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Korbmacher F, Bernabeu M. Induced pluripotent stem cell-based tissue models to study malaria: a new player in the research game. Curr Opin Microbiol 2025; 84:102585. [PMID: 40010012 DOI: 10.1016/j.mib.2025.102585] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/02/2024] [Revised: 01/27/2025] [Accepted: 02/07/2025] [Indexed: 02/28/2025]
Abstract
Most in vitro studies on parasite development and pathogenesis in the human host have been conducted using traditional primary or immortalized cells, despite their inherent limitations. Breakthroughs in the field of induced pluripotent stem cells (iPSCs) are revolutionizing disease modeling, offering alternatives to traditional in vivo and in vitro infection models. Human iPSCs differentiate into all cell types, proliferate indefinitely, and offer experimental advantages, like genome editing and donor control. iPSCs can be engineered into complex 3D tissue models that closely mimic morphology and function of their in vivo counterparts and allow for precise experimental manipulation. The physiological complexity of iPSC-based tissue models has improved rapidly. Given Plasmodium's systemic impact across multiple organs, these models provide an invaluable resource for studying parasite-tissue interactions. This opinion article focuses on recent developments of iPSC-based models for Plasmodium research. We describe the main highlights and potential use of these systems while acknowledging current limitations.
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Affiliation(s)
- François Korbmacher
- European Molecular Biology Laboratory (EMBL) Barcelona, Carrer del Doctor Aiguader 88, 08003 Barcelona, Spain
| | - Maria Bernabeu
- European Molecular Biology Laboratory (EMBL) Barcelona, Carrer del Doctor Aiguader 88, 08003 Barcelona, Spain.
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2
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Sima N, Ayllon-Hermida A, Fernández-Becerra C, del Portillo HA. Extracellular vesicles in malaria: proteomics insights, in vitro and in vivo studies indicate the need for transitioning to natural human infections. mBio 2025; 16:e0230424. [PMID: 39868784 PMCID: PMC11898581 DOI: 10.1128/mbio.02304-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2025] Open
Abstract
Globally, an estimated 2.1 billion malaria cases and 11.7 million malaria deaths were averted in the period 2000-2022. Noticeably, despite effective control measurements, in 2022 there were an estimated 249 million malaria cases in 85 malaria-endemic countries and an increase of 5 million cases compared with 2021. Further understanding the biology, epidemiology, and pathogenesis of human malaria is therefore essential for achieving malaria elimination. Extracellular vesicles (EVs) are membrane-enclosed nanoparticles pivotal in intercellular communication and secreted by all cell types. Here, we will review what is currently known about EVs in malaria, from biogenesis and cargo to molecular insights of pathophysiology. Of relevance, a meta-analysis of proteomics cargo, and comparisons between in vitro and in vivo human studies revealed striking differences with those few studies reported from patients. Thus, indicating the need for rigor standardization of methodologies and for transitioning to human infections to elucidate their physiological role. We conclude with a focus on translational aspects in diagnosis and vaccine development and highlight key gaps in the knowledge of EVs in malaria research.
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Affiliation(s)
- Núria Sima
- ISGlobal, Barcelona, Spain
- IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
| | - Alberto Ayllon-Hermida
- ISGlobal, Barcelona, Spain
- IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
- School of Medicine and Health Sciences, University of Barcelona, Barcelona, Spain
| | - Carmen Fernández-Becerra
- ISGlobal, Barcelona, Spain
- IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
- CIBERINFEC, ISCIII-CIBER de Enfermedades Infecciosas, Instituto de Salud Carlos III, Madrid, Spain
| | - Hernando A. del Portillo
- ISGlobal, Barcelona, Spain
- IGTP, Germans Trias i Pujol Research Institute, Badalona, Barcelona, Spain
- Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Spain
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3
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Rodríguez-Obediente K, Yepes-Pérez Y, Benavides-Ortiz D, Díaz-Arévalo D, Reyes C, Arévalo-Pinzón G, Patarroyo MA. Invasion-inhibitory peptides chosen by natural selection analysis as an antimalarial strategy. Mol Immunol 2023; 163:86-103. [PMID: 37769577 DOI: 10.1016/j.molimm.2023.09.013] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2023] [Revised: 09/06/2023] [Accepted: 09/14/2023] [Indexed: 10/03/2023]
Abstract
Plasmodium vivax's biological complexity has restricted in vitro culture development for characterising antigens involved in erythrocyte invasion and their immunological relevance. The murine model is proposed as a suitable alternative in the search for therapeutic candidates since Plasmodium yoelii uses homologous proteins for its invasion. The AMA-1 protein is essential for parasite invasion of erythrocytes as it is considered an important target for infection control. This study has focused on functional PyAMA-1 peptides involved in host-pathogen interaction; the protein is located in regions under negative selection as determined by bioinformatics analysis. It was found that pyama1 has two highly conserved regions amongst species (>70%) under negative selection. Fourteen synthetic peptides spanning both conserved regions were evaluated; 5 PyAMA-1 peptides having high specific binding (HABP) to murine erythrocytes were identified. The parasite's invasion inhibition capability was analysed through in vitro assays, suggesting that peptides 42681 (43-ENTERSIKLINPWDKYMEKY-62), 42903 (206-RYSSNDANNENQPFSFTPEK-225) and 42904 (221-FTPEKIENYKDLSYLTKNLR-240) had greater than 50% inhibition profile and restricted P. yoelii intra-erythrocyte development. This work proposes that the screening of conserved HABPs under negative selective pressure might be good candidates for developing a synthetic anti-malarial vaccine since they share functionally-relevant characteristics, such as interspecies conservation, specific RBC binding profile, invasion and parasite development inhibition capability, and the predicted B-epitopes within were recognised by sera obtained from experimentally-infected mice.
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Affiliation(s)
- Kewin Rodríguez-Obediente
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia; MSc programme in Microbiology, Biotechnology Institute, Universidad Nacional de Colombia, Bogotá, Colombia
| | - Yoelis Yepes-Pérez
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia
| | - Daniel Benavides-Ortiz
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia; School of Health Sciences, Universidad Colegio Mayor de Cundinamarca, Bogotá, Colombia
| | - Diana Díaz-Arévalo
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia; Animal Science Faculty, Universidad de Ciencias Aplicadas y Ambientales (U.D.C.A), Bogotá, Colombia
| | - César Reyes
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia
| | - Gabriela Arévalo-Pinzón
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia; Microbiology Department, Science Faculty, Pontificia Universidad Javeriana, Bogotá, Colombia
| | - Manuel Alfonso Patarroyo
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Bogotá, Colombia; Microbiology Department, Faculty of Medicine, Universidad Nacional de Colombia, Bogotá, Colombia.
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Kumari S, Sinha A. Culture and transfection: Two major bottlenecks in understanding Plasmodium vivax biology. Front Microbiol 2023; 14:1144453. [PMID: 37082177 PMCID: PMC10110902 DOI: 10.3389/fmicb.2023.1144453] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2023] [Accepted: 02/28/2023] [Indexed: 04/07/2023] Open
Abstract
The long term in vitro culture of Plasmodium falciparum was successfully established by Trager and Jensen in 1976; however it largely remains unachieved for P. vivax. The major obstacle associated with Plasmodium vivax in vitro culture is its predilection for invading younger reticulocytes and the complex remodelling of invaded reticulocytes. There are many factors under exploration for this predilection and host–parasite interactions between merozoites and invaded reticulocytes. These include various factors related to parasite, host and environment such as compromised reticulocyte osmotic stability after invasion, abundance of iron in the reticulocytes which makes them favourable for P. vivax growth and propagation and role of a hypoxic environment in P. vivax in vitro growth. P. vivax blood stage transfection represents another major hurdle towards understanding this parasite’s complex biology. Efforts in making this parasite amenable for molecular investigation by genetic modification are limited. Newer approaches in sustaining a longer in vitro culture and thereby help advancing transfection technologies in P. vivax are urgently needed that can be explored to understand the unique biology of this parasite.
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Roobsoong W, Yadava A, Draper SJ, Minassian AM, Sattabongkot J. The challenges of Plasmodium vivax human malaria infection models for vaccine development. Front Immunol 2023; 13:1006954. [PMID: 36685545 PMCID: PMC9849360 DOI: 10.3389/fimmu.2022.1006954] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2022] [Accepted: 12/09/2022] [Indexed: 01/07/2023] Open
Abstract
Controlled Human Malaria Infection models (CHMI) have been critical to advancing new vaccines for malaria. Stringent and safe preparation of a challenge agent is key to the success of any CHMI. Difficulty producing the Plasmodium vivax parasite in vitro has limited production of qualified parasites for CHMI as well as the functional assays required to screen and down-select candidate vaccines for this globally distributed parasite. This and other challenges to P. vivax CHMI (PvCHMI), including scientific, logistical, and ethical obstacles, are common to P. vivax research conducted in both non-endemic and endemic countries, with additional hurdles unique to each. The challenges of using CHMI for P. vivax vaccine development and evaluation, lessons learned from previous and ongoing clinical trials, and the way forward to effectively perform PvCHMI to support vaccine development, are discussed.
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Affiliation(s)
- Wanlapa Roobsoong
- Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand
| | - Anjali Yadava
- Biologics Research & Development, Walter Reed Army Institute of Research, Silver Spring, MD, United States
| | - Simon J. Draper
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | | | - Jetsumon Sattabongkot
- Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand
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Satchwell TJ. Generation of red blood cells from stem cells: Achievements, opportunities and perspectives for malaria research. Front Cell Infect Microbiol 2022; 12:1039520. [PMID: 36452302 PMCID: PMC9702814 DOI: 10.3389/fcimb.2022.1039520] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2022] [Accepted: 10/21/2022] [Indexed: 06/22/2024] Open
Abstract
Parasites of the genus Plasmodium that cause malaria survive within humans by invasion of, and proliferation within, the most abundant cell type in the body, the red blood cell. As obligate, intracellular parasites, interactions between parasite and host red blood cell components are crucial to multiple aspects of the blood stage malaria parasite lifecycle. The requirement for, and involvement of, an array of red blood cell proteins in parasite invasion and intracellular development is well established. Nevertheless, detailed mechanistic understanding of host cell protein contributions to these processes are hampered by the genetic intractability of the anucleate red blood cell. The advent of stem cell technology and more specifically development of methods that recapitulate in vitro the process of red blood cell development known as erythropoiesis has enabled the generation of erythroid cell stages previously inaccessible in large numbers for malaria studies. What is more, the capacity for genetic manipulation of nucleated erythroid precursors that can be differentiated to generate modified red blood cells has opened new horizons for malaria research. This review summarises current methodologies that harness in vitro erythroid differentiation of stem cells for generation of cells that are susceptible to malaria parasite invasion; discusses existing and emerging approaches to generate novel red blood cell phenotypes and explores the exciting potential of in vitro derived red blood cells for improved understanding the broad role of host red blood cell proteins in malaria pathogenesis.
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Leong YW, Russell B, Malleret B, Rénia L. Erythrocyte tropism of malarial parasites: The reticulocyte appeal. Front Microbiol 2022; 13:1022828. [PMID: 36386653 PMCID: PMC9643692 DOI: 10.3389/fmicb.2022.1022828] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2022] [Accepted: 10/07/2022] [Indexed: 10/28/2023] Open
Abstract
Erythrocytes are formed from the enucleation of erythroblasts in the bone marrow, and as erythrocytes develop from immature reticulocytes into mature normocytes, they undergo extensive cellular changes through their passage in the blood. During the blood stage of the malarial parasite life cycle, the parasite sense and invade susceptible erythrocytes. However, different parasite species display varying erythrocyte tropisms (i.e., preference for either reticulocytes or normocytes). In this review, we explore the erythrocyte tropism of malarial parasites, especially their predilection to invade reticulocytes, as shown from recent studies. We also discuss possible mechanisms mediating erythrocyte tropism and the implications of specific tropisms to disease pathophysiology. Understanding these allows better insight into the role of reticulocytes in malaria and provides opportunities for targeted interventions.
