Abstract
Inhibiting Plasmodium development in mosquitoes will block malaria transmission. Fibrinogen-related protein 1 (FREP1) is critical for parasite infection in Anopheles gambiae and facilitates Plasmodium invasion in mosquitoes through interacting with gametocytes and ookinetes. To test the hypothesis that small molecules that disrupt this interaction will prevent parasites from infecting mosquitoes, we developed an ELISA-based method to screen a fungal extract library. We obtained a candidate fungal extract of Aspergillus niger that inhibited the interaction between FREP1 and P. falciparum infected cells by about 92%. The inhibition specificity was confirmed by immunofluorescence assays. Notably, feeding mosquitoes with the candidate fungal extract significantly inhibited P. falciparum infection in the midgut without cytotoxicity or inhibition of the development of P. falciparum gametocytes or ookinetes. A bioactive natural product that prevents FREP1 from binding to gametocytes or ookinetes was isolated and identified as P-orlandin. Importantly, the nontoxic orlandin significantly reduced P. falciparum infection intensity in mosquitoes. Therefore, disruption of the interaction between FREP1 and parasites effectively reduces Plasmodium infection in mosquitoes. Targeting FREP1 with small molecules is thus an effective novel approach to block malaria transmission.
Highlights
Because fibrinogen-related protein 1 (FREP1) expressed in E. coli forms inclusion bodies and lacks post-translational modification, we used an insect cell expression system to generate functional FREP1 that is similar to endogenous FREP1
We found that FREP1 was detected in the culture medium of the insect High Five cells transfected with pIB-FREP1 DNA but not in the cell lysate (Fig. 1a), confirming that FREP1 is a secreted protein
We previously reported that the mosquito protein FREP1 mediates Plasmodium infection in An. gambiae[16] through its interaction with parasites in mosquito midguts[24]
Summary
Targeting FREP1 with small molecules is an effective novel approach to block malaria transmission. Research efforts have focused on drugs that kill parasites in the blood stage[11,12,13], while no compounds have been developed that target mosquito proteins and block malaria transmission without killing the mosquitoes. The sporozoites in salivary glands are injected into the host to start another cycle of infection This Plasmodium invasion of mosquitoes depends on the interactions between parasites and mosquito molecules. Antibodies against anopheline alanyl aminopeptidase N (AnAPN1)[21] and carboxypeptidases B22 have been reported to block Plasmodium infection[23] Targeting these mosquito proteins with small molecules to block malaria transmission has not yet been reported. If such candidate compounds were to be identified, they could be administered in combination with anti-malaria drugs to block malaria transmission or alternatively the compounds could be sprayed outdoors or on bednets to block malaria transmission
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