OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024

OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 1 OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 Acknowledgements This report was produced by the joint World Organisation for Animal Health (WOAH) and Food and Agricultural Organisation (FAO) network of scientific expertise on animal influenzas, OFFLU’s Avian Influenza Vaccination technical activity, Ian Brown (The Pirbright Institute), Ashley Banyard, Joe James, Joshua Linton Jenkins (Animal Health and Plant Agency), Francesco Bonefante (Istituto Zooprofilattico Sperimentale delle Venezie), Erica Spackman (US National Poultry Research Center, USDA-ARS), Frank Wong (CSIRO Australian Centre for Disease Preparedness), Nicola Lewis (Royal Veterinary College), Fabien Filaire and Amelia Coggon (FAO). Antigenic cartography was carried out by Amelia Coggon and Nicola Lewis (Royal Veterinary College). Sera and hemagglutination inhibition (HI) data were generated by the International Reference Laboratory for Avian Influenza team led by Joshua Lynton-Jenkins at the Animal Health and Plant Agency (United Kingdom) and Francesco Bonfante and Silvia Maniero at the Istituto Zooprofilattico Sperimentale delle Venezie (Italy). Haemagluttination inhibition (HI) data was provided by Erica Spackman and Jiho Lee at the US National Poultry Research Center, USDA-ARS (United States of America) and Frank Wong and Joanne Grimsey at the CSIRO Australian Centre for Disease Preparedness (Australia). Viral isolates were kindly shared with the International Reference Laboratories for Avian Influenza from the following countries: Bangladesh, Belgium, Bulgaria, Burkina Faso, Cambodia, Cameroon, Croatia, Czech Republic, Falkland Islands, Finland, Georgia, Ghana, from the Indonesian Archipelago, Iran, Ireland, Israel, Italy, Ivory Coast, Kosovo, Latvia, Moldova, Nepal, Niger, Nigeria, Norway, Philippines, Poland, Romania, Slovakia, Slovenia, Spain, Switzerland, South Georgia and South Sandwich Islands, Togo, Turkey, Ukraine, Viet Nam and the United Kingdom. CVVs were kindly shared by the GISRS network. We would like to thank all the OFFLU network experts, FAO and WOAH for their support with this project and Les Sims for his extensive contributions to this project. OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 2 Disclaimer: This technical report uses both published nomenclature (widely used by many countries at present (i.e. “GISAID clade”) 1 but not recently updated) AND unpublished nomenclature proposed by the WHO/FAO/WOAH H5 Nomenclature Working Group 2 (i.e. provisional Nextclade/LABEL clade). Throughout the report, the use of the different nomenclatures is discriminated by footnotes. This report provides the point of view of independent OFFLU experts and does not necessarily reflect the position of the parent organisations FAO and WOAH. Summary OFFLU has developed a system for providing information to countries on the antigenic characteristics of circulating high pathogenicity avian influenza (HPAI) A(H5Nx) viruses of the Goose/Guangdong/1/96-lineage. This process uses reference antisera produced in chickens against either representative isolates of relevant circulating clades or isolates that are genetic representatives of vaccine seed strains (hereinafter referred to as surrogate vaccine seed strains). Contemporary viruses are tested against these sera using the HI assay and antigenic cartography is applied on the HI titers to establish the antigenic relationship between the virus antigens and available antisera. This provides a numerical value for how closely matched the circulating viruses are to the antigens used to raise the