Highly pathogenic avian influenza H5N1 virus outbreak among common terns (Sterna hirundo) in Namibia, 2025-2026
DOI:
https://doi.org/10.12834/VetIt.4031.40792.4Keywords:
Avian influenza, Namibia, Common terns, H5N1, Clade 2.3.4.4bAbstract
Highly pathogenic avian influenza A(H5N1) viruses of clade 2.3.4.4b continue to spread globally, causing major outbreaks in wild birds and poultry. In Africa, however, genomic data remain limited, restricting understanding of viral introduction routes and circulation patterns. Here, we report the whole-genome characterisation of an HPAI A(H5N1) virus detected in a common tern (Sterna hirundo) found dead on the Namibian coast during the most recent avian influenza outbreak recorded in the country. Viral RNA was subjected to whole-genome sequencing using the Illumina Viral Surveillance Panel v2 on a NextSeq 1000 platform. Complete or near-complete sequences were obtained for all eight genome segments and deposited in GenBank. Phylogenetic analyses, performed using African clade 2.3.4.4b H5Nx sequences and the closest related sequences identified through database searches, showed that the Namibian virus belonged to clade 2.3.4.4b and clustered within the EA-2024-DI.2 subgenotype. Across all segments, the virus grouped with contemporary European EA-2024-DI.2 viruses circulating during the 2024–2025 epidemic wave, supporting a likely Eurasian origin. For six of the eight segments, it also clustered closely with an EA-2024-DI.2 virus detected in a gull-billed tern in Uganda in December 2024. Molecular analysis identified a polybasic haemagglutinin cleavage site consistent with high pathogenicity and a mutational profile broadly similar to contemporary EA-2024-DI.2 viruses. The HA substitution, associated in previous studies with increased binding to mammalian-type α2-6 receptors, may warrant further investigation. These findings highlight the role of migratory seabirds in H5N1 dissemination and reinforce the need for strengthened genomic surveillance in African wild birds and poultry.References
Abolnik C. (2025). Avian influenza situation report-Africa. Canadian journal of microbiology, 71, 1–4. https://doi.org/10.1139/cjm-2025-0199
Ayllon, J., Domingues, P., Rajsbaum, R., Miorin, L., Schmolke, M., Hale, B. G., & García-Sastre, A. (2014). A single amino acid substitution in the novel H7N9 influenza A virus NS1 protein increases CPSF30 binding and virulence. Journal of virology, 88(20), 12146–12151. https://doi.org/10.1128/JVI.01567-14
EFSA AHAW Panel (EFSA Panel on Animal Health and Animal Welfare), ECDC, Alvarez, J., Boklund, A., Dippel, S., Dórea, F., Figuerola, J., Herskin, M. S., Michel, V., Miranda Chueca, M. Á., Nannoni, E., Nielsen, S. S., Nonno, R., Riber, A. B., Stegeman, J. A., Ståhl, K., Thulke, H.-H., Tuyttens, F., Winckler, C., Brugerolles, C., … Melidou, A. (2025). Preparedness, prevention and control related to zoonotic avian influenza. EFSA Journal, 23(1), e9191. https://doi.org/10.2903/j.efsa.2025.9191
