single CD4 T cell gene expression and T cell receptor profile of human volunteers experimentally infected with malaria

Human volunteers were experimentally infected with Plasmodium falciparum blood-stage malaria parasites in the context of the VAC063C experimental medicine study (ClinicalTrials.gov NCT03906474). The gene expression profile and T cell receptor alpha and beta chains of CD4 T cells were analysed at single cell resolution at baseline (before malaria infection) and six days after treatment (at the peak of the circulating T cell response). We are comparing the T cell response to the first malaria infection of life (with the highest risk of severe disease) and the third infection where disease tolerance is established. This single cell data is complemented by bulk RNAseq data of flow sorted CD4 T cell subsets (naive, memory and regulatory) available through the GEO SuperSeries GSE172481. Overall design: CD4 T cells were flow sorted from cryopreserved PBMCs and barcoded using oligo-tagged antibodies. Six individually barcoded samples were pooled (one pool for first malaria infection and a separate pool for third infection). We then super-loaded the 10X Genomics Chromium Controller based on the workflow described by Stoeckius et al. 2018 (doi.org/10.1186/s13059-018-1603-1) to capture approx 30,000 singlets per pool across 2 wells of an A chip. We generated three libraries: i) cell surface barcode, ii) 5' gene expression and iii) T cell receptor alpha and beta chains (after amplification of the V(D)J regions). All libraries were separately indexed and pooled at a ratio of 8% cell surface, 84% gene expression and 8% V(D)J for sequencing on a NovaSeq 6000 Illumina platform to yield at least 66,000 150 bp paired end reads per cell. In total we obtained 5931 million paired end reads (2848 and 3083 million paired end reads for first and third infection, respectively) at the expected ratios.