We were interested in identifying genes in the dipteran flies Calliphora erythrocephala and Drosophila melanogaster with a role in oogenesis and early embryogenesis. A biochemical screen was used to complement the extensive genetic screens that have been performed to identify such genes in Drosophila. Radio-labelled cDNA probes were made using poly(A)$\sp+$ RNA preparations from staged Calliphora ovaries and embryos. These probes were used to isolate clones which were strongly expressed during oogenesis but not during embryogenesis. Four Calliphora genes which are absolutely "oogenesis-specific" in their expression pattern, as defined by our screening protocol, were identified. These are called A10B, B8I, C7F and GG7K. Three of these clones are expressed in the somatically derived follicle cells of the ovary and have been identified as being homologous to the Drosophila yolk protein 1 (A10B and B8I) and a vitelline membrane protein (GG7K). Interestingly, the yolk protein homologs are expressed in a specialized subset of follicle cells known as the border cells in Calliphora. The fourth gene, (C7F) is expressed in the nurse cells, the transcripts are translocated to the oocyte proper and maintained throughout the first four hours of embryogenesis. C7F is also expressed in late pupae and adult male flies. A number of Calliphora genes were identified which, although not oogenesis-specific, were more strongly expressed in the oocyte than the embryo. These were classified as "oogenesis-differentials". C7F and the Calliphora oogenesis-differential genes were used to screen Drosophila cDNA and genomic libraries for homologs. Some characterization of these Drosophila homologs is described here. ME31B, a maternally expressed Drosophila gene from the 31B region of the second chromosome was isolated by directly screening Drosophila libraries with Calliphora cDNA probes. ME31B is expressed throughout oogenesis and the transcript is maintained in the mature egg until four hours after fertilization. The ME31B transcript is evenly distributed throughout the oocyte and egg. A 1.5 kb cDNA for ME31B was completely sequenced. Comparison of the coding sequence with a protein data bank allowed us to show that ME31B is a member of a family of NTP-dependent helicases. The possible mutant phenotype of ME31B is discussed.

The housefly, Musca domestica, was used to test the short-term and long-term environmental effects of high population density. The first phase addresses the issue that there are two main selection forces that drive mating behavior, inter- and intrasexual selection. In intersexual selection the females are actively discriminating when choosing a mate and, thus, these male-female interactions are what predominantly define that population. However, in intrasexual selection, males potentially compete against each other over the pool of receptive females. I tested the hypothesis that a less complex courtship would be optimal in a high-density environment, short-term (i.e., one generation). Specifically, I videotaped the mating behavior of individuals subjected to one of two treatments: high-density or low-density (i.e., 200 virgin male-female pairs in a 2 L or 114 L cage, respectively). In both treatments, the flies were allowed to mate for 30 minutes while being videotaped. The proportion of time spent in three male courtship behaviors (HOLD, FORWARD, BUZZ) and one female courtship behavior (WINGOUT) were determined. I found that the mating propensity (percent of mated pairs) was significantly greater in the high-density environment. The courtships in the high-density environment were also significantly less complex (i.e., less FORWARD, less HOLD). My findings suggest that high-density environments stimulated competition among males causing the intrasexual selection processes to outweigh the intersexual processes. The second phase tests the prediction that long-term (i.e., eight generations), high population density will drive the evolution of courtship repertoire towards decreased complexity. I applied the previously outlined methods. Additionally, only those pairs that mated within the allotted time were allowed to contribute to the following generation. The courtship behavior assays suggest that the synergistic effects of high density on the males and inbreeding depression drove the evolution of increased courtship complexity and exaggerated inbreeding depression, therefore, not supporting the prediction or the results of first phase. In the low-density treatment, courtship became less complex and mating propensity increased. These results are applicable to populations with unnaturally high densities and potential for inbreeding such as those in laboratory agricultural pest control, and conservation projects.

The Drosophila melanogaster mutation bicaudal (bic) is the founding member of the group of maternal effect mutations that disrupt anterior-posterior axis formation by creating mirror image duplications of posterior segments in the anterior half of the embryo. The gene affected by the bic mutation was cloned, and a recessive embryonic lethal mutation, $vr22\sp{P3},$ was shown to be an allele of the same gene, demonstrating that the gene is essential. During embryogenesis, bic gene expression was detected in mesoderm and anterior and posterior midgut, and persistent expression was found in the somatic musculature. Late in embryogenesis, transcripts were detected in the central nervous system. The bic gene was shown to be expressed in two distinct locations in ovaries: transcripts were detected in somatic cells of the germarium, and a second activation of transcription was found exclusively in the germ-line derived nurse cells of ovarian follicles. The maternal effect of the bic mutation was shown to originate in failure to express transcripts of the gene in the germ-line nurse cells during oogenesis. The bic gene was found to encode the Drosophila homolog of beta NAC (Nascent polypeptide Associated Complex), a recently discovered ribosome associated protein of eukaryotes. Beta NAC is a subunit of a heterodimer that associates with nascent polypeptides as they emerge from the ribosome. In vitro, NAC regulates the specificity of co-translational targeting of nascent chains to the Endoplasmic Reticulum. The studies of the bic mutant effects presented here demonstrated that beta NAC has other unpredicted ribosome-associated functions, being required to ensure localized expression of proteins via translational control of mRNA. In oocytes/embryos of bic mutant females, negative regulation of translation of the mRNA of nanos, the posterior determinant, fails; NANOS protein is aberrantly expressed in the anterior compartment of the early embryo, and consequently, bicaudal embryos are created. Negative regulation of nanos was thus shown to include ribosome associated events. Furthermore, these first in vivo studies of the activity of beta NAC (BIC) identified a new ribosome associated function for the protein, and established a physiologically relevant and informative molecular genetic model system for future studies of NAC.

Social insect workers often sacrifice their own reproduction so that they may help relatives to reproduce more successfully. However, genetically dissimilar colony-mates may prefer to aid different kin, and this may result in disruptive conflicts. I developed polymorphic microsatellite genetic loci for the African swarm-founding wasp Polybioides tabidus to examine mechanisms to reduce such conflicts. Swarm-founding wasps have many reproductive queens in their colonies which should lower relatedness, increasing the potential for conflicts. I found that even though P. tabidus colonies contained many queens, relatedness was elevated because new queens were only produced after the number of old queens had been reduced to one, or nearly one. Queens were thus highly related, elevating relatedness in the colony as a whole, and promoting sociality. This unique pattern of queen production is consistent with a worker manipulation of the sex ratios known as cyclical oligogyny. Under cyclical oligogyny, new queens are produced when the colony has few queens, while males are produced when the colony has many queens. The males were indeed produced when queen number was higher, and I found evidence that the workers collectively controlled male production. Colonies which produced normal haploid males also produced diploid males, which have a diploid genome but are homozygous at the sex determining locus. P. tabidus does not appear to effectively distinguish between diploid and haploid males, and diploid males should have occurred in colonies without haploid males too. Their absence indicates that they were actively eliminated from colonies in which the workers did not favor male production. Workers also may have controlled who produced the males. Each worker should prefer to produce the males herself. However, I found that the queens produced the males. This may be explained by collective worker policing because the workers would be more highly related to queen-derived males than to the sons of other workers and should prevent reproduction by other workers. Alternatively, each worker might restrain herself if worker reproduction was costly to the success of the colony. The results of this study indicate that collective worker control is an important mechanism stabilizing cooperation.

pushover (push) is a mutation that causes neuronal hyperexcitability, uncoordination and male sterility. Here it is also shown that push mutant wandering third instar larvae exhibit segmental nerves that are slightly thickened compared to wild type controls. The additional thickness of these nerves is due to a thicker perineural glia layer surrounding the axon bundle of the nerve. From investigation of push mutant testes, it was determined that mature sperm are produced, but are immotile. The push gene was cloned and sequenced and encodes a 5322 amino acid protein with 12 putative transmembrane helices. Sequencing of the push gene in the push 1 mutant line revealed a premature stop codon at amino acid position 728 and in push2 a premature stop codon at position 883. In situ hybridization experiments show push is expressed in the embryonic CNS and in primary spermatocytes in the adult testes and suggest that push is not expressed in the perineural glia of wandering third instar larvae. Because push affects but is not expressed in these glia, it is hypothesized that push is involved in a signaling pathway connecting the motor neuron to the perineural glia.

