ResearchPad - gametocytes https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Single-cell transcription analysis of <i>Plasmodium vivax</i> blood-stage parasites identifies stage- and species-specific profiles of expression]]> https://www.researchpad.co/article/elastic_article_14651 Analysis of individual Plasmodium vivax parasites reveals the tight control of the expression of most genes during the intra-erythrocytic cycle and the differentiation of male and female gametocytes, and highlights differences between the development of P. vivax and P. falciparum.

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<![CDATA[Plasmodium male gametocyte development and transmission are critically regulated by the two putative deadenylases of the CAF1/CCR4/NOT complex]]> https://www.researchpad.co/article/5c5ca300d5eed0c48441efa2

With relatively few known specific transcription factors to control the abundance of specific mRNAs, Plasmodium parasites may rely more on the regulation of transcript stability and turnover to provide sufficient gene regulation. Plasmodium transmission stages impose translational repression on specific transcripts in part to accomplish this. However, few proteins are known to participate in this process, and those that are characterized primarily affect female gametocytes. We have identified and characterized Plasmodium yoelii (Py) CCR4-1, a putative deadenylase, which plays a role in the development and activation of male gametocytes, regulates the abundance of specific mRNAs in gametocytes, and ultimately increases the efficiency of host-to-vector transmission. We find that when pyccr4-1 is deleted or its protein made catalytically inactive, there is a loss in the initial coordination of male gametocyte maturation and a reduction of parasite infectivity of the mosquito. Expression of only the N-terminal CAF1 domain of the essential CAF1 deadenylase leads to a similar phenotype. Comparative RNA-seq revealed that PyCCR4-1 affects transcripts important for transmission-related functions that are associated with male or female gametocytes, some of which directly associate with the immunoprecipitated complex. Finally, circular RT-PCR of one of the bound, dysregulated transcripts showed that deletion of the pyccr4-1 gene does not result in gross changes to its UTR or poly(A) tail length. We conclude that the two putative deadenylases of the CAF1/CCR4/NOT complex play critical and intertwined roles in gametocyte maturation and transmission.

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<![CDATA[Adaptive plasticity in the gametocyte conversion rate of malaria parasites]]> https://www.researchpad.co/article/5c256c7dd5eed0c484474e45

Sexually reproducing parasites, such as malaria parasites, experience a trade-off between the allocation of resources to asexual replication and the production of sexual forms. Allocation by malaria parasites to sexual forms (the conversion rate) is variable but the evolutionary drivers of this plasticity are poorly understood. We use evolutionary theory for life histories to combine a mathematical model and experiments to reveal that parasites adjust conversion rate according to the dynamics of asexual densities in the blood of the host. Our model predicts the direction of change in conversion rates that returns the greatest fitness after perturbation of asexual densities by different doses of antimalarial drugs. The loss of a high proportion of asexuals is predicted to elicit increased conversion (terminal investment), while smaller losses are managed by reducing conversion (reproductive restraint) to facilitate within-host survival and future transmission. This non-linear pattern of allocation is consistent with adaptive reproductive strategies observed in multicellular organisms. We then empirically estimate conversion rates of the rodent malaria parasite Plasmodium chabaudi in response to the killing of asexual stages by different doses of antimalarial drugs and forecast the short-term fitness consequences of these responses. Our data reveal the predicted non-linear pattern, and this is further supported by analyses of previous experiments that perturb asexual stage densities using drugs or within-host competition, across multiple parasite genotypes. Whilst conversion rates, across all datasets, are most strongly influenced by changes in asexual density, parasites also modulate conversion according to the availability of red blood cell resources. In summary, increasing conversion maximises short-term transmission and reducing conversion facilitates in-host survival and thus, future transmission. Understanding patterns of parasite allocation to reproduction matters because within-host replication is responsible for disease symptoms and between-host transmission determines disease spread.

