Vaccine effectiveness against malaria is dramatically reduced in malaria-exposed compared to malaria-naïve populations, potentially due to altered immune responses in secondary lymphoid organs following repeated infection. Newly developed human tonsil and spleen organoids, which replicate key features of B and T cell immunity, provide an exciting opportunity to overcome challenges of other models and to improve our understanding of innate-adaptive interactions in lymphoid tissue. The objectives of this study were to use these organoids to investigate the impact of malaria parasites on 1) cells within lymphoid tissues and 2) responses to a heterologous antigen.

To better characterize the impact of iRBC stimulation on human lymphoid reponses, we cocultured three human spleen organoids with iRBC and uninfected red blood cells for 10 days. We then enriched for Vd2+ cells using fluorescence-activated cell sorting, and used the BD Rhapsody platform to generate multiomic single-cell data, combining transcriptomic, T cell receptor sequence, and surface protein expression data. Data contained in this repository represents the Seurat object generated after alignment and cell calling, but before quality control. We hope these data are useful for researchers investigating the utility of organoids as a means of enabling tissue-specific immunity in humans. Our experiments were done with post-autopsy human tissues, following established protocols and institutional ethics review. We report no conflicts of interest.

Spleens organoids were prepared as per Wager et al. 2021 (PMC7891554). Briefly, cryopreserved splenic cell suspensions were thawed and counted, 6 x 106 cells in 100 µl immune organoid media were added to each transwell (Millipore PICM01250) in 12-well plates. Immune organoid media containing 0.5 µl g/mL recombinant human B cell activation factor (BAFF, Biolegend 559608) was added to the outside of the well. Spleen tissue from 3 donors was used and an aliquot of cells from d0 was frozen for each. Organoids were set up and stimulated with uRBC or 3D7 iRBC (12 x 106 iRBC or uRBC/well) for 10 days with media changes every 3-4 days. Stimulated cells, as well as d0 cells, were then stained with an anti-Vδ2+ antibody (PerCP-conjugated, 1:50) and a cell viability dye (Live/Dead Aqua, 1:400) for 30 minutes, washed, and then sorted on a Sony cell sorter. For each sample, 8,000-40,000 Vδ2+ γδ T cells were sorted and depending on the number collected, 10,000-30,000 Vδ2- cells were sorted in order to reach a total of 40,000-50,000 cells per sample. Cells were sorted into cold PBS containing 2% fetal bovine serum. Fewer cells were collected for corresponding PBMC samples, which as would be expected from a blood sample following autopsy, had a lot of dead cells and debris. The percentage of total cells that were Vδ2+ T cells ranged from 20-80% per sample (80% for all iRBC-stimulated cells, lower for some uRBC-stimulated and d0). Samples were transferred to the Stanford Genomics Core, where they were then barcoded and stained with 13 BD AbSeq Antibody-Oligonucleotides Conjugates corresponding to cell markers of interest (see table below). Single cells (15,000-35,000 live cells per sample with the exception of the PBMC sample, which had 4925) were captured on the BD Rhapsody and mRNA was captured and sequenced. The BD Rhapsody Analysis Pipeline was used to generate sequencing data for transcriptomes, as well as T and B cell receptors.