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falciparum gametocyte culture

Helmi Pett, Bronner P Gonçalves, Alassane Dicko, Issa Nébié, Alfred B Tiono, Kjerstin Lanke, John Bradley, Ingrid Chen, Halimatou Diawara, Almahamoudou Mahamar, Harouna M Soumare, Sekou F Traore, Ibrahima Baber, Sodiomon B Sirima, Robert Sauerwein, Joelle Brown, Roly Gosling, Ingrid Felger, Chris Drakeley, Teun Bousema
BACKGROUND: Quantifying gametocyte densities in natural malaria infections is important to estimate malaria transmission potential. Two molecular methods (Pfs25 mRNA quantitative reverse transcriptase PCR (qRT-PCR) and Pfs25 mRNA quantitative nucleic acid sequence based amplification (QT-NASBA)) are commonly used to determine gametocyte densities in clinical and epidemiological studies and allow gametocyte detection at densities below the microscopic threshold for detection. Here, reproducibility of these measurements and the association between estimated gametocyte densities and mosquito infection rates were compared...
November 8, 2016: Malaria Journal
Mohammad Behram Khan, Jonathan Wee Kent Liew, Cherng Shii Leong, Yee-Ling Lau
BACKGROUND: Anopheles mosquitoes transmit malaria which is one of the world's most threatening diseases. Anopheles dirus (sensu stricto) is among the main vectors of malaria in South East Asia. The mosquito innate immune response is the first line of defence against malaria parasites during its development. The immune deficiency (IMD) pathway, a conserved immune signaling pathway, influences anti-Plasmodium falciparum activity in Anopheles gambiae, An. stephensi and An. albimanus. The aim of the study was to determine the role of Rel2, an IMD pathway-controlled NF-kappaβ transcription factor, in An...
September 29, 2016: Parasites & Vectors
Michael J Delves, Ursula Straschil, Andrea Ruecker, Celia Miguel-Blanco, Sara Marques, Jake Baum, Robert E Sinden
The prevention of parasite transmission from the human host to the mosquito has been recognized as a vital tool for malaria eradication campaigns. However, transmission-blocking antimalarial drug and/or vaccine discovery and development is currently hampered by the expense and difficulty of producing mature Plasmodium falciparum gametocytes in vitro-the parasite stage responsible for mosquito infection. Current protocols for P. falciparum gametocyte culture usually require complex parasite synchronization and addition of stimulating and/or inhibitory factors, and they may not have demonstrated the essential property of mosquito infectivity...
September 2016: Nature Protocols
Noemí Bahamontes-Rosa, María G Gomez-Lorenzo, Joël Lelièvre, Ane Rodriguez Alejandre, María Jesus Almela, Sonia Lozano, Esperanza Herreros, Francisco-Javier Gamo
BACKGROUND: Drugs that kill or inhibit Plasmodium gametocytes in the human host could potentially synergize the impact of other chemotherapeutic interventions by blocking transmission. To develop such agents, reliable methods are needed to study the in vitro activity of compounds against gametocytes. This study describes a novel assay for characterizing the activity of anti-malarial drugs against the later stages of Plasmodium falciparum gametocyte development using real-time PCR (qPCR)...
July 22, 2016: Malaria Journal
Elya Dekel, Anna Rivkin, Meta Heidenreich, Yotam Nadav, Yifat Ofir-Birin, Ziv Porat, Neta Regev-Rudzki
Malaria is the most devastating parasitic disease of humans, caused by the unicellular protozoa of the Plasmodium genus, such as Plasmodium falciparum (Pf) and is responsible for up to a million deaths each year. Pf life cycle is complex, with transmission of the parasite between humans via mosquitos involving a remarkable series of morphological transformations. In the bloodstream, the parasites undergo asexual multiplications inside the red blood cell (RBC), where they mature through the ring (R), trophozoite (T) and schizont (S) stages, and sexual development, resulting in gametocytes (G)...
June 25, 2016: Methods: a Companion to Methods in Enzymology
Katharina Ehrhardt, Christiane Deregnaucourt, Alice-Anne Goetz, Tzvetomira Tzanova, Valentina Gallo, Paolo Arese, Bruno Pradines, Sophie H Adjalley, Denyse Bagrel, Stephanie Blandin, Michael Lanzer, Elisabeth Davioud-Charvet
Previously, we presented the chemical design of a promising series of antimalarial agents, 3-[substituted-benzyl]-menadiones, with potent in vitro and in vivo activities. Ongoing studies on the mode of action of antimalarial 3-[substituted-benzyl]-menadiones revealed that these agents disturb the redox balance of the parasitized erythrocyte by acting as redox cyclers-a strategy that is broadly recognized for the development of new antimalarial agents. Here we report a detailed parasitological characterization of the in vitro activity profile of the lead compound 3-[4-(trifluoromethyl)benzyl]-menadione 1c (henceforth called plasmodione) against intraerythrocytic stages of the human malaria parasite Plasmodium falciparum We show that plasmodione acts rapidly against asexual blood stages, thereby disrupting the clinically relevant intraerythrocytic life cycle of the parasite, and furthermore has potent activity against early gametocytes...
