Background and Aim: Avian malaria is a tropical disease caused by protozoans of the genera Plasmodium and Haemoproteus. As a nonlethal disease, avian malaria can affect the lifespan and reproductive rate of birds. If there is a differential effect depending on bird species, then this disease might have a significant effect on avian biodiversity. The current study aimed to determine the incidence of Plasmodium in hummingbirds in humid premontane forest areas. Materials and Methods: Blood samples (n=60) were collected from hummingbirds from two areas (Santuario de Aves Milpe and Hacienda Puyucunapi) of Pichincha Province, Ecuador. Prevalence and parasitemia were determined by microscopic examination of blood smears stained with Giemsa reagent. Both study sites are part of a 1000 m elevational gradient; hence, elevation was used as a predictor variable for prevalence and parasitemia levels in a Mann–Whitney U-test. This test was also used to test for a sex bias. Results: This study reports on a total of 12 bird species that inhabit both study sites. At Milpe, the lower elevation site, a prevalence of 100% was recorded, whereas at Puyucunapi, the prevalence was 96%. The combined prevalence was 97%. Elevation and sex did not influence prevalence nor parasitemia in hummingbirds. Conclusion: This study does not suggest a significant elevation or sex bias on prevalence and parasitemia in hummingbirds.