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Background: Human African Trypanosomiasis (HAT) is a neglected tropical disease caused by infections due to Trypanosoma brucei subspecies. In addition to the well-established environmental and behavioural risks of becoming infected, there is evidence for a genetic component to the response to trypanosome infection. We undertook a candidate gene case-control study to investigate genetic associations further. Methodology: We genotyped one polymorphism in each of seven genes (IL1A, IL1RN, IL4RN, IL6, HP, HPR, and HLA-G) in 73 cases and 250 controls collected from 19 ethno-linguistic subgroups stratified into three major ethno-linguistic groups, 2 pooled ethno-linguistic groups and 11 ethno-linguistic subgroups from three Cameroonian HAT foci. The seven polymorphic loci tested consisted of three SNPs, three variable numbers of tandem repeat (VNTR) and one INDEL. Results: We found that the genotype (TT) and minor allele (T) of IL1A gene as well as the genotype 1A3A of IL1RN were associated with an increased risk of getting Trypanosoma brucei gambiense and develop HAT when all data were analysed together and also when stratified by the three major ethno-linguistic groups, 2 pooled ethno-linguistic subgroups and 11 ethno-linguistic subgroups. Conclusion: This study revealed that one SNP rs1800794 of IL1A and one VNTR rs2234663 of IL1RN were associated with the increased risk to be infected by Trypanosoma brucei gambiense and develop sleeping sickness in southern Cameroon. The minor allele T and the genotype TT of SNP rs1800794 in IL1A as well as the genotype 1A3A of IL1RN rs2234663 VNTR seem to increase the risk of getting Trypanosoma brucei gambiense infections and develop sleeping sickness in southern Cameroon. [ABSTRACT FROM AUTHOR]

All of our current knowledge of African trypanosome metabolism is based on results from trypanosomes grown in culture or in rodents. Drugs against sleeping sickness must however treat trypanosomes in humans. We here compare the transcriptomes of Trypanosoma brucei rhodesiense from the blood and cerebrospinal fluid of human patients with those of trypanosomes from culture and rodents. The data were aligned and analysed using new user-friendly applications designed for Kinetoplastid RNA-Seq data. The transcriptomes of trypanosomes from human blood and cerebrospinal fluid did not predict major metabolic differences that might affect drug susceptibility. Usefully, there were relatively few differences between the transcriptomes of trypanosomes from patients and those of similar trypanosomes grown in rats. Transcriptomes of monomorphic laboratory-adapted parasites grown in in vitro culture closely resembled those of the human parasites, but some differences were seen. In poly(A)-selected mRNA transcriptomes, mRNAs encoding some protein kinases and RNA-binding proteins were under-represented relative to mRNA that had not been poly(A) selected; further investigation revealed that the selection tends to result in loss of longer mRNAs. [ABSTRACT FROM AUTHOR]

Human African Trypanosomiasis (HAT) or sleeping sickness is a Neglected Tropical Disease. Long regarded as an invariably fatal disease, there is increasing evidence that infection by T. b. gambiense can result in a wide range of clinical outcomes, including latent infections, which are long lasting infections with no parasites detectable by microscopy. The determinants of this clinical diversity are not well understood but could be due in part to parasite or host genetic diversity in multiple genes, or their interactions. A candidate gene association study was conducted in Côte d’Ivoire using a case-control design which included a total of 233 subjects (100 active HAT cases, 100 controls and 33 latent infections). All three possible pairwise comparisons between the three phenotypes were tested using 96 SNPs in16 candidate genes (IL1, IL4, IL4R, IL6, IL8, IL10, IL12, IL12R, TNFA, INFG, MIF, APOL1, HPR, CFH, HLA-A and HLA-G). Data from 77 SNPs passed quality control. There were suggestive associations at three loci in IL6 and TNFA in the comparison between active cases and controls, one SNP in each of APOL1, MIF and IL6 in the comparison between latent infections and active cases and seven SNP in IL4, HLA-G and TNFA between latent infections and controls. No associations remained significant after Bonferroni correction, but the Benjamini Hochberg false discovery rate test indicated that there were strong probabilities that at least some of the associations were genuine. The excess of associations with latent infections despite the small number of samples available suggests that these subjects form a distinct genetic cluster different from active HAT cases and controls, although no clustering by phenotype was observed by principle component analysis. This underlines the complexity of the interactions existing between host genetic polymorphisms and parasite diversity. [ABSTRACT FROM AUTHOR]

Introduction: The incidence of gambiense human African trypanosomiasis (gHAT) in Uganda has been declining, from 198 cases in 2008, to only 20 in 2012. Interruption of transmission of the disease by early diagnosis and treatment is core to the control and eventual elimination of gHAT. Until recently, the format of available screening tests had restricted screening and diagnosis to central health facilities (passive screening). We describe a novel strategy that is contributing to elimination of gHAT in Uganda through expansion of passive screening to the entire population at risk. Methodology / Principal findings: In this strategy, patients who are clinically suspected of having gHAT at primary health facilities are screened using a rapid diagnostic test (RDT), followed by parasitological confirmation at strategically located microscopy centres. For patients who are positive with the RDT and negative by microscopy, blood samples undergo further testing using loop-mediated isothermal amplification (LAMP), a molecular test that detects parasite DNA. LAMP positive patients are considered strong suspects, and are re-evaluated by microscopy. Location and upgrading of facilities to perform microscopy and LAMP was informed by results of georeferencing and characterization of all public healthcare facilities in the 7 gHAT endemic districts in Uganda. Three facilities were upgraded to perform RDTs, microscopy and LAMP, 9 to perform RDTs and microscopy, and 200 to screen patients with RDTs. This reduced the distance that a sick person must travel to be screened for gHAT to a median distance of 2.5km compared to 23km previously. In this strategy, 9 gHAT cases were diagnosed in 2014, and 4 in 2015. Conclusions: This enhanced passive screening strategy for gHAT has enabled full coverage of the population at risk, and is being replicated in other gHAT endemic countries. The improvement in case detection is making elimination of the disease in Uganda an imminent possibility. [ABSTRACT FROM AUTHOR]

Background: Human African trypanosomiasis (HAT), a lethal disease induced by Trypanosoma brucei gambiense, has a range of clinical outcomes in its human host in West Africa: an acute form progressing rapidly to second stage, spontaneous self-cure and individuals able to regulate parasitaemia at very low levels, have all been reported from endemic foci. In order to test if this clinical diversity is influenced by host genetic determinants, the association between candidate gene polymorphisms and HAT outcome was investigated in populations from HAT active foci in Guinea. Methodology and results: Samples were collected from 425 individuals; comprising of 232 HAT cases, 79 subjects with long lasting positive and specific serology but negative parasitology and 114 endemic controls. Genotypes of 28 SNPs in eight genes passed quality control and were used for an association analysis. IL6 rs1818879 allele A (p = 0.0001, OR = 0.39, CI95 = [0.24–0.63], BONF = 0.0034) was associated with a lower risk of progressing from latent infection to active disease. MIF rs36086171 allele G seemed to be associated with an increased risk (p = 0.0239, OR = 1.65, CI95 = [1.07–2.53], BONF = 0.6697) but did not remain significant after Bonferroni correction. Similarly MIF rs12483859 C allele seems be associated with latent infections (p = 0.0077, OR = 1.86, CI95 = [1.18–2.95], BONF = 0.2157). We confirmed earlier observations that APOL1 G2 allele (DEL) (p = 0.0011, OR = 2.70, CI95 = [1.49–4.91], BONF = 0.0301) is associated with a higher risk and APOL1 G1 polymorphism (p = 0.0005, OR = 0.45, CI95 = [0.29–0.70], BONF = 0.0129) with a lower risk of developing HAT. No associations were found with other candidate genes. Conclusion: Our data show that host genes are involved in modulating Trypanosoma brucei gambiense infection outcome in infected individuals from Guinea with IL6 rs1818879 being associated with a lower risk of progressing to active HAT. These results enhance our understanding of host-parasite interactions and, ultimately, may lead to the development of new control tools. [ABSTRACT FROM AUTHOR]

