Genomic Surveillance of Influenza Virus Evolution in Wild Bird Populations
DOI:
https://doi.org/10.5281/zenodo.22688172Keywords:
avian influenza, genomic surveillance, wild waterfowl, reassortment, phylodynamics, H5N1 clade 2.3.4.4b, Scheldt-Meuse estuaryAbstract
Wild waterfowl remain the principal reservoir from which highly pathogenic avian influenza (HPAI) viruses
periodically spill into poultry and, less often, mammals, yet routine surveillance at wetland stopover sites still
under-samples the genomic diversity circulating in these hosts between outbreak seasons. We carried out an
eighteen-month active surveillance programme at four wetland complexes along the Scheldt-Meuse estuarine
system in Belgium, collecting 2,847 fresh faecal samples from mixed waterfowl roosts and combining real-time
RT-PCR screening with whole-genome nanopore sequencing of positive samples. Of 312 influenza A-positive
samples, 268 yielded genomes of sufficient completeness for phylogenetic and reassortment analysis. HPAI H5N1
clade 2.3.4.4b dominated detections during the fourth quarter of each surveillance year (October-December),
coinciding with the arrival of Arctic-breeding Anser and Anas species, while low-pathogenicity subtypes, principally
H3N8, H4N6 and H11N9, circulated at a low background prevalence throughout the study. Whole-genome
analysis identified eleven independent reassortment events involving exchange of internal gene segments
between clade 2.3.4.4b and co-circulating low-pathogenicity viruses, four of which produced viruses carrying a
PB2 segment with the mammalian-adaptation-associated marker M631L in a haemagglutinin and neuraminidase
backbone otherwise typical of wild-bird lineages. Bayesian phylodynamic reconstruction placed the most recent
common ancestor of the local H5N1 lineage substantially earlier than its first detection in regional poultry, implying
a period of cryptic circulation in wild hosts. These findings argue for surveillance designs that sample wild bird
reservoirs continuously across the annual cycle rather than reactively following poultry outbreaks, since the
genomic events most relevant to spillover risk -- reassortment and the accrual of mammalian-adaptation markers
-- were detectable in wild birds weeks to months before any associated event in domestic or captive animals.
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