reference antisera. Greater antigenic distances between surrogate vaccine antigens and currently circulating strains indicate evolution of viral sequences through mutations that lead to increased antigenic distance over time, known as antigenic drift. As poultry vaccines are primarily designed to provide protection against a specific antigen, this antigenic drift can impact the ability of vaccine derived antibodies to bind and/or neutralise contemporary viruses. Consequently, it can become necessary to update the antigens used in vaccine formulations. Results herein provide insight on this need. Further information on the background to the approach taken and the methods applied is available in Module 1: Background report and Module 2: Guidance report. This document provides the technical results. As of July 2024, the viruses causing the most outbreaks in poultry globally are subtype H5N1 within clade 2.3.4.4b. However, outbreaks have also been associated with other H5Nx clades, including clade 2.3.2.1a detected in South Asia, and clades 2.3.2.1c1/g2/e2 in Southeast Asia. Due to the dominating impact of these clades on poultry, the AIM assessment focused on defining the antigenic reactivity between the sampled field viruses in these clades and representative CVVs. Following HI assessment, antigenic cartography was undertaken using eight H5N1 clade 2.3.2.1a 1 Smith GJ, Donis RO, World Health Organization, World Organisation for Animal Health, Food Agriculture Organization, H5 Evolution Working Group. Nomenclature updates resulting from the evolution of avian influenza A(H5) virus clades 2.1.3.2a, 2.2.1, and 2.3.4 during 2013-2014. Influenza Other Respir Viruses 2015;9(5):271-6. Available at: http://onlinelibrary.wiley.com/doi/10.1111/irv.12324/epdf. Clade 2.3.2.1c viruses have evolved into different sublineages since they were last formally classified in 2014. An updated, standardised clade nomenclature for viruses that previously fell within clade 2.3.2.1c is being developed2 but has not yet been published. 2 Provisional nomenclature was taken from the WHO/FAO/WOAH H5 Nomenclature Working Group who define "clades" using HA gene sequences, and define clades as genetically distinct, monophyletic groups of viruses. This nomenclature splits clade 2.3.4.4 into eight additional sub-clades, named 2.3.4.4a through 2.3.4.4h due to high circulating diversity within the clade and. well as subclades 2.3.2.1a through 2.3.2.1g for the 2.3.2.1 split. OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 3 viruses collected between 2021 and 2023 from South Asia; one H5N1 clade 2.3.2.11 above1/g1 above virus collected in 2022 from Southeast Asia; 9 H5N8 clade 2.3.4.4b viruses collected since 2020 from Europe and Asia, 74 H5N1 clade 2.3.4.4b viruses collected since 2020 from Asia, Africa and Europe and the Americas; two H5N5 clade 2.3.4.4b virus collected in Europe since 2020 and one H5N3 virus collected in Europe in 2021. Results for viruses collected in 2023 are outlined and discussed in this report. Outcomes from the analyses undertaken within this period include the following points: • Vaccine antigens not within the clade 2.3.4.4 are antigenically distant from clade 2.3.4.4b viruses and would not be expected to be suitable candidate antigens for inactivated whole virus vaccines against these strains (figure 2). • Subtype-specific heterogeneity over previous years was noted in antigenic distances for clade 2.3.4.4b viruses (figure 3). • In geographically restricted regions, there is evidence of antigenic drift in viruses isolated from poultry (table 3). • Viruses tested from wild birds in Europe did not exhibit antigenic drift (table 3). • Clade 