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Background: Sleeping sickness caused by Trypanosoma brucei rhodesiense is a fatal disease and endemic in Southern and Eastern Africa. There is an urgent need to develop novel diagnostic and control tools to achieve elimination of rhodesiense sleeping sickness which might be achieved through a better understanding of trypanosome gene expression and genetics using endemic isolates. Here, we describe transcriptome profiles and population structure of endemic T. b. rhodesiense isolates in human blood in Malawi. Methodology: Blood samples of r-HAT cases from Nkhotakota and Rumphi foci were collected in PaxGene tubes for RNA extraction before initiation of r-HAT treatment. 100 million reads were obtained per sample, reads were initially mapped to the human genome reference GRCh38 using HiSat2 and then the unmapped reads were mapped against Trypanosoma brucei reference transcriptome (TriTrypDB54_TbruceiTREU927) using HiSat2. Differential gene expression analysis was done using the DeSeq2 package in R. SNP calling from reads that were mapped to the T. brucei genome was done using GATK in order to identify T.b. rhodesiense population structure. Results: 24 samples were collected from r-HAT cases of which 8 were from Rumphi and 16 from Nkhotakota foci. The isolates from Nkhotakota were enriched with transcripts for cell cycle arrest and stumpy form markers, whereas isolates in Rumphi focus were enriched with transcripts for folate biosynthesis and antigenic variation pathways. These parasite focus-specific transcriptome profiles are consistent with the more virulent disease observed in Rumphi and a less symptomatic disease in Nkhotakota associated with the non-dividing stumpy form. Interestingly, the Malawi T.b. rhodesiense isolates expressed genes enriched for reduced cell proliferation compared to the Uganda T.b. rhodesiense isolates. PCA analysis using SNPs called from the RNAseq data showed that T. b. rhodesiense parasites from Nkhotakota are genetically distinct from those collected in Rumphi. Conclusion: Our results suggest that the differences in disease presentation in the two foci is mainly driven by genetic differences in the parasites in the two major endemic foci of Rumphi and Nkhotakota rather than differences in the environment or host response. Author summary: A better understanding of Trypanosoma brucei rhodesiense gene expression profiles and population structure using endemic isolate may speed up the search for novel diagnostic and control tools for rhodesiense sleeping sickness. Here, we analysed T. b. rhodesiense transcriptome profiles from endemic isolated from peripheral blood in Nkhotakota and Rumphi foci in Malawi. In Nkhotakota focus, T. b. rhodesiense transcripts were enriched for cell cycle arrest and stumpy marker whereas in Rumphi focus, the isolates were enriched for antigenic variation and folate biosynthesis biological pathways. Furthermore, we also found that T. b. rhodesiense population structure in Nkhotakota focus is different from Rumphi focus. The differences in trypanosome gene expression profiles and population structure are consistent with a less severe and acute sleeping sickness clinical profiles in Nkhotakota and Rumphi foci respectively and suggest that the differences in pathology in the two foci are mainly due to differences in the parasite rather than in the host response. [ABSTRACT FROM AUTHOR]

T. b. rhodesiense is the causative agent of Rhodesian human African trypanosomiasis (r-HAT) in Malawi. Clinical presentation of r-HAT in Malawi varies between foci and differs from East African HAT clinical phenotypes. The purpose of this study was to gain more insights into the transcriptomic profiles of patients with early stage 1 and late stage 2 HAT disease in Malawi. Whole blood from individuals infected with T. b. rhodesiense was used for RNA-Seq. Control samples were from healthy trypanosome negative individuals matched on sex, age range, and disease foci. Illumina sequence FASTQ reads were aligned to the GRCh38 release 84 human genome sequence using HiSat2 and differential analysis was done in R Studio using the DESeq2 package. XGR, ExpressAnalyst and InnateDB algorithms were used for functional annotation and gene enrichment analysis of significant differentially expressed genes. RNA-seq was done on 23 r-HAT case samples and 28 healthy controls with 7 controls excluded for downstream analysis as outliers. A total of 4519 genes were significant differentially expressed (p adjusted <0.05) in individuals with early stage 1 r-HAT disease (n = 12) and 1824 genes in individuals with late stage 2 r-HAT disease (n = 11) compared to controls. Enrichment of innate immune response genes through neutrophil activation was identified in individuals with both early and late stages of the disease. Additionally, lipid metabolism genes were enriched in late stage 2 disease. We further identified uniquely upregulated genes (log2 Fold Change 1.4–2.0) in stage 1 (ZNF354C) and stage 2 (TCN1 and MAGI3) blood. Our data add to the current understanding of the human transcriptome profiles during T. b. rhodesiense infection. We further identified biological pathways and transcripts enriched than were enriched during stage 1 and stage 2 r-HAT. Lastly, we have identified transcripts which should be explored in future research whether they have potential of being used in combination with other markers for staging or r-HAT. Author summary: Clinical outcomes of Rhodesian sleeping sickness in endemic countries varies depending on disease foci. To gain more insight of r-HAT disease in Malawi, we analyzed gene expression profiles in blood from 23 individuals with early stage 1 and late stage 2 sleeping sickness versus 21 healthy controls. We found differences in gene expression profiles in individuals with stage 1 and stage 2 disease but not between Malawi's r-HAT foci. Furthermore, innate immune response transcripts were elevated in individuals with stage 1 sleeping sickness, and there was enrichment of proliferating neutrophil precursors in cases versus controls. We have also shown that wasting in individuals with late stage sleeping sickness might be linked with altered lipid metabolic process. Lastly, we have identified transcripts that in combination with other markers maybe explored in future for possibility of being used in staging of sleeping sickness without the need of the invasive lumber puncture. Our findings add to the current understanding of human response to T. b. rhodesiense infection in Malawian r-HAT patients. [ABSTRACT FROM AUTHOR]

Background: Individuals genetically susceptible to high schistosomiasis worm burden may contribute disproportionately to transmission and could be prioritized for control. Identifying genes involved may guide development of therapy. Methodology/Principal findings: A cohort of 606 children aged 10–15 years were recruited in the Albert Nile region of Uganda and assessed for Schistosoma mansoni worm burden using the Up-Converting Particle Lateral Flow (UCP-LF) test detecting circulating anodic antigen (CAA), point-of-care Circulating Cathodic Antigen (POC-CCA) and Kato-Katz tests. Whole genome genotyping was conducted on 326 children comprising the top and bottom 25% of worm burden. Linear models were fitted to identify variants associated with worm burden in preselected candidate genes. Expression quantitative trait locus (eQTL) analysis was conducted for candidate genes with UCP-LF worm burden included as a covariate. Single Nucleotide Polymorphism loci associated with UCP-LF CAA included IL6 rs2066992 (OR = 0.43, p = 0.0006) and rs7793163 (OR = 2.0, p = 0.0007); IL21 SNP kgp513476 (OR 1.79, p = 0.0025) and IL17B SNP kgp708159 (OR = 0.35, p = 0.0028). A haplotype in the IL10 locus was associated with lower worm burden (OR = 0.53, p = 0.015) and overlapped SNPs rs1800896, rs1800871 and rs1800872. Significant haplotypes (p<0.05, overlapping significant SNP) associated with worm burden were observed in IL6 and the Th17 pathway IL12B and IL17B genes. There were significant eQTL in the IL6, IL5, IL21, IL25 and IFNG regions. Conclusions: Variants associated with S. mansoni worm burden were in IL6, FCN2, RNASE3, IL10, IL12B and IL17B gene loci. However only eQTL associations remained significant after Bonferroni correction. In summary, immune balance, pathogen recognition and Th17 pathways may play a role in modulating Schistosoma worm burden. Individuals carrying risk variants may be targeted first in allocation of control efforts to reduce the burden of schistosomiasis in the community. Author summary: This study investigated genetic risk factors for high worm burden in a cohort of 606 children aged 10–15 years old in the Albert Nile region of Uganda. Genotyping was conducted on 326 children who comprised the top and bottom 25% of worm burden. Results showed single nucleotide polymorphisms (SNPs) associated with worm burden in the IL6, IL21 and IL17B genes, and a haplotype in the IL10 locus associated with lower worm burden. Expression quantitative trait locus (eQTL) analysis revealed significant gene and worm burden associations in the IL6 region, and expression associations in the IL5, IL21, IL25 and IFNG loci. These findings suggest that immune balance, pathogen recognition and Th17 pathways may be involved in modulating Schistosoma worm burden and that individuals carrying risk variants may be prioritized to improve control efforts. [ABSTRACT FROM AUTHOR]

Over 290 million people are infected by schistosomes worldwide. Schistosomiasis control efforts focus on mass drug treatment with praziquantel (PZQ), a drug that kills the adult worm of all Schistosoma species. Nonetheless, re-infections have continued to be detected in endemic areas with individuals living in the same area presenting with varying infection intensities. Our objective was to characterize the transcriptome profiles in peripheral blood of children between 10–15 years with varying intensities of Schistosoma mansoni infection living along the Albert Nile in Uganda. RNA extracted from peripheral blood collected from 44 S. mansoni infected (34 high and 10 low by circulating anodic antigen [CAA] level) and 20 uninfected children was sequenced using Illumina NovaSeq S4 and the reads aligned to the GRCh38 human genome. Differential gene expression analysis was done using DESeq2. Principal component analysis revealed clustering of gene expression by gender when S. mansoni infected children were compared with uninfected children. In addition, we identified 14 DEGs between S. mansoni infected and uninfected individuals, 56 DEGs between children with high infection intensity and uninfected individuals, 33 DEGs between those with high infection intensity and low infection intensity and no DEGs between those with low infection and uninfected individuals. We also observed upregulation and downregulation of some DEGs that are associated with fibrosis and its regulation. These data suggest expression of fibrosis associated genes as well as genes that regulate fibrosis in S. mansoni infection. The relatively few significant DEGS observed in children with schistosomiasis suggests that chronic S. mansoni infection is a stealth infection that does not stimulate a strong immune response. Author summary: Schistosomiasis is a neglected tropical disease transmitted via an intermediate snail host through contact with contaminated fresh water. Even with routine Mass Drug Administration for treatment of the infection, re-infections are still common and variations in infection intensity and pathology are still observed in individuals in the same location. These may be due to differences in individuals' response to S. mansoni infection. In this study, we used RNAseq to identify differentially expressed genes associated with S. mansoni infection in children between 10–15 years. We conducted comparisons between phenotypes including infection intensities measured by circulating anodic antigen, wasting by body mass index and stunting by height-for-age Z-score. Our data showed very low numbers of significantly differentially expressed genes in all comparisons. Some of the few differentially expressed genes that were observed were associated with fibrosis which is the cause of pathology in humans and has been observed in late stages of S. mansoni infection in murine studies. [ABSTRACT FROM AUTHOR]