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Affiliation(s)
- Yew Wai Leong
- A*STAR Infectious Diseases Labs, Agency for Science, Technology and Research, Singapore, Singapore
| | - Bruce Russell
- Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand
| | - Benoit Malleret
- Singapore Immunology Network, Agency for Science, Technology and Research, Singapore, Singapore
- Department of Microbiology and Immunology, Immunology Translational Research Program, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore
| | - Laurent Rénia
- A*STAR Infectious Diseases Labs, Agency for Science, Technology and Research, Singapore, Singapore
- Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore, Singapore
- School of Biological Sciences, Nanyang Technological University, Singapore, Singapore
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Dumarchey A, Lavazec C, Verdier F. Erythropoiesis and Malaria, a Multifaceted Interplay. Int J Mol Sci 2022; 23:ijms232112762. [PMID: 36361552 PMCID: PMC9657351 DOI: 10.3390/ijms232112762] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2022] [Revised: 10/19/2022] [Accepted: 10/20/2022] [Indexed: 01/24/2023] Open
Abstract
One of the major pathophysiologies of malaria is the development of anemia. Although hemolysis and splenic clearance are well described as causes of malarial anemia, abnormal erythropoiesis has been observed in malaria patients and may contribute significantly to anemia. The interaction between inadequate erythropoiesis and Plasmodium parasite infection, which partly occurs in the bone marrow, has been poorly investigated to date. However, recent findings may provide new insights. This review outlines clinical and experimental studies describing different aspects of ineffective erythropoiesis and dyserythropoiesis observed in malaria patients and in animal or in vitro models. We also highlight the various human and parasite factors leading to erythropoiesis disorders and discuss the impact that Plasmodium parasites may have on the suppression of erythropoiesis.
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Affiliation(s)
- Aurélie Dumarchey
- Inserm U1016, CNRS UMR8104, Université Paris Cité, Institut Cochin, 75014 Paris, France
- Laboratoire d’Excellence GR-Ex, 75015 Paris, France
| | - Catherine Lavazec
- Inserm U1016, CNRS UMR8104, Université Paris Cité, Institut Cochin, 75014 Paris, France
- Laboratoire d’Excellence GR-Ex, 75015 Paris, France
| | - Frédérique Verdier
- Inserm U1016, CNRS UMR8104, Université Paris Cité, Institut Cochin, 75014 Paris, France
- Laboratoire d’Excellence GR-Ex, 75015 Paris, France
- Correspondence:
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Aparici Herraiz I, Caires HR, Castillo-Fernández Ó, Sima N, Méndez-Mora L, Risueño RM, Sattabongkot J, Roobsoong W, Hernández-Machado A, Fernandez-Becerra C, Barrias CC, del Portillo HA. Advancing Key Gaps in the Knowledge of Plasmodium vivax Cryptic Infections Using Humanized Mouse Models and Organs-on-Chips. Front Cell Infect Microbiol 2022; 12:920204. [PMID: 35873153 PMCID: PMC9302440 DOI: 10.3389/fcimb.2022.920204] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2022] [Accepted: 06/06/2022] [Indexed: 11/13/2022] Open
Abstract
Plasmodium vivax is the most widely distributed human malaria parasite representing 36.3% of disease burden in the South-East Asia region and the most predominant species in the region of the Americas. Recent estimates indicate that 3.3 billion of people are under risk of infection with circa 7 million clinical cases reported each year. This burden is certainly underestimated as the vast majority of chronic infections are asymptomatic. For centuries, it has been widely accepted that the only source of cryptic parasites is the liver dormant stages known as hypnozoites. However, recent evidence indicates that niches outside the liver, in particular in the spleen and the bone marrow, can represent a major source of cryptic chronic erythrocytic infections. The origin of such chronic infections is highly controversial as many key knowledge gaps remain unanswered. Yet, as parasites in these niches seem to be sheltered from immune response and antimalarial drugs, research on this area should be reinforced if elimination of malaria is to be achieved. Due to ethical and technical considerations, working with the liver, bone marrow and spleen from natural infections is very difficult. Recent advances in the development of humanized mouse models and organs-on-a-chip models, offer novel technological frontiers to study human diseases, vaccine validation and drug discovery. Here, we review current data of these frontier technologies in malaria, highlighting major challenges ahead to study P. vivax cryptic niches, which perpetuate transmission and burden.
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Affiliation(s)
- Iris Aparici Herraiz
- Barcelona Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain
- Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Badalona, Spain
| | - Hugo R. Caires
- i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto, Portugal
| | - Óscar Castillo-Fernández
- Barcelona Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain
- Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Badalona, Spain
- Institute of Nanoscience and Nanotechnology (IN2UB), University of Barcelona, Barcelona, Spain
| | - Núria Sima
- Barcelona Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain
- Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Badalona, Spain
| | - Lourdes Méndez-Mora
- Department of Condensed Matter Physics, University of Barcelona (UB), Barcelona, Spain
| | - Ruth M. Risueño
- Josep Carreras Leukaemia Research Institute (IJC), Barcelona, Spain
| | - Jetsumon Sattabongkot
- Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand
| | - Wanlapa Roobsoong
- Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand
| | - Aurora Hernández-Machado
- Institute of Nanoscience and Nanotechnology (IN2UB), University of Barcelona, Barcelona, Spain
- Department of Condensed Matter Physics, University of Barcelona (UB), Barcelona, Spain
- Centre de Recerca Matemàtica (CRM), Barcelona, Spain
| | - Carmen Fernandez-Becerra
- Barcelona Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain
- Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Badalona, Spain
| | - Cristina C. Barrias
- i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto, Portugal
- INEB – Instituto de Engenharia Biomédica, Universidade do Porto, Porto, Portugal
- ICBAS – Instituto de Ciências Biomédicas de Abel Salazar, Universidade do Porto, Porto, Portugal
| | - Hernando A. del Portillo
- Barcelona Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain
- Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Badalona, Spain
- Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain
- *Correspondence: Hernando A. del Portillo,
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10
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Feldman TP, Egan ES. Uncovering a Cryptic Site of Malaria Pathogenesis: Models to Study Interactions Between Plasmodium and the Bone Marrow. Front Cell Infect Microbiol 2022; 12:917267. [PMID: 35719356 PMCID: PMC9201243 DOI: 10.3389/fcimb.2022.917267] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2022] [Accepted: 05/03/2022] [Indexed: 12/21/2022] Open
Abstract
The bone marrow is a critical site of host-pathogen interactions in malaria infection. The discovery of Plasmodium asexual and transmission stages in the bone marrow has renewed interest in the tissue as a niche for cellular development of both host and parasite. Despite its importance, bone marrow in malaria infection remains largely unexplored due to the challenge of modeling the complex hematopoietic environment in vitro. Advancements in modeling human erythropoiesis ex-vivo from primary human hematopoietic stem and progenitor cells provide a foothold to study the host-parasite interactions occurring in this understudied site of malaria pathogenesis. This review focuses on current in vitro methods to recapitulate and assess bone marrow erythropoiesis and their potential applications in the malaria field. We summarize recent studies that leveraged ex-vivo erythropoiesis to shed light on gametocyte development in nucleated erythroid stem cells and begin to characterize host cell responses to Plasmodium infection in the hematopoietic niche. Such models hold potential to elucidate mechanisms of disordered erythropoiesis, an underlying contributor to malaria anemia, as well as understand the biological determinants of parasite sexual conversion. This review compares the advantages and limitations of the ex-vivo erythropoiesis approach with those of in vivo human and animal studies of the hematopoietic niche in malaria infection. We highlight the need for studies that apply single cell analyses to this complex system and incorporate physical and cellular components of the bone marrow that may influence erythropoiesis and parasite development.
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Affiliation(s)
- Tamar P. Feldman
- Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, United States
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, United States
| | - Elizabeth S. Egan
- Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, United States
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, United States
- *Correspondence: Elizabeth S. Egan,
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11
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Ndegwa DN, Kundu P, Hostetler JB, Marin-Menendez A, Sanderson T, Mwikali K, Verzier LH, Coyle R, Adjalley S, Rayner JC. Using Plasmodium knowlesi as a model for screening Plasmodium vivax blood-stage malaria vaccine targets reveals new candidates. PLoS Pathog 2021; 17:e1008864. [PMID: 34197567 PMCID: PMC8279373 DOI: 10.1371/journal.ppat.1008864] [Citation(s) in RCA: 22] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2020] [Revised: 07/14/2021] [Accepted: 06/01/2021] [Indexed: 11/18/2022] Open
Abstract
Plasmodium vivax is responsible for the majority of malaria cases outside Africa. Unlike P. falciparum, the P. vivax life-cycle includes a dormant liver stage, the hypnozoite, which can cause infection in the absence of mosquito transmission. An effective vaccine against P. vivax blood stages would limit symptoms and pathology from such recurrent infections, and therefore could play a critical role in the control of this species. Vaccine development in P. vivax, however, lags considerably behind P. falciparum, which has many identified targets with several having transitioned to Phase II testing. By contrast only one P. vivax blood-stage vaccine candidate based on the Duffy Binding Protein (PvDBP), has reached Phase Ia, in large part because the lack of a continuous in vitro culture system for P. vivax limits systematic screening of new candidates. We used the close phylogenetic relationship between P. vivax and P. knowlesi, for which an in vitro culture system in human erythrocytes exists, to test the scalability of systematic reverse vaccinology to identify and prioritise P. vivax blood-stage targets. A panel of P. vivax proteins predicted to function in erythrocyte invasion were expressed as full-length recombinant ectodomains in a mammalian expression system. Eight of these antigens were used to generate polyclonal antibodies, which were screened for their ability to recognize orthologous proteins in P. knowlesi. These antibodies were then tested for inhibition of growth and invasion of both wild type P. knowlesi and chimeric P. knowlesi lines modified using CRISPR/Cas9 to exchange P. knowlesi genes with their P. vivax orthologues. Candidates that induced antibodies that inhibited invasion to a similar level as PvDBP were identified, confirming the utility of P. knowlesi as a model for P. vivax vaccine development and prioritizing antigens for further follow up.
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Affiliation(s)
- Duncan N. Ndegwa
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
- Department of Biological Sciences, University of Embu, Embu, Kenya
| | - Prasun Kundu
- Cambridge Institute for Medical Research, University of Cambridge, Cambridge Biomedical Campus, Hills Road, Cambridge, United Kingdom
| | - Jessica B. Hostetler
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | | | - Theo Sanderson
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Kioko Mwikali
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Lisa H. Verzier
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Rachael Coyle
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Sophie Adjalley
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Julian C. Rayner
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
- Cambridge Institute for Medical Research, University of Cambridge, Cambridge Biomedical Campus, Hills Road, Cambridge, United Kingdom
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12
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Thomson-Luque R, Bautista JM. Home Sweet Home: Plasmodium vivax-Infected Reticulocytes-The Younger the Better? Front Cell Infect Microbiol 2021; 11:675156. [PMID: 34055670 PMCID: PMC8162270 DOI: 10.3389/fcimb.2021.675156] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2021] [Accepted: 04/16/2021] [Indexed: 01/17/2023] Open
Abstract
After a century of constant failure to produce an in vitro culture of the most widespread human malaria parasite Plasmodium vivax, recent advances have highlighted the difficulties to provide this parasite with a healthy host cell to invade, develop, and multiply under in vitro conditions. The actual level of understanding of the heterogeneous populations of cells—framed under the name ‘reticulocytes’—and, importantly, their adequate in vitro progression from very immature reticulocytes to normocytes (mature erythrocytes) is far from complete. The volatility of its individual stability may suggest the reticulocyte as a delusory cell, particularly to be used for stable culture purposes. Yet, the recent relevance gained by a specific subset of highly immature reticulocytes has brought some hope. Very immature reticulocytes are characterized by a peculiar membrane harboring a plethora of molecules potentially involved in P. vivax invasion and by an intracellular complexity dynamically changing upon its quick maturation into normocytes. We analyze the potentialities offered by this youngest reticulocyte subsets as an ideal in vitro host cell for P. vivax.