2.3.2.1 and clade 2.3.4.4 viruses are antigenically distinct from older virus clades including antisera raised against clade 1, clade 2.2, or clade 2.3.4 viruses (figure 2). This report indicates that, despite extensive circulation in wild birds and global spread, HPAI A(H5Nx) viruses from wild birds don’t appear to have undergone significant changes in antigenic properties. However, within poultry populations in geographically restricted areas HPAI A(H5Nx) viruses are evolving and drifting antigenically. Continued monitoring for genetic and antigenic changes, especially in countries where the virus is enzootic in poultry is crucial for early detection of antigenic change and response to update vaccines where necessary. OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 4 Vaccine seed strains Table 1: Seed strains of vaccines which are currently understood to be in use or have been used in the past. Where available clade information and references have been included. Clade information is described in the literature. Where sequences for the seed strain are available is noted by * and the nomenclature according to2 included if different. Seed Strain / HA Gene Source Clade1 Reference Represented A/Goose/Guangdong/96 0* Shi et al., 2022 No A/chicken/Vietnam/C58/04 1* EFSA 2023 Yes A/Vietnam/1194/2004 1* EFSA 2023 Yes A/Chicken/Shanxi/2/2006 7.2* Shi et al., 2022 No A/Chicken/Liaoning/S4092/2011 7.2 Shi et al., 2022 No A/chicken/Legok/2003 2.1.1 EFSA 2023 Pending A/CK/Egypt/ME1010/2016 2.2.1.1 EFSA 2023 Pending A/Chicken/Egypt/Q1995D/2010 2.2.1.2* EFSA 2023 Pending A/Chicken/Egypt/RG-173 CAL/2017 2.2.1.2 EFSA 2023 Pending A/Duck/EGYPT/M2583D/2010 2.2.1.2* EFSA 2023 Pending A/chicken/West Java/Pwt-Wij/2006 2.3.1.2 EFSA 2023 No A/duck/Sukoharjo/BBVW-14289/2012 2.3.2.1g2* Indriani et al., 2014 Yes A/Hubei/1/2010 2.3.2.1a2* EFSA 2023 Pending A/duck/Guangdong/S1322/2010 2.3.2.1b2* Shi et al., 2022 Yes A/chicken/Vietnam/NCVD-KA435/13 2.3.2.1c1/e2* EFSA 2023 Pending A/chicken/Tanggamus/03171107665/2017 2.3.2.1c EFSA 2023 No A/chicken/Liaoning/SD007/2017 2.3.2.1d Shi et al., 2022 Pending rgCA2/2.3.2.1d 2.3.2.1d Kang et al., 2022 No A/duck/Anhui/SI246/2014 2.3.2.1 Shi et al., 2022 No A/Duck/Anhui/1/2006 2.3.4* Shi et al., 2022 Yes A/chicken/Guizhou/4/2013 2.3.4.4/g2* Shi et al., 2022 No A/duck/Korea/ES2/2016 2.3.4.4/e2* EFSA 2023 Pending A/Waterfowl/Korea/S57/2016 2.3.4.4 Kuruppuarachchi et al., 2022 No A/Gyrfalcon/WA/41088-6/2014 2.3.4.4c2* EFSA 2023 Yes A/chicken/Egypt/ME-2018/2018 2.3.4.4b2* Yes A/green-winged teal/Egypt/877/2016 2.3.4.4b2* EFSA 2023 Yes A/whooper swan/Shanxi/4–1/2020 2.3.4.4b2* Shi et al., 2022 Yes A/duck/Guizhou/S4184/2017 2.3.4.4h Shi et al., 2022 Pending A/duck/Fujian/S1424/2020 2.3.4.4h Shi et al., 2022 Pending rgES3/2.3.4.4h 2.3.4.4h Kang et al., 2022 No A/duck/Guanzou/S4184/2017 2.3.4.4h Shi et al., 2022 Pending A/Duck/VietNam/QB7412 unknown EFSA 2023 No OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 5 Panel of chicken antisera used in OFFLU AIM Table 2: Viruses which were used in OFFLU aim to generate sera. Clade* provisional nomenclature was taken from the WHO/FAO/WOAH H5 Nomenclature Working Group who define "clades" using HA gene sequences, and define clades as genetically distinct, monophyletic groups of viruses. This nomenclature splits clade 2.3.4.4 into eight additional sub-clades, named 2.3.4.4a through 2.3.4.4h due to high circulating diversity within the clade and. well as sub-clades 2.3.2.1a through 2.3.2.1g for the 2.3.2.1 split. Strain Subtype Clade1 Clade2 Similar vaccine seed strain A/Vietnam/1194/2004/1 H5N1 1 1 A/chicken/Vietnam/C58/04 A/Turkey/turkey/2005 H5N1 2.2 2.2.1 