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<![CDATA[Modeling the impact of Plasmodium falciparum sexual stage immunity on the composition and dynamics of the human infectious reservoir for malaria in natural settings]]> https://www.researchpad.co/article/5afd69a1463d7e7322194038

Malaria transmission remains high in Sub-Saharan Africa despite large-scale implementation of malaria control interventions. A comprehensive understanding of the transmissibility of infections to mosquitoes may guide the design of more effective transmission reducing strategies. The impact of P. falciparum sexual stage immunity on the infectious reservoir for malaria has never been studied in natural settings. Repeated measurements were carried out at start-wet, peak-wet and dry season, and provided data on antibody responses against gametocyte/gamete antigens Pfs48/45 and Pfs230 as anti-gametocyte immunity. Data on high and low-density infections and their infectiousness to anopheline mosquitoes were obtained using quantitative molecular methods and mosquito feeding assays, respectively. An event-driven model for P. falciparum sexual stage immunity was developed and fit to data using an agent based malaria model infrastructure. We found that Pfs48/45 and Pfs230 antibody densities increased with increasing concurrent gametocyte densities; associated with 55–70% reduction in oocyst intensity and achieved up to 44% reduction in proportions of infected mosquitoes. We showed that P. falciparum sexual stage immunity significantly reduces transmission of microscopic (p < 0.001) but not submicroscopic (p = 0.937) gametocyte infections to mosquitoes and that incorporating sexual stage immunity into mathematical models had a considerable impact on the contribution of different age groups to the infectious reservoir of malaria. Human antibody responses to gametocyte antigens are likely to be dependent on recent and concurrent high-density gametocyte exposure and have a pronounced impact on the likelihood of onward transmission of microscopic gametocyte densities compared to low density infections. Our mathematical simulations indicate that anti-gametocyte immunity is an important factor for predicting and understanding the composition and dynamics of the human infectious reservoir for malaria.

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<![CDATA[Epigenetic Roulette in Blood Stream Plasmodium: Gambling on Sex]]> https://www.researchpad.co/article/5989db21ab0ee8fa60bcf44c ]]> <![CDATA[Plasmodium vivax gametocytes in the bone marrow of an acute malaria patient and changes in the erythroid miRNA profile]]> https://www.researchpad.co/article/5989db5aab0ee8fa60bdf31f ]]> <![CDATA[Associations between Season and Gametocyte Dynamics in Chronic Plasmodium falciparum Infections]]> https://www.researchpad.co/article/5989d9f7ab0ee8fa60b70cb4

Introduction

In a markedly seasonal malaria setting, the transition from the transmission-free dry season to the transmission season depends on the resurgence of the mosquito population following the start of annual rains. The sudden onset of malaria outbreaks at the start of the transmission season suggests that parasites persist during the dry season and respond to either the reappearance of vectors, or correlated events, by increasing the production of transmission stages. Here, we investigate whether Plasmodium falciparum gametocyte density and the correlation between gametocyte density and parasite density show seasonal variation in chronic (largely asymptomatic) carriers in eastern Sudan.

Materials and Methods

We recruited and treated 123 malaria patients in the transmission season 2001. We then followed them monthly during four distinct consecutive epidemiological seasons: transmission season 1, transmission-free season, pre-clinical period, and transmission season 2. In samples collected from 25 participants who fulfilled the selection criteria of the current analysis, we used quantitative PCR (qPCR) and RT-qPCR to quantify parasite and gametocyte densities, respectively.

Results and Discussion

We observed a significant increase in gametocyte density and a significantly steeper positive correlation between gametocyte density and total parasite density during the pre-clinical period compared to the preceding transmission-free season. However, there was no corresponding increase in the density or prevalence of total parasites or gametocyte prevalence. The increase in gametocyte production during the pre-clinical period supports the hypothesis that P. falciparum may respond to environmental cues, such as mosquito biting, to modulate its transmission strategy. Thus, seasonal changes may be important to ignite transmission in unstable-malaria settings.

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<![CDATA[Therapeutic efficacy of artemether-lumefantrine against uncomplicated Plasmodium falciparum malaria in a high-transmission area in northwest Ethiopia]]> https://www.researchpad.co/article/5989db59ab0ee8fa60bdf11c

Malaria, particularly due to Plasmodium falciparum, remains a major public health threat in Ethiopia. Artemether-lumefantine (AL) has been the first-line antimalarial drug against uncomplicated P. falciparum malaria in the country since 2004. Regular monitoring of antimalarial drugs is recommended by the World Health Organization (WHO) to help early detection of drug resistant strains of the parasite and contain their rapid spread. The objective of this study was to assess the therapeutic efficacy of AL in a high-transmission setting in Ethiopia. The study site was Setit Humera, northwest Ethiopia. Single-arm prospective study of a 28-day follow-up was conducted from October 2014 to January 2015 according to the revised WHO 2009 drug efficacy study protocol. Study end-points were classified into primary end-point and secondary end-point. While the primary end-point was the day-28 adequate clinical and parasitological response the secondary end-points were clinical and parasitological evaluations (parasite, fever and gametocyte clearance rate, incidence of drug adverse events) and the relative increment in hemoglobin (Hb) level from baseline to day (D) 14 and D28. A total of 92 patients were enrolled and 79 had completed the 28-day follow-up period. The overall cure rate was 98.8% with 95% confidence interval of 0.915–0.998 without polymerase chain reaction correction. The parasite clearance rate was high with fast resolution of clinical symptoms; 100% of the study participants cleared parasitaemia and fever on D3. Gametocyte carriage was reduced from 7% on D0 to 1% on D3 and complete clearance was achieved on D14. Mean Hb concentration significantly increased on D28 compared to that on D14. There was no serious adverse event. AL was efficacious and safe in a high-transmission setting for treatment of uncomplicated falciparum malaria.

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<![CDATA[Quantifying Transmission Investment in Malaria Parasites]]> https://www.researchpad.co/article/5989d9f0ab0ee8fa60b6e21c

Many microparasites infect new hosts with specialized life stages, requiring a subset of the parasite population to forgo proliferation and develop into transmission forms. Transmission stage production influences infectivity, host exploitation, and the impact of medical interventions like drug treatment. Predicting how parasites will respond to public health efforts on both epidemiological and evolutionary timescales requires understanding transmission strategies. These strategies can rarely be observed directly and must typically be inferred from infection dynamics. Using malaria as a case study, we test previously described methods for inferring transmission stage investment against simulated data generated with a model of within-host infection dynamics, where the true transmission investment is known. We show that existing methods are inadequate and potentially very misleading. The key difficulty lies in separating transmission stages produced by different generations of parasites. We develop a new approach that performs much better on simulated data. Applying this approach to real data from mice infected with a single Plasmodium chabaudi strain, we estimate that transmission investment varies from zero to 20%, with evidence for variable investment over time in some hosts, but not others. These patterns suggest that, even in experimental infections where host genetics and other environmental factors are controlled, parasites may exhibit remarkably different patterns of transmission investment.

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<![CDATA[Monitoring the Prevalence of Leucocytozoon sabrazesi in Southern China and Testing Tricyclic Compounds against Gametocytes]]> https://www.researchpad.co/article/5989d9f7ab0ee8fa60b70c79

Leucocytozoon parasites infect many species of avian hosts, including domestic chicken, and can inflict heavy economic loss on the poultry industry. Two major species of Leucocytozoon parasites have been reported in China, L. sabrazesi and L. caulleryi, although L. sabrazesi appears to be more widespread than L. caulleryi in southern China. The traditional method for detecting Leucocytozoon infection is microscopic examination of blood smears for the presence of mature gametocytes in circulation, which may miss infections with low parasitemia (gametocytemia) or immature gametocytes. Here we developed a PCR-based method to monitor L. sabrazesi infections at seven sites in four provinces of China after testing two PCR primer pairs based on parasite mitochondrial cytochrome b (cytb) and cytochrome c oxidase III (coxIII) genes. We compared the results of PCR detection with those of microscopic observation. As expected, the PCR assays were more sensitive than microscope examination in detecting L. sabrazesi infection and were able to detect parasite DNA after gametocytes disappeared in the blood stream. Using these methods, we investigated monthly dynamics of L. sabrazesi in chickens from a free-range farm in Xiamen, Fujian province of China, over one year. Our results showed that chickens were infected with L. sabrazesi year-round in southern China. Finally, we tested several compounds for potential treatment of Leucocytozoon infections, including primaquine, ketotifen, clomipramine hydrochloride, desipramine hydrochloride, sulfaquinoxaline, and pyrimethamine. Only primaquine had activity against L. sabrazesi gametocytes. Our results provide important information for controlling parasite transmission in southern China and disease management.

