Thursday, May 16, 2019

ECDC: Influenza Virus Characterisation, April 2019


Every month during the Northern Hemisphere's flu season - and slightly less often over the summer - the ECDC  publishes a review of recently isolated seasonal flu viruses collected across the EU in their Influenza Virus Characterization Report.
There are 4 known types of flu viruses (A, B, C & D), but of these only A & B are considered significant contributors to our flu seasons. Influenza A - which is divided into two subtypes (H1N1 & H3N2) - and Influenza B, which has two Lineages (B/Yamagata & B/Victoria). 
Complicating matters, within each of these four categories there are a growing number of continually evolving variants and/or subclades - all juggling, not only for survival, but for dominance on the world stage.

While A/H1N1 and both Influenza B lineages remain reasonably stable in comparison, the A/H3N2 subtype - which has been in continual circulation since 1968 - has evolved into numerous co-circulating subclades.
H3N2 is not only the longest continually circulating Influenza A strain on record (50+ years), it has survived the arrival of two new H1N1 viruses (1977 and 2009), neither of which were able to supplant the H3 juggernaut. 
All flu seasons bring a certain degree uncertainty, but over the past few years this growing diversity in H3N2 viruses has added additional complexity to the twice annual selection of flu vaccine strains.

So much so, that last February - when the WHO normally decides on which strains to put in next fall's vaccine - they opted to delay their decision on the H3N2 component for 30 days (see WHO: (Partial) Recommended Composition Of 2019-2020 Northern Hemisphere Flu Vaccine).
At issue was the sudden rise of H3N2 clade 3C.3a reported in the United States (and other places), which had started last fall's season as a minor component of what appeared on track to being a relatively mild H1N1 season.
By early 2019 we'd switched into a moderately severe H3N2 season with clade 3C.3a leading the pack (see CDC HAN #0418: Influenza Season Continues with an Increase in Influenza A(H3N2) Activity).

Making matters worse, in other regions of the world, different antigenically distinct H3N2 clades continue strong, making the selection of next fall's H3N2 vaccine component even more difficult.

The most recent ECDC Influenza Characterisation (April 2019) report lists the current H3N2 players on the field:
HA gene sequences of the test viruses characterised antigenically in the March 2019 report are now available andthe genetic clades are shown in Tables 4-1 to 4-3 and most are included in the HA phylogenetic analysis (Figure 2). Viruses in clades 3C.2a and 3C.3a have been in circulation since the 2013–14 northern hemisphere influenza season, with clade 3C.2a viruses having been dominant since the 2014–15 influenza season, notably subclade 3C.2a2 viruses, though subgroup 3C.2a1b viruses have predominated in recent months (Figure 2).
The HA gene sequences of viruses in both clades continue to diverge. Notably, clade 3C.3a viruses have evolved to carry HA1 amino acid substitutions of L3I, S91N, N144K (loss of a N-linked glycosylation motif at residues 144-146), F193S and K326R, compared to A/Stockholm/6/2014, and levels of detection since January 2019 have been increasing in a number of WHO European region countries (Figure 2) and North America. New genetic groups have also emerged among the clade 3C.2a viruses, designated as subclades/subgroups. Amino acid substitutions that define these subclades/subgroups are:
Clade 3C.2a: L3I, N144S (resulting in the loss of a potential glycosylation site), F159Y, K160T (in the majority of viruses, resulting in the gain of a potential glycosylation site) and Q311H in HA1, and D160N in HA2, e.g. A/Hong Kong/7295/2014 a cell culture-propagated surrogate for A/Hong Kong/4801/2014 (a former vaccine virus)

Subclade 3C.2a1: those in clade 3C.2a plus: N171K in HA1 and I77V and G155E in HA2, most also carry N121K in HA1, e.g. A/Singapore/INFIMH-16-0019/2016 (2018–19 northern hemisphere vaccine virus)

Subgroup 3C.2a1a: those in subclade 3C.2a1 plus T135K in HA1, resulting in the loss of a potential glycosylation site, and also G150E in HA2, e.g. A/Greece/4/2017

Subgroup 3C.2a1b: those in subclade 3C.2a1 plus K92R and H311Q in HA1, e.g. A/La Rioja/2202/2018, with many viruses in this subgroup carrying additional HA1 amino acid substitutions

Subclade 3C.2a2: those in clade 3C.2a plus T131K, R142K and R261Q in HA1, e.g. A/Switzerland/8060/2017 (2019 southern hemisphere vaccine virus)

Subclade 3C.2a3: those in clade 3C.2a plus N121K and S144K in HA1, e.g. A/Cote d’Ivoire/544/2016

Subclade 3C.2a4: those in clade 3C.2a plus N31S, D53N, R142G, S144R, N171K, I192T, Q197H and A304T in HA1 and S113A in HA2, e.g. A/Valladolid/182/2017

Clade 3C.3a: T128A (resulting in the loss of a potential glycosylation site), R142G and N145S in HA1 which defined clade 3C.3 plus A138S, F159S and N225D in HA1, many with K326R, e.g. A/England/538/2018.
Globally, the great majority of viruses with collection dates from 1 September 2018 have HA genes that continue to fall into genetic groups within clade 3C.2a, with those in subgroup 3C.2a1b having been more numerous than those in subclade 3C.2a2 for the period September 2018 to March 2019 (Figure 2).
Notably, a significant number of the subgroup 3C.2a1b viruses have fallen in two recently emerged clusters. One defined by amino acid substitutions T131K in HA1 with V200I in HA2 and the other by T128A and T135K substitutions in HA1 (both resulting in loss of potential glycosylation sequons). Further, as indicated above, numbers of clade 3C.3a virus detections have been increasing in recent weeks in a number of countries/regions.
In late March the WHO decided to switch to the surging Clade 3C.3a H3N2  virus, betting that it will become the dominant H3 strain worldwide by next fall.  Meanwhile, Australia - which is already seeing record setting flu activity during their summer  (see chart below) - will be using last year's selected H3N2 vaccine candidate (Subclade 3C.2a2).

