Dissecting fatty acid metabolism in livestock trypanosomes

Animal African Trypanosomiasis (AAT), also known as Nagana, is a devastating disease affecting cattle and other livestock across sub-Saharan Africa, with more than 60 million cattle at risk and in excess of 3 million deaths per year, with a great socio-economic impact on agricultural communities. AAT is principally caused by the single-celled parasite Trypanosoma congolense. There are no vaccines to treat the disease, and treatment failure is common due to resistance against the drugs used to treat AAT. New drugs are sorely needed, but a major problem hindering their development is a lack of biological understanding of T. congolense. Instead, most of our current knowledge on trypanosomes derives from the related species T. brucei, variants of which cause the human disease Sleeping Sickness. One potential avenue to drug discovery against AAT is by gaining an understanding of parasite cell metabolism, and how drugs may affect this in ways that kill the pathogen without impacting the host.

African trypanosomes are unique in their metabolism, and whilst it was previously thought that T. congolense and T. brucei were very similar in their biology, recent data I have generated has highlighted several metabolic distinctions between these species. For example, whilst T. brucei is known to rely almost entirely on host-derived glucose for its energy demands, T. congolense appears to use less glucose, and metabolise it at a reduced rate. In addition, T. brucei is able to use some of the products of glucose metabolism to generate a class of metabolites called fatty acids, an important group of molecules for cellular structure. In contrast, we found no evidence of glucose-derived fatty acids in T. congolense.

I recently found that whilst T. brucei is able to synthesise fatty acids, T. congolense prefers to scavenge these from the bloodstream of the host. In particular, I have identified four fatty acids that T. congolense requires to grow in the laboratory. This requirement has not previously been reported for African trypanosomes, and the ways in which these four fatty acids are used by T. congolense are therefore novel to trypanosome biology. Indeed, fatty acid metabolism is unexplored in this parasite species. Crucially, these mechanisms of fatty acid usage may also differ from host metabolism, and this could be exploited for drug development.

In this study, I propose to investigate how these fatty acids are used, and metabolised by T. congolense. To do this, I will use cutting edge technologies such as mass spectrometry to map in detail how the four fatty acids are metabolised by T. congolense, and whether they are a potential source of energy for this parasite species. I will combine this approach with bioinformatic techniques and high-resolution mass spectrometry imaging to investigate the enzymes T. congolense possesses and where these are localised in the cell, in order to accurately determine the major metabolic pathways involved in fatty acid metabolism. I will then investigate how the activity in these pathways changes in response to the environment, depending on the presence and absence of exogenous fatty acids, in order to understand whether these pathways are essential to the parasite, and indeed a viable avenue for drug discovery. Finally, I will select key enzymes and use newly-available genetic tools to manipulate their presence in the cell in order to determine the overall effect on cell growth. Combined with the testing of known inhibitors, these objectives will give a very detailed picture of fatty acid metabolism in T. congolense, giving us an insight into the biology of this understudied pathogen and potentially identifying candidate drug targets.

Trypanosoma congolense is a major cause of Animal African Trypanosomiasis, a debilitating tsetse-transmitted disease in livestock, primarily cattle, across sub-Saharan Africa, with >60 million cattle at risk, and ~3 million deaths per year. New drugs are sorely needed but efforts are hindered by a paucity of knowledge regarding the biology of T. congolense. One effective strategy for drug development is the identification and targeting of metabolic pathways unique to a pathogen. My recent data has shown that T. congolense is highly resistant to inhibitors of fatty acid biosynthesis, a process carried out by trypanosomes via glucose- and threonine-derived acetate to meet their lipid demands. Combined with my data highlighting that four fatty acids - oleic acid, linoleic acid, propionic acid and 3-hydroxybutyric acid - are critical to T. congolense in vitro viability, I hypothesise that unlike T. brucei, T. congolense relies on scavenging of exogenous fatty acids, in lieu of synthesis. However, the processes involved in metabolism of these fatty acids are unknown. In this study I propose to address the hypothesis that fatty acid scavenging and metabolism in T. congolense are distinct from other eukaryotes, and critical to the parasite. I will ascertain the fate of the four essential fatty acids by carrying out stable-isotope-resolved metabolomics and lipidomics in addition to an in silico analysis of fatty acid metabolism and the application of mass spectrometry imaging. I will combine these data with a proteomics analysis of cellular response to different levels of exogenous fatty acids, in order to reveal essential proteins involved in fatty acid metabolism. Finally, I will apply gene manipulation in the form of RNA interference, as well as testing T. congolense susceptibility to metabolic inhibitors, in order to test the essentiality of fatty acid metabolism, with the ultimate aim of identifying novel drug targets for livestock trypanosomiasis.