Syngamid worms (Nematoda: Syngamidae) parasitizing birds of prey are considered cosmopolitan, but the efforts to understand their biology and systematics are restricted to the Holarctic region. However, in the Neotropical region there is only one recent record with no data about its molecular characterization or its significance to the health of its hosts. Thus, this study aimed to identify through an integrative approach the Syngamid worms parasitizing a native owl, and to describe its pathological consequences. A total of 14 barn owls (Tyto alba) were dissected between 2015 and 2021, from which one bird was found to be parasitized by a high parasitic burden (i.e. 185 worms). Considering light microscopy and SEM, these nematodes were morphologically identified as Cyathostoma americana, and then supported through molecular analyses of nuclear loci ITS and LSU. The pathological changes were described as severe airsacculitis and pneumonia, which probably were the cause of death of the bird. This study represents the first effort to characterize the parasitism by this nematode in a nocturnal bird of prey from the Neotropics, with a lethal outcome. Additional studies are required from other geographical areas and Neotropical countries to better understand the dynamics of this parasite and their hosts, considering both diurnal and nocturnal birds of prey.