Elgendy, E. M., Arai, Y., Kawashita, N., Daidoji, T., Takagi, T., Ibrahim, M. S., Nakaya, T., & Watanabe, Y. (2017). Identification of polymerase gene mutations that affect viral replication in H5N1 influenza viruses isolated from pigeons. The Journal of general virology, 98(1), 6–17. https://doi.org/10.1099/jgv.0.000674
European Food Safety Authority, European Union Reference Laboratory for Avian Influenza, Ducatez, M., Fusaro, A., Gonzales, J. L., Kuiken, T., Ståhl, K., Staubach, C., Terregino, C., & Kohnle, L. (2025). Unprecedented high level of highly pathogenic avian influenza in wild birds in Europe during the 2025 autumn migration. EFSA journal. European Food Safety Authority, 23(11), e9811. https://doi.org/10.2903/j.efsa.2025.9811
Fan, S., Deng, G., Song, J., Tian, G., Suo, Y., Jiang, Y., Guan, Y., Bu, Z., Kawaoka, Y., & Chen, H. (2009). Two amino acid residues in the matrix protein M1 contribute to the virulence difference of H5N1 avian influenza viruses in mice. Virology, 384(1), 28–32. https://doi.org/10.1016/j.virol.2008.11.044
Fasina, F. O., Abolnik, C., Meseko, C. A., Carapeto, S. O., Palamara, E., Brun, M., et al. (2025). Recent updates on high pathogenicity avian influenza in sub-Saharan Africa (2019–present). EMPRES-Animal Health, 360, 49. Food and Agriculture Organization of the United Nations. https://openknowledge.fao.org/server/api/core/bitstreams/4f3a6938-bb26-4450-b847-96027b1cfec0/content
Feng, X., Wang, Z., Shi, J., Deng, G., Kong, H., Tao, S., Li, C., Liu, L., Guan, Y., & Chen, H. (2015). Glycine at Position 622 in PB1 Contributes to the Virulence of H5N1 Avian Influenza Virus in Mice. Journal of virology, 90(4), 1872–1879. https://doi.org/10.1128/JVI.02387-15
Fusaro, A., Zecchin, B., Giussani, E., Palumbo, E., Agüero-García, M., Bachofen, C., Bálint, Á., Banihashem, F., Banyard, A. C., Beerens, N., Bourg, M., Briand, F. X., Bröjer, C., Brown, I. H., Brugger, B., Byrne, A. M. P., Cana, A., Christodoulou, V., Dirbakova, Z., Fagulha, T., … Monne, I. (2024). High pathogenic avian influenza A(H5) viruses of clade 2.3.4.4b in Europe-Why trends of virus evolution are more difficult to predict. Virus evolution, 10(1), veae027.
https://doi.org/10.1093/ve/veae027
Gao, W., Zu, Z., Liu, J., Song, J., Wang, X., Wang, C., Liu, L., Tong, Q., Wang, M., Sun, H., Sun, Y., Liu, J., Chang, K. C., & Pu, J. (2019). Prevailing I292V PB2 mutation in avian influenza H9N2 virus increases viral polymerase function and attenuates IFN-β induction in human cells. The Journal of general virology, 100(9), 1273–1281. https://doi.org/10.1099/jgv.0.001294
Gao, Y., Zhang, Y., Shinya, K., Deng, G., Jiang, Y., Li, Z., Guan, Y., Tian, G., Li, Y., Shi, J., Liu, L., Zeng, X., Bu, Z., Xia, X., Kawaoka, Y., & Chen, H. (2009). Identification of amino acids in HA and PB2 critical for the transmission of H5N1 avian influenza viruses in a mammalian host. PLoS pathogens, 5(12), e1000709. https://doi.org/10.1371/journal.ppat.1000709
Grubaugh ND, Gangavarapu K, Quick J, Matteson NL, De Jesus JG, Main BJ, Tan AL, Paul LM, Brackney DE, Grewal S, Gurfield N, Van Rompay KKA, Isern S, Michael SF, Coffey LL, Loman NJ, Andersen KG. An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. Genome Biol. 2019 Jan 8;20(1):8. doi: 10.1186/s13059-018-1618-7. PMID: 30621750; PMCID: PMC6325816.
