Anopheles merus distribution and TEP1 genotypes along the Kenyan coast
Authors: Bartilol Brian , Omuoyo Donwilliams , Karisa Jonathan , Ominde Kelly , Mbogo Charles , Mwangangi Joseph , Maia Marta , Rono Martin 
Published: May 2022

Background
Malaria remains one of the most important infectious diseases in Sub-Saharan Africa, responsible for approximately 228 million cases and 602,000 deaths in 2020. Malaria transmission is mainly driven by mosquitoes of the Anopheles gambiae and more recently Anopheles funestus complex. The gains made in malaria control are threatened by insecticide resistance and behavioural plasticity among these vectors. This, therefore, calls for the development of alternative approaches such as malaria transmission-blocking using gene drive systems that can lead to population replacement of infection-susceptible mosquitoes with mosquitoes that are refractory to Plasmodium spp. infection. One such gene that can be utilised is the Thioester-containing protein 1 (TEP1) gene which mediates the killing of Plasmodium falciparum in the mosquito midgut. Here we investigated the frequencies and distribution of TEP1 alleles in wild-caught malaria vectors along the Kenyan coast.
Methods
Mosquitoes were collected using CDC light traps both indoors and outdoors from 20 houses in Garithe village, along the Kenyan coast. The mosquitoes were dissected, and the different parts were used to determine their species, blood-meal source, and sporozoite status. The data was analysed and visualised using the R (v 4.0.1) and STATA (v 17.0).
Results
A total of 18,802 mosquitoes were collected consisting of 77.8% (n = 14,631) Culex spp, 21.4% (n = 4,026) An. gambiae s.l, 0.4% (n = 67) An. funestus and 0.4% (n = 78) other Anopheles (An. coustanii, An. pharoensis and An. pretoriensis). A subset of randomly selected An. gambiae s.l (n = 518) was identified by polymerase chain reaction (PCR), of these 77.2% were An. merus, 22% were An. arabiensis and the rest were not detected. Mosquitoes collected were predominantly exophilic with the outdoor catches being higher across all the species: Culex spp 93% (IRR = 11.6, 95% Cl [5.9–22.9] p < 0.001), An. gambiae s.l 92% (IRR = 7.2, 95% Cl [3.6–14.5]; p < 0.001), An. funestus 91% (IRR = 10.3, 95% Cl [3.3–32.3]; p < 0.001). We identified the following genotypes among An. merus: *R2/R2, *R3/R3, *R3/S2, *S1/S1 and *S2/S2. Among An. arabiensis, we identified *R2/R2, *S1/S1, *S2/S2. Tests on haplotype diversity showed that the most diverse allele was TEP1*S1 followed by TEP1*R2. Tajima’s D values were positive for TEP1*S1 indicating that there is a balancing selection, negative for TEP1*R2 indicating there is a recent selective sweep and as for TEP1*R3 there was no evidence of selection. Phylogenetic analysis showed two distinct clades exist: Refractory and susceptible alleles.
Conclusion
We find that the malaria vectors An. gambiae s.l and An. funestus are predominantly exophilic. TEP1 genotyping for An. merus revealed 5 allelic combinations: *R2/R2, *R3/R3, *R3/S2, *S1/S1 and *S2/S2 while in An. arabiensis, we only identified 3 allelic combinations: *R2/R2, *S1/S1, *S2/S2. The TEP1*R3 allele was restricted to only An. merus among these sympatric mosquito species and we find that there is no evidence of recombination or selection in this allele.