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Affiliation(s)
- Richard Thomson-Luque
- Center of Infectious Diseases, Parasitology, Heidelberg University Hospital, Heidelberg, Germany
| | - José M Bautista
- Department of Biochemistry and Molecular Biology and Research Institute Hospital 12 de Octubre (Imas12), Universidad Complutense de Madrid, Madrid, Spain
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13
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Asali S, Raz A, Turki H, Mafakher L, Razmjou E, Solaymani-Mohammadi S. Restricted genetic heterogeneity of the Plasmodium vivax transmission-blocking vaccine (TBV) candidate Pvs48/45 in a low transmission setting: Implications for the Plasmodium vivax malaria vaccine development. INFECTION GENETICS AND EVOLUTION 2021; 89:104710. [PMID: 33421653 DOI: 10.1016/j.meegid.2021.104710] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/13/2020] [Revised: 12/28/2020] [Accepted: 01/04/2021] [Indexed: 12/14/2022]
Abstract
Plasmodium vivax is the most widespread malaria species parasitizing humans outside Africa, with approximately 100 million cases reported per year. Most human cases of P. vivax are asymptomatic with low parasitemia, making active case detection-based elimination programme challenging and less effective. Despite the widespread distribution of P. vivax, no effective vaccines are currently available. Transmission blocking vaccines have recently emerged as potential vaccine candidates to reduce transmission rates to below the essential levels required for the maintenance of the parasite life cycle. Here, we demonstrated that P. vivax was the predominant species found in a malaria-endemic area, although P. vivax/P. falciparum co-infections were also common. Through genomic sequence analysis and neighbor-joining algorithms, we demonstrated limited genetic heterogeneity in the P. vivax transmission-blocking vaccine candidate Pvs48/45 among clinical isolates of P. vivax. Restricted genetic polymorphism occurred at both nucleotide and amino acid levels. The most frequent mutation was A → G at nucleotide position 77 (46.7%), whereas the least frequent was C → T at nucleotide position 1230 (3.3%). The occurrence of single nucleotide polymorphisms (SNPs) distribution at 6/8 positions (75%) led to changes in amino acid sequences in the Pvs48/45 loci, whereas 2/8 (25%) of SNPs resulted in no amino acid sequence variations. Consistently, the nucleotide diversity in the Pvs48/45 locus among the P. vivax population studied was extremely low (π = 0.000525). Changes in amino acid sequences in the Pvs48/45 protein did not result in substantial conformational modifications in the tertiary structures of these proteins. Unveiling the population genetic structure and genetic heterogeneity of vaccine target antigens are necessary for rational design of transmission-blocking antibody vaccines and to monitor the vaccine efficacy in clinical trials in endemic areas for malaria.
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Affiliation(s)
- Soheila Asali
- Department of Parasitology and Mycology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran
| | - Abbasali Raz
- Malaria and Vector Research Group, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
| | - Habibollah Turki
- Infectious and Tropical Diseases Research Center, Hormozgan Health Institute, Hormozgan University of Medical Sciences, Bandar Abbas, Iran
| | - Ladan Mafakher
- Medicinal Plant Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
| | - Elham Razmjou
- Department of Parasitology and Mycology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran; Microbial Biotechnology Research Center (MBiRC), Iran University of Medical Sciences, Tehran, Iran.
| | - Shahram Solaymani-Mohammadi
- Laboratory of Mucosal Immunology, Department of Biomedical Sciences, School of Medicine and Health Sciences, University of North Dakota, Grand Forks, ND, United States.
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14
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A Way Forward for Culturing Plasmodium vivax. Trends Parasitol 2020; 36:512-519. [PMID: 32360314 DOI: 10.1016/j.pt.2020.04.002] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2020] [Revised: 03/28/2020] [Accepted: 04/02/2020] [Indexed: 01/12/2023]
Abstract
Trager and Jensen established a method for culturing Plasmodium falciparum, a breakthrough for malaria research worldwide. Since then, multiple attempts to establish Plasmodium vivax in continuous culture have failed. Unlike P. falciparum, which can invade all aged erythrocytes, P. vivax is restricted to reticulocytes. Thus, a constant supply of reticulocytes is considered critical for continuous P. vivax growth in vitro. A critical question remains why P. vivax selectively invades reticulocytes? What do reticulocytes offer to P. vivax that is not present in mature erythrocytes? One possibility is protection from oxidative stress by glucose-6-phosphate dehydrogenase (G6PD). Here, we also suggest supplements to the media and procedures that may reduce oxidative stress and, as a result, establish a system for the continuous culture of P. vivax.
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15
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Ssemaganda A, Giddam AK, Zaman M, Skwarczynski M, Toth I, Stanisic DI, Good MF. Induction of Plasmodium-Specific Immune Responses Using Liposome-Based Vaccines. Front Immunol 2019; 10:135. [PMID: 30774635 PMCID: PMC6367261 DOI: 10.3389/fimmu.2019.00135] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2018] [Accepted: 01/16/2019] [Indexed: 12/30/2022] Open
Abstract
In the development of vaccines, the ability to initiate both innate and subsequent adaptive immune responses need to be considered. Live attenuated vaccines achieve this naturally, while inactivated and sub-unit vaccines generally require additional help provided through delivery systems and/or adjuvants. Liposomes present an attractive adjuvant/delivery system for antigens. Here, we review the key aspects of immunity against Plasmodium parasites, liposome design considerations and their current application in the development of a malaria vaccine.
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Affiliation(s)
| | | | - Mehfuz Zaman
- Institute for Glycomics, Griffith University, Southport, QLD, Australia
| | - Mariusz Skwarczynski
- School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD, Australia
| | - Istvan Toth
- School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD, Australia
- School of Pharmacy, The University of Queensland, Brisbane, QLD, Australia
- Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, Australia
| | | | - Michael F. Good
- Institute for Glycomics, Griffith University, Southport, QLD, Australia
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16
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Suphakhonchuwong N, Chaijaroenkul W, Rungsihirunrat K, Na-Bangchang K, Kuesap J. Evaluation of Plasmodium vivax isolates in Thailand using polymorphic markers Plasmodium merozoite surface protein (PvMSP) 1 and PvMSP3. Parasitol Res 2018; 117:3965-3978. [PMID: 30306265 DOI: 10.1007/s00436-018-6106-1] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2018] [Accepted: 10/01/2018] [Indexed: 12/27/2022]
Abstract
Malaria is a significant public health problem in several tropical countries including Thailand. The prevalence of Plasmodium vivax infection has been increasing in the past decades. Plasmodium vivax merozoite surface protein (PvMSP) gene encodes a malaria vaccine candidate antigen. Its polymorphic nature leads to antigenic variation, the barrier for vaccine development, drug resistance, and potential for multiple-clone infections within the malaria patients. The objective of this study was to investigate the genetic diversity of PvMSP1 and PvMSP3 gene in P. vivax populations in Thailand. A total of 100 P. vivax isolates collected from the western (Kanchanaburi and Tak Provinces) and southern (Ranong Provinces) regions along the Thai-Myanmar border were analyzed using polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). Analysis of the F1, F2, and F3 regions of PvMSP1 revealed 5, 2, and 3 allelic variants, respectively. Three major types of PvMSP3-α and two major types of PvMSP3-β were identified based on the PCR product sizes. After digestion with restriction enzymes, 29, 25, 26, and 18 patterns were distinguished by RFLP for PvMSP1 (F2, Alu I), PvMSP1 (F2, Mnl I), PvMSP3-α, and PvMSP3-β, respectively. Combination of each family variant (PvMSP1 and PvMSP3) resulted in high genetic polymorphism of P. vivax population. Additionally, using PvMSP1 polymorphic marker revealed a significant association between multiple-genotype infections and P. vivax parasitemia. The results strongly supported that P. vivax populations in the endemic areas along the Thai-Myanmar border are highly diverse.
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Affiliation(s)
| | - Wanna Chaijaroenkul
- Chulabhorn International College of Medicine, Thammasat University, Pathumthani, Thailand
| | | | - Kesara Na-Bangchang
- Chulabhorn International College of Medicine, Thammasat University, Pathumthani, Thailand
| | - Jiraporn Kuesap
- Faculty of Allied Health Sciences, Thammasat University, Pathumthani, Thailand.
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17
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Bermúdez M, Moreno-Pérez DA, Arévalo-Pinzón G, Curtidor H, Patarroyo MA. Plasmodium vivax in vitro continuous culture: the spoke in the wheel. Malar J 2018; 17:301. [PMID: 30126427 PMCID: PMC6102941 DOI: 10.1186/s12936-018-2456-5] [Citation(s) in RCA: 53] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2018] [Accepted: 08/13/2018] [Indexed: 01/01/2023] Open
Abstract
Understanding the life cycle of Plasmodium vivax is fundamental for developing strategies aimed at controlling and eliminating this parasitic species. Although advances in omic sciences and high-throughput techniques in recent years have enabled the identification and characterization of proteins which might be participating in P. vivax invasion of target cells, exclusive parasite tropism for invading reticulocytes has become the main obstacle in maintaining a continuous culture for this species. Such advance that would help in defining each parasite protein’s function in the complex process of P. vivax invasion, in addition to evaluating new therapeutic agents, is still a dream. Advances related to maintenance, culture medium supplements and the use of different sources of reticulocytes and parasites (strains and isolates) have been made regarding the development of an in vitro culture for P. vivax; however, only some cultures having few replication cycles have been obtained to date, meaning that this parasite’s maintenance goes beyond the technical components involved. Although it is still not yet clear which molecular mechanisms P. vivax prefers for invading young CD71+ reticulocytes [early maturation stages (I–II–III)], changes related to membrane proteins remodelling of such cells could form part of the explanation. The most relevant aspects regarding P. vivax in vitro culture and host cell characteristics have been analysed in this review to explain possible reasons why the species’ continuous in vitro culture is so difficult to standardize. Some alternatives for P. vivax in vitro culture have also been described.
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Affiliation(s)
- Maritza Bermúdez
- Receptor-ligand Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 No. 26-20, Bogotá, Colombia
| | - Darwin Andrés Moreno-Pérez
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 No. 26-20, Bogotá, Colombia.,Livestock Sciences Faculty, Universidad de Ciencias Aplicadas y Ambientales (U.D.C.A), Calle 222 No. 55-37, Bogotá, DC, Colombia
| | - Gabriela Arévalo-Pinzón
- Receptor-ligand Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 No. 26-20, Bogotá, Colombia
| | - Hernando Curtidor
- Receptor-ligand Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 No. 26-20, Bogotá, Colombia.,Basic Sciences Department, School of Medicine and Health Sciences, Universidad del Rosario, Carrera 24 No. 63C-69, Bogotá, DC, Colombia
| | - Manuel Alfonso Patarroyo
- Molecular Biology and Immunology Department, Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 No. 26-20, Bogotá, Colombia. .,Basic Sciences Department, School of Medicine and Health Sciences, Universidad del Rosario, Carrera 24 No. 63C-69, Bogotá, DC, Colombia.
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18
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Bourgard C, Albrecht L, Kayano ACAV, Sunnerhagen P, Costa FTM. Plasmodium vivax Biology: Insights Provided by Genomics, Transcriptomics and Proteomics. Front Cell Infect Microbiol 2018; 8:34. [PMID: 29473024 PMCID: PMC5809496 DOI: 10.3389/fcimb.2018.00034] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2017] [Accepted: 01/25/2018] [Indexed: 12/17/2022] Open
Abstract
During the last decade, the vast omics field has revolutionized biological research, especially the genomics, transcriptomics and proteomics branches, as technological tools become available to the field researcher and allow difficult question-driven studies to be addressed. Parasitology has greatly benefited from next generation sequencing (NGS) projects, which have resulted in a broadened comprehension of basic parasite molecular biology, ecology and epidemiology. Malariology is one example where application of this technology has greatly contributed to a better understanding of Plasmodium spp. biology and host-parasite interactions. Among the several parasite species that cause human malaria, the neglected Plasmodium vivax presents great research challenges, as in vitro culturing is not yet feasible and functional assays are heavily limited. Therefore, there are gaps in our P. vivax biology knowledge that affect decisions for control policies aiming to eradicate vivax malaria in the near future. In this review, we provide a snapshot of key discoveries already achieved in P. vivax sequencing projects, focusing on developments, hurdles, and limitations currently faced by the research community, as well as perspectives on future vivax malaria research.