As if things weren't complicated enough, over the past 5 years H3N2 viruses have become very difficult to analyze, as explained by the ECDC below:
As described in many previous reports, influenza A(H3N2) viruses have continued to be difficult to characterise antigenically by HI assay due to variable agglutination of red blood cells (RBCs) from guinea pigs, turkeys and humans, often with the loss of ability to agglutinate any of these RBCs. As was first highlighted in the November 2014 report 3 , this is a particular problem for most viruses that fall in genetic clade 3C.2a.
All of which brings us to the latest highly detailed ECDC characterization report (PDF File).  I've reproduced the Executive Summary below, but this is only a snapshot from a much larger report.
As you will see, H1N1 viruses in circulation continue to appear to be a good match to the vaccine strain, but most of the H3N2 viruses that they tested were poorly recognized by the existing flu vaccine. 
I'll have a bit more after the break. 
This is the sixth report for the 2018–19 influenza season. As of week 18/2019, 203 585 influenza detections across the WHO European Region had been reported. Detections were 99% type A viruses, with A(H1N1)pdm09 prevailing over A(H3N2), and 1% type B viruses, with 80 (65%) of 124 ascribed to a lineage being B/Yamagata-lineage.

Executive summary

Since the March 2019 characterisation report1, a further shipment of influenza-positive specimens from an EU/EEA country was received at the London WHO CC, the Francis Crick Worldwide Influenza Centre (WIC). A total of 1 057 virus specimens, with collection dates after 31 August 2018, have been received.
All 59 A(H1N1)pdm09 test viruses characterised antigenically since the March 2019 characterisation report showed good reactivity with antiserum raised against the 2018–19 vaccine virus, A/Michigan/45/2015 (clade 6B.1). The 391 test viruses with collection dates from week 40/2018 genetically characterised at the WIC, including an H1N2 reassortant, all fell in a 6B.1 subclade, designated 6B.1A, defined by HA1 amino acid substitutions of S74R, S164T and I295V. Of these recently circulating viruses, 355 also have HA1 S183P substitution, often with additional substitutions in HA1 and/or HA2.
Since the last report, only 26 A(H3N2) viruses successfully recovered had sufficient HA titre to allow antigenic characterisation by HI assay in the presence of oseltamivir. These viruses were poorly recognised by antisera raised against the currently used vaccine virus, egg-propagated A/Singapore/INFIMH-16-0019/2016, in HI assays. Of the 321 viruses with collection dates from week 40/2018 genetically characterised at the WIC, 267 were clade 3C.2a (with 32 3C.2a2, 13 3C.2a3, six 3C.2a4 and 216 3C.2a1b) and 54 were clade 3C.3a.
No B/Victoria-lineage viruses were characterised in this reporting period. All recent viruses carry HA genes that fall in clade 1A but encode HA1 amino acid substitutions of I117V, N129D and V146I compared to a previous vaccine virus, B/Brisbane/60/2008. Groups of viruses defined by deletions of two (Δ162-163, 1A(Δ2)) or three (Δ162-164, 1A(Δ3)) amino acids in HA1 have emerged, with the triple deletion group having subgroups of Asian and African origin. HI analyses with panels of post-infection ferret antisera have shown these virus groups to be antigenically distinguishable. Of a total of five viruses characterised from EU/EEA countries this season, one has been Δ162-163 and four Δ162-164 (three African and one Asian subgroup).
Including the two B/Yamagata-lineage viruses reported on here, a total of 11 from the 2018–19 season have been characterised. All have HA genes that fall in clade 3 and encode HA1 amino acid substitutions of L172Q and M251V compared to the vaccine virus B/Phuket/3073/2013 but remain antigenically similar to the vaccine virus recommended for use in quadrivalent vaccines for current and subsequent northern hemisphere influenza seasons.
As the number of viral players increase, so do the number of possible outcomes.  Among the known unknowns, we are waiting to see:
  • Which H3N2 strain dominates the Southern Hemisphere flu season, and how well the existing vaccine handles it
  • Whether clade 3C.3a continues to rise globally, as it has in the United States
  • Whether (or how much) the delay in picking a vaccine strain will impact the delivery of flu shots next fall
  • If the 2019-2020 flu season is dominated by H3N2 or H1N1
  • Whether H3N2 clade 3C.3a persists throughout next year's flu season, or - as we've seen this season - is overtaken by something else
  • And finally, how well next fall's vaccine will work against the (then) currently circulating strains
As our global society becomes increasingly mobile in this 21st century, so do the viruses we carry. While this has obvious pandemic implications, it also makes seasonal flu more volatile, complex, and unpredictable. 

As influenza viruses become more antigenically diverse, the job of picking which virus to include in next season's flu vaccine will only become more difficult.

Credit NIAID
All of which makes the development of a`universal' flu vaccine (see J.I.D.: NIAID's Strategic Plan To Develop A Universal Flu Vaccine) of greater importance than ever. 

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