Transcriptome profiles ofT.b. rhodesiensein Malawi reveal focus specific gene expression Profiles associated with pathology

Abstract

AbstractBackgroundSleeping sickness caused byT.b. rhodesienseis a fatal disease and endemic in Southern and Eastern Africa. There is an urgent need to develop novel diagnostic and control tools in order 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 endemicT. b. rhodesienseisolates in human blood in Malawi.MethodologyBlood 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 againstTrypanosoma bruceireference transcriptome (TriTrypDB54_TbruceiTREU927) using HiSat2. Differential gene expression analysis was done using the DeSeq2 package in R. SNPs calling from reads that were mapped to theT. bruceigenome was done using GATK in order to identifyT.b. rhodesiensepopulation structure.Results24 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 more silent disease in Nkhotakota associated with the non-dividing stumpy form. Interestingly, the MalawiT.b. rhodesienseisolates expressed genes enriched for reduced cell proliferation compared to the UgandaT.b. rhodesienseisolates. PCA analysis using SNPs called from the RNAseq data showed thatT. b. rhodesienseparasites from Nkhotakota are genetically distinct from those collected in Rumphi.ConclusionOur results have added new insights on how clinical phenotypes of r-HAT in Malawi might be associated with differences in gene expression profiles and population structure ofT.b. rhodesiensefrom its two major endemic foci of Rumphi and Nkhotakota.Author SummaryA better understanding ofT. b. rhodesiensegene expression profiles and population structure using endemic isolate may fast track the current search for novel diagnostic and control tools for rhodesiense sleeping sickness. Here, we analysedT. b. rhodesiensetranscriptome profiles from endemic isolated from peripheral blood in Nkhotakota and Rumphi foci in Malawi. In Nkhotakota focus,T. b. rhodesiensetranscripts 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 thatT. b. rhodesiensepopulation 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.