Thesis (Ph. D.)--University of Hawaii at Manoa, 2002. / Includes bibliographical references (leaves 133-142).

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A dissertation submitted to the Faculty of Science, University of the Witwatersrand, Johannesburg, in fulfilment of requirements for the degree of Master of Science. 2015. / SNAMA, the protein of interest in this thesis is found in the common model organism Drosophila melanogaster, also known as the fruit fly it is also found in all eukaryotic organisms but not in prokaryotes. SNAMA is a 1231 amino acid protein that belongs to the RbBP6 superfamily. Members of this family are characterized by a zinc finger motif, a DWNN domain (domain with no name) and a RING finger motif. The human RbBP6 contains the Rb-binding and p53-binding domains in addition. The mammalian RbBP6 hence interacts with p53 and Rb and it is important for the development and tumorigenesis as a negative regulator of p53. Bioinformatics studies show that transcription of the Snama gene is driven by a single TATA-less promoter which give rise to a single 3.9 kb transcript. However, experimental evidence confirming the promoter region has not being published. The main aim of this study was to examine the regulation of Snama by identifying the maximal promoter sequence that shows promoter activity in mammalian cell line. This was achieved by using specifically designed primers to amplify the putative Snama promoters, ligating promoters in reporter vector (pGL3 basic). The recombinant products used to transfect eukaryotic cells (Cos7, African green monkey cells) and determining the maximal promoter sequence that expresses luciferase activity. The promoter sequences were labelled with biotin attached to the primers and Electrophoretic mobility shift assay (EMSA) was conducted to confirm binding of proteins on the putative promoter fragments. The segment designated promoter 6 has maximal positive activity and many proteins in the cell extract bind to it shown by EMSA. Interestingly the longer fragment designated promoter 7 has less promoter activity. This may suggest that this fragment also contains some repressive elements.

Doctor of Philosophy(PhD), / Until recently, Northwestern Australia was thought to be relatively free of serious fruit fly pests. Although a noxious strain, present in Darwin since 1985, was widely believed to be an infestation of the Queensland fruit fly, Bactrocera tryoni, from the East coast, the fruit flies present outside this area were believed to be the benign endemic species, B. aquilonis. However, during the year 2000, infestations of fruit flies were discovered on major commercial crops in both Western Australia and the Northern Territory. It was not known whether these outbreaks were due to an invasion of the major pest species, Bactrocera tryoni, a change in the behaviour of B. aquilonis, or a hybridisation event between the two species. Finding the source of these outbreaks has been complicated by the fact that, since B. tryoni and B. aquilonis are virtually indistinguishable morphologically, it was not known which species are present in the region. Traditionally any tryoni complex fly caught in the Northwest was called B. aquilonis based solely on location. In order to get a good population profile of the region, an extensive trapping program was set up to include flies from urban areas, commercial crops and natural areas where the benign strain is thought to remain. Tests of genetic differentiation and clustering analyses revealed a high degree of homogeneity in the Northwest samples, suggesting that just one species is present in the region. The Northwest samples were genetically differentiated from the Queensland samples but only to a small degree (FST =0.0153). MtDNA sequencing results also showed a small degree of differentiation between these regions. A morphological study of wing shape indicated that there are some minor identifiable morphological differences between East coast and Northwest laboratory reared flies. This difference was greater than that seen between B. jarvisi populations across the same geographic range. The results suggest that the flies caught in the Northwest are a separate population of B. tryoni. Soon after pest flies were discovered in Darwin, a population became established in Alice Springs. This population had a low genetic diversity compared with Queensland and Darwin populations, and showed evidence of being heavily founded. In 2000, an outbreak was discovered in the nearby town of Ti Tree. Due to the geographic and genetic similarity of these populations, Alice Springs was determined to be the source of the Ti Tree outbreak. To investigate the founding of these populations, a program was developed to estimate the propagule size. Using a simulation method seven different statistics were tested for estimating the propagule size of an outbreak population. For outbreaks originating from populations with high genetic diversity, the number of alleles was a good estimator of propagule size. When, however, the genetic diversity of the source population was already reduced, allele frequency measures, particularly the likelihood of obtaining the outbreak population from the source population, gave more accurate estimates. Applying this information to the Alice Springs samples, it was estimated that just five flies were needed to found the major population in and around Alice Springs. For Ti Tree, the propagule size was estimated to be 27 flies (minimum 10). In 2000, a much larger outbreak occurred in the developing horticultural region of Kununurra in northern Western Australia. An important question for the management of the problem is whether there is an established fly population or the flies are reinvading each year. This population was found to have a large amount of gene flow from the Northern Territory. Within the Kununurra samples, one group of flies was genetically differentiated from all the other samples. This group came from a small geographic area on the periphery of Kununurra and appeared to be the result of an invasion into this area at the time when the population was building up following the dry season. A further threat to the Northwest horticultural regions comes from B. jarvisi. A recent increase in the host range of this species has lead to speculation that it may become a greater pest in Northwestern Australia. At the present time, protocols for the population monitoring and disinfestation of this species are not in place. Here it is shown that B. jarvisi eggs are more heat tolerant than B. tryoni eggs and that monitoring of B. jarvisi populations is possible using cue lure traps placed according to fruiting time and location of their favoured host, Planchonia careya.

