Animal influenza viruses - Impacts of influenza A virus in Danish swine herds

Abstract

Since the filtration experiments performed by Richard E. Shope in 1931, swine influenza A virus (swIAV) has been linked to influenza-like symptoms in pigs, including fever, coughing, anorexia and apathy. Since then, several different subtypes and variants of swIAV have appeared and continue to pose clinical problems in affected herds globally. For a proper control of swIAV, it is crucial to understand the dynamics of the virus. However, several aspects of the infection remain unclear. It is uncertain how the virus spreads in and between herds, which age groups are infected and what the clinical impact of swIAV is. Several aspects of the immunity towards swIAV are also unknown e.g. how does it evolve and which responses are needed to protect the pigs. Other unsolved questions relate to the evolution of the virus, where the level and impact of antigenic drift needs investigations. Finally, there is a lack of studies providing field data on the efficacy of different control measures. The main aim of this PhD project was to perform research activities in order to answer some of the above-mentioned questions. The thesis consists of four major parts. Part 1 presents the background for the studies carried out during the PhD project and introduces the general aims of the project as well as the thesis outline. Part 2 includes a literature review divided into two sections. The first section reviews the biology of influenza A virus (IAV), providing information on the taxonomy, virus lifecycle, subtypes, lineages and nomenclature, methods for IAV characterization, viral evolution and distribution of IAV in different species. The second section of the literature review focuses on swIAV and its pathogenesis, clinical signs, immunology, vaccines and maternally derived antibodies (MDAs). The epidemiology of swIAV is also reviewed both in regard to the distribution of swIAV globally and the transmission patterns within and between herds. Finally, a review of different control strategies is presented. Part 3 includes the four manuscripts written during the PhD project. An intensive overall discussion and conclusion completes the thesis in Part 4, which also includes future perspectives for swIAV research. Manuscript 1 describes and discusses the results of longitudinal field studies carried out in three Danish swine herds, with focus on the transmission dynamics and clinical impacts of swIAV. The results of the study revealed early presence of swIAV, which affected piglets as young as three days of age. The early infections were observed in all herds, despite the presence of IAV antibodies in the sows, indicating that the protection obtained by MDAs was incomplete. Moreover, a small number of sows tested positive for swIAV, indicating a possible role of the sows in the transmission dynamics. Interestingly, a number of pigs tested positive for swIAV over two consecutive or non-consecutive samplings, indicating a possibility for prolonged shedding and reinfection, respectively. Finally, the clinical impacts of swIAV were documented by observing the correlations between swIAV, nasal discharge and increased coughing index. Manuscript 2 describes a study performed in a single herd where it was decided to initiate vaccination of piglets at the time of castration with an inactivated swIAV vaccine. The results showed very limited effect of vaccination, as the total number of infected pigs and clinical signs were similar between the vaccinated- and the control group. Due to the higher frequency of sampling in the study, it was possible to investigate the presence of prolonged and recurrent shedding in details. Extensive genetic characterization of the swIAV sequences obtained from prolonged and recurrent shedders documented the presence of a single swIAV strain, and revealed evidence of antigenic drift in the recurrent shedders. Manuscript 3 provided unique data on an acute swIAV outbreak in an enzootic infected herd, where it was subsequently decided to implement mass sow vaccination, thereby allowing for investigation of the effects of this vaccination strategy. Several interesting observations were made in this study. The acute outbreak with a new swIAV subtype caused early infection of piglets, as swIAV was almost exclusively identified in the one-week-old piglets. The early infection suggested that the piglets were not protected by MDAs. Interestingly, viral characterization revealed that the HA gene, of the enzootic swIAV strain and outbreak swIAV strain, belonged to the same lineage, but also showed major differences, specifically in the region encoding the globular head. Subsequent serological tests confirmed a lack of cross-protection between the two swIAV strains. After vaccination, the infection time was delayed and the viral load was reduced. However, an increase in numbers of prolonged shedders was observed, which along with the delayed infection time resulted in the spread of swIAV to all age groups of the herd. Moreover, evidence of an increased substitution rate with positive selection in the HA gene was observed after the implementation of mass sow vaccination. Manuscript 4 was based on repeated cross-sectional studies with monthly sampling in a single herd over a one-year period. The enzootic nature of swIAV was confirmed as the virus persisted in the herds over the full project period. The results confirmed the observations made in Manuscript 1, as a large number of sows/gilts tested positive for swIAV during the study. Investigations into the viral evolution indicated that swIAV evolved in a similar manner to human seasonal influenza. These results provided a basis for explaining the vast diversity observed within the single swIAV lineages, and provoked a discussion of the possible implications of extensive antigenic drift on the control of swIAV. Ultimately, this PhD project has contributed to a deeper understanding of the transmission dynamics and impacts of swIAV. The studies carried out in this PhD are the first to show extensive swIAV infection in newborn piglets, and emphasize the role of sows and gilts in the transmission dynamics. All four studies document the enzootic nature of swIAV within the herds, and provide evidence of prolonged and recurrent swIAV shedding, which could enhance the herd-level persistence of swIAV. The results suggest that the presence of pre-existing immunity is likely to play a role in the generation of prolonged and recurrent shedders, and potentially drive the evolution of swIAV. In addition, the results reveal a high level of genetic diversity and a fast evolution of swIAV, which potentially has consequences for the transmission and the control of the virus. Finally, results on vaccine efficacy have been provided, which in combination with the increased knowledge of the swIAV dynamics, will help the industry in the implementation of optimal control measures in the future.