Giussani, E., Sartori, A., Salomoni, A., Cavicchio, L., De Battisti, C., Pastori, A., Varotto, M., Zecchin, B., Hughes, J., Monne, I., & Fusaro, A. (2025). FluMut: A tool for mutation surveillance in highly pathogenic H5N1 genomes. Virus Evolution, 11(1), veaf011. https://doi.org/10.1093/ve/veaf011
Guo, H., de Vries, E., McBride, R., Dekkers, J., Peng, W., Bouwman, K. M., Nycholat, C., Verheije, M. H., Paulson, J. C., van Kuppeveld, F. J., & de Haan, C. A. (2017). Highly Pathogenic Influenza A(H5Nx) Viruses with Altered H5 Receptor-Binding Specificity. Emerging infectious diseases, 23(2), 220–231. https://doi.org/10.3201/eid2302.161072
Hoang, D. T., Chernomor, O., Von Haeseler, A., Minh, B. Q., & Vinh, L. S. (2018). UFBoot2: Improving the Ultrafast Bootstrap Approximation. Molecular Biology and Evolution, 35(2), 518–522. https://doi.org/10.1093/molbev/msx281
Hu, M., Yuan, S., Zhang, K., Singh, K., Ma, Q., Zhou, J., Chu, H., & Zheng, B. J. (2017). PB2 substitutions V598T/I increase the virulence of H7N9 influenza A virus in mammals. Virology, 501, 92–101. https://doi.org/10.1016/j.virol.2016.11.008
Imai, H., Shinya, K., Takano, R., Kiso, M., Muramoto, Y., Sakabe, S., Murakami, S., Ito, M., Yamada, S., Le, M. T., Nidom, C. A., Sakai-Tagawa, Y., Takahashi, K., Omori, Y., Noda, T., Shimojima, M., Kakugawa, S., Goto, H., Iwatsuki-Horimoto, K., Horimoto, T., … Kawaoka, Y. (2010). The HA and NS genes of human H5N1 influenza A virus contribute to high virulence in ferrets. PLoS pathogens, 6(9), e1001106. https://doi.org/10.1371/journal.ppat.1001106
Li, J., Ishaq, M., Prudence, M., Xi, X., Hu, T., Liu, Q., & Guo, D. (2009). Single mutation at the amino acid position 627 of PB2 that leads to increased virulence of an H5N1 avian influenza virus during adaptation in mice can be compensated by multiple mutations at other sites of PB2. Virus research, 144(1-2), 123–129. https://doi.org/10.1016/j.virusres.2009.04.008
Li, J., Zhang, K., Chen, Q., Zhang, X., Sun, Y., Bi, Y., Zhang, S., Gu, J., Li, J., Liu, D., Liu, W., & Zhou, J. (2018). Three amino acid substitutions in the NS1 protein change the virus replication of H5N1 influenza virus in human cells. Virology, 519, 64–73. https://doi.org/10.1016/j.virol.2018.04.004
Li, Z., Jiang, Y., Jiao, P., Wang, A., Zhao, F., Tian, G., Wang, X., Yu, K., Bu, Z., & Chen, H. (2006). The NS1 gene contributes to the virulence of H5N1 avian influenza viruses. Journal of virology, 80(22), 11115–11123. https://doi.org/10.1128/JVI.00993-06
Lo, F. T., Zecchin, B., Diallo, A. A., Racky, O., Tassoni, L., Diop, A., Diouf, M., Diouf, M., Samb, Y. N., Pastori, A., Gobbo, F., Ellero, F., Diop, M., Lo, M. M., Diouf, M. N., Fall, M., Ndiaye, A. A., Gaye, A. M., Badiane, M., Lo, M., … Monne, I. (2022). Intercontinental Spread of Eurasian Highly Pathogenic Avian Influenza A(H5N1) to Senegal. Emerging infectious diseases, 28(1), 234–237. https://doi.org/10.3201/eid2801.211401
Jiao, P., Tian, G., Li, Y., Deng, G., Jiang, Y., Liu, C., Liu, W., Bu, Z., Kawaoka, Y., & Chen, H. (2008). A single-amino-acid substitution in the NS1 protein changes the pathogenicity of H5N1 avian influenza viruses in mice. Journal of virology, 82(3), 1146–1154. https://doi.org/10.1128/JVI.01698-07