The objective of this dissertation is the isolation of DNA sequences that are differentially represented in males and females of the Oriental fruit fly Bactrocera dorsalis, specifically by initiating a molecular characterization of Y chromosome sequences in this species. Cytological observations have established the presence of a diminutive Y chromosome in B. dorsalis males. To isolate DNA sequences from the Y chromosome, a special method of genomic DNA isolation known as Representational Difference Analysis (RDA) was utilized to obtain DNA sequences unique to the B. dorsalis male genome. Genomic DNA from B. dorsalis males served as the "tester" DNA and female genomic DNA as the "driver" DNA. Six distinct RDA products were obtained following two complete rounds of DNA hybridization and difference enrichment via the Polymerase Chain Reaction (PCR). One ofthese products (RDA product 1) was used to isolate a genomic DNA clone (3.1a) from a B. dorsalis male genomic DNA minilibrary. This sequence shows similarity to the reverse transcriptase of R1 retrotransposable elements. The presence of R1 elements in the Tephritid insects has heretofore been undescribed, although these elements have been previously described in the genomes of other Dipteran species. Oligonucleotide primers for PCR were designed for the 3.1a clone. These primers consistently produce different amplification patterns in PCRs ofgenomic DNA from B. dorsalis males vs. females. Amplification using male genomic DNA produces 325 bp and 2.6 kb products while only a 2.6 kb product is obtained from female DNA. The amplification products obtained with these primers are also produced in PCRs of genomic DNA from B. dorsalis embryos and third instar larvae, suggesting the ability of this method to infer sex at pre-adult stages ofthe B. dorsalis life cycle. Similar amplification products have also been obtained in other Bactrocera species. Both the 325 bp male PCR product and the 2.6 kb products have regions of sequence similarity to R1 elements. The 2.6 kb product contains a putative 1.7 kb open reading frame (ORF) encoding 583 amino acids. Three amino acid motifs found in Drosophila R1 element reverse transcriptases are present in comparable locations within the hypothetical ORF product. Both of these sequences are also repetitively represented in the B. dorsalis male and female genomes. However, the 325 bp male product produces some bands that are male specific when used as a probe for Southern blots of B. dorsalis male and female genomic DNA. The amplification pattern produced by the 3.1a primers is consistent with what would be expected if the 2.6 kb and 325 bp PCR products originated from the B. dorsalis X and Y chromosomes, respectively. Thus, the cloned male-specific sequence recovered here is potentially useful both as a gateway into the relatively uncharacterized B. dorsalis Y chromosome and as a tool for the characterization of other aspects of the B. dorsalis genome.