September 2016: Antimicrobial Agents and Chemotherapy
Sandra Duffy, Sasdekumar Loganathan, John P Holleran, Vicky M Avery
The tightly controlled induction of Plasmodium falciparum gametocytes in large-scale culture is a fundamental requirement for malaria drug discovery applications including, but not limited to, high-throughput screening. This protocol uses magnetic separation for isolation of hemozoin-containing parasites in order to (i) increase parasitemia, (ii) decrease hematocrit and (iii) introduce higher levels of young red blood cells in a culture simultaneously within 2-4 h. These parameters, along with red blood cell lysis products that are generated through schizont rupture, are highly relevant for enabling optimum induction of gametocytogenesis in vitro...
May 2016: Nature Protocols
Susana Campino, Ernest Diez Benavente, Samuel Assefa, Eloise Thompson, Laura G Drought, Catherine J Taylor, Zaria Gorvett, Celine K Carret, Christian Flueck, Al C Ivens, Dominic P Kwiatkowski, Pietro Alano, David A Baker, Taane G Clark
BACKGROUND: Transmission of the malaria parasite Plasmodium falciparum from humans to the mosquito vector requires differentiation of a sub-population of asexual forms replicating within red blood cells into non-dividing male and female gametocytes. The nature of the molecular mechanism underlying this key differentiation event required for malaria transmission is not fully understood. METHODS: Whole genome sequencing was used to examine the genomic diversity of the gametocyte non-producing 3D7-derived lines F12 and A4...
April 21, 2016: Malaria Journal
Andreea Waltmann, Stephan Karl, Chris Chiu, Ivo Mueller
An important aspect of many malaria molecular epidemiology and transmission studies is RNA-based detection of gametocytes. Ensuring RNA stability represents a challenge in tropical, resource-limited environments, as RNA may quickly degrade when samples are not preserved under adequate conditions. This study investigated the degradation of pfs25 messenger RNA (mRNA), the most widely used Plasmodium falciparum gametocyte marker, in whole blood spiked with cultured P. falciparum gametocytes, exposed to different temperatures for up to 48 hours, and collected with different anticoagulants...
April 2016: American Journal of Tropical Medicine and Hygiene
Will Stone, Bryan Grabias, Kjerstin Lanke, Hong Zheng, Emily Locke, Diadier Diallo, Ashley Birkett, Merribeth Morin, Teun Bousema, Sanjai Kumar
BACKGROUND: The infectivity of Plasmodium gametocytes is typically determined by microscopically examining the midguts of mosquitoes that have taken a blood meal containing potentially infectious parasites. Such assessments are required for the development and evaluation of transmission-reducing interventions (TRI), but are limited by subjectivity, technical complexity and throughput. The detection of circumsporozoite protein (CSP) by enzyme-linked immunosorbent assay (ELISA) and enhanced chemiluminescent slot-blot (ECL-SB) may be used as objective, scalable alternatives to microscopy for the determination of infection prevalence...
November 14, 2015: Malaria Journal
Linda Eva Amoah, Courage Kakaney, Bethel Kwansa-Bentum, Kwadwo Asamoah Kusi
BACKGROUND: Malaria still remains a major health issue in Ghana despite the introduction of Artemisinin-based combination therapy (ACT) coupled with other preventative measures such as the use of insecticide treated nets (ITNs). The global quest for eradication of malaria has heightened the interest of identifying drugs that target the sexual stage of the parasite, referred to as transmission-blocking drugs. This study aimed at assessing the efficacy and gametocydal effects of some commonly used herbal malaria products in Ghana...
2015: PloS One
Mynthia Cabrera, Liwang Cui
Currently, the World Health Organization recommends addition of a 0.25-mg base/kg single dose of primaquine (PQ) to artemisinin combination therapies (ACTs) for Plasmodium falciparum malaria as a gametocytocidal agent for reducing transmission. Here, we investigated the potential interactions of PQ with the long-lasting components of the ACT drugs for eliminating the asexual blood stages and gametocytes of in vitro-cultured P. falciparum strains. Using the SYBR green I assay for asexual parasites and a flow cytometry-based assay for gametocytes, we determined the interactions of PQ with the schizonticides chloroquine, mefloquine, piperaquine, lumefantrine, and naphthoquine...
December 2015: Antimicrobial Agents and Chemotherapy
Marta Tibúrcio, Matthew W A Dixon, Oliver Looker, Sumera Younis Younis, Leann Tilley, Pietro Alano
BACKGROUND: Plasmodium falciparum sexual development plays a fundamental role in the transmission and spread of malaria. The ability to generate gametocytes can be lost during culture in vitro, often associated with the loss of a subtelomeric region of chromosome 9. Gametocytogenesis starts with erythrocyte invasion by a sexually committed merozoite, but the first available specific marker of sexual differentiation appears only from 24 h post invasion. METHODS: Specific antibodies and gene fusions were produced to study the timing of expression and the sub-cellular localization of the P...
