| Influenza A virus(IAV) is negative strand RNA virus and belongs to Orthomyxoviridae. With a wide range of host, IAV can infect human, pigs, avian and dogs et al. Those IAVs in pigs, which can cause acute, highly contagious respiratory disease, are known as Swine Influenza virus(SIV). Pig is considered to playing as a “mixer” on the IAV transmission, such like the "bird- pig- human" circle, according to its special receptor distribution on the surface of respiratory epithelial cells. More and more IAVs from other host species, especially the avian influenza viruse, were reported to infect human and cause fatal cases, by using pigs as a intermediate host. Today IAVs are though to be posting a huge threat against the public health. Vaccination has been proven to be the best strategy in providing protection from the Influenza virus infection, however rapid virus evolution through genetic drift & shift remains a challenge and it limits the usefulness of current vaccines in providing protection from divergent Influenza viruses.Base on the problems showed above, in this study, I mainly focus on the following parts:1. Evaluate the risk of pigs population infected by subtypes of IAV in Guangdong.2. Evaluate the risk of human infected by subtypes of zoonotic IAV in Guangdong.3. Evaluate the immunogenicity and protective efficacy of a novel PICV vector Flu vaccine.For the first part, 6510 pig serum samples(4910 from pig farms and 1600 from live poultry market(LPM)) were collected from 6 cities of Guangdong province, from Mar 2013 to Mar 2014. Titers of the neutralizing antibodies, which were contained in those samples, against subtypes of IAVs from different host species(human, avian and pigs), were tested through hemagglutination inhibition assay(HI). Samples were classified into different categories according to different factors or variables, such as sample sites, growth stage, time stay in LPM and season. Finished the serum prevalence calculation followed by generating the Odds Ratio(OR value) through univariate analysis base on those variables. The results suggested that the main factors, which influence pigs infected by subtypes of IAV, were growth stages/time stay in LPM, season and avian contact. As longer as the pigs stay in LPM, the easier they got infected by the avian flu. Pigs were much easier to have flu virus infection in fall than in other seasons. Frequency and close contact with birds may play a main role during the process of pigs getting infected by avian flu viruses.For the second part, in order to evaluate the threat of viral infection faced by those working with animals, a cross-sectional, sero-epidemiological study was conducted in between December 2013 and January 2014. Individuals working with swine, at poultry farms, or live poultry markets(LPMs), and veterinarians, along with controls not exposed to animals were enrolled in this study and 11(4 human, 3 swine, 3 avian, and 1 canine) influenza A viruses were used in hemagglutination inhibition(HI) assays(7 strains) and the cross-reactivity test(9 strains) in which 5 strains were used in both tests. Univariate analysis was performed to identify which variables were significantly associated with seropositivity. Odds ratios(OR) revealed that swine workers had a significantly higher risk of elevated antibodies against A/swine/Guangdong/L6/2009(H1N1), a classical swine virus, and A/swine/Guangdong/SS1/2012(H1N1), a Eurasian avian-like swine virus than non-exposed controls. Poultry farm workers were at a higher risk of infection with avian influenza H7N9 and H9N2. LPM workers were at a higher risk of infection with 3 subtypes of avian influenza, H5N1, H7N9, and H9N2. Interestingly, the OR also indicated that LPM workers were at risk of H1N1 swine influenza virus infection, perhaps due to the presence of pigs in the LPM. Our data suggests that animal handlers are more likely to have antibodies against animal influenza viruses.For the third part, Pichinde virus(PICV) is a bi-segmented enveloped RNA virus in the Arenaviridae family that targets macrophages and dendritic cells(DCs) early in the infection and induces strong innate and adaptive immunity in mice. We have recently developed a tri-segmented PICV as a vaccine vector platform to express up to two foreign genes and to induce strong humoral and cell-mediated immunity. We have further optimized the vaccine vector to simultaneously express the hemagglutination(HA or H) and the nucleoprotein(NP or P) gene of the influenza A virus A/PR8, r P18tri-H/P. We show that the r P18tri-H/P vector can efficiently induce both HA-specific neutralizing antibodies(Neu Ab) and NP-specific CTLs via different immunization routes and that the vaccine-specific immune responses was significantly increased upon a booster dose and remained at high levels even after three booster doses. To understand the underlying immune mechanisms, we have measured the vector-specific Neu Abs and CTLs after each dose and determined the effects of pre-existing anti-PICV immunity on the development of vaccine-specific primary and secondary immune responses. Moreover, we have examined the ability of using this novel r P18tri-based vaccine vector to induce cross-reactive anti-influenza immunity in a prime-and-boost strategy. In summary, we report the development and in-depth characterization of a novel PICV-based live vaccine vector that can express dual foreign antigens to induce strong humoral and cell-mediated immunity and is ideal for a prime-and-boost vaccination strategy.In summury, we not only evaluated the risk of zoonotic influenza A viruses infecting human and pigs in Guangdong province which indicated that both of these two species were facing the threat posed by multiple subtypes of influenza A viruses, but also developed and tested a novel PICV vector flu vaccine which could carry at least two foreign proteins, such as flu HA and NP, and induce the sufficient immune response in mice for keeping them safe upon lethalvirus challenge. This novel virus vector vaccine is expected to be a new option for fighting against flu viruses in the future. |
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