The Population Genetic Structure of the Malaria Mosquito Anopheles melas Throughout Its West-African Range

Anopheles melas is a brackish water mosquito found along the coast of West-Africa where it can be the dominant malaria vector locally. In order to facilitate genetic studies of this species and to examine the usefulness of microsatellite markers when used in a sibling species, 45 microsatellite loci originally developed for Anopheles gambiae were sequenced in An. melas. These loci were evaluated on their suitability as polymorphic markers based on repeat structure, length, and polymorphism in wild An. melas populations. Of the 45 loci, 18 were not considered promising markers in An. melas. A total of 48 out of 90 An. gambiae primers contained at least one mismatch with the An. melas annealing site. An. melas-specific primers were designed for 27 loci, and their variability was examined in two wild populations from Equatorial Guinea. Based on a low level of polymorphism, Hardy-Weinberg disequilibrium, or poor amplification, a further 12 loci were excluded. The remaining fifteen loci were screened in four additional wild populations from a wider geographic region including Equatorial Guinea, Cameroon, The Gambia, and Guinea Bissau. These loci showed an average heterozygosity ranging from 0.18 to 0.79, with 2.5 to 15 average alleles per locus, yielding 13 highly polymorphic markers and two loci with more limited variability in a wide geographic region. To examine the effects of cross species amplification, five of the original An. gambiae markers were also amplified in the An. melas populations. Null alleles were found for one of these An. gambiae markers. We discuss the pitfalls of using microsatellite loci even in a very closely related species, and conclude that in addition to the well-known problem of null alleles associated with this practice, many loci may prove to be of very limited use as polymorphic markers even when used in a sibling species. Fifteen An. melas-specific markers were subsequently amplified and analyzed in 11 wild An. melas populations from throughout the range of this species, including Bioko Island, Equatorial Guinea. We analyzed pair-wise population differentiation between all populations, and found that all but two comparisons were significant (p-val.