Interferon (IFN) and phosphatidylinositol 3-kinase (PI3K) are apoptosis regulators that are targeted by viruses to promote survival of infected cells. Significant crosstalk exists between IFN and PI3K, and this study sought to investigate the relationships between IFN, PI3K and apoptosis during virus infection. Parainfluenza virus 5 (PIV5) and influenza A virus (IAV) are both negative-sense single-stranded RNA viruses that encode multifunctional proteins in order to maximise their genome coding capacity. The PIV5 V and IAV NS1 proteins are well-studied as IFN antagonists and, in addition, both are reported to modulate PI3K signalling. Less well-studied is the role of these proteins in apoptosis regulation; the ability of V and NS1 to inhibit apoptosis was therefore investigated. PIV5/V was found to limit cell death in response to a number of apoptosis inducers in a manner that required its STAT1- degradative activity and also inhibited activation of the PI3K downstream target, Akt. IAV/NS1 binds directly to PI3K to stimulate its activity, and this is reported to mediate anti-apoptotic signalling during IAV infection. However, a virus expressing an NS1 unable to bind PI3K did not induce more apoptosis than wt virus. NS1 expression, either in a stable cell-line or during virus infection, was also unable to protect cells from pro-apoptotic stimuli. NS1-mediated PI3K activation similarly had no effect on IFN production or ISG expression in infected cells. In contrast, other NS1 mutant viruses induced large amounts of apoptosis. These viruses also induced significant levels of IFN and were unable to cause apoptosis in IFN-deficient cells, indicating that NS1 limits apoptosis induction through its IFN antagonist functions. The implications of this work for anti-cancer and anti-viral therapies are discussed.

As formulações vacinais contra o Vírus Respiratório Sincicial Humano, HRSV, estão associadas à indução de eosinofilia pulmonar mediada por uma resposta de células TCD4+ Th2, após exposição ao HRSV selvagem. Foi identificado um peptídeo da proteína viral G, que modificado perde a capacidade de predispor à eosinofilia, tornando-o um imunógeno atraente. Células T CD8+ específicas para HRSV reduzem a resposta Th2, mediam resistência a desafio com o vírus, e estão relacionadas à redução dos sintomas. Assim, neste trabalho buscamos e identificamos epítopos de células TCD8+ na polimerase viral, utilizando programas de predição, imunização com peptídeos e avaliação da resposta celular. Também construímos vacinas de DNA contendo a seqüência nucleotídica do peptídeo da proteína G modificado. A caracterização da resposta imune estimulada por essas vacinas e por peptídeos purificados revelou que o plasmídio pTGMCTB, bem como os peptídeos GM e GMCTB, foram capazes de induzir anticorpos que, porém, não se mostraram neutralizantes de HRSV e protetores frente a desafio. / Vaccines against human respiratory syncytial virus (HRSV) are associated with pulmonary eosinophilia induction mediated by a TCD4+ Th2 response, after exposition to wild HRSV. A peptide from the viral protein G was identified to predispose to eosinophilia and loses this ability when mutated, making it an interesting immunogen. CD8+ T cells specific to HRSV reduce the Th2 response, mediate resistance to virus challenge, and are related to symptom-reduction. Thus, in the present work, we searched for and identified CD8+ T cell epitopes in the viral polymerase; using prediction programs, peptide immunization and evaluation of the cellular response. We also constructed DNA vaccines containing the nucleotide sequence of the mutated peptide from G protein mentioned above. The characterization of the immune response elicited by these vaccines and purified peptides showed that the pTGMCTB plasmid, as well as GM and GMCTB peptides were able to induce antibody response; however they are not neutralizing and protective against HRSV challenge.

Fast-evolving RNA viruses are a leading cause of morbidity and mortality among human and animal populations, contributing significantly to both global health and economic burden. The advent and revolution of high-throughput sequencing has empowered phylogenetic analyses with increasing amounts of temporally and spatially sampled viral data. Moreover, the parallel advancement in molecular evolution and phylogenetic methods has provided investigators with a unique opportunity to gain detailed insight into the evolutionary and epidemiological dynamics of emerging viral pathogens. Using state-of-the-art statistical approaches, this thesis addresses some of the important but controversial questions in viral emergence. Chapter 2 introduces a new framework to quantify and investigate reassortment events in influenza A viruses. By developing a computationally efficient algorithm to calculate the largest common subtree for a pair of tree sets, which are estimated from diffe rent parts of the genome for the same taxa set, the level of phylogenetic incongruency due to reassortment can be appropriately ascertained. Chapters 3, 4 and 5 investigate the evolutionary origins of three diff erent viruses: the novel emergence and cross-species transmission of SARSCoV, the genesis and dissemination of the unique HCV circulating recombinant form, and the ancient divergence of all influenza viruses, respectively. Moreover, Chapter 4 presents an improved statistical framework, which provides more precise evolutionary estimates, by utilizing the hierarchical bayes approach to investigate recombination events in emerging RNA viruses. The last empirical study, presented in Chapter 6, applies the recently developed Bayesian phylogeography models to a large viral sequence dataset sampled from southern Viet Nam to examine the fine-scale spatiotemporal dynamics of endemic dengue in Southeast Asia. The work presented here reflects both the advancements made in sequencing technology and statistical phylogenetics, along with some of the challenges that remain in studying the emergence of fast-evolving RNA viruses. This thesis proposes new and improved solutions to these evolutionary problems, such as incorporating non-vertical evolution (i.e. homologous recombination and reassortment) into the phylodynamic framework, with the aim of facilitating future investigations of emerging viral diseases.

Includes Appendix listing other publications by the author. Includes bibliographical references (leaves 134-181). Alfalfa mosaic virus was isolated from lucerne (Medicago sativa) plants with a variety of disease symptoms. Experiments showed that each isolate was biologically distinct and that the host range and symptomatology of each isolate was affected by the environmental condition.

Includes appendix containing journal publications co-authored by the author. Includes bibliographical references (leaves 130-151). Eighteen strains of Cucumber mosaic virus, including forteen from Australia, two from the USA, and two from Japan were used in this study.

Includes bibliographical references (leaves 82-92). Five isolates of pea seed-borne mosaic virus were compared by host range and symptomatology on 16 pisum sativum cultivars lines, 21 lines of Lathyrus and Lens spp. and several indicator species

Includes bibliographical references (leaves [102]-120). Studies the structure of the cucumber mosaic virus genome and the control of its expression.

Includes appendix containing journal publications co-authored by the author. / Includes bibliographical references (leaves 130-151). / xiii, 151 leaves : ill., photos ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Eighteen strains of Cucumber mosaic virus, including forteen from Australia, two from the USA, and two from Japan were used in this study. / Thesis (Ph.D.)--University of Adelaide, Dept. of Crop Protection, 1992

Includes bibliographical references (leaves 82-92). / xii, 92 leaves : ill., photos ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Five isolates of pea seed-borne mosaic virus were compared by host range and symptomatology on 16 pisum sativum cultivars lines, 21 lines of Lathyrus and Lens spp. and several indicator species / Thesis (Ph.D.)--University of Adelaide, Dept. of Crop Protection, 1993

Bibliography: leaves 127-138. This thesis studies the minor double stranded RNAs (dsRNA) and single stranded RNAs (ssRNA) which are consistently associated with plants infected with Q strain of cucumber mosaic virus (Q-CMV). The investigations are focused on the structural elucidation of new RNAs which have been observed in single stranded and double stranded RNA profiles of Q strain of CMV.