Koopmans, M. P. G., Barton Behravesh, C., Cunningham, A. A., Adisasmito, W. B., Almuhairi, S., Bilivogui, P., Bukachi, S. A., Casas, N., Cediel Becerra, N., Charron, D. F., Chaudhary, A., Ciacci Zanella, J. R., Dar, O., Debnath, N., Dungu, B., Farag, E., Gao, G. F., Khaitsa, M., Machalaba, C., Mackenzie, J. S., … One Health High-Level Expert Panel (2024). The panzootic spread of highly pathogenic avian influenza H5N1 sublineage 2.3.4.4b: a critical appraisal of One Health preparedness and prevention. The Lancet. Infectious diseases, 24(12), e774–e781. https://doi.org/10.1016/S1473-3099(24)00438-9
Kralj, J., Martinović, M., Jurinović, L., Szinai, P., Sütő, S., & Preiszner, B. (2020). Geolocator study reveals east African migration route of Central European Common Terns. Avian Research, 11, Article 6. https://doi.org/10.1186/s40657-020-00191-z
Kuiken, T., & Cromie, R. (2022). Protect wildlife from livestock diseases. Science (New York, N.Y.), 378(6615), 5. https://doi.org/10.1126/science.adf0956
Kuo, R. L., & Krug, R. M. (2009). Influenza a virus polymerase is an integral component of the CPSF30-NS1A protein complex in infected cells. Journal of virology, 83(4), 1611–1616. https://doi.org/10.1128/JVI.01491-08
Minh, B. Q., Schmidt, H. A., Chernomor, O., Schrempf, D., Woodhams, M. D., Von Haeseler, A., & Lanfear, R. (2020). IQ-TREE 2: New Models and Efficient Methods for Phylogenetic Inference in the Genomic Era. Molecular Biology and Evolution, 37(5), 1530–1534. https://doi.org/10.1093/molbev/msaa015
Molini, U., Aikukutu, G., Roux, J. P., Kemper, J., Ntahonshikira, C., Marruchella, G., Khaiseb, S., Cattoli, G., & Dundon, W. G. (2020). Avian Influenza H5N8 Outbreak in African Penguins (Spheniscus demersus), Namibia, 2019. Journal of wildlife diseases, 56(1), 214–218.
Molini, U., Yabe, J., Meki, I. K., Ouled Ahmed Ben Ali, H., Settypalli, T. B. K., Datta, S., Coetzee, L. M., Hamunyela, E., Khaiseb, S., Cattoli, G., Lamien, C. E., & Dundon, W. G. (2023). Highly pathogenic avian influenza H5N1 virus outbreak among Cape cormorants (Phalacrocorax capensis) in Namibia, 2022. Emerging microbes & infections, 12(1), 2167610. https://doi.org/10.1080/22221751.2023.2167610
Monne, I. (2025). Monitoring the genetic evolution of highly pathogenic avian influenza viruses in Europe [Conference presentation]. World Organisation for Animal Health (WOAH) Regional Representation for Europe. https://rr-europe.woah.org/app/uploads/2025/02/Isabella-Monne02_-SGE_HPAI-2.pdf
Nao, N., Kajihara, M., Manzoor, R., Maruyama, J., Yoshida, R., Muramatsu, M., Miyamoto, H., Igarashi, M., Eguchi, N., Sato, M., Kondoh, T., Okamatsu, M., Sakoda, Y., Kida, H., & Takada, A. (2015). A Single Amino Acid in the M1 Protein Responsible for the Different Pathogenic Potentials of H5N1 Highly Pathogenic Avian Influenza Virus Strains. PloS one, 10(9), e0137989. https://doi.org/10.1371/journal.pone.0137989
Pohlmann, A., Stejskal, O., King, J., Bouwhuis, S., Packmor, F., Ballstaedt, E., Hälterlein, B., Hennig, V., Stacker, L., Graaf, A., Hennig, C., Günther, A., Liang, Y., Hjulsager, C., Beer, M., & Harder, T. (2023). Mass mortality among colony-breeding seabirds in the German Wadden Sea in 2022 due to distinct genotypes of HPAIV H5N1 clade 2.3.4.4b. The Journal of general virology, 104(4), 10.1099/jgv.0.001834. https://doi.org/10.1099/jgv.0.001834
Song, J., Feng, H., Xu, J., Zhao, D., Shi, J., Li, Y., Deng, G., Jiang, Y., Li, X., Zhu, P., Guan, Y., Bu, Z., Kawaoka, Y., & Chen, H. (2011). The PA protein directly contributes to the virulence of H5N1 avian influenza viruses in domestic ducks. Journal of virology, 85(5), 2180–2188. https://doi.org/10.1128/JVI.01975-10