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<![CDATA[Effectiveness of Seasonal Malaria Chemoprevention in Children under Ten Years of Age in Senegal: A Stepped-Wedge Cluster-Randomised Trial]]> https://www.researchpad.co/article/5989db23ab0ee8fa60bcfa95

Background

Seasonal Malaria Chemoprevention (SMC) with sulfadoxine-pyrimethamine (SP) plus amodiaquine (AQ), given each month during the transmission season, is recommended for children living in areas of the Sahel where malaria transmission is highly seasonal. The recommendation for SMC is currently limited to children under five years of age, but, in many areas of seasonal transmission, the burden in older children may justify extending this age limit. This study was done to determine the effectiveness of SMC in Senegalese children up to ten years of age.

Methods and Findings

SMC was introduced into three districts over three years in central Senegal using a stepped-wedge cluster-randomised design. A census of the population was undertaken and a surveillance system was established to record all deaths and to record all cases of malaria seen at health facilities. A pharmacovigilance system was put in place to detect adverse drug reactions. Fifty-four health posts were randomised. Nine started implementation of SMC in 2008, 18 in 2009, and a further 18 in 2010, with 9 remaining as controls. In the first year of implementation, SMC was delivered to children aged 3–59 months; the age range was then extended for the latter two years of the study to include children up to 10 years of age. Cluster sample surveys at the end of each transmission season were done to measure coverage of SMC and the prevalence of parasitaemia and anaemia, to monitor molecular markers of drug resistance, and to measure insecticide-treated net (ITN) use. Entomological monitoring and assessment of costs of delivery in each health post and of community attitudes to SMC were also undertaken. About 780,000 treatments were administered over three years. Coverage exceeded 80% each month. Mortality, the primary endpoint, was similar in SMC and control areas (4.6 and 4.5 per 1000 respectively in children under 5 years and 1.3 and 1.2 per 1000 in children 5-9 years of age; the overall mortality rate ratio [SMC: no SMC] was 0.90, 95% CI 0.68–1.2, p = 0.496). A reduction of 60% (95% CI 54%–64%, p < 0.001) in the incidence of malaria cases confirmed by a rapid diagnostic test (RDT) and a reduction of 69% (95% CI 65%–72%, p < 0.001) in the number of treatments for malaria (confirmed and unconfirmed) was observed in children. In areas where SMC was implemented, incidence of confirmed malaria in adults and in children too old to receive SMC was reduced by 26% (95% CI 18%–33%, p < 0.001) and the total number of treatments for malaria (confirmed and unconfirmed) in these older age groups was reduced by 29% (95% CI 21%–35%, p < 0.001). One hundred and twenty-three children were admitted to hospital with a diagnosis of severe malaria, with 64 in control areas and 59 in SMC areas, showing a reduction in the incidence rate of severe disease of 45% (95% CI 5%–68%, p = 0.031). Estimates of the reduction in the prevalence of parasitaemia at the end of the transmission season in SMC areas were 68% (95% CI 35%–85%) p = 0.002 in 2008, 84% (95% CI 58%–94%, p < 0.001) in 2009, and 30% (95% CI -130%–79%, p = 0.56) in 2010. SMC was well tolerated with no serious adverse reactions attributable to SMC drugs. Vomiting was the most commonly reported mild adverse event but was reported in less than 1% of treatments. The average cost of delivery was US$0.50 per child per month, but varied widely depending on the size of the health post. Limitations included the low rate of mortality, which limited our ability to detect an effect on this endpoint.

Conclusions

SMC substantially reduced the incidence of outpatient cases of malaria and of severe malaria in children, but no difference in all-cause mortality was observed. Introduction of SMC was associated with an overall reduction in malaria incidence in untreated age groups. In many areas of Africa with seasonal malaria, there is a substantial burden in older children that could be prevented by SMC. SMC in older children is well tolerated and effective and can contribute to reducing malaria transmission.

