Previous studies of Plasmodium vivax transmission to Anopheles spp. mosquitoes have not been able to predict mosquito infectivity on the basis of microscopic or molecular quantification of parasites (total parasites in the sample or total number of gametocytes) in infected blood. Two methods for production of P. vivax ookinete cultures in vitro, with yields of 106 macrogametocytes, 104 zygotes, and 103 ookinetes, respectively, per 10 mL of P. vivax-infected patient blood with approximately 0.01% parasitemia, were used to study P. vivax sexual stage development. The quantity of gametocytes, determined by counting Giemsa-stained blood smears, and quantity and type of gametocyte as determined by quantitative reverse transcriptase-polymerase chain reaction for Pvalpha tubulin II and macrogametocyte-specific pvg377 did not predict ookinete yield. Factors that affect the efficiency of in vitro P. vivax ookinete transformation remain poorly understood.