Although social hymenopteran colonies show a high level of cooperation among their members, colony members can have conflicts among themselves as well. One of these intra-colonial conflicts is who produces the males. I studied the resolution of conflict in stingless bees, a species-rich group with a tropical distribution. In the majority of stingless bee species both workers and queens are able to produce males. Therefore intracolonial conflict over male production is predicted. Because stingless bee queens mate only once, workers are more related to their own and to each other's sons than to the sons of the queen. Thus on genetic grounds, worker production of males is expected. However, workers might not reproduce if the costs of reproduction are high, or if the queen is able to suppress workers. The decision could have been made in the bygone times and the current pattern does not serve adaptive functions at the present. To test my predictions of conflict over male production I looked at three levels: within colonies, within species, and between species. On the colony and species level, I hypothesized that current conflict is expressed by behavioral antagonism between the workers and their queen. Furthermore, I predicted behavioral conflict to be higher in the periods when males are produced compared to periods with only female production. On the level of comparison between species I expected more signs of conflict in species where both workers and queen produce males than in species where males are all queen derived. The conclusions of this study concerning conflict over male production in stingless bees are: (1) Genetic tools confirmed that workers reproduce in some, but not in other species. (2) The costs involved with worker reproduction could explain why in some species workers reproduce and in others not. (3) There is not only a variance of worker reproduction between, but also within species. Demographical factors might be essential determining the amount of worker reproduction within species. (4) The pattern of worker reproduction could be explained by costs although phylogenetic relationships could explain the pattern also.

When considering how insecticides impact beneficial insects, in addition to direct mortality, we need to assess sublethal effects on population numbers which reduce the efficacy of the predators and parasitoids in reducing pest insects. The physiological sublethal effects can result in reduced longevity and reproduction. Understanding the sublethal effects of insecticides on beneficial insects is important in agroecosystems, especially in the context of natural biological control which aims to augment endemic natural enemies. The objective of this research study was to evaluate the environmental effects of insecticides on non target predator and parasitoid insects, combining the review of the literature and the synthesis of both a laboratory and a field research study. The lab study considered the effects of chlorpyrifos and deltamethrin on three natural enemies of Russian wheat aphid; the field study considered the effects of chlorpyrifos, deltamethrin, cyhalothrin-lambda and imidacloprid on ground dwelling arthropods in a Canola field.

This study comprises an examination of over 4000 male specimens, including nearly all type material, a detailed study of the genitalia, and a key to the majority of the species of Tricholabiodes Radoszkowski. Thirty species and subspecies are redescribed and 22 new species described. The 22 newly described species are: T. acer, T. alveolus, T. brothersi, T. concavus, T. convexus, T. denticidatus, T. disgregus, T. femoralis, T. ferrugineus, T. indistinctus, T. inornatus, T. longicarinatus, T. liiridus, T. parallel™, T. paulocellatiis, T. petiolatus, T. protitberans, T. recurvatus, T. sinuatus, T. thisboides, T. tortilis and T. trochantalis. Tricholabiodes semisthataeformis Bischoff and T. pathzii Invrea are synonyms of T. stigmaticus Bischoff and T. pallidicornis Bischoff, respectively. Phenograms and principal component plots were derived to clarify species status, make decisions on species limits and used to determine the morphological similarity between the species. The phenetic analysis was used only as a tool, and not a final product. For the determination of species limits, which included an analysis of 447 specimens, the continuous quantitative and coded characters were analysed separately. Forty-three continuous quantitative characters were analysed either as standardized measurements (against mesosoma length) or as ratios (32), since it was not possible, even via gap coding, to code these characters. Scatterplots and a phenogram from the principal components and cluster analyses respectively, are presented. Size and shape were not particularly helpful characters in determining species limits. One hundred and twenty five coded characters were analysed in a cluster analysis and part of the final phenogram is presented. For the determination of morphological similarity between the species, a hypothetical specimen, typical of each species, was derived. Again, one hundred and twenty-five coded characters were analysed in a cluster analysis and the final phenogram is presented. Representatives from each of the species and subspecies were examined with respect to 93 coded characters. The character states were polarised using the outgroup Dasylabroides Andre. Where Tricholabiodes had all states occurring in Dasylabroides, and the primitive state could not be identified, these characters, and their states, were considered for the entire tribe, and the sister tribe of Dasylabrini, Sphaeropthalmini, was taken as the outgroup. The cladograms were constructed with the software Hennig86. The most variable characters were eliminated from the analysis. Selection of the cladogram representing the most likely phylogeny of the genus was based on parsimony, resolution of the tree, character placement on the tree, comparison of the tree with weighted/unweighted consensus trees and biogeography. The phylogeny presented, which is to be regarded only as a hypothesis, suggests that Tricholabiodes underwent nine separate radiations. The southern African species are divided into two lineages: the first divergence stems from the base of the tree while the more recent lineage stems from the apex. Evidence suggests that the genus arose in central Africa, spreading south (twice) into southern Africa, north into North Africa, west across central North Africa and east into southeastern Asia. The study has also shown that the majority of the species are restricted in their distribution, with none of Palaearctic species occurring in southern Africa, and vice versa. It is hypothesised that the present distribution of the genus is partially restricted by dispersal ability and climate. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 1998.