Trial Registration

ClinicalTrials.gov NCT00712374

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<![CDATA[Simulating within-vector generation of the malaria parasite diversity]]> https://www.researchpad.co/article/5989db5cab0ee8fa60bdff4a

Plasmodium falciparum, the most virulent human malaria parasite, undergoes asexual reproduction within the human host, but reproduces sexually within its vector host, the Anopheles mosquito. Consequently, the mosquito stage of the parasite life cycle provides an opportunity to create genetically novel parasites in multiply-infected mosquitoes, potentially increasing parasite population diversity. Despite the important implications for disease transmission and malaria control, a quantitative mapping of how parasite diversity entering a mosquito relates to diversity of the parasite exiting, has not been undertaken. To examine the role that vector biology plays in modulating parasite diversity, we develop a two-part model framework that estimates the diversity as a consequence of different bottlenecks and expansion events occurring during the vector-stage of the parasite life cycle. For the underlying framework, we develop the first stochastic model of within-vector P. falciparum parasite dynamics and go on to simulate the dynamics of two parasite subpopulations, emulating multiply infected mosquitoes. We show that incorporating stochasticity is essential to capture the extensive variation in parasite dynamics, particularly in the presence of multiple parasites. In particular, unlike deterministic models, which always predict the most fit parasites to produce the most sporozoites, we find that occasionally only parasites with lower fitness survive to the sporozoite stage. This has important implications for onward transmission. The second part of our framework includes a model of sequence diversity generation resulting from recombination and reassortment between parasites within a mosquito. Our two-part model framework shows that bottlenecks entering the oocyst stage decrease parasite diversity from what is present in the initial gametocyte population in a mosquito’s blood meal. However, diversity increases with the possibility for recombination and proliferation in the formation of sporozoites. Furthermore, when we begin with two parasite subpopulations in the initial gametocyte population, the probability of transmitting more than two unique parasites from mosquito to human is over 50% for a wide range of initial gametocyte densities.

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<![CDATA[Characterization of Novel Antimalarial Compound ACT-451840: Preclinical Assessment of Activity and Dose–Efficacy Modeling]]> https://www.researchpad.co/article/5989db45ab0ee8fa60bd84bb

Background

Artemisinin resistance observed in Southeast Asia threatens the continued use of artemisinin-based combination therapy in endemic countries. Additionally, the diversity of chemical mode of action in the global portfolio of marketed antimalarials is extremely limited. Addressing the urgent need for the development of new antimalarials, a chemical class of potent antimalarial compounds with a novel mode of action was recently identified. Herein, the preclinical characterization of one of these compounds, ACT-451840, conducted in partnership with academic and industrial groups is presented.

Method and Findings

The properties of ACT-451840 are described, including its spectrum of activities against multiple life cycle stages of the human malaria parasite Plasmodium falciparum (asexual and sexual) and Plasmodium vivax (asexual) as well as oral in vivo efficacies in two murine malaria models that permit infection with the human and the rodent parasites P. falciparum and Plasmodium berghei, respectively. In vitro, ACT-451840 showed a 50% inhibition concentration of 0.4 nM (standard deviation [SD]: ± 0.0 nM) against the drug-sensitive P. falciparum NF54 strain. The 90% effective doses in the in vivo efficacy models were 3.7 mg/kg against P. falciparum (95% confidence interval: 3.3–4.9 mg/kg) and 13 mg/kg against P. berghei (95% confidence interval: 11–16 mg/kg). ACT-451840 potently prevented male gamete formation from the gametocyte stage with a 50% inhibition concentration of 5.89 nM (SD: ± 1.80 nM) and dose-dependently blocked oocyst development in the mosquito with a 50% inhibitory concentration of 30 nM (range: 23–39). The compound’s preclinical safety profile is presented and is in line with the published results of the first-in-man study in healthy male participants, in whom ACT-451840 was well tolerated. Pharmacokinetic/pharmacodynamic (PK/PD) modeling was applied using efficacy in the murine models (defined either as antimalarial activity or as survival) in relation to area under the concentration versus time curve (AUC), maximum observed plasma concentration (Cmax), and time above a threshold concentration. The determination of the dose–efficacy relationship of ACT-451840 under curative conditions in rodent malaria models allowed prediction of the human efficacious exposure.