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Affiliation(s)
- Catarina Bourgard
- Laboratory of Tropical Diseases, Department of Genetics, Evolution, Microbiology and Immunology, University of Campinas - UNICAMP, Campinas, Brazil
| | - Letusa Albrecht
- Laboratory of Tropical Diseases, Department of Genetics, Evolution, Microbiology and Immunology, University of Campinas - UNICAMP, Campinas, Brazil.,Laboratory of Regulation of Gene Expression, Instituto Carlos Chagas, Curitiba, Brazil
| | - Ana C A V Kayano
- Laboratory of Tropical Diseases, Department of Genetics, Evolution, Microbiology and Immunology, University of Campinas - UNICAMP, Campinas, Brazil
| | - Per Sunnerhagen
- Department of Chemistry and Molecular Biology, University of Gothenburg, Gothenburg, Sweden
| | - Fabio T M Costa
- Laboratory of Tropical Diseases, Department of Genetics, Evolution, Microbiology and Immunology, University of Campinas - UNICAMP, Campinas, Brazil
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19
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Plasmodium falciparum malaria skews globin gene expression balance in in-vitro haematopoietic stem cell culture system: Its implications in malaria associated anemia. Exp Parasitol 2018; 185:29-38. [DOI: 10.1016/j.exppara.2018.01.003] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2017] [Revised: 11/01/2017] [Accepted: 01/02/2018] [Indexed: 01/02/2023]
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20
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Abstract
Systemic inflammation mediated by Plasmodium parasites is central to malaria disease and its complications. Plasmodium parasites reside in erythrocytes and can theoretically reach all host tissues via the circulation. However, actual interactions between parasitized erythrocytes and host tissues, along with the consequent damage and pathological changes, are limited locally to specific tissue sites. Such tissue specificity of the parasite can alter the outcome of malaria disease, determining whether acute or chronic complications occur. Here, we give an overview of the recent progress that has been made in understanding tissue-specific immunopathology during Plasmodium infection. As knowledge on tissue-specific host-parasite interactions accumulates, better treatment modalities and targets may emerge for intervention in malaria disease.
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21
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Armistead JS, Adams JH. Advancing Research Models and Technologies to Overcome Biological Barriers to Plasmodium vivax Control. Trends Parasitol 2017; 34:114-126. [PMID: 29153587 DOI: 10.1016/j.pt.2017.10.009] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/16/2017] [Revised: 10/25/2017] [Accepted: 10/25/2017] [Indexed: 02/06/2023]
Abstract
Malaria prevalence has declined in the past 10 years, especially outside of sub-Saharan Africa. However, the proportion of cases due to Plasmodium vivax is increasing, accounting for up to 90-100% of the malaria burden in endemic regions. Nonetheless, investments in malaria research and control still prioritize Plasmodium falciparum while largely neglecting P. vivax. Specific biological features of P. vivax, particularly invasion of reticulocytes, occurrence of dormant liver forms of the parasite, and the potential for transmission of sexual-stage parasites prior to onset of clinical illness, promote its persistence and hinder development of research tools and interventions. This review discusses recent advances in P. vivax research, current knowledge of its unique biology, and proposes priorities for P. vivax research and control efforts.
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Affiliation(s)
- Jennifer S Armistead
- Center for Global Health and Infectious Diseases Research, Department of Global Health, College of Public Health, University of South Florida, Tampa, FL 33612, USA
| | - John H Adams
- Center for Global Health and Infectious Diseases Research, Department of Global Health, College of Public Health, University of South Florida, Tampa, FL 33612, USA.
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22
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Mehlotra RK, Blankenship D, Howes RE, Rakotomanga TA, Ramiranirina B, Ramboarina S, Franchard T, Linger MH, Zikursh-Blood M, Ratsimbasoa AC, Zimmerman PA, Grimberg BT. Long-term in vitro culture of Plasmodium vivax isolates from Madagascar maintained in Saimiri boliviensis blood. Malar J 2017; 16:442. [PMID: 29100506 PMCID: PMC5670718 DOI: 10.1186/s12936-017-2090-7] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2017] [Accepted: 10/27/2017] [Indexed: 02/06/2023] Open
Abstract
Background Plasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportionally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum. Despite the prominence of P. vivax as a major human pathogen, vivax malaria qualifies as a neglected and under-studied tropical disease. Significant challenges bringing P. vivax into the laboratory, particularly the capacity for long-term propagation of well-characterized strains, have limited the study of this parasite’s red blood cell (RBC) invasion mechanism, blood-stage development, gene expression, and genetic manipulation. Methods and results Patient isolates of P. vivax have been collected and cryopreserved in the rural community of Ampasimpotsy, located in the Tsiroanomandidy Health District of Madagascar. Periodic, monthly overland transport of these cryopreserved isolates to the country’s National Malaria Control Programme laboratory in Antananarivo preceded onward sample transfer to laboratories at Case Western Reserve University, USA. There, the P. vivax isolates have been cultured through propagation in the RBCs of Saimiri boliviensis. For the four patient isolates studied to-date, the median time interval between sample collection and in vitro culture has been 454 days (range 166–961 days). The median time in culture, continually documented by light microscopy, has been 159 days; isolate AMP2014.01 was continuously propagated for 233 days. Further studies show that the P. vivax parasites propagated in Saimiri RBCs retain their ability to invade human RBCs, and can be cryopreserved, thawed and successfully returned to productive in vitro culture. Conclusions/significance Long-term culture of P. vivax is possible in the RBCs of Saimiri boliviensis. These studies provide an alternative to propagation of P. vivax in live animals that are becoming more restricted. In vitro culture of P. vivax in Saimiri RBCs provides an opening to stabilize patient isolates, which would serve as precious resources to apply new strategies for investigating the molecular and cellular biology of this important malaria parasite. Electronic supplementary material The online version of this article (10.1186/s12936-017-2090-7) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Rajeev K Mehlotra
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA
| | - D'Arbra Blankenship
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA
| | - Rosalind E Howes
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA.,Oxford Big Data Institute, Nuffield Department of Medicine, University of Oxford, Oxford, UK
| | - Tovonahary A Rakotomanga
- National Malaria Control Programme, Ministry of Health, Antananarivo, Madagascar.,Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
| | - Brune Ramiranirina
- Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
| | - Stephanie Ramboarina
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA.,Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
| | - Thierry Franchard
- National Malaria Control Programme, Ministry of Health, Antananarivo, Madagascar.,Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
| | - Marlin H Linger
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA
| | - Melinda Zikursh-Blood
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA
| | - Arsène C Ratsimbasoa
- National Malaria Control Programme, Ministry of Health, Antananarivo, Madagascar.,Faculty of Sciences, University of Antananarivo, Antananarivo, Madagascar
| | - Peter A Zimmerman
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA.
| | - Brian T Grimberg
- Center for Global Health and Diseases, Case Western Reserve University School of Medicine, Cleveland, OH, 44106-4983, USA.
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23
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Sinha S, Sarma P, Sehgal R, Medhi B. Development in Assay Methods for in Vitro Antimalarial Drug Efficacy Testing: A Systematic Review. Front Pharmacol 2017; 8:754. [PMID: 29123481 PMCID: PMC5662882 DOI: 10.3389/fphar.2017.00754] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/03/2017] [Accepted: 10/04/2017] [Indexed: 11/13/2022] Open
Abstract
The emergence and spread of drug resistance are the major challenges in malaria eradication mission. Besides various strategies laid down by World Health Organization, such as vector management, source reduction, early case detection, prompt treatment, and development of new diagnostics and vaccines, nevertheless the need for new and efficacious drugs against malaria has become a critical priority on the global malaria research agenda. At several screening stages, millions of compounds are screened (1,000–2,000,000 compounds per screening campaign), before pre-clinical trials to select optimum lead. Carrying out in vitro screening of antimalarials is very difficult as different assay methods are subject to numerous sources of variability across different laboratories around the globe. Despite this, in vitro screening is an essential part of antimalarial drug development as it enables to resource various confounding factors such as host immune response and drug–drug interaction. Therefore, in this article, we try to illustrate the basic necessity behind in vitro study and how new methods are developed and subsequently adopted for high-throughput antimalarial drug screening and its application in achieving the next level of in vitro screening based on the current approaches (such as stem cells).
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Affiliation(s)
- Shweta Sinha
- Department of Medical Parasitology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Phulen Sarma
- Department of Pharmacology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Rakesh Sehgal
- Department of Medical Parasitology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
| | - Bikash Medhi
- Department of Pharmacology, Postgraduate Institute of Medical Education and Research, Chandigarh, India
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24
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Abstract
Plasmodium vivax is the second most prevalent cause of malaria worldwide and the leading cause of malaria outside of Africa. Although infections are seldom fatal clinical disease can be debilitating and imposes significant health and economic impacts on affected populations. Estimates of transmission and prevalence intensity can be problematic because many episodes of vivax originate from hypnozoite stages in the liver that have remained dormant from previous infections by an unknown mechanism. Lack of treatment options to clear hypnozoites and the ability to infect mosquitoes before disease symptoms present represent major challenges for control and eradication of vivax malaria. Compounding these challenges is the unique biology of P. vivax and limited progress in development of experimental research tools, thereby hindering development of new drugs and vaccines. Renewed emphasis on vivax malaria research is beginning to make progress in overcoming some of these challenges.
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Affiliation(s)
- John H Adams
- Center for Global Health and Infectious Diseases, Department of Global Health, University of South Florida, Tampa, Florida 33612
| | - Ivo Mueller
- Population Health & Immunity Division, Walter & Eliza Hall Institute, Parkville, Victoria 3052, Australia
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25
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Arévalo-Pinzón G, Bermúdez M, Hernández D, Curtidor H, Patarroyo MA. Plasmodium vivax ligand-receptor interaction: PvAMA-1 domain I contains the minimal regions for specific interaction with CD71+ reticulocytes. Sci Rep 2017; 7:9616. [PMID: 28855657 PMCID: PMC5577344 DOI: 10.1038/s41598-017-10025-6] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2017] [Accepted: 08/02/2017] [Indexed: 12/18/2022] Open
Abstract
The malarial parasite’s invasion is complex, active and coordinated, involving many low and high affinity interactions with receptors on target cell membrane. Proteomics analysis has described around 40 proteins in P. vivax which could be involved in reticulocyte invasion; few have been studied with the aim of elucidating how many of them establish specific interactions with their respective host cells. Given the importance of knowing which of the parasite’s protein regions are functionally important for invasion, minimum regions mediating specific interaction between Plasmodium vivax apical membrane antigen 1 (PvAMA-1) and its host cell were here elucidated. The region covering PvAMA-1 domains I and II (PvAMA-DI-II) specifically bound to the CD71+ red blood cell subpopulation. A 20 residue-long region (81EVENAKYRIPAGRCPVFGKG100) located in domain I was capable of inhibiting PvAMA-DI-II recombinant protein binding to young reticulocytes (CD71+CD45−) and rosette formation. This conserved peptide specifically interacted with high affinity with reticulocytes (CD71+) through a neuraminidase- and chymotrypsin-treatment sensitive receptor. Such results showed that, despite AMA-1 having universal functions during late Plasmodium invasion stages, PvAMA-1 had reticulocyte-preferring binding regions, suggesting that P. vivax target cell selection is not just restricted to initial interactions but maintained throughout the erythrocyte invasion cycle, having important implications for designing a specific anti-P. vivax vaccine.
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Affiliation(s)
- Gabriela Arévalo-Pinzón
- Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 # 26-20, Bogotá, Colombia.,PhD Program in Biomedical and Biological Sciences, Universidad del Rosario, Carrera 24 #, 63C-69, Bogotá, Colombia
| | - Maritza Bermúdez
- Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 # 26-20, Bogotá, Colombia.,MSc Program in Biological Sciences, Pontificia Universidad Javeriana, Carrera 7 # 40-62, Bogotá, Colombia
| | - Diana Hernández
- Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 # 26-20, Bogotá, Colombia
| | - Hernando Curtidor
- Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 # 26-20, Bogotá, Colombia.,School of Medicine and Health Sciences, Universidad del Rosario, Carrera 24 #, 63C-69, Bogotá, Colombia
| | - Manuel Alfonso Patarroyo
- Fundación Instituto de Inmunología de Colombia (FIDIC), Carrera 50 # 26-20, Bogotá, Colombia. .,School of Medicine and Health Sciences, Universidad del Rosario, Carrera 24 #, 63C-69, Bogotá, Colombia.
