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The effects of natural Wolbachia bacteria on their Anopheles mosquito hosts and implications for malaria biocontrol strategies

Principal Supervisor: Prof Thomas WalkerLink opens in a new window

Co-supervisor: Erin Gorsich

PhD project title: The effects of natural Wolbachia bacteria on their Anopheles mosquito hosts and implications for malaria biocontrol strategies

University of Registration: University of Warwick

Project outline:

Wolbachia is an endosymbiotic bacterium being used for mosquito biocontrol strategies. Wolbachia transinfected Aedes (Ae.) aegypti mosquitoes which both replace wild populations and block transmission of major arboviruses, such as dengue virus (DENV), are now being applied for disease control. The wMel Ae. aegypti line (Walker et al. Nature 2011) has been released and established in more than ten DENV-endemic countries and has significantly reduced DENV incidence including by 77% in the latest randomised controlled trial in Indonesia. Globally, DENV causes ~10,000 deaths/year but this is signficantly lower than ~409,000 deaths caused by malaria. Despite a decrease in malaria since 2000 and encouraging progress in vaccine development, emergence of insecticide resistance has reduced the effectiveness of traditional vector control, therefore novel approaches are now critical.

Recently studies have reported the potential presence of Wolbachia strains in wild An. gambiae and An. funestus complex populations although mostly with highly sensitive nested-PCR of only limited genes and report low prevalence rates. In contrast, we have discovered strains in An. moucheti (wAnM) and An. demeilloni (wAnD) at significantly higher densities which can be visualised in mosquito ovaries by microscopy (Walker et at. Current Biology 2021). There are several reasons why wAnM and wAnD should be further characterised in natural hosts. Firstly, Wolbachia density is positively correlated with arboviral inhibition suggesting strains would significantly inhibit malaria parasites. Secondly, genome analysis provides evidence for the presence of CI factor (cif) gene homologs and high prevalence rates would be consistent with CI induction. Our preliminary analysis also suggests the wAnD and wAnM strains dominate the host microbiome when present in both wild and early colony generations of An. demeilloni and An. moucheti.

In this PhD project proposal, the candidate would firstly look to further investigate the mosquito microbiome using targeted barcoded high-throughput amplicon sequencing of the bacterial 16S rRNA gene to determine if Wolbachia is the dominant bacterial species in different life stages including the immature stages. This is important given Anopheles mosquitoes can acquire other bacteria from their environment – some of which (eg. Asaia) have shown to compete with Wolbachia in laboratory populations. Secondly, the project would look to investigate insecticide resistance in An. moucheti and An. demeilloni. Insecticide resistance has been shown to be a key reason why releases of Wolbachia-infected Ae. aegypti strains have failed to establish. Furthermore, there is growing evidence of an association between the Anopheles mosquito microbiota and insecticide resistance. This Phd project would look to measure phenotypic insecticide resistance in Wolbachia-infected and equivalent uninfected individuals. CDC resistance intensity bioassays for three pyrethroid insecticides will be conducted and RNA-seq will be carried out on naturally infected individuals to determine any interaction between Wolbachia strains and expression of key enzymes/pathways associated with insecticide resistance. RNA-seq reads will be screened for target site mutations associated with eg. DDT, pyrethroid resistance and known voltage-gated sodium channel mutations. qRT-PCR validation of RNA-seq data will be undertaken on differentially transcribed insecticide resistance genes. Finally, the effect of Wolbachia strains on mosquito immunity will be assessed through RNA-Seq to determine differential expression levels of mosquito immune genes such as cecropin D, defensin and transferrin. qRT-PCR validation of RNA-seq data will also be undertaken on differentially transcribed insect immune genes.


Walker, T. et al & Hughes, G.L. (2021). Stable high-density and maternally inherited Wolbachia infections in Anopheles moucheti and Anopheles demeilloni mosquitoes. Current Biology, S0960-9822(21)00429-2. 

Walker, T. et al & Hoffmann, A.A. (2011). The wMel Wolbachia strain blocks dengue and invades caged Aedes aegypti populations. Nature, 476, 450-453.

BBSRC Strategic Research Priority: Understanding the rules of life Immunology, and Microbiology.


Techniques that will be undertaken during the project:

mosquito field collection and preservation

nucleic acid extraction

quantitative PCR

barcoded high throughput amplicon sequencing and analysis

phenotypic insecticide resistance assays

RNA sequencing and analysis


Contact: Prof Thomas WalkerLink opens in a new window