Monthly sampling of 71 laboratory nests, each with a pair of adults, revealed that eggs and first-instar larvae were observed in the third month, minor workers in the fourth month and minor soldiers in the seventh month. Mortality of the pairs was high, with only five pairs surviving over the 10 month period. Laboratory nests did not develop further than the copularium. Excavation of 30 nests, of differing sizes, revealed that five were juvenile nests, consisting of only thin shelving with a few flattened fungus combs scattered throughout, and all, even the youngest nest (3 to 5 years), had a small mound. The queens from these young nests were small and had white pleural and intersegmental membranes. Twenty mature nests had a medium to large mound with large air passages and a medium to large hive with a well defined fungus garden containing large fungus combs. The queens from these nests were medium- to large sized, with white to brown pleural and intersegmental membranes. The remaining five nests had mounds often covered with grass, and a hive that contained less fungus comb than expected. The mounds of these nests were classified using their sandy pediment or crumbly texture. These were senescent or declining nests. The queens had pale brown pleural membranes and brown intersegmental membranes, and were often flaccid. The royal cell was commonly found in the middle to upper part of the nursery (20 nests), but sometimes was found at the edge of the nursery (five nests). The royal cells of five nests were not found or had been destroyed during excavation. The "youngest" mound was one to two years old and the "oldest" was more than 25 years old. The youngest queen was estimated to be three to five years old and the oldest queen more than 27 years. The nest seems to remain subterranean for two or less years before producing a mound. Thirteen nests were vigorous and five declining. The remaining 12 nests could not be classified as no fungus comb was collected from the nests. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1997.

Insecticide resistance in malaria vector mosquitoes reduces the efficacy of insecticide in killing and can therefore cause a major problem for malaria vector control by insecticides. In Mozambique, pyrethroid resistance in Anopheles funestus was first detected in December 1999 in the southern corner of Maputo Province. Since then, various collections have been made at selected sites throughout the country and WHO standard susceptibility tests and biochemical assays were conducted to determine the susceptibility status and the major resistance mechanisms, in the Fl generation of field collected mosquitoes. Three malaria vector species: Anopheles funestus s.s., Anopheles gambiae s.s. and Anopheles arabiensis were identified in this study by Polymerase Chain Reaction (PCR) and their distributions plotted. The susceptibility data indicate that the Anopheles funestus s.s population in southern Mozambique is widely resistant to pyrethroid and with low levels of carbamate resistance evident at six localities. No resistance to organophosphate and DDT was observed at any study sites. Biochemical tests indicate the presence of an altered acetlylcholinesterase in all collection localities with the exception of Massinga district. Elevated esterase activity with substrate a-naphthyl acetate were detect in Boane with a probable role in organophosphate resistance. Elevated GST were detected in Boane, Moamba and Catembe. Very low levels monooxygenase titres were registered in all the localities in Mozambique, which suggest that this resistance mechanism is not operating in these areas. Pyrethroid resistance in the Anopheles gambiae complex was detected only in Anopheles arabiensis from one locality. No resistant to other groups of insecticide were observed. Altered acetlylcholinesterases were registered in all collection localities and in both species: Anopheles gambiae s.s. and Anopheles arabiensis. Elevated esterase with substrate a-naphthyl acetate were detected in Anopheles arabiensis at only one locality. Elevated GSTs were detected at all localities and in both species. The implications of the findings for malaria vector control in Mozambique are discussed. / Thesis (M.Sc.)-University of Natal, Durban, 2003.