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26
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Pornthanakasem W, Riangrungroj P, Chitnumsub P, Ittarat W, Kongkasuriyachai D, Uthaipibull C, Yuthavong Y, Leartsakulpanich U. Role of Plasmodium vivax Dihydropteroate Synthase Polymorphisms in Sulfa Drug Resistance. Antimicrob Agents Chemother 2016; 60:4453-63. [PMID: 27161627 PMCID: PMC4958149 DOI: 10.1128/aac.01835-15] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2015] [Accepted: 04/19/2016] [Indexed: 11/20/2022] Open
Abstract
Dihydropteroate synthase (DHPS) is a known sulfa drug target in malaria treatment, existing as a bifunctional enzyme together with hydroxymethyldihydropterin pyrophosphokinase (HPPK). Polymorphisms in key residues of Plasmodium falciparum DHPS (PfDHPS) have been characterized and linked to sulfa drug resistance in malaria. Genetic sequencing of P. vivax dhps (Pvdhps) from clinical isolates has shown several polymorphisms at the positions equivalent to those in the Pfdhps genes conferring sulfa drug resistance, suggesting a mechanism for sulfa drug resistance in P. vivax similar to that seen in P. falciparum To characterize the role of polymorphisms in the PvDHPS in sulfa drug resistance, various mutants of recombinant PvHPPK-DHPS enzymes were expressed and characterized. Moreover, due to the lack of a continuous in vitro culture system for P. vivax parasites, a surrogate P. berghei model expressing Pvhppk-dhps genes was established to demonstrate the relationship between sequence polymorphisms and sulfa drug susceptibility and to test the activities of PvDHPS inhibitors on the transgenic parasites. Both enzyme activity and transgenic parasite growth were sensitive to sulfadoxine to different degrees, depending on the number of mutations that accumulated in DHPS. Ki values and 50% effective doses were higher for mutant PvDHPS enzymes than the wild-type enzymes. Altogether, the study provides the first evidence of sulfa drug resistance at the molecular level in P. vivax Furthermore, the enzyme inhibition assay and the in vivo screening system can be useful tools for screening new compounds for their activities against PvDHPS.
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Affiliation(s)
| | | | - Penchit Chitnumsub
- National Center for Genetic Engineering and Biotechnology, Pathum Thani, Thailand
| | - Wanwipa Ittarat
- National Center for Genetic Engineering and Biotechnology, Pathum Thani, Thailand
| | | | - Chairat Uthaipibull
- National Center for Genetic Engineering and Biotechnology, Pathum Thani, Thailand
| | - Yongyuth Yuthavong
- National Center for Genetic Engineering and Biotechnology, Pathum Thani, Thailand
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Shaw-Saliba K, Thomson-Luque R, Obaldía N, Nuñez M, Dutary S, Lim C, Barnes S, Kocken CHM, Duraisingh MT, Adams JH, Pasini EM. Insights into an Optimization of Plasmodium vivax Sal-1 In Vitro Culture: The Aotus Primate Model. PLoS Negl Trop Dis 2016; 10:e0004870. [PMID: 27463518 PMCID: PMC4963040 DOI: 10.1371/journal.pntd.0004870] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2016] [Accepted: 06/30/2016] [Indexed: 01/03/2023] Open
Abstract
Malaria is one of the most significant tropical diseases, and of the Plasmodium species that cause human malaria, P. vivax is the most geographically widespread. However, P. vivax remains a relatively neglected human parasite since research is typically limited to laboratories with direct access to parasite isolates from endemic field settings or from non-human primate models. This restricted research capacity is in large part due to the lack of a continuous P. vivax in vitro culture system, which has hampered the ability for experimental research needed to gain biological knowledge and develop new therapies. Consequently, efforts to establish a long-term P. vivax culture system are confounded by our poor knowledge of the preferred host cell and essential nutrients needed for in vitro propagation. Reliance on very heterogeneous P. vivax field isolates makes it difficult to benchmark parasite characteristics and further complicates development of a robust and reliable culture method. In an effort to eliminate parasite variability as a complication, we used a well-defined Aotus-adapted P. vivax Sal-1 strain to empirically evaluate different short-term in vitro culture conditions and compare them with previous reported attempts at P. vivax in vitro culture Most importantly, we suggest that reticulocyte enrichment methods affect invasion efficiency and we identify stabilized forms of nutrients that appear beneficial for parasite growth, indicating that P. vivax may be extremely sensitive to waste products. Leuko-depletion methods did not significantly affect parasite development. Formatting changes such as shaking and static cultures did not seem to have a major impact while; in contrast, the starting haematocrit affected both parasite invasion and growth. These results support the continued use of Aotus-adapted Sal-1 for development of P. vivax laboratory methods; however, further experiments are needed to optimize culture conditions to support long-term parasite development.
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Affiliation(s)
- Kathryn Shaw-Saliba
- Department of Immunology and Infectious Diseases, Harvard T. H. Chan School of Public Health, Boston, Massachusetts, United States of America
| | - Richard Thomson-Luque
- Center for Global Health & Infectious Diseases Research, Department of Global Health, College of Public Health, University of South Florida, Tampa, Florida, United States of America
| | - Nicanor Obaldía
- Center for the Evaluation of Antimalarial Drugs and Vaccines, Tropical Medicine Research / Instituto Conmemorativo Gorgas de Estudios de la Salud, Panamá, Panamá
| | - Marlon Nuñez
- Center for the Evaluation of Antimalarial Drugs and Vaccines, Tropical Medicine Research / Instituto Conmemorativo Gorgas de Estudios de la Salud, Panamá, Panamá
| | - Sahir Dutary
- Center for the Evaluation of Antimalarial Drugs and Vaccines, Tropical Medicine Research / Instituto Conmemorativo Gorgas de Estudios de la Salud, Panamá, Panamá
| | - Caeul Lim
- Department of Immunology and Infectious Diseases, Harvard T. H. Chan School of Public Health, Boston, Massachusetts, United States of America
| | - Samantha Barnes
- Center for Global Health & Infectious Diseases Research, Department of Global Health, College of Public Health, University of South Florida, Tampa, Florida, United States of America
| | | | - Manoj T. Duraisingh
- Department of Immunology and Infectious Diseases, Harvard T. H. Chan School of Public Health, Boston, Massachusetts, United States of America
- * E-mail: (MTD); (JHA); (EMP)
| | - John H. Adams
- Center for Global Health & Infectious Diseases Research, Department of Global Health, College of Public Health, University of South Florida, Tampa, Florida, United States of America
- * E-mail: (MTD); (JHA); (EMP)
| | - Erica M. Pasini
- Biomedical Primate Research Centre, Rijswijk, The Netherlands
- * E-mail: (MTD); (JHA); (EMP)
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Noulin F. Malaria modeling: In vitro stem cells vs in vivo models. World J Stem Cells 2016; 8:88-100. [PMID: 27022439 PMCID: PMC4807312 DOI: 10.4252/wjsc.v8.i3.88] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/24/2015] [Revised: 12/07/2015] [Accepted: 01/29/2016] [Indexed: 02/06/2023] Open
Abstract
The recent development of stem cell research and the possibility of generating cells that can be stably and permanently modified in their genome open a broad horizon in the world of in vitro modeling. The malaria field is gaining new opportunities from this important breakthrough and novel tools were adapted and opened new frontiers for malaria research. In addition to the new in vitro systems, in recent years there were also significant advances in the development of new animal models that allows studying the entire cell cycle of human malaria. In this paper, we review the different protocols available to study human Plasmodium species either by using stem cell or alternative animal models.
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Erythropoiesis in Malaria Infections and Factors Modifying the Erythropoietic Response. Anemia 2016; 2016:9310905. [PMID: 27034825 PMCID: PMC4789361 DOI: 10.1155/2016/9310905] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2015] [Accepted: 02/02/2016] [Indexed: 12/13/2022] Open
Abstract
Anemia is the primary clinical manifestation of malarial infections and is responsible for the substantial rate of morbidity. The pathophysiology discussed till now catalogued several causes for malarial anemia among which ineffective erythropoiesis being remarkable one occurs silently in the bone marrow. A systematic literature search was performed and summarized information on erythropoietic response upon malaria infection and the factors responsible for the same. This review summarizes the clinical and experimental studies on patients, mouse models, and in vitro cell cultures reporting erythropoietic changes upon malaria infection as well as factors accountable for the same. Inadequate erythropoietic response during malaria infection may be the collective effect of various mediators generated by host immune response as well as parasite metabolites. The interplay between various modulators causing the pathophysiology needs to be explored further. Globin gene expression profiling upon malaria infection should also be looked into as abnormal production of globin chains could be a possible contributor to ineffective erythropoiesis.
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Improvement of culture conditions for long-term in vitro culture of Plasmodium vivax. Malar J 2015; 14:297. [PMID: 26243280 PMCID: PMC4524445 DOI: 10.1186/s12936-015-0815-z] [Citation(s) in RCA: 38] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2015] [Accepted: 07/20/2015] [Indexed: 12/25/2022] Open
Abstract
BACKGROUND The study of the biology, transmission and pathogenesis of Plasmodium vivax is hindered due to the lack of a robustly propagating, continuous culture of this parasite. The current culture system for P. vivax parasites still suffered from consistency and difficulties in long-term maintenance of parasites in culture and for providing sufficient biological materials for studying parasite biology. Therefore, further improvement of culture conditions for P. vivax is needed. METHODS Clinical samples were collected from patients diagnosed with P. vivax in western Thailand. Leukocyte-depleted P. vivax infected blood samples were cultured in a modified McCoy's 5A medium at 5% haematocrit under hypoxic condition (5% O2, 5% CO2, and 90% N2). Reticulocytes purified from adult peripheral blood were added daily to maintain 4% reticulocytes. Parasites were detected by microscopic examination of Giemsa-stained smears and molecular methods. RESULTS The effects of culture variables were first analysed in order to improve the culture conditions for P. vivax. Through analysis of the sources of host reticulocytes and nutrients of culture medium, the culture conditions better supporting in vitro growth and maturation of the parasites were identified. Using this system, three of 30 isolates could be maintained in vitro for over 26 months albeit parasite density is low. CONCLUSIONS Based on the analysis of different culture variables, an improved and feasible protocol for continuous culture of P. vivax was developed.
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Campo B, Vandal O, Wesche DL, Burrows JN. Killing the hypnozoite--drug discovery approaches to prevent relapse in Plasmodium vivax. Pathog Glob Health 2015; 109:107-22. [PMID: 25891812 PMCID: PMC4455353 DOI: 10.1179/2047773215y.0000000013] [Citation(s) in RCA: 77] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
The eradication of malaria will only be possible if effective, well-tolerated medicines kill hypnozoites in vivax and ovale malaria, and thus prevent relapses in patients. Despite progress in the 8-aminoquinoline series, with tafenoquine in Phase III showing clear benefits over primaquine, the drug discovery challenge to identify hypnozoiticidal or hypnozoite-activating compounds has been hampered by the dearth of biological tools and assays, which in turn has been limited by the immense scientific and logistical challenges associated with accessing relevant human tissue and sporozoites. This review summarises the existing drug discovery series and approaches concerning the goal to block relapse.
