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1074 The Darc Side of Vivax Malaria in Africa: Unveiling Invasion Pathways into Duffy-Negative Erythroblasts

Program: Oral and Poster Abstracts
Session: 101. Red Cells and Erythropoiesis, Excluding Iron: Poster I
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Translational Research
Saturday, December 9, 2023, 5:30 PM-7:30 PM

Isabelle Bouyssou, PhD1,2*, Sara El Hoss, PhD3*, Cécile Doderer-Lang4*, Matthieu Schoenhals, PhD5*, Lova Tsikiniaina Rasoloharimanana5*, Inès Vigan-Womas6*, Arsène Ratsimbasoa7*, David C Rees, MBBS3, Andargie Abate8*, Lemu Golassa, PhD8*, Solenne Mabilotte4*, Micheline Guillotte2*, Francisco J. Martinez Blazquez2*, Chetan E. Chitnis2*, John Strouboulis, PhD3* and Didier Ménard, PhD1,2,4,9*

1Malaria Genetics and Resistance Unit, INSERM U1201, Institut Pasteur, Université Paris Cité, Paris, France
2Malaria Parasite Biology and Vaccines Unit, Institut Pasteur, Université Paris Cité, Paris, France
3Red Cell Haematology Lab, School of Cancer and Pharmaceutical Sciences, King's College London, London, United Kingdom
4Institute of Parasitology and Tropical Diseases, UR7292 Dynamics of Host-Pathogen Interactions, University of Strasbourg, Strasbourg, France
5Immunology of Infectious Diseases, Institut Pasteur of Madagascar, Antananarivo, Madagascar
6Institut Pasteur de Dakar, Dakar, Senegal
7Faculté de Médecine, Université de Fianarantsoa, Fianarantsoa, Madagascar
8Aklilu Lemma Institute of Pathobiology, Addis Ababa University, Addis Ababa, Ethiopia
9Laboratory of Parasitology and Medical Mycology, CHU Strasbourg, Strasbourg, France

Background & Objectives: Plasmodium vivax malaria was long thought to be absent from sub-Saharan Africa owing to the high prevalence of people lacking the Duffy antigen receptor for chemokines (DARC) on their erythrocytes. The interaction between P. vivax Duffy binding protein (PvDBP) and DARC is assumed to be the main mechanism used by P. vivax merozoites to invade human erythrocytes. However, the increasing numbers of vivax malaria cases in Duffy-negative African individuals has raised questions about alternative P. vivax invasion pathway(s) other than PvDBP-DARC interaction. Since P. vivax has a tropism for CD71+ immature reticulocytes and the hematopoietic niches of the bone marrow may be a hidden reservoir of P. vivax parasites, we hypothesized that P. vivax merozoites may be able to invade erythroblasts derived from Duffy-positive (DP) and Duffy-negative (DN) individuals. Therefore, our objectives were to (i) investigate the expression of DARC during DP and DN erythropoiesis and (ii) examine the infection of DP and DN erythroblasts with P. vivax merozoites in vitro to study and identify host receptors used by P. vivax to invade erythroblasts.

Methods: We performed in vitro erythropoiesis assays (two-phase culture) of genotypically DN and DP erythroid progenitors. Erythroid differentiation, CD71 and DARC expression profiles were monitored using flow cytometry, microscopy, and western blot over time during phase 2. In vitro invasion assays were conducted using DP and DN erythroblasts to investigate whether genotypically DN erythroblasts could be invaded by P. vivax. At D7 of phase 2, we thawed and cultured parasite isolates obtained from two Malagasy and seven Ethiopian patients infected with P. vivax. After approximately 24-30 hours of in vitro maturation (at D8 of phase 2), each P. vivax parasite culture was placed in co-culture with either DP or DN erythroblasts for 24-48 hours. We examined the infection of DP and DN erythroblasts by P. vivax parasites by light and confocal microscopy.

Results: Using in vitro erythropoiesis, we show that both genotypically DP and DN erythroblasts have similar kinetics of terminal erythroid differentiation. Moreover, we found similar CD71 expression profiles in both DP and DN erythroblasts. Interestingly, we observed that a subset of DN erythroblasts transiently expressed DARC during erythropoiesis. This was confirmed using both flow cytometry and western blot analysis. In vitro invasion assays conducted on both DP and DN erythroblasts using parasite isolates, showed that P. vivax merozoites were able to invade both DP and DN erythroblasts (Figure 1). We found that all the DP and DN erythroblasts that were infected with P. vivax were CD71+ and DARC+. These results confirm that P. vivax can invade DP and DN erythroblasts expressing both DARC and CD71, irrespective of the origin of the isolates.

Conclusion: This study provides the first biological evidence for the ability of P. vivax to invade genotypically DN erythroblasts and highlights that African populations may represent a silent reservoir of P. vivax infections, with major implications for the control and elimination of vivax malaria in sub-Saharan Africa.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH