Studies on the host genetic resistance and susceptibility to influenza A virus

Influenza infections represent a serious threat to human health and are causing respiratory disease in human and animals. Many of the biological mechanisms of the host-pathogen interactions after influenza A virus (IAV) infection are still unclear. Both intrinsic and extrinsic factors determine the severity of influenza disease. To study the importance of host and virus virulence factors, I investigated in my thesis how different genetic backgrounds of the host influenced the function of orthomyxoviridae resistance gene 1 (Mx1). It was shown previously that congenic B6.A2G-Mx1r/r (B6-Mx1r/r) mice carrying a functional Mx1 allele are highly resistant against IAV. In particular, the type I interferon induced protein MX1 has been shown to represent a strong effector of innate immunity and to act by blocking nuclear import of viral RNA and inhibits viral replication. To assess the influence of genetic background on Mx1 function, I compared the host response to IAV infection in congenic mice B6-Mx1r/r to congenic D2(B6). A2G-Mx1r/r (D2-Mx1r/r) mice. Most surprisingly, congenic D2-Mx1r/r mice harboring a functional Mx1 wild type allele were highly susceptible to H1N1 virus infection. I observed that the survival proportion was related to both the genetic background and the copy numbers of functional Mx1 alleles. Furthermore, the genetic background influenced Mx1 antiviral activity by regulating virus titer in the lungs after infection. Infection of D2-Mx1r/r mice led to a 100-fold higher viral load lungs compared to infection of B6-Mx1r/r. In contrast, congenic B6-Mx1r/r mice started to reduce virus earlier and did not develop severe disease. Additionally, D2-Mx1r/r mice which were pretreated with interferon α were fully protected from lethal infections. These observations suggest that B6 mice carry genetic factors which initiate Mx1 function and which are absent in D2 mice. Furthermore, in a D2 background, IAV replicates very rapidly at early time points and Mx1 protective functions are activated too late to prevent the severe outcomes. In the second part of my thesis I investigated in the importance of fibrinolysis and the influence of vascular permeability in the context of IAV infection. Recent studies have shown that IAV alters the plasminogen conversion pathway and hemostasis after infection. Here, I studied infections with two IAV strains, PR8 and WSN. The WSN strain has a unique neuraminidase which can convert plasminogen into plasmin and allows cleavage of the hemagglutinin via activated plasmin in the absence of other host serine proteases. In mice, WSN can disseminate to extra-pulmonary organs while PR8 is restricted to the respiratory tract. In my studies, I investigated if dissemination of WSN was related to hyperfibrionlysis from plasminogen conversion. For this, I studied the host response and virus replication in a mouse knockout mutant of the Serpine1 gene. I showed that Serpine1 mutants were more susceptible to H1N1 infections compared to wild type mice. The increase in susceptibility was caused by an enhancement of fibrinolysis, resulting in leaking of red blood cells into the alveolar space and dissemination of virus into the kidneys. Also, WSN-infected wild type mice showed higher levels of protein leakage into the bronchoalveolar space fluid compared to PR8-infected mice. In conclusion, my results strongly suggest that dysregulation of the plasminogen activation pathway contributes to the enhanced severity in IAV-infected mouse lungs. Hyperfibrinolysis increases lung hemorrhage and enhances vascular permeability which may lead to virus dissemination. Furthermore, I investigated the kinetics of cellular changes in the peripheral blood after IAV infection of B6 and D2 mice. We observed that the ratio of granulocytes to lymphocytes in the peripheral blood and lung correlated well with disease severity. By fluorometric analysis, a massive infiltration of Ly6G+CD11b+ cells in the lung was detected which was strongly associated with the lethal infection in B6 mice, whereas D2 mice showed an increase in proinflammatory cells in the lungs after IAV infections. These immune cell infiltrates of the lung reflected quantitative and qualitative differences in the periphery. In addition, I investigated genetic variants of the NK cell gene Klrd1. It was reported that Klrd1 is deleted in DBA/2J mice. After performing fluorometric analyses of NK cell from D2 mice, I could further show that the sub-strain DBA/2Rj stock from the Janvier Breeding Centre in France expresses the CD94 protein. On the other hand, I identified a spontaneous deletion spanning the last coding exon of the Klrd1 gene in DBA/2J mice from the Jackson laboratory by high throughput sequencing. An additional deletion in the intronic region between exons 2 and 3 was also identified in both D2 sub-strains. These results revealed the presence of different Klrd1 alleles in these two D2 sub-strains.