Song, J., Xu, J., Shi, J., Li, Y., & Chen, H. (2015). Synergistic Effect of S224P and N383D Substitutions in the PA of H5N1 Avian Influenza Virus Contributes to Mammalian Adaptation. Scientific reports, 5, 10510. https://doi.org/10.1038/srep10510
Sonnberg, S., Webby, R. J., & Webster, R. G. (2013). Natural history of highly pathogenic avian influenza H5N1. Virus research, 178(1), 63–77. https://doi.org/10.1016/j.virusres.2013.05.009
Spesock, A., Malur, M., Hossain, M. J., Chen, L. M., Njaa, B. L., Davis, C. T., Lipatov, A. S., York, I. A., Krug, R. M., & Donis, R. O. (2011). The virulence of 1997 H5N1 influenza viruses in the mouse model is increased by correcting a defect in their NS1 proteins. Journal of virology, 85(14), 7048–7058. https://doi.org/10.1128/JVI.00417-11
Suttie, A., Deng, Y.-M., Greenhill, A. R., Dussart, P., Horwood, P. F., & Karlsson, E. A. (2019). Inventory of molecular markers affecting biological characteristics of avian influenza A viruses. Virus Genes, 55(6), 739–768. https://doi.org/10.1007/s11262-019-01700-z
Wang, W., Lu, B., Zhou, H., Suguitan, A. L., Jr, Cheng, X., Subbarao, K., Kemble, G., & Jin, H. (2010). Glycosylation at 158N of the hemagglutinin protein and receptor binding specificity synergistically affect the antigenicity and immunogenicity of a live attenuated H5N1 A/Vietnam/1203/2004 vaccine virus in ferrets. Journal of virology, 84(13), 6570–6577. https://doi.org/10.1128/JVI.00221-10
Wasilenko, J. L., Sarmento, L., & Pantin-Jackwood, M. J. (2009). A single substitution in amino acid 184 of the NP protein alters the replication and pathogenicity of H5N1 avian influenza viruses in chickens. Archives of virology, 154(6), 969–979. https://doi.org/10.1007/s00705-009-0399-4
Wessels, U., Abdelwhab, E. M., Veits, J., Hoffmann, D., Mamerow, S., Stech, O., Hellert, J., Beer, M., Mettenleiter, T. C., & Stech, J. (2018). A Dual Motif in the Hemagglutinin of H5N1 Goose/Guangdong-Like Highly Pathogenic Avian Influenza Virus Strains Is Conserved from Their Early Evolution and Increases both Membrane Fusion pH and Virulence. Journal of virology, 92(17), e00778-18. https://doi.org/10.1128/JVI.00778-18
Xiao, C., Ma, W., Sun, N., Huang, L., Li, Y., Zeng, Z., Wen, Y., Zhang, Z., Li, H., Li, Q., Yu, Y., Zheng, Y., Liu, S., Hu, P., Zhang, X., Ning, Z., Qi, W., & Liao, M. (2016). PB2-588 V promotes the mammalian adaptation of H10N8, H7N9 and H9N2 avian influenza viruses. Scientific reports, 6, 19474. https://doi.org/10.1038/srep19474
Xie, R., Edwards, K. M., Wille, M., Wei, X., Wong, S. S., Zanin, M., El-Shesheny, R., Ducatez, M., Poon, L. L. M., Kayali, G., Webby, R. J., & Dhanasekaran, V. (2023). The episodic resurgence of highly pathogenic avian influenza H5 virus. Nature, 622(7984), 810–817. https://doi.org/10.1038/s41586-023-06631-2
Yamayoshi, S., Yamada, S., Fukuyama, S., Murakami, S., Zhao, D., Uraki, R., Watanabe, T., Tomita, Y., Macken, C., Neumann, G., & Kawaoka, Y. (2014). Virulence-affecting amino acid changes in the PA protein of H7N9 influenza A viruses. Journal of virology, 88(6), 3127–3134. https://doi.org/10.1128/JVI.03155-13
Yang, Z.-Y., Wei, C.-J., Kong, W.-P., Wu, L., Xu, L., Smith, D. F., & Nabel, G. J. (2007). Immunization by Avian H5 Influenza Hemagglutinin Mutants with Altered Receptor Binding Specificity. Science, 317(5839), 825–828. https://doi.org/10.1126/science.1135165
