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Larvae of the common green lacewing (GLW), Chrysopa carnea Stephens, have been reported tolerant to synthetic pyrethroid insecticides including permethrin (C/T) (3-phenoxybenzyl-3-(2,2-dichlorovinyl)-2,2-dimethyl-cyclopropanecarboxylate), the trans isomer being more toxic than cis. An investigation was performed to determine the possible role of metabolism in this tolerance. Following topical application, GLW metabolized 80% of cis and 71% of trans within two hrs. About 95% of both cis and trans were metabolized by 50 hrs. Metabolism of C/T in vitro was compared to a susceptible insect, the tobacco budworm (TBW), Heliothis virescens (F.). GLW degraded cis 1.7-fold faster than trans, while TBW metabolized trans at a slightly higher rate than cis. When esterases and oxidases were active together or alone, cis and trans were metabolized faster by GLW than TBW. Metabolism of C/T by GLW was primarily oxidative with hydrolysis as a secondary mechanism. Trans is more toxic to GLW apparently because of this isomer's lower rate of detoxication. Several metabolites of C/T, cis-, and trans-permethrin were identified in studies with GLW in vivo and in vitro. It appeared that cis was metabolized more intensively than trans in vivo. The roles of esterases and oxidases in metabolizing C/T, cis-, and trans-isomers were studied in vitro and the following number of metabolites were identified: with C/T--five with esterases plus oxidases, six with esterases, and seven with oxidases alone. With cis--six metabolites were produced when esterases plus oxidases were active, five with esterases, and four with oxidases alone. With trans--esterases plus oxidases produced four metabolites, three with esterases alone, and seven with the oxidases alone. A few unknowns were exhibited in each case. Hydroxylation at the 2'-position of the phenoxybenzyl group seems to be important for GLW tolerance to C/T. Hydroxylation could be the first step in detoxifying C/T and its isomers. The toxicity of trans to GLW could be explained by the limited routes by which esterases acting alone can degrade this isomer; only three metabolites were produced with esterases while seven were produced with oxidases.

Call number: LD2668 .T4 1986 J86 / Master of Science / Biology

Malaria is the leading cause of mortality and morbidity in Sudan. The annual malaria cases and deaths are estimated at 7.5 million and 35, 000 respectively. One of the possible factors that have led to this situation is the development of insecticide resistance in the main malaria vector in Sudan, Anopheles arabiensis. This study therefore, was initiated to identify the malaria vectors in Gezira and Sennar states of central Sudan, determine their susceptibility levels to the different classes of insecticides used for malaria vector control, identify mechanisms of resistance, and determine the sporozoite infection rate and the blood meal sources in these populations. The polymerase chain reaction (PCR) for species identification revealed that An. arabiensis was the only member of the An. gambiae complex present in the study area. The blood meal analysis using ELISA showed high anthropophily with 89.2% feeding on humans. The overall sporozoite infection rate was 2.3 %. WHO susceptibility tests showed complete susceptibility of An. arabiensis to bendiocarb (100% mortality) and multiple resistance to permethrin (54-78%), DDT (55-66%) and malathion (76-78%). The kdr mutation analysis revealed the presence of the West African kdr allele with the majority of specimens being heterozygous (RS). The kdr in DDT/permethrin susceptible specimens were: 15% homozygous for the kdr mutation (RR), 64.2% heterozygous (RS) and 20.8% homozygous for the susceptible allele (SS). Amongst the DDT/permethrin resistant specimens, 13% were SS, 48.7% RS and 38.3% RR. The apparent lack of correlation between kdr and resistant phenotype strongly suggests that other resistance mechanisms are playing a role.

