Malaria continues to expand as a major public health threat throughout the world. The spread of drug resistant malaria and the lack of an effective vaccine make alternative and complementary approaches to malaria control increasingly important. The overall theme of this research program has been to develop mechanistic concepts of how to block malaria transmission by interfering with P/as/Doc//um-mosquito molecular interactions within the mosquito midgut, particular at the ookinete-peritrophix matrix interface. The last project period showed that interfering with the function of Plasmodium chitinase(s) by antibody or chitinase gene deletion interfered with parasite infectivity for the mosquito. As part of ongoing efforts to continue delineating molecular mechanisms by which the Plasmodium ookinete invades the mosquito midguCwe have identified a second P. gallinaceum ookinete-secreted chitinase, the only known ortholog of the P. falciparum chitinase, PfCHTI, and an ookinete-secreted plasmepsin, an aspartic protease that we hypothesize is involved working together with chitinase(s) to allow the ookinete to penetrate the peritrophic matrix within the mosquito midgut. The hypothesis underlying this proposal is that Plasmodium ookinetes secrete chitinases and plasmepsins that are both involved in invading the mosquito midgut, either additively or synergistically. This competitive renewal proposal h^is three specific aims: 1) To determine mechanisms by which the P. falciparum PfCHTI chitinase orthologi^e in P. gallinaceum, PgCHT2, facilitates ookinete penetration of the peritrophic matrix;2) To identify thejP. gallinaceum ookinete-produced protease that mediates activation of the P. gallinaceum chitinase PgCHTI;and 3) To determine the role of Plasmodium aspartic proteases (plasmepsins) in ookinete invasion1of the mosquito midgut. This project will yield insight into basic biological processes of P/asmod/um-mosquito midgut interactions, including the function of the P. falciparum chitinase, PfCHTI, and provide the scientific basis for developing novel potential approaches to interrupting malaria transmission at the vertebrate host-mosquito vector interface.