Adaptive divergence between incipient species of Anopheles gambiae increases resistance to Plasmodium

Abstract

The African malaria mosquito Anopheles gambiae is diversifying into ecotypes known as M and S forms. This process is thought to be promoted by adaptation to different larval habitats, but its genetic underpinnings remain elusive. To identify candidate targets of divergent natural selection in M and S, we performed genomewide scanning in paired population samples from Mali, followed by resequencing and genotyping from five locations in West, Central, and East Africa. Genome scans revealed a significant peak of M-S divergence on chromosome 3L, overlapping five known or suspected immune response genes. Resequencing implicated a selective target at or near the TEP1 gene, whose complement C3-like product has antiparasitic and antibacterial activity. Sequencing and allele-specific genotyping showed that an allelic variant of TEP1 has been swept to fixation in M samples from Mali and Burkina Faso and is spreading into neighboring Ghana, but is absent from M sampled in Cameroon, and from all sampled S populations. Sequence comparison demonstrates that this allele is related to, but distinct from, TEP1 alleles of known resistance phenotype. Experimental parasite infections of advanced mosquito intercrosses demonstrated a strong association between this TEP1 variant and resistance to both rodent malaria and the native human malaria parasite Plasmodium falciparum. Although malaria parasites may not be direct agents of pathogen-mediated selection at TEP1 in nature--where larvae may be the more vulnerable life stage--the process of adaptive divergence between M and S has potential consequences for malaria transmission.

Fig. 1.

Genomic microarray analysis of M-S divergence on chromosome 3L in Mali. A peak of significant differentiation spanning ∼100 kb was detected at ∼11 Mb from the centromere (cen), below which five genes are predicted, four belonging to the TEP immune gene family. Orientation of the coding strand (5′–3′) is indicated by boxes, above or below the horizontal line, which enclose the number of SFPs detected per gene.

Fig. 2.

Geographically restricted natural selection in the M form. (A) Country-specific plots of within-form nucleotide diversity (π, Right y-axis) and between-form divergence (F_ST, Left y-axis), at surveyed candidate and reference genes. (B) Within-form divergence between population pairs, at candidate and reference genes. Brackets and asterisks denote statistically significant contrasts between candidate and reference genes in the M form.

Fig. 3.

Three allelic classes at the TEP1 locus. Amino acid alignment based on the TED region of TEP1 shows only variable positions, numbered following ref. for a subset of available sequences. Shading highlights amino acid differences within and between allelic classes, which are separated by a horizontal black line and labeled at Right (s, TEP1s; r^A, TEP1r^A; and r^B, TEP1r^B). Allele designations at Left refer to sequences from laboratory strains (L3-5, Mali-NIH, 4Arr, PEST, and G3) or from the field. Alleles from the field are labeled by geographic origin (GH, Ghana; BF, Burkina Faso; ML, Mali; CM, Cameroon; and KN, Kenya) followed by molecular form (M, M form and S, S form) or by author (Obb, Obbard et al.; ref. 27).

Fig. 4.

Geographic structure of allelic variation at TEP1. Frequency of TEP1s (s, blue), TEP1r^A (r^A, red), and TEP1r^B (r^B, green) alleles of 1,370 chromosomes sampled in M and S from the indicated countries in Africa. Numbers within pie graphs represent chromosomes sampled per form in each country.

Fig. 5.

The TEP1r^B allele increases mosquito resistance to Plasmodium. (A) An. gambiae–P. berghei: Number of live oocysts per midgut and mean (central bar) ± SE, P < 0.0001. (B) An. gambiae–P. berghei: Number of melanized ookinetes per midgut and mean (central bar) ± SE, P < 0.0001. (C) An. gambiae–P. falciparum: Number of live oocysts per midgut and mean (central bar) ± SE, P < 0.01.