Proteome characterization in various biological fluids of Trypanosoma brucei gambiense-infected subjects

Human African trypanosomiasis (HAT) is a neglected tropical disease that is endemic in sub-Saharan Africa. Control of the disease has been recently improved by better screening and treatment strategies, and the disease is on the WHO list of possible elimination. However, some physiopathological aspects of the disease transmission and progression remain unclear. We propose a new proteomic approach to identify new targets and thus possible new biomarkers of the disease. We also focused our attention on fluids classically associated with HAT (serum and cerebrospinal fluid (CSF)) and on the more easily accessible biological fluids urine and saliva. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) established the proteomic profile of patients with early and late stage disease. The serum, CSF, urine and saliva of 3 uninfected controls, 3 early stage patients and 4 late stage patients were analyzed. Among proteins identified, in CSF, urine and saliva, respectively, 37, 8 and 24 proteins were differentially expressed and showed particular interest with regards to their function. The most promising proteins (Neogenin, Neuroserpin, secretogranin 2 in CSF; moesin in urine and intelectin 2 in saliva) were quantified by enzyme-linked immunosorbent assay in a confirmatory cohort of 14 uninfected controls, 23 patients with early stage disease and 43 patients with late stage disease. The potential of two proteins, neuroserpin and moesin, with the latter present in urine, were further characterized. Our results showed the potential of proteomic analysis to discover new biomarkers and provide the basis of the establishment of a new proteomic catalogue applied to HAT-infected subjects and controls. SIGNIFICANCE: Sleeping sickness, also called Human African Trypanosomiasis (HAT), is a parasitic infection caused by a parasitic protozoan, Trypanosoma brucei gambiense or T. b. rhodesiense which are transmitted via an infected tsetse fly: Glossina. For both, the haemolymphatic stage (or first stage) signs and symptoms are intermittent fever, lymphadenopathy, hepatosplenomegaly, headaches, pruritus, and for T. b. rhodesiense infection a chancre is often formed at the bite site. Meningoencephalitic stage (or second stage) occurs when parasites invade the CNS, it is characterised by neurological signs and symptoms such as altered gait, tremors, neuropathy, somnolence which can lead to coma and death if untreated. first stage of the disease is characterizing by fevers, headaches, itchiness, and joint pains and progressive lethargy corresponding to the second stage with confusion, poor coordination, numbness and trouble sleeping. Actually, diagnosing HAT requires specialized expertise and significant resources such as well-equipped health centers and qualified staff. Such resources are lacking in many endemic areas that are often in rural locales, so many individuals with HAT die before the diagnosis is established. In this study, we analysed by mass spectrometry the entire proteome of serum, CSF, urine and saliva samples from infected and non-infected Angolan individuals to define new biomarkers of the disease. This work of proteomics analysis is a preliminary stage to the characterization of the whole proteome, of these 4 biological fluids, of HAT patients. We have identified 69 new biomarkers. Five of them have been thoroughly investigated by ELISA quantification. Neuroserpine and Moesin are respectively promising new biomarkers in CSF and urine's patient for a better diagnosis.