Transmission-blocking vaccines (TBVs) targeting ookinete surface proteins expressed on sexual-stage malaria parasites are considered one promising strategy for malaria control. To evaluate the prospect of developing non-invasive and easy-to-administer mucosal malaria transmission-blocking vaccines, mice were immunized intranasally with a Plasmodium vivax ookinete surface protein, Pvs25 with a mucosal adjuvant cholera toxin (CT). Immunization induced significant serum IgG with high IgG1/IgG2a ratio (indicative of Th-2 type immune response). Feeding Anopheles dirus mosquitoes with mixtures of immune sera and gametocytemic blood derived from vivax-infected volunteer patients in Thailand significantly reduced both the number of midgut oocysts as well as the percentage of infected mosquitoes. The observed transmission-blocking effect was dependent on immune sera dilution. This study demonstrates for the first time that the mucosally induced mouse immune sera against a human malaria ookinete surface protein can completely block parasite transmission to vector mosquitoes, suggesting the possibility of non-invasive mucosal vaccines against mucosa-unrelated important pathogens like malaria.