The evolution and spread of influenza virus impose great impact on the society in the past century. Humans are still facing the zoonotic threat arising from animal influenza viruses; however, the existing knowledge is not sufficient to provide an accurate prediction of the next pandemic strain. My objective is to put forward the preparedness for influenza pandemics in two ways: molecular diagnosis and cross-protective vaccine. Influenza gene reassortment can take place during co-infection and produce novel viruses. While the pig is regarded as the potential mixing vessel for influenza viruses, it is important to develop rapid diagnostic assays to identify reassortment events in swine surveillance. In this study, rapid diagnostic assays were developed for genotyping pandemic H1N1/2009 and closely related swine influenza viruses. Eight real-time SYBR green-based reverse transcription-polymerase chain reactions (RT-PCR) containing locked nuclei acid (LNA) probes differentiated segments derived from pandemic H1N1/2009, Eurasian avian-like (EA), and triple reassortant (TR) swine virus lineages. With the help of these assays, 41 swine isolates collected during January 2009 to January 2010 in an ongoing swine surveillance programme in Hong Kong were successfully genotyped. Ten of these viruses were pandemic H1N1/2009 viruses, suggesting zoonotic transmissions of the virus from humans to pigs. A novel pandemic reassortant was also discovered during the course of surveillance. This study is the first to show pandemic H1N1/2009 virus has reassorted with other endemic swine viruses in the pig population. The genotyping assays provide a rapid and high throughput screening method to select viruses of interesting segment combination for downstream sequencing analysis and characterization. They will be useful for understanding viral reassortment and complex evolutionary dynamics in routine swine surveillance activities. Current influenza vaccines are strain-specific and the production schedule shows significant delay to reach the general population. The study aims at engineering a novel live attenuated virus vaccine which offers cross-subtypic protection. The feasibility for the conserved hemagglutinin (HA) stalk domain to induce broadly neutralizing antibodies was investigated. By manipulating the segment-specific packaging sequences, a nine-segment A/Puerto Rico/8/1934 (H1N1) (PR8) influenza virus carrying the headless HA of a different subtype (A/Hong Kong/1/1968 (H3N2)) (HK68) was generated by reverse genetics. The virus showed attenuated growth in in vitro cell culture and reduced pathogenicity in mice. Although mice vaccinated with this vaccine were better protected in the challenge of HK68 virus in comparison to unvaccinated mice, there is not enough evidence to verify cross-reactive immunity conferred by the HK68 headless HA immunogen, due to the heterosubtypic response induced by live virus administration. The absence of cross-neutralizing antibodies in immune sera may be explained by sub-optimal folding of the HK68 headless HA. Further studies are needed to modify the immunogen into its native conformation and elucidate the underlying vaccine-host interactions. These two studies have resulted in a useful diagnostic tool to increase influenza surveillance capacity at the swine-human interface, and the foundation for better universal vaccine design based on the conserved HA stalk domain. / published_or_final_version / Public Health / Master / Master of Philosophy

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy