Comparative lipidomic profiling of African trypanosomes reveals metabolic distinctions impacting pan-species drug development

African trypanosomes are a group of pathogens that cause disease in humans (human African trypanosomiasis; HAT) and animals (African animal trypanosomosis; AAT) across sub-Saharan Africa, leading to profound impacts on human health as well as economic instability. Achieving sustainable disease control requires a One Health approach, yet the development of urgently required pan-species chemotherapeutics is severely hindered by a lack of unified biological understanding of the causative species. A major bottleneck in drug discovery is the distinct in vitro requirements of the clinically relevant species. Whilst Trypanosoma brucei is typically cultured in FBS-supplemented culture, T. congolense , the primary cause of AAT, requires the use of goat serum. The differing serum supplementation requirements of these two trypanosome species point to metabolic distinctions, which may be important considerations in developing experimental systems to enable the identification and design of novel, pan-species therapies. In this study, untargeted LC-MS lipidomics analyses were conducted to determine the relative lipidomic profiles of T. brucei and T. congolense bloodstream form parasites. The use of a novel medium formulation that permits effective in vitro culture of both species revealed that their lipidomic profiles are distinct. Notably, ether phospholipids, key molecules in T. brucei lipid biology, are relatively absent in T. congolense . Furthermore, the T. brucei lipidome is enriched with shorter chain saturated lipids, whilst T. congolense exhibits a preference for longer-chain polyunsaturated fatty acids (PUFAs). These observations indicate that there are significant differences in the ways these parasites synthesise and remodel their lipid complement, highlighting an evolutionary divergence between the species that likely carries implications for host-pathogen interactions as well as trypanosome membrane biology. Crucially, defining the species-specific lipid dependencies provides the foundation for pan-species culture systems, thereby removing a critical barrier to the discovery and design of novel therapeutics effective against all African trypanosome species.