In vitro cultivation of Trypanosoma congolense bloodstream forms: State of the art and advances

Animal African Trypanosomosis (AAT or Nagana) is a severe vector-borne disease caused by protozoan parasites belonging to the Trypanosomatidae family and is usually cyclically transmitted by blood-sucking tsetse flies. AAT remains a major problem in sub-Saharan Africa. Among the main AAT causative agents, Trypanosoma congolense (T. congolense or Tc) is one of the most important trypanosome species, in terms of economic and animal health impacts, infecting cattle and a wide range of animal hosts as well. To advance in AAT prevention and control, it is essential to better understand trypanosome biology and pathogenesis using bloodstream form (BSF) in vitro culture. The in vitro cultivation of T. congolense IL3000 BSF strain is already well established and widely used in research studies and drug activity assays. However, it may probably no longer truly reflect the reality of field trypanosome strains, due to decades of use and subsequent modifications. Here, we propose a novel culture protocol that supports the long-term in vitro growth of the animal-infective BSFs of three Savannah and Forest types of T. congolense strains, including T. congolense clone IL1180, which is not only a field strain but also a commonly-used reference strain in experimental animal assays. We established a homemade culture medium which made it possible to sustain T. congolense IL1180 growth from infected mouse blood for 18 days in axenic conditions. Moreover, we developed an efficient freezing/thawing system that allowed, for the first time, T. congolense IL1180 BSF growth within 30 days after thawing. Our results on T. congolense adaptation to in vitro culture are encouraging for future gene studies using new molecular tools or for new therapeutic drug assays.