Malaria transmission entails development of the Plasmodium parasite in its insect vector, the Anopheles mosquito. Parasite invasion of the mosquito midgut is the critical first step and involves adhesion to host epithelial cell ligands. Partial evidence suggests that midgut oligosaccharides are important ligands for parasite adhesion; however, the identity of these glycans remains unknown. We have identified a population of chondroitin glycosaminoglycans along the apical midgut microvilli of Anopheles gambiae and further demonstrated ookinete recognition of these glycans in vitro. By repressing the expression of the peptide-Oxylosyltransferase homolog of An. gambiae by means of RNA interference, we blocked glycosaminoglycan chain biosynthesis, diminished chondroitin sulfate levels in the adult midgut, and substantially inhibited parasite development. We provide evidence for the in vivo role of chondroitin sulfate proteoglycans in Plasmodium falciparum invasion of the midgut and insight into the molecular mechanisms mediating parasite-mosquito interactions.Anopheles ͉ glycosaminoglycans ͉ glycosytransferase ͉ malaria ͉ RNAi M alaria is caused by Plasmodium parasites, among which, Plasmodium falciparum inflicts the severest infection on human populations. The complex parasite life cycle includes developmental stages within both mammals and its obligatory insect vector, the Anopheles mosquito (1). Plasmodium gametocytes that are taken up in an infected blood meal transform into ookinetes in the mosquito midgut lumen. Ookinetes then migrate to the gut periphery where they are thought to recognize and adhere to midgut ligands. These steps directly precede cell invasion, traversal, and the differentiation of ookinetes into oocysts beneath the basal lamina. Each oocyst produces thousands of sporozoites that subsequently invade the mosquito salivary glands and are delivered to a vertebrate host during a succeeding blood meal. Clearly, ookinete invasion of midgut epithelia is the critical step for parasite establishment in the mosquito and, therefore, represents the best paradigm to develop novel transmission-blocking strategies (i.e., approaches that prevent parasite passage through the mosquito and therefore impede the subsequent cascade of secondary infections in humans).Plasmodium ookinete molecules belonging to the Thrombospondin-related adhesive protein (TRAP) family, which includes the sporozoite surface protein, TRAP, and two ookinete proteins, the circumsporozoite and TRAP-related protein (CTRP) and the von Willebrand Factor A domain protein (WARP), have been demonstrated to bind to the glycosaminoglycan (GAG), heparin, in vitro (2). The major circumsporozoite protein (CSP) and TRAP both recognize heparan sulfate (HS) proteoglycans on the liver sinusoid, a critical step toward the establishment of parasite infection in humans (3, 4). Similarly, ookinete CTRP and WARP gene knockouts were shown to be incapable of invading the mosquito midgut, strongly implying their essential role in mosquito midgut cell invasion ...
