Thesis (M.S.)--University of Texas at El Paso, 2009. / Title from title screen. Vita. CD-ROM. Includes bibliographical references. Also available online.

Thesis supervisor: C. Martin Stoltzfus. Includes bibliographic references (p. 121-143).

Thesis (M.S.)--University of Tennessee, Knoxville, 2009. / Title from title page screen (viewed on Oct. 23, 2009). Thesis advisor: Reza Hajimorad. Vita. Includes bibliographical references.

Beet yellows virus (BYV) is a filamentous, positive-strand RNA virus that belongs to the family Closteroviridae. BYV particles encapsidate a 15.5 kb RNA and posses complex polar architecture. A long virion body is formed by the major capsid protein(CP), whereas the minor capsid protein (CPm) assembles a short tail that encapsidates the 5'-terminal region of BYV RNA. In addition to proteins required for viral RNA replication and encapsidation, BYV encodes four proteins whose role in the virus life cycle was unknown. These proteins include a small, 6-kDa, hydrophobic protein (p6), a homolog of the cellular 70-kDa heat shock proteins (Hsp7Oh), a 64-kDa protein (p64), and a 20-kDa protein (p20). It was found recently that Hsp7Oh, p64, and p20 are incorporated into BYV virions, and that Hsp7Oh is required for the virus movement from cell to cell. In this study, we characterized genetic requirements for BYV assembly and cell-to-cell movement, and determined relationships between these two processes. It was demonstrated that in addition to Hsp7Oh, p6, p64, CP, and CPm are each essential, but not sufficient for virus movement. These results indicated that five-component movement machinery of BYV is the most complex among plant viruses. Extensive mutational analysis of CP and CPm revealed strong correlation between abilities of BYV to assemble tailed virions and to move from cell to cell, suggesting that formation of functional virions is a prerequisite for virus translocation. We have found that CPm, Hsp7Oh, and p64 are necessary for the efficient virion tail formation. Assembly of the virion tails and bodies was shown to occur independent of each other and likely to involve two separate packaging signals within the genomic RNA. Our work demonstrated that BYV encodes one conventional movement protein, p6, whose only known function is to mediate virus movement. The other four movement associated proteins of BYV, CP, CPm, Hsp7Oh, and p64 are the virion components, each of which is required for assembly of the tailed, movement-competent virions. Based on these and other data, we propose that BYV and other closteroviruses evolved virion tails as a specialized device for the directional cell-to-cell movement of large RNA genomes. / Graduation date: 2005 / Best scan available.

Tetraviruses are a family of small non-enveloped positive sense RNA viruses that exclusively infect members of the order Lepidoptera. Their replication biology is poorly studied because, with the exception of Providence virus (PrV), tetraviruses are unable to replicate in tissue culture cells. The overall aim of the research described in this thesis was to develop a fundamental understanding of the replication of tetraviruses, focussing on the site of replication within host cells and in particular, the subcellular localisation of the viral replicase. Helicoverpa armigera stunt virus (HaSV, Genus: Omegatetravirus) was chosen for this study because it is the only tetravirus for which the cDNAs have been shown to be infectious. In the absence of tissue culture cell lines susceptible to HaSV infection, the approach was to use confocal fluorescence microscopy to examine the subcellular localisation of the HaSV replicase fused to enhanced green fluorescent protein (EGFP) in mammalian and insect tissue culture cells. The replicase (with EGFP fused at its C-terminus) localised to punctate structures throughout the cytoplasm of transfected HeLa and Sf9 cells. These structures were then shown – using live cell imaging and time lapse photography – to behave similarly to cellular endocytic organelles and fluorescence partially overlapped with membranes containing the late endosomal marker protein CD63. Biochemical fractionation of Sf9 cells expressing the replicase via a recombinant baculovirus (as well as transfected HeLa and Sf9 cells expressing EGFP-replicase fusion proteins) demonstrated that the replicase was strongly associated with detergentresistant membranes (DRMs) in these cells. Deletion analysis of the replicase coding sequence revealed two regions involved in the generation of the punctuate structures. Firstly, the C-terminal half of the replicase RNAdependant RNA polymerase domain was found to be essential for targeting and the tight association with DRMs while the second region, within the Nterminal 44 amino acids, enhanced localisation through a combination of secondary structural elements and sequence-specific functions. A comparative immunofluorescence study on PrV, which replicates as a persistent infection in an insect midgut cell line, showed that the PrV replicase also localised to punctate structures in the cytoplasm. Biochemical fractionation showed that the replicase was also strongly associated with DRMs. This thesis describes the development of new experimental systems for the study of tetravirus replication biology and the data lead to the conclusion that the HaSV replicase associates with DRMs derived from alternate endocytic pathway organelles.

Reciprocal interference experiments between Potato virus Y (PVY) and Pepper mottle virus (PeMV) in Capsicum annuum L. 'Tabasco' and 'Special pepper' (a selection of Anaheim chilli peppers), showed suppression of local lesion production in both directions but incomplete suppression of challenge virus replication (incomplete cross protection). However, suppression was reduced by increasing the concentration of the challenge virus. The source of inoculum of the viruses did not have a significant effect on the subsequent interference between the viruses. A direct relationship was established between counts of local lesions and virus particles counted using the electron microscope. Tobacco etch virus (TEV) was transmitted from one Capsicum annuum L. 'Tabasco' plant to another in the same pot within 4 days after infection of the test plant. Transmission probably took place via root grafts, as the necrotic roots of the test plants intertwined with the uninoculated Tabasco plants. Mixed infections of PVY and PeMV resulted in the production of PVY-N, a new strain of PVY. The new strain, which could be recognized by changes in biological and serological properties, was produced only in mixed infections and was stable after six serial transfers in several hosts. Evidence suggests that the development of the new strain is host dependent. Potato Virus Y in mixed infections with PeMV or TEV in C. annuum L. 'Anaheim' did not induce local lesions, systemic necrosis nor death of Special pepper, while PeMV alone induced the death of Tabasco. The behavior of TEV in Tabasco in a mixed infection was temperature dependent, while TEV alone induced wilt and death of Tabasco irrespective of the greenhouse temperature and season of the year.

Copy of author's seven page article in pocket inside back cover. / Bibliography: leaves 118-143. / x, 143, [52] leaves, [23] leaves of plates : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Microbiology and Immunology, 1995?

Includes bibliography / 108 leaves, [15] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Biochemistry, 1986

Thesis (Ph. D.)--University of Adelaide, Dept. of Plant Science, 1997. / Includes bibliographical references (leaves 127-138).

Thesis (Ph.D.)--University of Delaware, 2008. / Principal faculty advisor: K. Eric Wommack, Dept. of Plant & Soil Sciences. Includes bibliographical references.