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Affiliation(s)
- Brice Campo
- Medicines for Malaria Venture, Geneva, Switzerland
| | - Omar Vandal
- The Bill and Melinda Gates Foundation, Seattle, WA, USA
| | - David L. Wesche
- The Bill and Melinda Gates Foundation, Seattle, WA, USA
- Great Lakes Drug Development/Certara, Princeton, NJ, USA
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A Pharmacokinetic Study of Antimalarial 3,5-Diaryl-2-aminopyridine Derivatives. Malar Res Treat 2015; 2015:405962. [PMID: 25893131 PMCID: PMC4393935 DOI: 10.1155/2015/405962] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2015] [Revised: 03/16/2015] [Accepted: 03/18/2015] [Indexed: 11/17/2022] Open
Abstract
Malaria caused by Plasmodium falciparum is responsible for approximately 80% of the incidence and 90% of deaths which occur in the World Health Organization (WHO) African region, with children and pregnant women having the highest incidence. P. falciparum has developed resistance, and therefore new effective candidate antimalarial drugs need to be developed. Previous studies identified 3,5-diaryl-2-aminopyridines as potential antimalarial drug candidates; therefore, derivatives of these compounds were synthesized in order to improve their desired properties and pharmacokinetic (PK) properties of the derivatives were investigated in a mouse model which was dosed orally and intravenously. Collected blood samples were analyzed using liquid chromatography coupled to mass spectrometer (LC-MS/MS). The mean peak plasma level of 1.9 μM was obtained at 1 hour for compound 1 and 3.3 μM at 0.5 hours for compound 2. A decline in concentration was observed with a half-life of 2.53 and 0.87 hours for compound 1 in mice dosed orally and intravenously, respectively. For compound 2 a half-life of 2.96 and 0.68 hours was recorded. The bioavailability was 69% and 59.7% for compound 1 and compound 2, respectively.
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Panichakul T, Ponnikorn S, Roytrakul S, Paemanee A, Kittisenachai S, Hongeng S, Udomsangpetch R. Plasmodium vivax inhibits erythroid cell growth through altered phosphorylation of the cytoskeletal protein ezrin. Malar J 2015; 14:138. [PMID: 25889165 PMCID: PMC4392472 DOI: 10.1186/s12936-015-0648-9] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2014] [Accepted: 03/15/2015] [Indexed: 12/27/2022] Open
Abstract
BACKGROUND The underlying causes of severe malarial anaemia are multifactorial. In previously reports, Plasmodium vivax was found to be able to directly inhibited erythroid cell proliferation and differentiation. The molecular mechanisms underlying the suppression of erythropoiesis by P. vivax are remarkably complex and remain unclear. In this study, a phosphoproteomic approach was performed to dissect the molecular mechanism of phosphoprotein regulation, which is involved in the inhibitory effect of parasites on erythroid cell development. METHODS This study describes the first comparative phosphoproteome analysis of growing erythroid cells (gECs), derived from human haematopoietic stem cells, exposed to lysates of infected erythrocytes (IE)/uninfected erythrocytes (UE) for 24, 48 and 72 h. This study utilized IMAC phosphoprotein isolation directly coupled with LC MS/MS analysis. RESULTS Lysed IE significantly inhibited gEC growth at 48 and 72 h and cell division resulting in the accumulation of cells in G0 phase. The relative levels of forty four phosphoproteins were determined from gECs exposed to IE/UE for 24-72 h and compared with the media control using the label-free quantitation technique. Interestingly, the levels of three phosphoproteins: ezrin, alpha actinin-1, and Rho kinase were significantly (p < 0.05) altered. These proteins display interactions and are involved in the regulation of the cellular cytoskeleton. Particularly affected was ezrin (phosphorylated at Thr567), which is normally localized to gEC cell extension peripheral processes. Following exposure to IE, for 48-72 h, the ezrin signal intensity was weak or absent. This result suggests that phospho-ezrin is important for actin cytoskeleton regulation during erythroid cell growth and division. CONCLUSIONS These findings suggest that parasite proteins are able to inhibit erythroid cell growth by down-regulation of ezrin phosphorylation, leading to ineffective erythropoiesis ultimately resulting in severe malarial anaemia. A better understanding of the mechanisms of ineffective erythropoiesis may be beneficial in the development of therapeutic strategies to prevent severe malarial anaemia.
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Affiliation(s)
- Tasanee Panichakul
- Faculty of Science and Technology, Suan Dusit Rajabhat University, 204/3 Sirindhorn Rd. Bangplat, 10700, Bangkok, Thailand.
| | - Saranyoo Ponnikorn
- Chulabhorn International College of Medicine, Thammasat University, 2nd Floor, Piyachart Building, Thammasat University, Rungsit campus, 12120, Patumthani, Thailand.
| | - Sittiruk Roytrakul
- Proteomics Research Laboratory, National Center for Genetic and Engineering and Biotechnology, National Science and Technology Development Agency, 113 Thailand Science Park, Phahonyothin Rd., Klong1, 12120, Klong Luang, Pathumthani, Thailand.
| | - Atchara Paemanee
- Proteomics Research Laboratory, National Center for Genetic and Engineering and Biotechnology, National Science and Technology Development Agency, 113 Thailand Science Park, Phahonyothin Rd., Klong1, 12120, Klong Luang, Pathumthani, Thailand.
| | - Suthathip Kittisenachai
- Proteomics Research Laboratory, National Center for Genetic and Engineering and Biotechnology, National Science and Technology Development Agency, 113 Thailand Science Park, Phahonyothin Rd., Klong1, 12120, Klong Luang, Pathumthani, Thailand.
| | - Suradej Hongeng
- Department of Pediatrics, Faculty of Medicine Ramathibodi Hospital, Mahidol University, 272 Rama VI Rd., Ratchathewi District, 10400, Bangkok, Thailand.
| | - Rachanee Udomsangpetch
- Department of Pathobiology, Faculty of Science, Mahidol University, 272 Rama VI Rd., Ratchathewi District, 10400, Bangkok, Thailand.
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Barber BE, William T, Grigg MJ, Parameswaran U, Piera KA, Price RN, Yeo TW, Anstey NM. Parasite biomass-related inflammation, endothelial activation, microvascular dysfunction and disease severity in vivax malaria. PLoS Pathog 2015; 11:e1004558. [PMID: 25569250 PMCID: PMC4287532 DOI: 10.1371/journal.ppat.1004558] [Citation(s) in RCA: 115] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2014] [Accepted: 11/06/2014] [Indexed: 12/05/2022] Open
Abstract
Plasmodium vivax can cause severe malaria, however its pathogenesis is poorly understood. In contrast to P. falciparum, circulating vivax parasitemia is low, with minimal apparent sequestration in endothelium-lined microvasculature, and pathogenesis thought unrelated to parasite biomass. However, the relationships between vivax disease-severity and total parasite biomass, endothelial autocrine activation and microvascular dysfunction are unknown. We measured circulating parasitemia and markers of total parasite biomass (plasma parasite lactate dehydrogenase [pLDH] and PvLDH) in adults with severe (n = 9) and non-severe (n = 53) vivax malaria, and examined relationships with disease-severity, endothelial activation, and microvascular function. Healthy controls and adults with non-severe and severe falciparum malaria were enrolled for comparison. Median peripheral parasitemia, PvLDH and pLDH were 2.4-fold, 3.7-fold and 6.9-fold higher in severe compared to non-severe vivax malaria (p = 0.02, p = 0.02 and p = 0.015, respectively), suggesting that, as in falciparum malaria, peripheral P. vivax parasitemia underestimates total parasite biomass, particularly in severe disease. P. vivax schizonts were under-represented in peripheral blood. Severe vivax malaria was associated with increased angiopoietin-2 and impaired microvascular reactivity. Peripheral vivax parasitemia correlated with endothelial activation (angiopoietin-2, von-Willebrand-Factor [VWF], E-selectin), whereas markers of total vivax biomass correlated only with systemic inflammation (IL-6, IL-10). Activity of the VWF-cleaving-protease, ADAMTS13, was deficient in proportion to endothelial activation, IL-6, thrombocytopenia and vivax disease-severity, and associated with impaired microvascular reactivity in severe disease. Impaired microvascular reactivity correlated with lactate in severe vivax malaria. Findings suggest that tissue accumulation of P. vivax may occur, with the hidden biomass greatest in severe disease and capable of mediating systemic inflammatory pathology. The lack of association between total parasite biomass and endothelial activation is consistent with accumulation in parts of the circulation devoid of endothelium. Endothelial activation, associated with circulating parasites, and systemic inflammation may contribute to pathology in vivax malaria, with microvascular dysfunction likely contributing to impaired tissue perfusion.
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Affiliation(s)
- Bridget E. Barber
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
- Department of Infectious Diseases, Queen Elizabeth Hospital, Sabah, Malaysia
- Infectious Diseases Society Sabah-Menzies School of Health Research Clinical Research Unit, Kota Kinabalu, Sabah, Malaysia
| | - Timothy William
- Department of Infectious Diseases, Queen Elizabeth Hospital, Sabah, Malaysia
- Infectious Diseases Society Sabah-Menzies School of Health Research Clinical Research Unit, Kota Kinabalu, Sabah, Malaysia
| | - Matthew J. Grigg
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
- Department of Infectious Diseases, Queen Elizabeth Hospital, Sabah, Malaysia
- Infectious Diseases Society Sabah-Menzies School of Health Research Clinical Research Unit, Kota Kinabalu, Sabah, Malaysia
| | - Uma Parameswaran
- Department of Infectious Diseases, Queen Elizabeth Hospital, Sabah, Malaysia
- Infectious Diseases Society Sabah-Menzies School of Health Research Clinical Research Unit, Kota Kinabalu, Sabah, Malaysia
| | - Kim A. Piera
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
| | - Ric N. Price
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
- Centre for Tropical Medicine, Nuffield Department of Clinical Medicine, University of Oxford, Oxford, United Kingdom
| | - Tsin W. Yeo
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
- Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore
- Insitute of Infectious Disease and Epidemiology, Tan Tock Seng Hospital, Singapore
| | - Nicholas M. Anstey
- Global and Tropical Health Division, Menzies School of Health Research and Charles Darwin University, Darwin, Northern Territory, Australia
- Department of Infectious Diseases, Royal Darwin Hospital, Darwin, Northern Territory, Australia
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Cornejo OE, Fisher D, Escalante AA. Genome-wide patterns of genetic polymorphism and signatures of selection in Plasmodium vivax. Genome Biol Evol 2014; 7:106-19. [PMID: 25523904 PMCID: PMC4316620 DOI: 10.1093/gbe/evu267] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
Plasmodium vivax is the most prevalent human malaria parasite outside of Africa. Yet, studies aimed to identify genes with signatures consistent with natural selection are rare. Here, we present a comparative analysis of the pattern of genetic variation of five sequenced isolates of P. vivax and its divergence with two closely related species, Plasmodium cynomolgi and Plasmodium knowlesi, using a set of orthologous genes. In contrast to Plasmodium falciparum, the parasite that causes the most lethal form of human malaria, we did not find significant constraints on the evolution of synonymous sites genome wide in P. vivax. The comparative analysis of polymorphism and divergence across loci allowed us to identify 87 genes with patterns consistent with positive selection, including genes involved in the “exportome” of P. vivax, which are potentially involved in evasion of the host immune system. Nevertheless, we have found a pattern of polymorphism genome wide that is consistent with a significant amount of constraint on the replacement changes and prevalent negative selection. Our analyses also show that silent polymorphism tends to be larger toward the ends of the chromosomes, where many genes involved in antigenicity are located, suggesting that natural selection acts not only by shaping the patterns of variation within the genes but it also affects genome organization.
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Affiliation(s)
- Omar E Cornejo
- School of Biological Sciences, Washington State University
| | - David Fisher
- Center for Evolutionary Medicine and Informatics, the Biodesign Institute, Arizona State University
| | - Ananias A Escalante
- Center for Evolutionary Medicine and Informatics, the Biodesign Institute, Arizona State University School of Life Sciences, Arizona State University Present address: Institute for Genomics and Evolutionary Medicine, Temple University, Philadelphia, PA.
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Nacer A, Movila A, Sohet F, Girgis NM, Gundra UM, Loke P, Daneman R, Frevert U. Experimental cerebral malaria pathogenesis--hemodynamics at the blood brain barrier. PLoS Pathog 2014; 10:e1004528. [PMID: 25474413 PMCID: PMC4256476 DOI: 10.1371/journal.ppat.1004528] [Citation(s) in RCA: 77] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2014] [Accepted: 10/17/2014] [Indexed: 12/16/2022] Open
Abstract
Cerebral malaria claims the lives of over 600,000 African children every year. To better understand the pathogenesis of this devastating disease, we compared the cellular dynamics in the cortical microvasculature between two infection models, Plasmodium berghei ANKA (PbA) infected CBA/CaJ mice, which develop experimental cerebral malaria (ECM), and P. yoelii 17XL (PyXL) infected mice, which succumb to malarial hyperparasitemia without neurological impairment. Using a combination of intravital imaging and flow cytometry, we show that significantly more CD8(+) T cells, neutrophils, and macrophages are recruited to postcapillary venules during ECM compared to hyperparasitemia. ECM correlated with ICAM-1 upregulation on macrophages, while vascular endothelia upregulated ICAM-1 during ECM and hyperparasitemia. The arrest of large numbers of leukocytes in postcapillary and larger venules caused microrheological alterations that significantly restricted the venous blood flow. Treatment with FTY720, which inhibits vascular leakage, neurological signs, and death from ECM, prevented the recruitment of a subpopulation of CD45(hi) CD8(+) T cells, ICAM-1(+) macrophages, and neutrophils to postcapillary venules. FTY720 had no effect on the ECM-associated expression of the pattern recognition receptor CD14 in postcapillary venules suggesting that endothelial activation is insufficient to cause vascular pathology. Expression of the endothelial tight junction proteins claudin-5, occludin, and ZO-1 in the cerebral cortex and cerebellum of PbA-infected mice with ECM was unaltered compared to FTY720-treated PbA-infected mice or PyXL-infected mice with hyperparasitemia. Thus, blood brain barrier opening does not involve endothelial injury and is likely reversible, consistent with the rapid recovery of many patients with CM. We conclude that the ECM-associated recruitment of large numbers of activated leukocytes, in particular CD8(+) T cells and ICAM(+) macrophages, causes a severe restriction in the venous blood efflux from the brain, which exacerbates the vasogenic edema and increases the intracranial pressure. Thus, death from ECM could potentially occur as a consequence of intracranial hypertension.
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Affiliation(s)
- Adéla Nacer
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
| | - Alexandru Movila
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
| | - Fabien Sohet
- Department of Anatomy, University of California San Francisco, San Francisco, California, United States of America
| | - Natasha M. Girgis
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
| | - Uma Mahesh Gundra
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
| | - P'ng Loke
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
| | - Richard Daneman
- Department of Anatomy, University of California San Francisco, San Francisco, California, United States of America
| | - Ute Frevert
- Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America
- * E-mail:
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Noulin F, Manesia JK, Rosanas-Urgell A, Erhart A, Borlon C, Van Den Abbeele J, d'Alessandro U, Verfaillie CM. Hematopoietic stem/progenitor cell sources to generate reticulocytes for Plasmodium vivax culture. PLoS One 2014; 9:e112496. [PMID: 25393299 PMCID: PMC4231068 DOI: 10.1371/journal.pone.0112496] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2014] [Accepted: 10/17/2014] [Indexed: 01/31/2023] Open
Abstract
The predilection of Plasmodium vivax (P. vivax) for reticulocytes is a major obstacle for its establishment in a long-term culture system, as this requires a continuous supply of large quantities of reticulocytes, representing only 1-2% of circulating red blood cells. We here compared the production of reticulocytes using an established in vitro culture system from three different sources of hematopoietic stem/progenitor cells (HSPC), i.e. umbilical cord blood (UCB), bone marrow (BM) and adult peripheral blood (PB). Compared to CD34+-enriched populations of PB and BM, CD34+-enriched populations of UCB produced the highest amount of reticulocytes that could be invaded by P. vivax. In addition, when CD34+-enriched cells were first expanded, a further extensive increase in reticulocytes was seen for UCB, to a lesser degree BM but not PB. As invasion by P. vivax was significantly better in reticulocytes generated in vitro, we also suggest that P. vivax may have a preference for invading immature reticulocytes, which should be confirmed in future studies.
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Affiliation(s)
- Florian Noulin
- Unit of Malariology, Institute of Tropical Medicine, Antwerp, Belgium
- * E-mail:
| | - Javed Karim Manesia
- Department of development and regeneration, Stem Cell Institute, Leuven, Belgium
| | | | - Annette Erhart
- Unit of Malariology, Institute of Tropical Medicine, Antwerp, Belgium
| | - Céline Borlon
- Unit of Malariology, Institute of Tropical Medicine, Antwerp, Belgium
| | - Jan Van Den Abbeele
- Unit of Veterinary Protozoology, Institute of Tropical Medicine, Antwerp, Belgium
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Frevert U, Nacer A. Fatal cerebral malaria: a venous efflux problem. Front Cell Infect Microbiol 2014; 4:155. [PMID: 25414834 PMCID: PMC4222339 DOI: 10.3389/fcimb.2014.00155] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2014] [Accepted: 10/13/2014] [Indexed: 12/20/2022] Open
Abstract
Most Plasmodium falciparum-infected children with cerebral malaria (CM) die from respiratory arrest, but the underlying pathology is unclear. Here we present a model in which the ultimate cause of death from CM is severe intracranial hypertension. Dynamic imaging of mice infected with P. berghei ANKA, an accepted model for experimental CM, revealed that leukocyte adhesion impairs the venous blood flow by reducing the functional lumen of postcapillary venules (PCV). The resulting increase in intracranial pressure (ICP) exacerbates cerebral edema formation, a hallmark of both murine and pediatric CM. We propose that two entirely different pathogenetic mechanisms-cytoadherence of P. falciparum-infected erythrocytes in pediatric CM and leukocyte arrest in murine CM-result in the same pathological outcome: a severe increase in ICP leading to brainstem herniation and death from respiratory arrest. The intracranial hypertension (IH) model unifies previous hypotheses, applies to human and experimental CM alike, eliminates the need to explain any selective recognition mechanism Plasmodium might use to target multiple sensitive sites in the brain, and explains how an intravascular parasite can cause so much neuronal dysfunction.
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Affiliation(s)
- Ute Frevert
- Division of Medical Parasitology, Department of Microbiology, New York University School of Medicine New York, NY, USA
| | - Adéla Nacer
- Unité de Biologie des Interactions Hôte-Parasite, Département de Parasitologie et Mycologie, Institut Pasteur Paris, France
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Fernandez-Becerra C, Lelievre J, Ferrer M, Anton N, Thomson R, Peligero C, Almela MJ, Lacerda MV, Herreros E, Del Portillo HA. Red blood cells derived from peripheral blood and bone marrow CD34⁺ human haematopoietic stem cells are permissive to Plasmodium parasites infection. Mem Inst Oswaldo Cruz 2014; 108:801-3. [PMID: 24037205 PMCID: PMC3970681 DOI: 10.1590/0074-0276108062013019] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2013] [Accepted: 07/12/2013] [Indexed: 01/17/2023] Open
Abstract
The production of fully functional human red cells in vitro from haematopoietic
stem cells (hHSCs) has been successfully achieved. Recently, the use of hHSCs
from cord blood represented a major improvement to develop the continuous
culture system for Plasmodium vivax. Here, we demonstrated that
CD34+hHSCs from peripheral blood and bone marrow can be expanded and
differentiated to reticulocytes using a novel stromal cell. Moreover, these
reticulocytes and mature red blood cells express surface markers for entrance of
malaria parasites contain adult haemoglobin and are also permissive to invasion
by P. vivax and Plasmodium falciparum
parasites.
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Affiliation(s)
- Carmen Fernandez-Becerra
- Barcelona Centre for International Health Research, Hospital Clinic, Universitat de Barcelona, BarcelonaSpain, Spain
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In vitro susceptibility of Plasmodium vivax to antimalarials in Colombia. Antimicrob Agents Chemother 2014; 58:6354-9. [PMID: 25114141 DOI: 10.1128/aac.03191-14] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023] Open
Abstract
The in vitro susceptibilities of 30 isolates of Plasmodium vivax to a number of antimalarials (chloroquine [CQ], mefloquine, amodiaquine, quinine, and artesunate [AS]) were evaluated. The isolates came from the region of Urabá in Colombia, in which malaria is endemic, and were evaluated by the schizont maturation test. The 50% inhibitory concentration (IC50) was 0.6 nM (95% confidence interval [CI], 0.3 to 1.0 nM) for artesunate, 8.5 nM (95% CI, 5.6 to 13.0 nM) for amodiaquine, 23.3 nM (95% CI, 12.4 to 44.1 nM) for chloroquine, 55.6 nM (95% CI, 36.8 to 84.1 nM) for mefloquine, and 115.3 nM (95% CI, 57.7 to 230.5 nM) for quinine. The isolates were classified according to whether the initial parasites were mature or immature trophozoites (Tfz). It was found that the IC50s for chloroquine and artesunate were significantly different in the two aforementioned groups (P < 0.001). The IC50s of CQ and AS were higher in the isolates from mature Tfz (CQ, 39.3 nM versus 17 nM; AS, 1.4 nM versus 0.3 nM), and 10% of the isolates showed lower susceptibilities to one of the antimalarial drugs, 13.3% to two antimalarial drugs, and 3.3% to more than three antimalarial drugs. It should be highlighted that despite the extensive use of chloroquine in Colombia, P. vivax continues to be susceptible to antimalarials. This is the first report, to our knowledge, showing in vitro susceptibilities of P. vivax isolates to antimalarials in Colombia.
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Antibody-conjugated paramagnetic nanobeads: kinetics of bead-cell binding. Int J Mol Sci 2014; 15:8821-34. [PMID: 24852940 PMCID: PMC4057761 DOI: 10.3390/ijms15058821] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/03/2014] [Revised: 04/23/2014] [Accepted: 04/24/2014] [Indexed: 01/28/2023] Open
Abstract
Specific labelling of target cell surfaces using antibody-conjugated paramagnetic nanobeads is essential for efficient magnetic cell separation. However, studies examining parameters determining the kinetics of bead-cell binding are scarce. The present study determines the binding rates for specific and unspecific binding of 150 nm paramagnetic nanobeads to highly purified target and non-target cells. Beads bound to cells were enumerated spectrophotometrically. Results show that the initial bead-cell binding rate and saturation levels depend on initial bead concentration and fit curves of the form A(1 − exp(−kt)). Unspecific binding within conventional experimental time-spans (up to 60 min) was not detectable photometrically. For CD3-positive cells, the probability of specific binding was found to be around 80 times larger than that of unspecific binding.
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42
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Patarroyo MA, Calderón D, Moreno-Pérez DA. Vaccines againstPlasmodium vivax: a research challenge. Expert Rev Vaccines 2014; 11:1249-60. [DOI: 10.1586/erv.12.91] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
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Reticulocytes from cryopreserved erythroblasts support Plasmodium vivax infection in vitro. Parasitol Int 2013; 63:278-84. [PMID: 24291603 DOI: 10.1016/j.parint.2013.11.011] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2013] [Revised: 11/19/2013] [Accepted: 11/21/2013] [Indexed: 11/20/2022]
Abstract
Plasmodium vivax is the most widely distributed human malaria parasite. Despite its importance, both clinical research and basic research have been hampered by lack of a convenient in vitro culture system, in part due to the parasite's infection preference of reticulocytes rather than mature erythrocytes. The use of reticulocyte-producing hematopoietic stem cell culture has been proposed for the maintenance of the parasite, but good numbers of reticulocytes and P. vivax parasites sufficient for practical use in research have been difficult to produce from this system. Here, we report an improved method of hematopoietic stem cell culture for P. vivax infection, which requires less time and produces higher or equivalent percentage of reticulocytes than previously reported systems. Reticulocytes were cultured from cryopreserved erythroblasts that were frozen after 8day-cultivation of purified CD34+ cells from human umbilical cord blood. This method of production allowed the recovery of reticulocytes in a shorter time than with continuous stem cell culture. We obtained a relatively high percentage of peak reticulocyte production by using co-cultivation with a mouse stromal cell line. Using P. vivax mature stage parasites obtained from infected Aotus monkeys, we observed substantial numbers (up to 0.8% of the total number of the cells) of newly invaded reticulocytes 24h after initial cultivation. The addition of fresh reticulocytes after 48h culture, however, did not result in significant increase of second cycle reticulocyte invasion. Assays of invasion inhibition with specific antibodies were successful with this system, demonstrating potential for study of biological processes as well as the conditions necessary for long-term maintenance of P. vivax in vitro.
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Held J, Kreidenweiss A, Mordmüller B. Novel approaches in antimalarial drug discovery. Expert Opin Drug Discov 2013; 8:1325-37. [PMID: 24090219 DOI: 10.1517/17460441.2013.843522] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
INTRODUCTION The development of new antimalarial drugs remains of the utmost importance, since Plasmodium falciparum has developed resistance against nearly all chemotherapeutics in clinical use. In an effort to contain the resistance of P. falciparum against artemisinins and to further eradication efforts, studies are ongoing to identify novel and more efficacious approaches to develop antimalarials. AREAS COVERED The authors review the classical and new approaches to antimalarial drug discovery, with a special emphasis on the various stages of the parasite's life cycle and the different Plasmodium species. The authors discuss the methodologies and strategies for early efficacy testing that aim to narrow down the portfolio of promising compounds. EXPERT OPINION The increased efforts in the discovery and development of new antimalarial compounds have led to the recognition of new promising hits. However, there is still major roadblock of selecting the most promising compounds and then further testing them in early clinical trials, especially in the current restricted economy. Controlled human malaria infection has much potential for speeding-up the early development process of many drug candidates including those which target the pre-erythrocytic stages.
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Affiliation(s)
- Jana Held
- University of Tübingen, Institute of Tropical Medicine , Wilhelmstraße 27, D-72074 Tübingen , Germany +49 7071 29 82364 ; +49 7071 295189 ;
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45
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Imai T, Ishida H, Suzue K, Hirai M, Taniguchi T, Okada H, Suzuki T, Shimokawa C, Hisaeda H. CD8(+) T cell activation by murine erythroblasts infected with malaria parasites. Sci Rep 2013; 3:1572. [PMID: 23535896 PMCID: PMC3610137 DOI: 10.1038/srep01572] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2012] [Accepted: 03/11/2013] [Indexed: 01/30/2023] Open
Abstract
Recent studies show that some human malaria parasite species Plasmodium falciparum and P.vivax parasitize erythroblasts; however, the biological and clinical significance of this is unclear. To investigate further, we generated a rodent malaria parasite (P. yoelii 17XNL) expressing GFP-ovalbumin (OVA). Its infectivity to erythroblasts was confirmed, and parasitized erythroblasts were capable of initiating malaria infections. Experiments showed that MHC class I molecules were highly expressed on parasitized erythroblasts. As CD8+ T cells recognize MHC class I and peptide complexes on target cells, and are involved in protection or pathology against malaria, we examined whether erythroblasts are targeted by CD8+ T cells. Purified non-parasitized erythroblasts pulsed with OVA peptides were recognized by OVA-specific CD8+ T cells. Crucially, parasitized erythroblasts isolated from GFP-OVA-, but not GFP- infected-mice, activated OT-I CD8+ T cells, indicating that CD8+ T cells recognize parasitized erythroblasts in an antigen-specific manner.
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Affiliation(s)
- Takashi Imai
- Department of Parasitology, Graduate School of Medicine, Gunma University, Gunma 371-8511, Japan
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46
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Moreno-Pérez DA, Ruíz JA, Patarroyo MA. Reticulocytes: Plasmodium vivax target cells. Biol Cell 2013; 105:251-60. [PMID: 23458497 DOI: 10.1111/boc.201200093] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2012] [Accepted: 02/22/2013] [Indexed: 02/05/2023]
Abstract
Reticulocytes represent the main invasion target for Plasmodium vivax, the second most prevalent parasite species around the world causing malaria in humans. In spite of these cells' importance in research into malaria, biological knowledge related to the nature of the host has been limited, given the technical difficulties present in working with them in the laboratory. Poor reticulocyte recovery from total blood, by different techniques, has hampered continuous in vitro P. vivax cultures being developed, thereby delaying basic investigation in this parasite species. Intense research during the last few years has led to advances being made in developing methodologies orientated towards obtaining enriched reticulocytes from differing sources, thereby providing invaluable information for developing new strategies aimed at preventing infection caused by malaria. This review describes the most recent studies related to obtaining reticulocytes and discusses approaches which could contribute towards knowledge regarding molecular interactions between target cell proteins and their main infective agent, P. vivax.
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47
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Noulin F, Borlon C, Van Den Abbeele J, D'Alessandro U, Erhart A. 1912-2012: a century of research on Plasmodium vivax in vitro culture. Trends Parasitol 2013; 29:286-94. [PMID: 23623759 DOI: 10.1016/j.pt.2013.03.012] [Citation(s) in RCA: 73] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2012] [Revised: 03/22/2013] [Accepted: 03/29/2013] [Indexed: 01/17/2023]
Abstract
The development of a continuous Plasmodium vivax blood cycle in vitro was first attempted 100 years ago. Since then, and despite the use of different methods, only short-term cultures have been achieved so far. The available literature has been reviewed in order to provide a critical overview of the currently available knowledge on P. vivax blood cycle culture systems and identify some unexplored ways forward. Results show that data accumulated over the past century remain fragmented and often contradictory, making it difficult to draw conclusions. There is the need for an international consortium on P. vivax culture able to collect, update, and share new evidence, including negative results, and thus better coordinate current efforts towards the establishment of a continuous P. vivax culture.
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Affiliation(s)
- Florian Noulin
- Unit of Malariology, Institute of Tropical Medicine, Antwerp, 2000, Belgium.
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48
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Galinski MR, Meyer EVS, Barnwell JW. Plasmodium vivax: modern strategies to study a persistent parasite's life cycle. ADVANCES IN PARASITOLOGY 2013; 81:1-26. [PMID: 23384620 DOI: 10.1016/b978-0-12-407826-0.00001-1] [Citation(s) in RCA: 44] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Plasmodium vivax has unique attributes to support its survival in varying ecologies and climates. These include hypnozoite forms in the liver, an invasion preference for reticulocytes, caveola-vesicle complex structures in the infected erythrocyte membrane and rapidly forming and circulating gametocytes. These characteristics make this species very different from P. falciparum. Plasmodium cynomolgi and other related simian species have identical biology and can serve as informative models of P. vivax infections. Plasmodium vivax and its model parasites can be grown in non-human primates (NHP), and in short-term ex vivo cultures. For P. vivax, in the absence of in vitro culture systems, these models remain highly relevant side by side with human clinical studies. While post-genomic technologies allow for greater exploration of P. vivax-infected blood samples from humans, these come with restrictions. Two advantages of NHP models are that infections can be experimentally tailored to address hypotheses, including genetic manipulation. Also, systems biology approaches can capitalise on computational biology combined with set experimental infection periods and protocols, which may include multiple sampling times, different types of samples, and the broad use of "omics" technologies. Opportunities for research on vivax malaria are increasing with the use of existing and new methodological strategies in combination with modern technologies.
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Affiliation(s)
- Mary R Galinski
- Department of Medicine, Division of Infectious Diseases, Emory University School of Medicine, Emory University, Atlanta, Georgia, USA.
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49
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Baird JK. Evidence and implications of mortality associated with acute Plasmodium vivax malaria. Clin Microbiol Rev 2013; 26:36-57. [PMID: 23297258 PMCID: PMC3553673 DOI: 10.1128/cmr.00074-12] [Citation(s) in RCA: 280] [Impact Index Per Article: 23.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Vivax malaria threatens patients despite relatively low-grade parasitemias in peripheral blood. The tenet of death as a rare outcome, derived from antiquated and flawed clinical classifications, disregarded key clinical evidence, including (i) high rates of mortality in neurosyphilis patients treated with vivax malaria; (ii) significant mortality from zones of endemicity; and (iii) the physiological threat inherent in repeated, very severe paroxysms in any patient, healthy or otherwise. The very well-documented course of this infection, with the exception of parasitemia, carries all of the attributes of "perniciousness" historically linked to falciparum malaria, including severe disease and fatal outcomes. A systematic analysis of the parasite biomass in severely ill patients that includes blood, marrow, and spleen may ultimately explain this historic misunderstanding. Regardless of how this parasite is pernicious, recent data demonstrate that the infection comes with a significant burden of morbidity and associated mortality. The extraordinary burden of malaria is not heavily weighted upon any single continent by a single species of parasite-it is a complex problem for the entire endemic world, and both species are of fundamental importance. Humanity must rally substantial resources, intellect, and energy to counter this daunting but profound threat.
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Affiliation(s)
- J Kevin Baird
- Eijkman-Oxford Clinical Research Unit, Jakarta, Indonesia, and the Centre for Tropical Medicine, Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom
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50
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Jangpatarapongsa K, Xia H, Fang Q, Hu K, Yuan Y, Peng M, Gao Q, Sattabongkot J, Cui L, Li B, Udomsangpetch R. Immunity to malaria in Plasmodium vivax infection: a study in central China. PLoS One 2012; 7:e45971. [PMID: 23049909 PMCID: PMC3457974 DOI: 10.1371/journal.pone.0045971] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2012] [Accepted: 08/23/2012] [Indexed: 12/03/2022] Open
Abstract
Background P. vivax infection is characterised by relapsing fever, indicating reinfection by previously hidden parasites in the host. Relapsed infection can lead to the activation of the memory T cell pool, which may lead to protective immunity. This study aims to characterise immune responses in acute P. vivax-infected patients living in an area of central China characterised by only P. vivax infection. Methodology/Principal Findings We conducted a cross-sectional immune-phenotypic analysis of adults using the following inclusion criteria: acute P. vivax infection (N = 37), a history of P. vivax infection (N = 17), and no known history of P. vivax infection (N = 21). We also conducted a 2-week longitudinal analysis following acute P. vivax infection, in which PBMC proliferation was measured in response to P. vivax and P. falciparum blood stage lysates. Using flow cytometry, we showed elevated memory T cells in the blood during acute P. vivax infection. The levels of γδ T cells were two-fold higher than those measured in naive controls. This result suggested that in the two populations, memory and γδ T cells promptly responded to P. vivax parasites. Interestingly, P. falciparum antigens stimulated T cells obtained from P. vivax-infected patients during a day 14-convalescence, whereas lymphocytes from the naïve control group responded to a lower degree of convalescence. Conclusions/Significance Cell-mediated immunity during the convalescent period of the P. vivax-infected hosts was comprised of T cells that were specifically able to recognise P. falciparum antigens. Although the magnitude of the response was only half that measured after stimulation with P. vivax antigens, the matter of cross-antigenic stimulation is of great interest.
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Affiliation(s)
- Kulachart Jangpatarapongsa
- Center for Innovation Development and Technology Transfer, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand
- Department of Clinical Microbiology and Applied Technology, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand
| | - Hui Xia
- Department of Parasitology, Bengbu Medical College, Anhui, China
- Anhui Key Laboratory of Infection and Immunity at Bengbu Medical College, Anhui, China
| | - Qiang Fang
- Department of Parasitology, Bengbu Medical College, Anhui, China
- Anhui Key Laboratory of Infection and Immunity at Bengbu Medical College, Anhui, China
| | - Kaiming Hu
- Department of Parasitology, Bengbu Medical College, Anhui, China
| | - Yuanying Yuan
- Department of Parasitology, Bengbu Medical College, Anhui, China
| | - Meiyu Peng
- Department of Immunology, Bengbu Medical College, Anhui, China
| | - Qi Gao
- Jiangsu Institute of Parasitic Disease, Wuxi, China
| | - Jetsumon Sattabongkot
- Mahidol Vivax Research Center, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand
| | - Liwang Cui
- Department of Entomology, Pennsylvania State University, State College, Pennsylvania, United States of America
| | - Baiqing Li
- Anhui Key Laboratory of Infection and Immunity at Bengbu Medical College, Anhui, China
- Department of Immunology, Bengbu Medical College, Anhui, China
- * E-mail: (RU); (BL)
| | - Rachanee Udomsangpetch
- Department of Pathobiology, Faculty of Science, Mahidol University, Bangkok, Thailand
- Center of Neglected Infectious Diseases, Mahidol University, Bangkok, Thailand
- * E-mail: (RU); (BL)
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