BackgroundTsetse-transmitted African animal trypanosomiasis is recognised as an important disease of ruminant livestock in sub-Saharan Africa, but also affects domestic pigs, withTrypanosoma simiaenotable as a virulent suid pathogen that can rapidly cause death.Trypanosoma simiaeis widespread in tsetse-infested regions, but its biology has been little studied compared toT. bruceiandT. congolense.MethodsTrypanosoma simiaeprocyclics were cultured in vitro and transfected using protocols developed forT. brucei. Genetically modified lines, as well as wild-type trypanosomes, were transmitted through tsetse flies,Glossina pallidipes, to studyT. simiaedevelopment in the tsetse midgut, proventriculus and proboscis. The development of proventricular trypanosomes was also studied in vitro. Image and mensural data were collected and analysed.ResultsAPFR1::YFPline successfully completed development in tsetse, but aYFP::HOP1line failed to progress beyond midgut infection. Analysis of image and mensural data confirmed that the vector developmental cycles ofT. simiaeandT. congolenseare closely similar, but we also found putative sexual stages inT. simiae, as judged by morphological similarity to these stages inT. brucei. Putative meiotic dividers were abundant amongT. simiaetrypanosomes in the proboscis, characterised by a large posterior nucleus and two anterior kinetoplasts. Putative gametes and other meiotic intermediates were also identified by characteristic morphology. In vitro development of proventricular forms ofT. simiaefollowed the pattern previously observed forT. congolense: long proventricular trypanosomes rapidly attached to the substrate and shortened markedly before commencing cell division.ConclusionsTo date,T. bruceiis the only tsetse-transmitted trypanosome with experimentally proven capability to undergo sexual reproduction, which occurs in the fly salivary glands. By analogy, sexual stages ofT. simiaeorT. congolenseare predicted to occur in the proboscis, where the corresponding portion of the developmental cycle takes place. While no such stages have been observed inT. congolense, forT. simiaeputative sexual stages were abundant in the tsetse proboscis. Although our initial attempt to demonstrate expression of a YFP-tagged, meiosis-specific protein was unsuccessful, the future application of transgenic approaches will facilitate the identification of meiotic stages and hybrids inT. simiae.Graphical AbstractSupplementary InformationThe online version contains supplementary material available at 10.1186/s13071-023-05847-5.