
Figure 1: A map showing the two sampling locations on Bioko Island
Malaria control depends crucially on two widespread vector control methods; long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS). Both methods rely on the indoor application of insecticides, and can successfully target vectors that feed and rest indoors. However, the efficacy of LLINs and IRS is threatened by insecticide resistance and changes in the mosquito’s behavior. This behavioral resistance involves vectors shifting towards outdoor biting and resting. While the biological basis of insecticide resistance is well-studied, nothing is known about the genetic basis of behavioral resistance. The malaria vector Anopheles gambiae (M) on Bioko Island has been subjected to IRS under the Bioko Island Malaria Control Project (BIMCP) since 2004. This has resulted in a shift of this mosquito’s behavior towards outdoor host seeking (Fig 2 and 3, Reddy et al 2010).

Figure 2: The proportion of indoor and outdoor host seeking An. gambiae and An. melas during 2009 in Mongola, Bioko Island. Reports prior to the start of the BIMCP in 2004 note the absence of outdoor feeding on the Island (from Reddy et al 2010).
Various pieces of data suggest that the behavioral resistance of these populations has a strong genetic basis and that it was selected for by insecticide use. We are applying a whole genome sequencing approach to map loci involved in outdoor biting in An. gambiae (M) in two locations on Bioko Island. Outdoor and indoor biting mosquitoes were collected using human landing catches as part of the vector monitoring of the BIMCP from 2009 to 2013. These samples will sequenced, providing > 2,000,000 SNPs that will be examined to identify alleles that are associated with feeding preference.

Figure 3: Proportion of outdoor host seeking An. gambiae during the night, indicating that outdoor host seeking is most common in the evening hours, when many people are outside (from Reddy et al 2010).
We have developed a simulation approach for establishing the false discovery rate (=significance threshold) for our analyses. Preliminary results show that we will be able to detect alleles that differ in frequency by 0.069 between the indoor and outdoor feeding population. In addition, we will identify loci that increased in frequency following the vector control by comparing pre-intervention samples collected in 2004 to post-control 2009 and 2013 samples. Simulations that incorporate genetic drift and sampling effects will provide the false discovery rate, allowing the identification of loci under positive selection.
These two approaches will provide a list of candidate loci underlying behavioral resistance in a major malaria vector. This will be the first study into the biological basis of behavioral resistance and takes the first step towards developing the tools to monitor or counteract this major threat to our vector control tools.
Publications from this work:
Increasing Outdoor Host-seeking in Anopheles gambiae over 6 years of vector control on Bioko Island (2016) Meyers JI, S Pathikonda, ZR Popkin-Hall, MC Medeiros, G Fuseini, A Matias, G Garcia, HJ Overgaard, V Kulkarni, VP Reddy, C Schwabe, J Lines, I Kleinschmidt, MA Slotman. Malaria Journal 15:239.
Outdoor host-seeking behaviour of Anopheles gambiae mosquitoes following initiation of malaria vector control on Bioko Island, Equatorial Guinea (2011) Reddy MR, HJ Overgaard, S Abaga, VP Reddy, A Caccone, AE Kiszewski, MA Slotman. Malaria Journal 10:184
This research is made possible by the vector monitoring efforts on Bioko Island supported by Marathon Oil Corporation and its partners.