Thesis (Ph. D.)--University of Queensland. / Includes bibliographical references.

Helicoverpa zea (Boddie), a pest of cotton that also occurs in field corn, is commonly controlled through the use of foliar-applied insecticides or transgenic crops expressing Bt genes. To prevent the selection of resistant populations, refuge systems have been implemented into the agroecosystem. Historically, structured refuge compliance among growers has been low, leading to the commercialization of seed blended refugia. To test the viability of seed blended refugia in the southern U.S., field studies were conducted in Mississippi and Georgia during the 2016, 2017, and 2018 growing seasons. To quantify adult H. zea emergence from structured and seed blended refuge options, emergence traps were utilized. Kernel damage and moth emergence timings were recorded. Various percentages of stand loss ranging from 0% to 50% were also simulated to determine yield effects in unprotected seed blended refugia. Lastly, H. zea feeding and emergence in a two-gene field corn variety expressing Cry1A.105 and Cry2Ab2 were compared to non-Bt field corn. When compared to a structured refuge, H. zea adult moth emergence from seed blended refugia did not significantly differ. Kernel damage was not different between seed blended treatments and structured refuge treatments. Moth emergence timings were not significantly delayed between the structured refuge and seed blended refuge treatments. Significant yield losses were observed when stand loss was simulated at various levels in field corn, suggesting that there is an opportunity to see yield losses in an unprotected seed blended refuge field corn landscape. Kernel damage did not significantly differ between field corn expressing Cry1A.105 and Cry2Ab2 and non-Bt field corn, possibly due to H. zea resistance to the Cry genes. However, there was a significant difference in emergence from two-gene expressing field corn and non-Bt field corn. This suggests that there may be high pupal mortality in two-gene corn plots. Based on these data, seed blended refuge could be a viable option to replace structured refuge strategies in the southern U.S., however, if left unprotected, yield loss could be observed in a case of high boring insect pressure. The significant loss of refuge plants can also compromise refuge effectiveness.

No description available.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1965. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Thesis (Ph.D.) - University of Queensland, 2004. / Includes bibliography.

Thesis (M. Sc. (Biochemistry, Microbiology & Biotechnology))--Rhodes University, 2005.

Crop pests are known to be adversely affected by rain. Because limited information on this aspect was available for Heliothis spp. occurring on cotton, this study was undertaken to examine the effect of water droplets from overhead irrigation and simulated rain on the survival of H. armiger eggs on cotton. Three aspects were examined: the effects of submersion, the physical impact of droplets on the eggs and the washing off of the eggs from cotton plants in the field. The first two aspects were examined in terms of the structure and respiratory system of the eggs, confirming that H. armiger eggs are able to survive initial wetting in the field. The wash-off of H. armiger eggs from cotton plants is explained in the light of the selection of oviposition sites by the moths, adhesion of the eggs to the plant parts and the dislodging and wash-off by water from simulated rain and overhead irrigation. The data are discussed in terms of the other mortality factors which occur in commercial cotton fields. All in all, it was found that while overhead irrigation should be taken into account in surveys of H. armiger for pest management purposes, it does not offer a viable control strategy and should not be investigated further

