An Evolution-Based Screen for Genetic Differentiation between Anopheles Sister Taxa Enriches for Detection of Functional Immune Factors

Abstract

Nucleotide variation patterns across species are shaped by the processes of natural selection, including exposure to environmental pathogens. We examined patterns of genetic variation in two sister species, Anopheles gambiae and Anopheles coluzzii, both efficient natural vectors of human malaria in West Africa. We used the differentiation signature displayed by a known coordinate selective sweep of immune genes APL1 and TEP1 in A. coluzzii to design a population genetic screen trained on the sweep, classified a panel of 26 potential immune genes for concordance with the signature, and functionally tested their immune phenotypes. The screen results were strongly predictive for genes with protective immune phenotypes: genes meeting the screen criteria were significantly more likely to display a functional phenotype against malaria infection than genes not meeting the criteria (p = 0.0005). Thus, an evolution-based screen can efficiently prioritize candidate genes for labor-intensive downstream functional testing, and safely allow the elimination of genes not meeting the screen criteria. The suite of immune genes with characteristics similar to the APL1-TEP1 selective sweep appears to be more widespread in the A. coluzzii genome than previously recognized. The immune gene differentiation may be a consequence of adaptation of A. coluzzii to new pathogens encountered in its niche expansion during the separation from A. gambiae, although the role, if any of natural selection by Plasmodium is unknown. Application of the screen allowed identification of new functional immune factors, and assignment of new functions to known factors. We describe biochemical binding interactions between immune proteins that underlie functional activity for malaria infection, which highlights the interplay between pathogen specificity and the structure of immune complexes. We also find that most malaria-protective immune factors display phenotypes for either human or rodent malaria, with broad specificity a rarity.

Preferential suppression of Anopheles gambiae host sequences allows detection of the mosquito eukaryotic microbiome

“…gambiae S and M molecular forms, respectively 2 . The two species occupy different larval ecological niches, and are probably adapted to distinct microbial communities 3–5 . Multiple studies have catalogued the prokaryotic microbiota composition of different Anopheles mosquito populations by sequencing of 16s rRNA hypervariable regions 6–13 .…”

The APL1 immune factor is encoded by a single ancestral gene in mostAnophelesspecies and expanded to three paralogs with distinct function in theAnopheles gambiaecomplex

“…These results are in contrast to silencing of the three APL1 paralogs In A. coluzzii , which consistently leads to elevated Plasmodium infection levels (18, 19) but not elevated mortality (Figure 3). Therefore, the three APL1 paralogs confer protection against Plasmodium infection independently of the need to protect against bacteria.…”

“…This observation suggests that the bacteria leading to elevated mortality in APL1-depleted A. stephensi are likely of enteric origin. The three APL1 paralogs found in the Gambiae complex are known to mediate protection from Plasmodium infection (19), but their involvement in protection against other pathogens including bacteria has not been reported.…”

“…Both strains are housed in the insectaries of the CEPIA platform at the Institut Pasteur, Paris. Mosquitoes were reared under standard conditions at 26°C and 80% relative humidity, with a 12 h light/dark cycle and continuous access to 10% sucrose solution in cotton pads (19).…”

“…The paralogs display distinct spectra of protection for different pathogen classes (18–20). APL1A activity inhibits development of the human parasite P. falciparum , while APL1C activity inhibits rodent malaria species (16), and APL1B modulates protection against both P. falciparum and the rodent parasites (19).…”