Cis-regulatory CYP6P9b P450\xa0variants associated with loss of insecticide-treated bed net efficacy against Anopheles funestus

Leon M J Mugenzi,Jack Hearn,Micareme Tchoupo,Jacob M Riveron,Gareth D Weedall,Magellan Tchouakui,Charles S Wondji,Benjamin D Menze,Murielle J Wondji,Helen Irving

doi:10.1038/s41467-019-12686-5

Abstract

Elucidating the genetic basis of metabolic resistance to insecticides in malaria vectors is crucial to prolonging the effectiveness of insecticide-based control tools including long lasting insecticidal nets (LLINs). Here, we show that cis-regulatory variants of the cytochrome P450 gene, CYP6P9b, are associated with pyrethroid resistance in the African malaria vector Anopheles funestus. A DNA-based assay is designed to track this resistance that occurs near fixation in southern Africa but not in West/Central Africa. Applying this assay we demonstrate, using semi-field experimental huts, that CYP6P9b-mediated resistance associates with reduced effectiveness of LLINs. Furthermore, we establish that CYP6P9b combines with another P450, CYP6P9a, to additively exacerbate the reduced efficacy of insecticide-treated nets. Double homozygote resistant mosquitoes (RR/RR) significantly survive exposure to insecticide-treated nets and successfully blood feed more than other genotypes. This study provides tools to track and assess the impact of multi-gene driven metabolic resistance to pyrethroids, helping improve resistance management.

Highlights

Elucidating the genetic basis of metabolic resistance to insecticides in malaria vectors is crucial to prolonging the effectiveness of insecticide-based control tools including long lasting insecticidal nets (LLINs)
When the southern Africa population (Malawi) is compared to all other regions directly, the striking difference is the high upregulation of two P450 genes CYP6P9a and CYP6P9b in Malawi with fold change (FC) of 28.3, 22.7 and 9.5 greater than in Cameroon, Uganda and Ghana for CYP6P9a (Supplementary Data 1)
To detect potential genetic variants associated with pyrethroid resistance, we focused our attention on the CYP6P9b P450 because this gene was highly over-expressed in resistant mosquitoes especially in southern Africa and after the recent characterisation of CYP6P9a, the other over-expressed gene in southern Africa[13]

Summary

Introduction

Elucidating the genetic basis of metabolic resistance to insecticides in malaria vectors is crucial to prolonging the effectiveness of insecticide-based control tools including long lasting insecticidal nets (LLINs). Metabolic resistance, considered to be more likely to cause control failure[7], remains—despite recent progress8–11— poorly characterised due to the complexity of this resistance mechanism[12] This is the reason why several cytochrome P450 genes associated with pyrethroid resistance have been reported, only a single DNA-based marker has been detected to date[13] limiting the design of field applicable diagnostic tools to detect and track this resistance. There is a need to characterise the molecular basis of pyrethroid resistance Africawide and detect related genetic variants so that DNA-based diagnostic assays can be designed to enhance our ability to detect resistance and rigorously characterise the impact on insecticidebased interventions

Methods

Results

Conclusion