2015: Malaria Journal
Omaira Vera, Paula Brelas de Brito, Letusa Albrecht, Keillen Monick Martins-Campos, Paulo F P Pimenta, Wuelton M Monteiro, Marcus V G Lacerda, Stefanie C P Lopes, Fabio T M Costa
Significant progress toward the control of malaria has been achieved, especially regarding Plasmodium falciparum infections. However, the unique biology of Plasmodium vivax hampers current control strategies. The early appearance of P. vivax gametocytes in the peripheral blood and the impossibility of culturing this parasite are major drawbacks. Using blood samples from 40 P. vivax-infected patients, we describe here a methodology to purify viable gametocytes and further infect anophelines. This method opens new avenues to validate transmission-blocking strategies...
October 2015: Antimicrobial Agents and Chemotherapy
Louis-Jérôme Leba, Lise Musset, Stéphane Pelleau, Yannick Estevez, Caroline Birer, Sébastien Briolant, Benoit Witkowski, Didier Ménard, Michael J Delves, Eric Legrand, Christophe Duplais, Jean Popovici
BACKGROUND: A major requirement for malaria elimination is the development of transmission-blocking interventions. In vitro transmission-blocking bioassays currently mostly rely on the use of very few Plasmodium falciparum reference laboratory strains isolated decades ago. To fill a piece of the gap between laboratory experimental models and natural systems, the purpose of this work was to determine if culture-adapted field isolates of P. falciparum are suitable for in vitro transmission-blocking bioassays targeting functional maturity of male gametocytes: exflagellation...
June 4, 2015: Malaria Journal
Tao Li, Abraham G Eappen, Adam M Richman, Peter F Billingsley, Yonas Abebe, Minglin Li, Debbie Padilla, Isabel Rodriguez-Barraquer, B Kim Lee Sim, Stephen L Hoffman
BACKGROUND: A vaccine that interrupts malaria transmission (VIMT) would be a valuable tool for malaria control and elimination. One VIMT approach is to identify sexual erythrocytic and mosquito stage antigens of the malaria parasite that induce immune responses targeted at disrupting parasite development in the mosquito. The standard Plasmodium falciparum membrane-feeding assay (SMFA) is used to assess transmission-blocking activity (TBA) of antibodies against candidate immunogens and of drugs targeting the mosquito stages...
2015: Malaria Journal
Karell G Pelle, Keunyoung Oh, Kathrin Buchholz, Vagheesh Narasimhan, Regina Joice, Danny A Milner, Nicolas Mb Brancucci, Siyuan Ma, Till S Voss, Ken Ketman, Karl B Seydel, Terrie E Taylor, Natasha S Barteneva, Curtis Huttenhower, Matthias Marti
BACKGROUND: During intra-erythrocytic development, late asexually replicating Plasmodium falciparum parasites sequester from peripheral circulation. This facilitates chronic infection and is linked to severe disease and organ-specific pathology including cerebral and placental malaria. Immature gametocytes - sexual stage precursor cells - likewise disappear from circulation. Recent work has demonstrated that these sexual stage parasites are located in the hematopoietic system of the bone marrow before mature gametocytes are released into the bloodstream to facilitate mosquito transmission...
2015: Genome Medicine
Sabrina D Lamour, Ursula Straschil, Jasmina Saric, Michael J Delves
BACKGROUND: Gametocytes are the Plasmodium life stage that is solely responsible for malaria transmission. Despite their important role in perpetuating malaria, gametocyte differentiation and development is poorly understood. METHODS: To shed light on the biochemical changes that occur during asexual and gametocyte development, metabolic characterization of media from in vitro intra-erythrocytic Plasmodium falciparum cultures was performed throughout gametocyte development by applying 1H nuclear magnetic spectroscopy, and using sham erythrocyte cultures as controls...
2014: Malaria Journal
Hangjun Ke, Paul A Sigala, Kazutoyo Miura, Joanne M Morrisey, Michael W Mather, Jan R Crowley, Jeffrey P Henderson, Daniel E Goldberg, Carole A Long, Akhil B Vaidya
Heme is an essential cofactor for aerobic organisms. Its redox chemistry is central to a variety of biological functions mediated by hemoproteins. In blood stages, malaria parasites consume most of the hemoglobin inside the infected erythrocytes, forming nontoxic hemozoin crystals from large quantities of heme released during digestion. At the same time, the parasites possess a heme de novo biosynthetic pathway. This pathway in the human malaria parasite Plasmodium falciparum has been considered essential and is proposed as a potential drug target...
December 12, 2014: Journal of Biological Chemistry
Natalie G Sanders, David J Sullivan, Godfree Mlambo, George Dimopoulos, Abhai K Tripathi
Discovery of transmission blocking compounds is an important intervention strategy necessary to eliminate and eradicate malaria. To date only a small number of drugs that inhibit gametocyte development and thereby transmission from the mosquito to the human host exist. This limitation is largely due to a lack of screening assays easily adaptable to high throughput because of multiple incubation steps or the requirement for high gametocytemia. Here we report the discovery of new compounds with gametocytocidal activity using a simple and robust SYBR Green I- based DNA assay...
2014: PloS One
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