3. Electronic Theses and Dissertations (ETDs) - All submissions
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Item Comparative insecticide resistance intensity studies between the pyrethroid resistant wild Zambian and the laboratory reared Mozambique Anopheles funestus(2015) Segoea, GodiraMalaria continues to be a significant cause of morbidity and mortality in the developing world with Africa being the most affected. Malaria vector control with chemical insecticides is the primary intervention to curb transmission. However, the success of chemical based interventions is threatened by the escalating development of insecticide resistance in the major anopheline mosquito vectors.Item Insecticide resistance and vector status of Anopheles funestus and An. gambiae populations at a sugar estate in Mozambique(2010-04-12T09:55:50Z) Kloke, Ronald GrahamMalaria is on the increase in Mozambique since 2001 and impacts primarily on children < 5 years of age. Insecticide resistance in the malaria vector mosquitoes is on the increase in Mozambique and Africa and is cause for serious concern. Maragra sugar estate is situated in close proximity to the nKomati river floodplain in a rural area in Mozambique and requires intense irrigation for cane growing and as a result provides extensive breeding sites for An. funestus and other mosquitoes. In the areas surrounding the estate there are two important vectors of malaria, Anopheles funestus group and An. gambiae complex. There is intense malaria transmission in the areas surrounding the sugar estate and the last entomological study on the vectors in the Manhica area was done in 1998. It was becoming increasingly urgent to identify to species level the vectors in this area and to monitor the insecticide resistance status of these vectors. Due to leakage (theft) of insecticides and a change by the National Malaria Control Programme (NMCP) to an insecticide to which the predominant vector is resistant, an entomological survey was carried out in this area from January 2009 to March 2009 to ascertain by Polymerase chain reaction (PCR) what species of malaria vectors were present inside and outside of the Maragra vector control area, their population levels and their vectorial status in these two areas. Insecticide resistance studies by insecticide exposure and the synergist piperonyl butoxide (pbo) were carried out using the World Health Organisation (WHO) bioassay method on collected An. funestus mosquitoes. This was done to establish this species resistance status to the four classes of insecticides recommended by the WHO for malaria vector control. The collections of An. arabiensis and An. merus that were identified were too few to carry out insecticide resistance tests on these two species. Enzyme linked v immuno-sorbent assay (ELISA) tests were undertaken to establish the vectorial capacity of Anopheles funestus and An. gambiae complex in this area. The predominant malaria vector species in this area is An. funestus s.s., with the secondary vector being An. arabiensis. An. funestus has a high vectorial capacity in this area and found to have a Plasmodium falciparum sporozoite rate of 6.02%. This is an increase in the sporozoite rate of 1.2% from 1998 when the last survey in this regard was carried out. Coupled with this increase is an increase in the An. funestus populations in this area since this time. One An. gambiae complex sample was found to be positive but the species is not known as this particular sample did not amplify on PCR. Anopheles funestus is highly resistant to synthetic pyrethroids and exhibits a lower level of resistance to bendiocarb, a carbamate insecticide in use at Maragra sugar estate. The synergist pbo mediates the resistance mechanism in both these insecticides indicating that the metabolic resistance mechanism present in this mosquito is strongly mediated by monooxygenase detoxification. The role of the medical entomologist is increasingly necessary and important in the monitoring of this resistance phenomenon in malaria vector mosquitoes, as is the role of the vector control programme manager in implementing and managing vector control programmes. The implication of cane sugar farming and its impact on vector production and malaria transmission is discussed. Insecticide resistance and the change by the NMCP to a synthetic pyrethroid to which the predominant vector of malaria is resistant is discussed.Item Role of monooxygenases in insecticide resistant anopheles funestus(diptera: culicidae)(2007-02-26T10:20:08Z) Amenya, Dolphine Achieng'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%.