A/swan/Hungary/4999/2006 A/Anhui/1/2005 H5N1 2.3.4 2.3.4 A/Duck/Anhui/1/2006 A/mynah/Indonesia/13064792-010/2013 H5N1 2.3.2.1c 2.3.2.1g A/duck/Sukoharjo/BBVW-14289/2012 A/chicken/Nepal/T360/2014 H5N1 2.3.2.1a 2.3.2.1a A/duck/Guangdong/S1322/2010 A/Mute_Swan/Croatia/102/2016 H5N8 2.3.4.4b 2.3.4.4b A/green-winged teal/Egypt/877/2016 A/mallard/Georgia/DT09382/2017 H5N8 2.3.4.4b 2.3.4.4b A/chicken/ME-2018 A/chicken/Czech Republic/1175-1/2020 H5N8 2.3.4.4b 2.3.4.4b A/chicken/Bulgaria/722-1_22VIR778-1/2021 H5N1 2.3.4.4b 2.3.4.4b A/duck/Cambodia/f4k241D3/2021 H5N8 2.3.4.4b 2.3.4.4b A/whooper swan/Shanxi/4–1/2020 A/great_skua/Scotland/B07779/2021 H5N1 2.3.4.4b 2.3.4.4b A/gyrfalcon/Washington/41088/6/2014 H5N8 2.3.4.4c 2.3.4.4c A/Gyrfalcon/WA/41088-6/2014 OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 6 Phylogenetic relationships of viruses included in the OFFLU AIM study Figure 1: A maximum likelihood phylogenetic tree of the HA1 of Gs/Gd lineage H5 viruses included in this project and vaccine seed strains where sequences were available. Leaves are coloured by test antigens in this study, vaccine seed strains, antisera used in this study and antisera and vaccine seed strains used in this study. Major clades are annotated along the branches according to nomenclature, minor clade are not annotated2. This image was generated in Auspice using Nextclade. OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 7 Antigenic cartography Antigenic cartography was undertaken to quantify and visualise the antigenic distances between viruses representative of vaccine seed strains and contemporary circulating viruses as described in the AIM pilot project. Maps were analysed and visualised using R Studio version 2023.03.1+446 and the Racmacs package version 1.2.9 built under R version 4.3.1 as described by Smith et al., (2004) and previously used in Lewis et al., (2021). For information regarding map generation and interpretation please contact an appropriate reference laboratory. Complementary genetic analysis was carried out for map testing using manually curated datasets with methods described as in the OFFLU avian data package for zoonotic influenza component of the VCM. Amino acid changes in the HA1 were visualised by reconstructing ancestry using treetime version 0.11.1 (Sagulenko et al., 2018) and were compared between within-clade and within-subtype test antigens. Antigenic maps were colored according to unpublished viral clade nomenclature and subtype using the H5Nx dataset in Nextclade (Aksamentov et al., 2021). OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 8 Figure 2: A 3-dimensional antigenic map showing the evolutionary relationships of H5 HPAI Gs/Gd lineage viruses. Each antiserum is represented by a red cube, for vaccine seed strains or surrogates and black cube for other. Antigens are represented as balls and are coloured by clade 1 or 2 according to the key. Each square represents one antigenic unit. One antigenic unit is representative of a 2-fold difference in HA assay titer. The corresponding table of antigenic distances can be found in table 3. OFFLU Avian Influenza Matching (AIM) Technical Report, July 2024 9 Figure 3: A 3-dimensional antigenic map showing the evolutionary relationships of H5 HPAI Gs/Gd lineage viruses of the clade 2.3.4.4. Each antiserum is represented by a red cube, for vaccine seed strains or surrogates and black cube for other. Antigens are represented as balls and are coloured by clade 1 or 2 according to the key. Each square represents one antigenic unit. One antigenic unit is representative of a 2-fold difference in HA assay titer. The corresponding table of antigenic distances can be found in table 3.