Background: Identification of species-specific trypanosome molecules is important for laboratory- and field-based research into epidemiology and disease diagnosis. Although Trypanosoma congolense is the most important trypanosome pathogen of cattle in Africa, no species-specific molecules found in infective bloodstream forms (BSF) of the parasites have been identified, thus limiting development of diagnostic tests. Methods: Immuno-mass spectrometric methods were used to identify a protein that is recognized by a T. congolense-specific monoclonal antibody (mAb) Tc6/42.6.4. The identified molecule was expressed as a recombinant protein in E. coli and was tested in several immunoassays for its ability to interact with the mAb. The three dimensional structure of the protein was modeled and compared to crystal- and NMR-structures of the homologous proteins from T. cruzi and T. brucei respectively, in order to examine structural differences leading to the different immunoreactivity of the T. congolense molecule. Enzyme-linked immunosorbent assays (ELISA) were used to measure antibodies produced by trypanosome-infected African cattle in order to assess the potential for use of T. congolense calflagin in a serodiagnostic assay. Results: The antigen recognized by the T. congolense-specific mAb Tc6/42.6.4 was identified as a flagellar calcium-binding protein, calflagin. The recombinant molecule showed immunoreactivity with the T. congolense-specific mAb confirming that it is the cognate antigen. Immunofluorescence experiments revealed that Ca2+ modulated the localization of the calflagin molecule in trypanosomes. Structural modelling and comparison with calflagin homologues from other trypanosomatids revealed four non-conserved regions on the surface of the T. congolense molecule that due to differences in surface chemistry and structural topography may form species-specific epitopes. ELISAs using the recombinant calflagin as antigen to detect antibodies in trypanosome-infected cattle showed that the majority of cattle had antibody responses. Area under the Receiver-Operating Characteristic (ROC) curves, associated with host IgG and IgM, were calculated to be 0.623 and 0.709 respectively, indicating a positive correlation between trypanosome infection and the presence of anti-calflagin antibodies. Conclusions: While calflagin is conserved among different species of African trypanosomes, our results show that T. congolense calflagin possesses unique epitopes that differentiate this protein from homologues in other trypanosome species. MAb Tc6/42.6.4 has clear utility as a laboratory tool for identifying T. congolense. T. congolense calflagin has potential as a serodiagnostic antigen and should be explored further for its utility in antigen-detection assays for diagnosis of cattle infections. [ABSTRACT FROM AUTHOR]

Background: During natural Trypanosoma brucei infections, the parasites differentiate spontaneously into a non-dividing "stumpy" form when a certain level of parasitaemia is attained. This form is metabolically adapted for rapid further differentiation into procyclic forms upon uptake by Tsetse flies. Results: We describe here four central Ugandan isolates of Trypanosoma brucei rhodesiense that have undergone only three rodent passages since isolation from human patients. As expected, SNP analysis shows that these isolates are more closely related to each other than to the commonly used strains Lister 427, Antat1.1, and TREU927. TREU927 generally has smaller copy numbers of repeated genes than the other strains, while Lister 427 trypanosomes with a 30-year history of in vitro culture and cloning have more histone genes than the other isolates. The recently isolated trypanosomes were grown in rats, and their transcriptomes characterised. In comparison with cultured procyclic and bloodstream forms, there were increases in mRNAs encoding the stumpy-form markers ESAG9 and PIP39, with coordinated alterations in the levels of over 600 additional mRNAs. Numerous mRNAs encoding proteins of no known function were either increased or decreased. The products of the mRNAs that were increased in parallel with PIP39 included not only enzymes of procyclic-form metabolism, but also components of the translational and RNA control machineries. Many of the mRNAs that were decreased in cells with elevated PIP39 reflected reduced cell division. Conclusions: These transcriptomes suggest new avenues for research into the regulation of trypanosome differentiation. [ABSTRACT FROM AUTHOR]

Background: Human African trypanosomiasis (HAT), also known as sleeping sickness, is a parasitic tropical disease. It progresses from the first, haemolymphatic stage to a neurological second stage due to invasion of parasites into the central nervous system (CNS). As treatment depends on the stage of disease, there is a critical need for tools that efficiently discriminate the two stages of HAT. We hypothesized that markers of brain damage discovered by proteomic strategies and inflammation-related proteins could individually or in combination indicate the CNS invasion by the parasite. Methods: Cerebrospinal fluid (CSF) originated from parasitologically confirmed Trypanosoma brucei gambiense patients. Patients were staged on the basis of CSF white blood cell (WBC) count and presence of parasites in CSF. One hundred samples were analysed: 21 from stage 1 (no trypanosomes in CSF and ⩽5 WBC/μL) and 79 from stage 2 (trypanosomes in CSF and/or .5 WBC/μL) patients. The concentration of H-FABP, GSTP-1 and S100β in CSF was measured by ELISA. The levels of thirteen inflammation-related proteins (IL-1ra, IL-1β, IL-6, IL-9, IL-10, G-CSF, VEGF, IFN-&#x03B3, TNF-α, CCL2, CCL4, CXCL8 and CXCL10) were determined by bead suspension arrays. Results: CXCL10 most accurately distinguished stage 1 and stage 2 patients, with a sensitivity of 84% and specificity of 100%. Rule Induction Like (RIL) analysis defined a panel characterized by CXCL10, CXCL8 and H-FABP that improved the detection of stage 2 patients to 97% sensitivity and 100% specificity. Conclusion: This study highlights the value of CXCL10 as a single biomarker for staging T. b. gambiense-infected HAT patients. Further combination of CXCL10 with H-FABP and CXCL8 results in a panel that efficiently rules in stage 2 HAT patients. As these molecules could potentially be markers of other CNS infections and disorders, these results should be validated in a larger multi-centric cohort including other inflammatory diseases such as cerebral malaria and active tuberculosis. [ABSTRACT FROM AUTHOR]