Spatial imaging and transcriptomic analysis of immune cells and malaria parasites in Plasmodium-infected human spleens

Spatial imaging and transcriptomic analysis of immune cells and malaria parasites in Plasmodium-infected human spleens. The human spleen is an immune sentinel and controls red blood cell quality. In malaria, the spleen has predominantly been considered a clearing filter for parasites and is a major lymphoid tissue for immune responses to blood-stage pathogens. However, recent studies suggest the existence of a more complex role, not limited to parasite clearance and destruction. Our paradigm-shifting study in chronic malaria has revealed that the spleen is the major niche for asexual-stage development of P. falciparum (Pf) and P. vivax (Pv), and points to the presence of an endosplenic lifecycle where malaria parasites can maintain infection in the reticulocyte-rich human spleen. This is particularly intense for Pv, with the spleen estimated to harbor >98% of total asexual biomass. While we have shown the spleen to be the major parasite reservoir in natural infections, an increased understanding of the mechanisms by which malaria parasites accumulate, avoid drug-killing, and suppress/evade defense systems in the spleen will be critical to the development of effective elimination strategies.

We now have unique access to malarial spleen tissue and blood samples from untreated chronic infections in Indonesia to understand the biology of the splenic parasite reservoir. We hypothesize that malaria parasites drive unique cellular, immunological and architectural changes to support an endosplenic lifecycle and the suppression/evasion of host immune responses. Here, we will leverage our unique access to the world’s only library of human spleens and blood samples obtained from semi-immune individuals with chronic malaria and aim to uncover; a) how malaria parasites take out host defense systems within the spleen; b) how the parasites have adapted to survive and replicate in this tissue, and; c) the mechanisms that lead to malaria immunity, which is thought to develop primarily within this organ.

Healthy human spleen tissue blocks and sections (unfixed and fixed) will be purchased from AMSbio, a commercial certified supplier of human tissue for research. CI Kho will travel to QIMRB and support CIs Loughland and Andrew in constructing five colour multi-parameter histocytometry panels to identify the phenotype, function and spatial distribution of specific splenic host cell populations. In particular, we will focus on T follicular helper cells (Tfh), B cells and follicular DCs (FDCs) within B cell follicles and germinal centers, as these populations are predicted to play key roles in malarial immune responses and vaccine responsiveness.

Human spleen samples will also be used to optimize wet and dry lab protocols and analytical tools for spatial transcriptomics using the cutting-edge 10x genomics platform available in QIMRB’s network. This includes the careful preparation of tissue on to visium spatial tissue slides, histological and immunofluorescent tissue staining, followed by tissue permeabilization and library construction, all based on the 10x Visium Spatila tissue preparation protocol. CI Chabikwa, a bioinformatician with expertise in RNA sequencing and analysis of large datasets, will establish an analytical pipeline to combine all datasets and generate high-resolution cellular and transcriptomic profiles of large tissue sections, ultimately producing a detailed cell atlas of human spleen tissue. CI Chabikwa, will be supported by CI Boyle, who is a leading expert in human immune responses in malaria and single-cell transcriptomics and will provide key leadership, support data generation, interpretation and publication.

Enabling Technologies & Assays

Mar 2021 — Oct 2023

$15,000

Australia

Australian Centre for Research Excellence in Malaria Elimination (ACREME)