4. Temporal monitoring of the 4.3kb SV allele frequency suggests it has evolved under strong positive selection
The high frequencies of this structural variant observed in Central (Cameroon) and Eastern Africa (Uganda) suggested that it is under a strong positive selection. To investigate this possibility, we measured the frequencies of this SV in An. funestus s.s. populations of Cameroon collected in 2014, 2016, 2017, 2018, 2020 and 2021. Samples were available for 3 locations: Tibati and Mibellon, both from the Adamawa region and Gounougou (Northern region).
In Tibati, a temporal comparison of the frequency of the 4.3kb structural variant in samples collected in 2018 revealed a 76% SV+ allele frequency and complete fixation of SV+ in 2021 Figure 2B. In addition, all three genotypes (SV+/SV+, SV+/SV- and SV-/SV-) were detected in 2018 at frequencies of 79%, 9% and 19%, respectively, while in 2021, only the SV+/SV+ genotype was detected (Figure 2C), indicating positive selection on the 4.3kb structural variant in this location.
A similar pattern was observed in Mibellon, with the structural variant present at a low allelic frequency (26%) in 2016, increasing to 98% in less than 2 years (2018) and then to 100% in 2020 (Figure 2D). In 2016, mosquitoes without the 4.3kb SV (SV-/SV) were more common (59%) than those with the structural variant in either the homozygous or heterozygous state (39% SV+/SV+; 11% SV+/SV-) (Figure 2E). By 2018 and 2020, the SV-/SV- homozygote was no longer detected, indicating that this 4.3kb structural variant was also driven to fixation under positive selection in this region.
Genotyping of this SV in samples collected in Gounougou in 2014, 2017, 2020 and 2021 revealed a similar selection pattern to those observed in Tibati and Mibellon, with 3% in 2014, 31% SV+ in 2017, 63% in 2020 and 98% in 2021 (Figure S3A). The SV+/SV+ homozygous genotype was at a very low frequency of 3% in 2014, reached a frequency of 26% in 2017 which doubled to 54% by 2020 in about 3 years and then almost got to fixation in 2021 (Figure 2F).
The rapid increases in allele frequency of the 4.3kb SV in these 3 regions indicate strong selection acting on this structural variant (or a tightly linked allele) in these mosquito populations.
Analysis of recent samples collected in Ghana and Benin (West Africa) in 2022 showed that this insertion is now present in these localities. In Ghana, this SV was identified at a frequency of 5% with only 3 heterozygotes (Figure S3B & S3C), while in the Benin 2022 samples, a higher frequency of 33% was observed (SV+). The genotypes were present at frequencies of 16% (SV+/SV+), 34% (SV+/SV-) and 50% (SV-/SV-) (Figure S3D & S3E).