DOI: 10.25442/hku.29979175.v1
title: supporting data for Thesis "<b><i>Dynamics of Influenza A Virus Emergence and Transmission from Hong Kong</i></b>"

description: Influenza A viruses (IAVs) pose a persistent global health threat, necessitating a comprehensive understanding of their dynamics, diversity, evolution, and transmission to inform pandemic preparedness. This thesis provides a multi-scale investigation into the IAVs, establishing Hong Kong as an important place for zoonotic risk and global viral spread.Firstly, surveillance in Hong Kong's animal populations (wild birds, swine, and poultry) identified significant threats. Highly pathogenic avian influenza viruses of clade 2.3.4.4b H5 were detected in wild birds, with viruses from two spoonbills from late 2022 being genetically related to a virus from a human in China. In swine, an Eurasian avian-like H1N1 swine influenza virus with PB1 and M segments derived from the H9N2 subtype, suggesting that H9N2 viruses are infecting pigs and reassorting with swine influenza viruses in China. Novel H3N8 viruses were identified in chicken in live poultry markets and chicken farms in Hong Kong, that are genetically similar to the zoonotic H3N8 viruses reported in mainland China. These findings highlight the region as a dangerous place for the emergence of high-risk strains.Secondly, a decade-long phylogeographic analysis of low pathogenetic avian influenza viruses in wild birds conclusively demonstrated Hong Kong's important role in global AIV dissemination. Significant viral migrations were observed from Hong Kong to adjacent regions (Japan/Korea, East China, Bangladesh) and to geographically distant continents (Australia, Europe, North America). The study identified Hong Kong as a source for the PA gene segment during an early stage (2010-2015) and documented extended circulation (longer persistence) of polymerase genes (PA, PB2, PB1). Furthermore, H6 viruses were pinpointed as "genetic hubs" due to their consistent role as donors for PB2 and PA segments, while H3 viruses served as primary sources for NP and M segments, underscoring subtype-specific contributions to reassortment.Thirdly, investigating genetic drift at a finer scale, intra-host analysis in human patients revealed significantly greater genetic diversity in H3N2 viruses compared to H1N1 within infected individuals. Evidence of anatomical compartmentalisation was found, with the throat hosting more diverse H3N2 viral populations than the nose, particularly in PA, PB2, and neuraminidase genes. Furthermore, a codon-deoptimized '8-mut' influenza virus, engineered with 373 silent mutations to mimic avian codon usage, demonstrated accelerated genetic diversification (predominantly synonymous mutations) over 11 passages in human A549 cells, compared to the wild-type strain. Critically, secondary often non-synonymous mutations consistently emerged directly adjacent to engineered silent mutations, especially when short dinucleotide repeats were introduced, suggesting genome destabilisation and potential compensatory adaptation mechanisms. These findings provide crucial lessons for the rational design of influenza vaccines with enhanced safety and efficacy profiles.Collectively, this thesis highlights the dynamic and complex evolutionary landscape of IAVs, affirming the continuous pandemic threat and the indispensable role of integrated surveillance and genetic characterisation in Hong Kong. <br><br><br>