Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.722034
Title: Mechanisms of insecticide resistance in Indian malaria vector Anopheles stephensi
Author: Dykes, C. L.
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2017
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Abstract:
To understand the mechanisms of insecticide resistance in Anopheles stephensi, DDTand pyrethroid-resistant lines were selected in the laboratory. Selection process showed fixation of L1014S-kdr mutation in DDT- and pyrethroid resistant colonies. Similar fixation was recorded in unselected mosquitoes (control) as well, suggesting low fitness cost of L1014S in laboratory conditions. Synergistic assays using piperonyl butoxide (PBO), a mixed function oxidase and esterase inhibitor, led to reversal of resistance in pyrethroid resistant mosquitoes but no reversal in DDT resistant mosquitoes was recorded. Elevated GST activity in laboratory strains led us to characterize the DDT resistance implicated GST epsilon genes. Quantitative-PCR showed elevated expression of AsGSTe2 and AsGSTe4 in DDT-resistant mosquitoes in both laboratory selected DDT-resistant strain and field caught mosquitoes. These observations prompted further investigations to molecularly characterize these genes examining mutational changes and the possible roles of allelic variation. Cloning and sequencing of the full genes revealed polymorphism which resulted in four variants in AsGSTe2 and three variants in AsGSTe4. Of the four variants of AsGSTe2, two variants (AsGSTe2.1 and AsGSTe2.2) found in DDT-resistant individuals were expressed in vitro in E.coli. Recombinant expression and DDTase assays of AsGSTe2.1 and AsGSTe2.2 showed them to efficiently metabolise DDT. DDTase activity examined for recombinant AsGSTe4 (AsGSTe4.1 and AsGSTe4.2 and AsGSTe4.3) showed that they did not metabolise DDT. Enzyme thermostability tests showed AsGste2 variants to be highly unstable compared to the orthologues in An. gambiae, Aedes aegypti and its corresponding AsGSTe4 variants. Further examination into the GST epsilon array provided evidence of tandem coduplication of AsGSTe2 and AsGSTe4 together in the GST-epsilon array of the laboratory DDT-resistant colony.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.722034  DOI: Not available
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