ABSTRACT
Melanins are structurally complex pigments produced by organisms in all domains of life from bacteria to animals, including humans. In insects, melanins are essential for survival and have key roles in cuticle sclerotization, wound healing and innate immunity. In this study, we used a diverse set of molecular, biochemical, and imaging approaches to characterize mosquito melanin involved in innate immune defense (melanotic capsules). We observed that melanotic capsules enclosing Plasmodium berghei ookinetes were composed of an acid-resistant and highly hydrophobic material with granular appearance, which are characteristic properties of melanins. Spectroscopical analyses reveal chemical signatures of eumelanins and pheomelanin. Furthermore, using an enrichment approach for the proteomic analysis, we identified a set of 14 of acid-resistant mosquito proteins embedded within the melanin matrix possibly related to an anti-Plasmodium response. Among these, AgMesh, a highly conserved protein among insect species that contains domains suggesting a role in immune recognition and function. We targeted AgMesh for further study using a RNAi-based gene silencing approach in mosquitos and challenged them with two Plasmodium spp. Surprisingly, AgMesh gene silencing in mosquitos was associated with reduced parasite infection, implying an important role in facilitating vector infection by Plasmodium spp. Our results provide a new approach to study aspects of insect melanogenesis that revealed proteins associated with melanotic capsule, one of which was strongly implicated in the pathogenesis of Plasmodium spp. mosquito infection. Given the conservation of AgMesh among disease-transmitting insect vector species, future analysis of this protein could provide fertile ground for the identification of strategies that block transmission of disease to humans.
AUTHOR SUMMARY Vector-borne diseases account for 17% of the global burden of all infection diseases and only malaria kills nearly 500,000 individuals per year, most of them children. Climate change is impacting the geographical distribution and incidence of these diseases, therefore novel ecological-friendly strategies for vector control are needed. Here, we adapted methodologies to study fungal melanogenesis and used them to explore the melanin-based immune response of Anopheles gambiae against malaria parasites. We revealed that pheomelanin together with eumelanin is incorporated in the melanotic capsules against Plasmodium. We also found that melanin-encapsulated Plasmodium is associated to acid-resistant mosquito gut proteins and identify several putative factors of the melanin-mediated immunity. Disruption of a surface-associated protein, found highly conserved among other mosquito vectors, demonstrates its ability to impaired parasite development within the mosquito gut. Our study provides a new approach to investigate the melanin-based defense mechanism in insects, which identified a new host molecule as a potential avenue for developing novel universal pest management schemes.
Competing Interest Statement
The authors have declared no competing interest.