically closely related to the vaccine virus B/Brisbane/60/2008. All the test viruses analysed in Table 4 showed good reactivity with antisera raised against these three reference viruses, hence they are considered to be antigenically similar to the vaccine virus.
Figure 3 shows a phylogenetic tree based on the HA1-coding region of the HA gene of selected viruses of the B/Victoria-lineage. Amino acid substitutions N75K, N165K and S172P define the B/Brisbane/60/2008 genetic clade. All recently collected influenza B viruses of the B/Victoria-lineage from EU and EU-affiliated countries come within the B/Brisbane/60/2008 genetic clade. The majority of viruses carry the amino acid substitution I146V in the HA compared with the vaccine virus B/Brisbane/60/2008 and many also carry the substitution L58P. Neither substitution has a marked affect on the antigenicity of the viruses.
B/Yamagata-lineage viruses
No new antigenic results have been generated for B/Yamagata-lineage viruses since our last report. Figure 4 shows a phylogenetic tree based on the HA1-coding region of the HA gene of selected viruses of the B/Yamagata-lineage. Amino acid substitutions S150I, N165Y and G229D define the B/Bangladesh/3333/2007 genetic clade.
The majority of recently collected viruses are in the B/Bangladesh/3333/2007 genetic clade and can be divided into several genetic sub-groups. A cluster of viruses from Sweden, Finland and Estonia, coming within a distinct genetic clade with B/Brisbane/3/2007 as the clade-prototype virus, was described in our previous report.
Note to the figures
The phylogenetic trees were constructed using neighbour-join in MEGA4. The bars indicate the proportion of nucleotide changes in the sequence. Reference strains are viruses against which post-infection ferret antisera have been raised. The colours indicate the date of sample collection. Isolates from ECDC countries are highlighted in yellow. Sequences for some of the viruses from non-European countries were recovered from GISAID and we acknowledge all laboratories who submitted sequences directly to the London WHO CC.
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