Avian trichomonosis is a parasitic disease that affects wild birds, The objective of this work was to determine the importance of avian trichomonosis in Bonelli's eagles to improve conservation measures in this population. One hundred and eighty-eight birds were studied: 181 chicks,, two juveniles, one subadult and four adults. The birds were externally examined and gross lesions at the oropharynx registered. Samples from the oropharyngeal cavity were obtained for Trichomonas spp. detection by culture and PCR, and positive samples were subjected to a multi-locus sequence typing approach, including the ITS1/5.8S/ITS2 region (ITS), ribosomal RNA small subunit (18S) and Fe-hydrogenase gene (FeHyd). Global prevalence for T. gallinae infection was 37.8% in total, 45.5% in nestlings. Thirty three percent of the birds developed lesions that ranged from mild (n=41) to moderate (n=14) or severe (n=7). MLST analysis showed five different MLS types, being ITS-A/18S-VI/FeHyd-A1 and ITS-D/18S-II/Fe-C4 the most frequent. An association between ITS-A/18S-VI/FeHyd-A1 and moderate or severe lesions was observed, but birds with type ITS-A/18S-VI/FeHyd-A2 also developed lesions. On the contrary, birds with MLS type ITS-D/18S-II/FeHyd-C4 displayed only a low proportion of mild lesions. Chicks raised in nests were at higher risk for T. gallinae infection and development of lesions than chicks raised in captivity. Disconrdances between samples cultured in TYM and samples subjected to PCR from oropharyngeal swabs were observed, being swab-ITS-PCR more sensitive.