The subgenus Megatrypanum Hoare, 1964, with the type species Trypanosoma theileri Laveran, 1902, comprises stercorarian trypanosomes of mammals. A substantial portion of this subgenus consists of T. theileri-like trypanosomes parasitizing cervids and bovids worldwide. Similar to most other members of the genus Trypanosoma that lack obvious economic importance, the biology of T. theileri-like trypanosomes remains poorly understood. In particular, fundamental aspects such as their host specificity, host–parasite interactions, and the morphology of developmental stages have been studied only to a limited extent. In this work, we provide a detailed description of the development and cellular organization of T. theileri-like trypanosomes in the horsefly Hybomitra montana using transmission electron microscopy (TEM). We show for the first time that T. theileri-like trypanosomes possess a well-developed cytostome–cytopharyngeal complex, morphologically similar to those in other stercorarian trypanosomes. This complex is present in the studied trypanosomes at the epimastigote stage and degrades during metacyclogenesis. In the host ileum, epimastigotes and trypomastigotes at different stages of metacyclogenesis are embedded in a fibrillar matrix that isolates them from the gut lumen. This promotes their accumulation in the vector, thereby increasing the efficiency of future infection of the vertebrate host, which occurs via contamination of the oral mucosa.