Gene mutation allows tracking of insecticide resistance in mosquitoes

Africa accounts for 85% of malaria cases and 90% of malaria deaths worldwide. Control strategies rely heavily on insecticide-based interventions such as Indoor Residual Spraying (IRS) and Long Lasting Insecticide Nets (LLINs). However, resistance to insecticides in mosquitoes such as Anopheles funestus is threatening the effectiveness of these tools. Wellcome Trust Senior Fellow Dr Charles Wondji from the Liverpool School of Tropical Medicine has discovered a genetic marker to track this resistance through a population. We asked him to explain his findings… 

Insecticide resistance is threatening the global control of malaria. The WHO Global Plan for Insecticide Resistance Management (GPIRM) has now recognised that if suitable management strategies are not developed, this resistance will have devastating public health consequences. The race is on to preserve the potency of current insecticides and the key to winning could lie in the genes of the mosquito.

Genetic information on insecticide resistance genes, and associated markers to spot them in mosquito populations, could enable us to track and anticipate the spread of resistance. We could even assess the impact of resistance on malaria transmission and design management strategies accordingly.  

Currently, we cannot track insecticide resistance in this way and this is a major obstacle when designing management strategies. Our team decided to look into the molecular basis of resistance to key insecticides in the mosquito Anopheles funestus, to see if we could overcome this problem.

It’s all in the genes

In our study published in the journal Genome Biology, we found that a single genetic mutation is associated with resistance to a range of insecticides including two widely used ones: DDT and pyrethroids. This is the first time such a marker for resistance has been identified in a mosquito species.

Our aim had been to find out how mosquitoes withstand exposure to the fatal insecticide DDT without dying and it turns out this is due to a single mutation in the GSTe2 gene. This mutation helps insects break down DDT molecules more efficiently until they are no longer toxic. There also appears to be an association between the mutated GSTe2 gene and resistance to other major insecticides used in public health.

The Study

The study started in Pahou in Benin when we found a population of mosquitoes that was fully resistant to both DDT and pyrethroids. We wanted to elucidate the molecular basis of that resistance in the population and design a simple diagnostic tool which could be used for detection and monitoring.

We took mosquitoes from that population in Pahou, as well as laboratory mosquitoes that were fully susceptible to DDT and pyrethroids, and compared their genomes via a genome-wide comparison study.

Double – Protection

We found that mosquitoes with resistance had significantly elevated levels of the GSTe2 protein, revealing an association between the up-regulation of the protein and resulting resistance.

In addition, a single mutation in the GSTe2 gene in mosquitoes from Benin, enabled the resulting GSTe2 protein to break down DDT more efficiently.

Our subsequent DNA-based diagnostic test for this type of resistance confirmed that this mutation strongly correlates with resistance in mosquitoes not only from Benin, but also other regions with DDT resistance in Africa.

How it works

Using X-ray crystallography to visualise the GSTe2 protein showed us that the mutation conferred resistance by opening up the ‘active site’ where DDT molecules bind to the protein to enable more DDT to attach and be broken down. The mosquitoes were surviving by breaking down more of their poison into non-toxic substances.

To take this one step further, we also introduced the gene into the common fruit fly, Drosophila melanogaster, and found they also became resistant to DDT and pyrethroids. This confirmed the association between this single mutation and insecticide resistance. We had unlocked the keys to resistance and have now used this to develop a way to identify this mutation in the field.

Tracking resistance in the field

Our diagnostic assays will allow control programs to detect and track resistance at an early stage in the field, which is an essential requirement to successfully tackle the growing problem of insecticide resistance. The TaqMan assays use fluorescence tagging to spot resistance in-country by readily identifying this mutation in a mosquitoes’ DNA at room-temperature in the lab.

My recently awarded Wellcome Trust Senior Fellowship in biomedical sciences will enable us to build on this work. We hope to detect more molecular markers to track resistance and understand the evolution of resistance and its impact on control interventions, as well as develop new diagnostic tools. If we can discover this, we could monitor the spread of resistance and anticipate its movements to tailor control programmes and slow down the global spread of this type of resistance.

The full paper is available to now: A single mutation in the GSTe2 gene allows tracking of metabolically-based insecticide resistance in a major malaria vector 

Image credits:  Chris Nurse, Wellcome Images