Dissecting the Genetic Basis of Resistance to Malaria Parasites in Anopheles gambiae

Abstract

The ability of Anopheles gambiae mosquitoes to transmit Plasmodium parasites is highly variable between individuals. However, the genetic basis of this variability has remained unknown. We combined genome-wide mapping and reciprocal allele-specific RNA interference (rasRNAi) to identify the genomic locus that confers resistance to malaria parasites, and demonstrate that polymorphisms in a single gene encoding the antiparasitic thioester-containing protein 1 (TEP1) explain a substantial part of the variability in parasite killing. The link between TEP1 alleles and resistance to malaria may offer new tools for controlling malaria transmission. The successful application of rasRNAi in Anopheles suggests that it could also be applied to other organisms where RNA interference is feasible to dissect complex phenotypes to the level of individual quantitative trait alleles.Anopheles gambiae mosquitoes are major vectors of Plasmodium falciparum, a protozoan parasite causing the most severe form of human malaria in Africa. The fact that mosquito strains completely wide variety of parasite species (1). Previous genetic analyses of crosses between this strain and the susceptible 4Arr strain infected with two simian parasite species focused on resistant to malaria parasites can be selected (1,2) indicates that genetic factors in mosquitoes control the level of parasite transmission. Understanding the genetic basis of this resistance has been a long-standing question. The L3-5 resistant strain kills and melanises a the melanotic encapsulation phenotype and identified several quantitative trait loci (QTLs), whose relative contributions varied with parasite species and between F2 generation families (3,4). Recently it became clear that melanisation occurs after parasite killing, as a means to dispose of dead parasites in some strains, while in others, killed parasites are only cleared by lysis (Fig. S2A) (5-7). In this study, we aimed at mapping the genomic regions and identifying genes that control resistance (i.e., the absence of live parasites) of mosquitoes to the rodent malaria parasite Plasmodium berghei.We set up reciprocal crosses of the resistant L3-5 and susceptible 4Arr strains. F1 mosquitoes were intercrossed and individual females were isolated to lay eggs, yielding 10 F2 families. ‡ To whom correspondence should be addressed. (9)), indicating that the killing of P. berghei and the mode of clearance of dead parasites are complex traits that are each likely to result from the segregation of several alleles.To map loci controlling resistance to parasites, we genotyped 39 informative markers spanning the entire genome in 206 selected F2 individuals with extreme phenotypes (Fig. S1 and supporting online text). Linkage analysis comparing resistant and non-resistant mosquitoes identified a single region on chromosome 3L (Fig. 1C). We interpreted this region, covering approximately 19Mb, as a major locus responsible for resistance to P. berghei, and named it Pbres1, for P. berghei resistance locus 1. We further c...

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

“…Recently, allelic variants of TEP1 from An. gambiae laboratory strains refractory to P. berghei were directly implicated in enhanced lysis and melanization of parasites (22), owing to faster binding kinetics relative to alleles in susceptible strains (25). Corresponding TEP1 alleles (TEP1r and TEP1s) also segregate in natural populations of An.…”

“…Like its C3 vertebrate homolog, TEP1 is a complement-like opsonin whose binding triggers killing, via phagocytosis of bacteria or lysis/melanization of the malaria parasites Plasmodium berghei and Plasmodium falciparum (17,(22)(23)(24). Recently, allelic variants of TEP1 from An.…”

Efficient ΦC31 integrase–mediated site-specific germline transformation of Anopheles gambiae

“…gambiae. In addition to gain-of-function studies, such a system would also permit functional analysis of alleles by a combination of stable RNAi and in vivo complementation to alleviate the otherwise laborious process of allele-specific transient RNAi 18 .…”

“…gambiae 16 . This methodology has proved successful for silencing gene expression in mosquito tissues encountered by Plasmodium parasites during its development in the mosquito [16][17][18][19][20] . However, this method only permits loss-of-function studies, is labour intensive, and offers little selectivity over which tissues are targeted for knockdown.…”

The molecular epidemiology of parasite infections: Tools and applications

“…This approach has been used successfully to map QTLs for traits of epidemiological significance in a number of parasite species; for example drug resistance in Plasmodium falciparum [195], virulence in Toxoplasma gondii [196] and resistance to malaria parasites in Anopheles gambiae [197]. Linkage analyses have been so successful because, as closely related individuals have large portions of their genome in common, a relatively small number of polymorphic neutral markers (usually 500 or less) are usually sufficient to detect linked regions [198].…”