Trypanosomes have long been recognised as being amongst the most important protozoan parasites of verte-brates, from both medical and veterinary perspectives. Whilst numerous insect species have been identified as vectors, the role of ticks is less well understood. Here we report the isolation and partial molecular character-isation of a novel trypanosome from questing Ixodes ricinus ticks collected in Slovakia. The trypanosome was isolated in tick cell culture and then partially characterised by microscopy and amplification of fragments of the 18S rRNA and 24S alpha rDNA genes. Analysis of the resultant sequences suggests that the trypanosome designated as Trypanosoma sp. Bratislava1 may be a new species closely related to several species or strains of trypanosomes isolated from, or detected in, ticks in South America and Asia, and to Trypanosoma caninum isolated from dogs in Brazil. This study highlights the potential involvement of ixodid ticks in the epidemiology of trypanosomes, as well as the use of tick cell lines for isolation of such tick-borne protozoa. Further studies are required to in-vestigate the epidemiology, transmission and life cycle of this putative novel species.