Sexual conflict can occur whenever the evolutionary interests of males and females differ, and when sexually antagonistic selection acts upon traits shared between the sexes, one or both sexes can be constrained from reaching their phenotypic optima. This intralocus sexual conflict can be characterised by a tug-of-war of allelic replacement until it is resolved, but examples of well-characterised sexually antagonistic loci are rare. This thesis investigates the basis and dynamics of intralocus sexual conflict over insecticide resistance at the Cyp6g1 locus in Drosophila melanogaster, and wing colouration in Drosophila simulans. In D. melanogaster, the Cyp6g1 locus is the site of a series of insecticide resistance alleles, one of which is sexually antagonistic when back-crossed to the old isogenic lab strain Canton-S. I investigated the presence of sexual conflict over this same allele in a recently collected and genetically heterogeneous population. I found evidence of balancing selection on resistance (Ch. 2) that could not be explained by overdominance or sex-specific dominance (Ch. 3). However, balancing selection could be explained by resistance conferring increased fecundity to females (Ch. 2-4), and decreased reproductive success to males (Ch. 4). This male cost can in turn be explained by a negative genetic correlation between reproductive success and Cyp6g1 expression (Ch. 4), possibly influencing levels of reproductive investment (Ch. 2). Additionally, I explored the dynamics of the sex-specific fitness effects of resistance across three Cyp6g1 alleles back-crossed to a single genetic background. I found no evidence of sexual antagonism, but revealed that the cost of resistance increased with more derived alleles, and that all alleles were more costly to females (Ch. 5). After decades of strong selection imposed by insecticide use an unresolved sexual conflict persists at the Cyp6g1 locus despite sexual dimorphism in resistance, and it does not appear that more derived Cyp6g1 alleles are necessarily involved in mediating this conflict. Wing interference patterns (WIPs) are a newly discovered trait subject to female mate choice in Drosophila. I explored the potential for intralocus sexual conflict over WIPs by measuring WIP traits from males and females from populations of D. simulans evolved under relaxed or elevated sexual selection. In response to sexual selection male WIPs evolved to be brighter, higher contrast, and shifted to longer wavelengths of light, but there was no associated response to selection in females (Ch. 6). While WIPs did not appear to be constrained from detectably responding to selection by acute intralocus sexual conflict, male WIPs from the relaxed selection regime were similar to female WIPs, suggesting a cost to sexually selected WIPs that may be indicative of sexually antagonistic selection. IASC is pervasive and can influence a wide range of fundamental evolutionary processes including sexual selection, speciation, and extinction. The research presented in this thesis adds to a body of evidence that sexual dimorphism does not necessarily resolve IASC, and documents the first evidence that WIPs do not appear to be subject to acute IASC and can evolve in response to sexual selection.

Student Number : 0318930A - PhD thesis - School of Animal, Plant and Environmental Studies - Faculty of Science / The widespread use of pyrethroid insecticides has led to the emergence of significant insecticide resistance in various parts of the world. An unprecedented increase in the number of annual malaria cases reported in Kwazulu Natal, South Africa in 1999 to 2000 was attributed to the re-emergence of pyrethroid-resistant Anopheles funestus Giles. Resistance was metabolic-based with increased monooxygenase (P450) metabolising the pyrethroid insecticides. This emphased the need to understand the molecular mechanisms conferring pyrethroid resistance in An. funestus. The present study aimed to firstly isolate P450 genes in An. funestus and secondly, to identify P450 gene over-expressed in a resistant (FUMOZ-R) strain compared to a susceptible (FANG) strain. A third aim was to construct an An. funestus cDNA library to lay the foundation for future studies on P450 monooxygenses. Degenerate primers based on conserved regions of three An. gambiae P450 families were used to amplify cDNAs from An. funestus. Eleven CYP4, four CYP6 and five CYP9 partial genes were isolated and sequenced. BLAST results revealed that An. funestus P450s have a high sequence similarity to An. gambiae with above 75% identity at the amino acid level. The exception was CYP9J14. The An. gambiae P450 with the closest similarity to CYP9J14 exhibited only 55% identity suggesting a recent duplication event in CYP9J14. Molecular phylogenetic analysis also supported this hypothesis. Intron positions were highly conserved between the two species. Expression studies using blot analysis implicated CYP6P9, an ortholog of CYP6P3 in An. gambiae, as the over-expressed P450. Dot blot analysis revealed a 500-fold expression higher in FUMOZ-R strain compared with FANG strain. Semiquantitative PCR revealed that CYP6P9 was developmentally regulated. Expression was not detected in eggs and was higher in larvae compared to pupae. Quantitative real time PCR showed that CYP6P9 expression was 4.5-fold higher in 3-day old FUMOZ-R males than females and 3.5-fold higher in the 14-day old males than 14- day old females. Statistically, this difference was not significant suggesting that CYP6P9 expression is not sex specific. The An. funestus cDNA library construction in λTriplEx2 vector was successful with a titre of 4.9 x108 pfu/ml and a transformation efficiency of 98%.

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