Lepidopteran insects like Helicoverpa armigera, more commonly known as the cotton bollworm, are economically important pests of a wide variety of crops throughout the world. The Helicoverpa armigera stunt virus (HaSV), a tetravirus with a bipartite single-stranded positive-sense RNA genome, has great potential as a biological pesticide against H. armigera. The larger genomic strand of this virus (RNA1) encodes the viral replicase, while the other (RNA2) encodes the 71 kDa capsid protein precursor (p71). 240 copies of p71 assemble into a procapsid with the concomitant encapsidation of the viral RNA. This is followed by a complex maturation event that is characterized by the autoproteolytic cleavage of p71 into the 64 kDa capsid protein (P64) and a 7 kDa peptide (p7). The rearrangements that occur during maturation results in the formation of mature HaSV capsids that can thereupon deliver RNA to other susceptible host cells. The principal objective of the research described in this study was to demonstrate that this virus could be assembled in Saccharomyces cerevisiae. S. cerevisiae expression vectors were constructed for the production of p71. This protein was detected in cell lysates from two different strains of S. cerevisiae, both containing either chromosomal or episomal copies of an expression cassette for P71. A number of factors relating to the expression of P71 (e.g. strains used, expression loci and expression rate) and the preparation of protein extracts from S. cerevisiae (e.g. the presence of various protease inhibitors and salt concentrations) were examined to attain optimal levels of soluble p71. A small fraction of the optimized soluble p71 was shown to be in the form of virus-like particles (VLPs), with a yield of ≤10⁷ VLPs from a 1.5l culture of P71⁺ cells. These particles were exclusively in the procapsid form, had a similar buoyant density to that of wild-type HaSV and could undergo maturation when the pH was reduced to 5. S. cerevisiae vectors were constructed for the episomal expression of the HaSV genomic RNAs. These vectors directed the transcription of RNA1 and RNA2 transcripts, which had similar sizes to those of the HaSV genomic RNAs. Mature HaSV particles were purified from cells, transgenic for P71, RNA1 and RNA2, by way of two different virus purification protocols that were developed during this study. RT-PCR analyses on RNA-extracts from these particles demonstrated that RNA transcripts, which were produced in trans with p71, could be encapsidated by HaSV capsids in S. cerevisiae. A droplet-feed bioassay on H. armigera larvae demonstrated that the S. cerevisiae-derived HaSV particles caused impaired larval development. This response was correlated with the detection of HaSV RNA2 in RNA extractions from larvae that were used in this bioassay. The results that were generated through the course of this study, provided proof for the concept of the non-host production of infectious HaSV particles from S. cerevisiae. This work could serve as a foundation for future research on the development of an expression system for the large-scale production of this virus as a biopesticide.

Tetraviruses are entomopathogenic viruses that propagate solely in lepidopteran hosts. Viruses of this group possess non-enveloped 38- to 40-nm capsids arranged in T = 4 surface symmetry. The viral genome consists of one or two single stranded positive sense RNA strands, which define the two genera of this family, the monopartite betatetraviruses and the bipartite omegatetraviruses. Two extensively studied members of the tetraviruses are the omegatetraviruses, Helicoverpa armigera stunt virus (HaSV) and the closely related Nudaurelia capensis ω virus (NωV). The larger genomic strand of HaSV (RNA1) encodes the viral replicase, while the other (RNA2) encodes the 71-kDa capsid precursor protein (p71). The pro-capsid is assembled from 240 copies of p71, which undergo a maturation auto-catalytic cleavage into the 64-kDa (p64) capsid protein and a 7-kDa peptide (p7) forming the capsid shell. The mechanism for the recognition and packaging of the viral genome is poorly understood for these viruses. The principle objective of the research described in this study was to develop in vitro and in vivo experimental systems to investigate interactions between the N terminal domain of HaSV p71 and viral RNAs. More specifically, the two positively charged clusters of predominantly arginine residues that are conserved amongst tetraviruses and the structurally analologous nodaviruses capsid protomers’ N terminal domains were investigated. An in vitro RNA-protein “pull down” system was developed using the rapid protein purification technique of the IMPACTTM-CN system. The coding sequence of the N terminal domain of p71 was fused to that of a chitin binding affinity tag (intein). This fusion protein was used as protein bait for the viral RNA. It was proposed that if RNA interacted with the fusion protein, it would be pulled down by the mass of affinity matrix and be precipitated and fluoresce when analysed by agarose gel electrophoresis using ethidium bromide. Despite optimisation of the in vitro assay, results were affected by the interaction between the intein-tag and nucleic acids, the state of the expressed fusion protein (in particular self-cleavage) and the excessive fluorescence present on the gels. The ADH2-GAPDH yeast expression system was used to investigate the in vivo assembly of p71 containing deletions of either one or both clusters within N terminal domain. It was found that all p71 mutants were expressed with the exception of the mutant containing a deletion of the second cluster. The reasons for this still require further investigation. The expressed p71 mutants were not processed into p64 and were degraded in vivo. In addition, an experimental attempt to purify assembled p71 mutant VLPs was unsuccessful. The assembly defect of p71 mutants emphasised the significance of the clusters, which are possibly required for interaction with viral RNAs for efficient VLP assembly. The results of this study suggest that an alternative tag or in vitro RNA-protein interaction assay be used. In addition, further experiments are required to investigate whether the co-expression of full length viral RNAs are required to rescue the in vivo assembly defect of p71 mutants into VLPs.

Thesis (Ph.D.) - University of Queensland, 2004. / Includes bibliography.