Investigating the role of glutathione and glutathione biosynthetic genes in the adaption of Anopheles gambiae to insecticides

Malaria remains a serious public health challenge in the tropical world, with 584,000 deaths globally in 2013, of which 90% occurred in Africa, and mostly in pregnant women and children under the age of five. <i>Anopheles gambiae</i> (<i>An</i>. <i>gambiae</i>) is the principal malaria vector in Africa, where vector control measures involve the use of insecticides in the forms of long-lasting insecticide-treated nets (LLINs) and indoor residual spraying (IRS). The development of insecticides resistance mitigates these approaches. Glutathione (GSH) is widely distributed among all living organisms, and is associated with detoxification pathways, especially the Glutathione S-transferases (GSTs). Its direct involvement and relevance in insecticide resistance in <i>An</i>. <i>gambiae</i> has not been determined. Thus, this work examines the contribution of GSH, its biosynthetic genes (<i>GCLM</i>, <i>GCLC</i>) and their possible transcriptional regulator <i>Nrf</i>2 in insecticide resistance in <i>An</i>. <i>gambiae</i> sampled from agricultural setting (areas of intensive agriculture) and residential setting (domestic area). Bioinformatics analysis, W.H.O. adult susceptibility bioassays and molecular techniques were employed to investigate. Total RNA was first isolated from the adults <i>An</i>. <i>gambiae</i> mosquitoes raised from agricultural and residential field-caught larvae which had been either challenged or unchallenged with insecticides. Semi-quantitative RT-PCR using gel image densitometry was used to determine the expression levels of <i>GCLM</i>, <i>GCLC</i> genes and <i>Nrf</i>2. Bioinformatics’ results established the presence of putative AGAP010259 (<i>AhR</i>) and AGAP005300 (<i>Nf</i>2e1) transcription factor binding sites in <i>An</i>. g<i>ambiae</i> GCLC and GCLM promoters <i>in silico</i>. <i>An</i>. <i>gambiae</i> <i>s.l</i>. studied here were highly resistant to DDT and permethrin but less resistant to bendiocarb. Both knockdown resistance (kdr) mutation variants L1014S and L1014F that confers resistance to pyrethroid insecticides were identified in both An. coluzzii and <i>An. arabiensis</i> sampled from northern Nigeria. The L1014F was much associated with <i>An</i>. <i>coluzzii</i>. A significant positive correlation (P=0.04) between the frequency of the L1014F point mutation and resistance to DDT and permethrin was observed. However, a weak or non-significant correlation (P=0.772) between the frequency of the L1014S point mutation and resistance was also found. L1014S and L1014F mutations co-occurred in both agricultural and residential settings with high frequencies. However, the frequencies of the two mutations were greater in the agricultural settings than in the residential settings. The levels of total, reduced and oxidized GSH were significantly higher in mosquitoes from agricultural sites than those from residential sites. Increased oxidized GSH levels appears to correlate with higher DDT resistance. The expression levels of <i>GCLM</i>, <i>GCLC</i> and Nrf2 were also significantly up-regulated in adults <i>An</i>. <i>gambiae</i> raised from agricultural and residential field-caught larvae when challenged with insecticide. However, there was higher constitutive expression of <i>GCLM</i>, <i>GCLC</i> and <i>Nrf</i>2 in mosquitoes from agricultural setting. The increased expression levels of these genes and also GSH levels in this population suggest their roles in the response and adaptation of <i>An. gambiae</i> to insecticide challenges. There exists the feasibility of using GSH status in <i>An. gambiae</i> to monitor adaptation and resistance to insecticides.

Name dc:type.qualificationname

PhD

Level dc:type.qualificationlevel

Doctoral Thesis

Grantor dc:publisher.institution

Abertay University

Year dc:date.issued

2015

Author dc:creator

• Abdu, Habibu

Advisors dc:contributor.advisor

• Deeni, Yusuf
• Spiers, Andrew

• Anopheles gambiae
• GCLM
• Glutathione
• Insecticide
• Nigeria

Language dc:language

eng

Identifier

oai:rke.abertay.ac.uk:studenttheses/d30eb4df-64c0-43fe-ba6d-0fe778ff6b80

OAI identifier oai:identifier

oai:rke.abertay.ac.uk:studenttheses/d30eb4df-64c0-43fe-ba6d-0fe778ff6b80