Thesis (Ph. D.)--University of Hawaii at Manoa, 1991. / Includes bibliographical references (leaves 71-87) / Microfiche. / x, 87 leaves, bound ill. 29 cm

Thesis (Ph. D.)--University of Hawaii at Manoa, 1991. / Includes bibliographical references (leaves 167-173) / Microfiche. / xxiv, 212 leaves, bound 29 cm

Doctor of Philosophy(PhD), / Until recently, Northwestern Australia was thought to be relatively free of serious fruit fly pests. Although a noxious strain, present in Darwin since 1985, was widely believed to be an infestation of the Queensland fruit fly, Bactrocera tryoni, from the East coast, the fruit flies present outside this area were believed to be the benign endemic species, B. aquilonis. However, during the year 2000, infestations of fruit flies were discovered on major commercial crops in both Western Australia and the Northern Territory. It was not known whether these outbreaks were due to an invasion of the major pest species, Bactrocera tryoni, a change in the behaviour of B. aquilonis, or a hybridisation event between the two species. Finding the source of these outbreaks has been complicated by the fact that, since B. tryoni and B. aquilonis are virtually indistinguishable morphologically, it was not known which species are present in the region. Traditionally any tryoni complex fly caught in the Northwest was called B. aquilonis based solely on location. In order to get a good population profile of the region, an extensive trapping program was set up to include flies from urban areas, commercial crops and natural areas where the benign strain is thought to remain. Tests of genetic differentiation and clustering analyses revealed a high degree of homogeneity in the Northwest samples, suggesting that just one species is present in the region. The Northwest samples were genetically differentiated from the Queensland samples but only to a small degree (FST =0.0153). MtDNA sequencing results also showed a small degree of differentiation between these regions. A morphological study of wing shape indicated that there are some minor identifiable morphological differences between East coast and Northwest laboratory reared flies. This difference was greater than that seen between B. jarvisi populations across the same geographic range. The results suggest that the flies caught in the Northwest are a separate population of B. tryoni. Soon after pest flies were discovered in Darwin, a population became established in Alice Springs. This population had a low genetic diversity compared with Queensland and Darwin populations, and showed evidence of being heavily founded. In 2000, an outbreak was discovered in the nearby town of Ti Tree. Due to the geographic and genetic similarity of these populations, Alice Springs was determined to be the source of the Ti Tree outbreak. To investigate the founding of these populations, a program was developed to estimate the propagule size. Using a simulation method seven different statistics were tested for estimating the propagule size of an outbreak population. For outbreaks originating from populations with high genetic diversity, the number of alleles was a good estimator of propagule size. When, however, the genetic diversity of the source population was already reduced, allele frequency measures, particularly the likelihood of obtaining the outbreak population from the source population, gave more accurate estimates. Applying this information to the Alice Springs samples, it was estimated that just five flies were needed to found the major population in and around Alice Springs. For Ti Tree, the propagule size was estimated to be 27 flies (minimum 10). In 2000, a much larger outbreak occurred in the developing horticultural region of Kununurra in northern Western Australia. An important question for the management of the problem is whether there is an established fly population or the flies are reinvading each year. This population was found to have a large amount of gene flow from the Northern Territory. Within the Kununurra samples, one group of flies was genetically differentiated from all the other samples. This group came from a small geographic area on the periphery of Kununurra and appeared to be the result of an invasion into this area at the time when the population was building up following the dry season. A further threat to the Northwest horticultural regions comes from B. jarvisi. A recent increase in the host range of this species has lead to speculation that it may become a greater pest in Northwestern Australia. At the present time, protocols for the population monitoring and disinfestation of this species are not in place. Here it is shown that B. jarvisi eggs are more heat tolerant than B. tryoni eggs and that monitoring of B. jarvisi populations is possible using cue lure traps placed according to fruiting time and location of their favoured host, Planchonia careya.

Thesis (Ph. D.)--University of Hawaii at Manoa, 2002. / Includes bibliographical references (leaves 178-190).

<p> Social learning, defined as learning from other individuals has been well studied in vertebrates and social insect species. I used fruit fly egg laying site selecting as a paradigm to test for social learning in a non-social insect. Focal females that experienced novel food together with mated females (models), who had laid eggs on that food, subsequently exhibited a stronger preference for laying eggs on that food over another novel food than focal females that experienced the same food alone. This socially influenced learning was also observed when focal females experienced both foods, one with mated models and their eggs and one alone. In contrast to the strong effect of a food with mated models and their eggs, neither a food with virgin models nor the aggregation pheromone (cVA) alone generated socially influenced learning. These results provide the first evidence for social learning about egg laying substrate in fruit flies. Further research utilizing fruit flies as a model system may help us gain a better understanding of the evolution and neurogenetics of social learning.</p> / Thesis / Master of Science (MSc)