We have demonstrated that gametocytes, the only stage capable of malaria transmission to the mosquito vector, develop in the extravascular environment of the human bone marrow. Preliminary studies in the mouse model have confirmed this finding and suggest existence of an asexual parasite reservoir in the bone marrow. In this innovative multidisciplinary proposal, we aimed to investigate the host pathogen interactions at the interface between infected red blood cell and bone marrow vasculature. Specifically, we focused on the following questions: how do parasites home to bone marrow? What are the changes in the bone marrow endothelium upon infection? How do parasites adhere with and transmigrate across the vascular endothelium in the bone marrow?
In aim 1 we proposed to define the specificity of parasite sequestration and endothelial activation in bone marrow. We have concluded several studies to address this aim. Specifically, we have used stage-specific parasite reporter lines and quantified their distribution across tissues, including bone marrow in the mouse model, demonstrating gametocyte enrichment in bone marrow, spleen and liver. We have also measured the homing of individual parasite stages to specific tissues, including bone marrow and spleen. These experiments further confirmed that bone marrow and spleen are the major sequestration sites for P. berghei gametocytes, in particular the extravascular environment. They also demonstrated that ring and merozoite stages are the stages preferentially homing to bone marrow. Indeed, a subset of merozoites were found within bone marrow resident erythroid cells, demonstrating their homing and subsequent invasion in bone marrow. This work has been published in 2018 (De Niz et al, Sci Adv, 2018). In a next series of experiments, we have performed a systematic study to investigate parasite infection in blood and haematopoietic tissues of spleen and bone marrow, in the rodent malaria model Plasmodium berghei. Specifically, we have used a combination of flow cytometry and single cell RNAseq to quantify the distribution of parasite stages and host cell types across the three locales and over time. These studies defined the host response upon parasite infection and parasite transcriptional signatures according to host cell. Specifically, we defined a host cell tropism for parasite invasion, metabolism and stage conversion in P. berghei. This work has been published in 2022 (Hentzschel et al, Sci Adv, 2022).
In aim 2 we proposed to define the signature of vascular activation in bone marrow upon malaria infection. In the mouse model, we have been able to visualize the extent of vascular leakage across tissues using fluorescent dextran, and demonstrated that it is limited to bone marrow, spleen and liver. This work has been published (De Niz et al., Sci Adv 2018). In in vitro assays with P. falciparum we were able to identify a series of candidate antigens on the gametocyte-infected RBC surface, most of them shared with asexual parasites. This work is also published (Dantzler et al., Sci Trans Med, 2019). Using the mouse model P. berghei we have also concluded a study focusing on parasite vascular sequestration and host responses, revealing a link between sequestration in adipose tissue, activation of the adipokine leptin and cerebral malaria. This work was performed in collaboration with Prof. Jay Mitchell and published recently (Mejia et al., Sci Adv 2021).
In aim 3 we proposed to elucidate mechanisms by which malaria parasites enter and exit the bone marrow parenchyma. We defined a series of mobility phenotypes of mature gametocytes in bone marrow and spleen, including their deformability and passage across the endothelial barrier. We also demonstrated that specific receptor-ligand interactions are required for merozoite homing to bone marrow, in particular P-selectin. This work has been published (De Niz et al., Sci Adv 2018).
Finally, we have published a review article (Venugopal et al., Nat Rev Micro 2020), providing the first overview of the new paradigm of haematopoietic infection in malaria.