Conclusion

The dual activity of ACT-451840 against asexual and sexual stages of P. falciparum and the activity on P. vivax have the potential to meet the specific profile of a target compound that could replace the fast-acting artemisinin component and harbor additional gametocytocidal activity and, thereby, transmission-blocking properties. The fast parasite reduction ratio (PRR) and gametocytocidal effect of ACT-451840 were recently also confirmed in a clinical proof-of-concept (POC) study.

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<![CDATA[A Putative Small Solute Transporter Is Responsible for the Secretion of G377 and TRAP-Containing Secretory Vesicles during Plasmodium Gamete Egress and Sporozoite Motility]]> https://www.researchpad.co/article/5989da5eab0ee8fa60b9085a

Regulated protein secretion is required for malaria parasite life cycle progression and transmission between the mammalian host and mosquito vector. During transmission from the host to the vector, exocytosis of highly specialised secretory vesicles, such as osmiophilic bodies, is key to the dissolution of the red blood cell and parasitophorous vacuole membranes enabling gamete egress. The positioning of adhesins from the TRAP family, from micronemes to the sporozoite surface, is essential for gliding motility of the parasite and transmission from mosquito to mammalian host. Here we identify a conserved role for the putative pantothenate transporter PAT in Plasmodium berghei in vesicle fusion of two distinct classes of vesicles in gametocytes and sporozoites. PAT is a membrane component of osmiophilic bodies in gametocytes and micronemes in sporozoites. Despite normal formation and trafficking of osmiophilic bodies to the cell surface upon activation, PAT-deficient gametes fail to discharge their contents, remain intraerythrocytic and unavailable for fertilisation and further development in the mosquito. Sporozoites lacking PAT fail to secrete TRAP, are immotile and thus unable to infect the subsequent rodent host. Thus, P. berghei PAT appears to regulate exocytosis in two distinct populations of vesicles in two different life cycle forms rather than acting as pantothenic transporter during parasite transmission.

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<![CDATA[PLoS Pathogens Issue Image | Vol. 13(10) October 2017]]> https://www.researchpad.co/article/5ab1686e463d7e570730a8e1

Going bananas!

Plasmodium falciparum sexual stage gametocytes undergo remarkable transformations as they develop to maturity inside the red blood cell of their host. This dramatic metamorphosis is highlighted in these 3D rendered images from Serial Block Face Scanning Electron Microscopy (SBF-SEM). The formation of a membrane structure called the inner membrane complex (magenta) which drives the gametocyte elongation process. The nucleus (yellow), mitochondria (red) and red blood cell (dark red) are also rendered. Dixon et al.

Image Credit: Boyin Liu and Matthew Dixon, Department of Biochemistry and Molecular Biology, Bio21 Institute, University of Melbourne.

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<![CDATA[Plant-Mediated Effects on Mosquito Capacity to Transmit Human Malaria]]> https://www.researchpad.co/article/5989d9dfab0ee8fa60b69110

The ecological context in which mosquitoes and malaria parasites interact has received little attention, compared to the genetic and molecular aspects of malaria transmission. Plant nectar and fruits are important for the nutritional ecology of malaria vectors, but how the natural diversity of plant-derived sugar sources affects mosquito competence for malaria parasites is unclear. To test this, we infected Anopheles coluzzi, an important African malaria vector, with sympatric field isolates of Plasmodium falciparum, using direct membrane feeding assays. Through a series of experiments, we then examined the effects of sugar meals from Thevetia neriifolia and Barleria lupilina cuttings that included flowers, and fruit from Lannea microcarpa and Mangifera indica on parasite and mosquito traits that are key for determining the intensity of malaria transmission. We found that the source of plant sugar meal differentially affected infection prevalence and intensity, the development duration of the parasites, as well as the survival and fecundity of the vector. These effects are likely the result of complex interactions between toxic secondary metabolites and the nutritional quality of the plant sugar source, as well as of host resource availability and parasite growth. Using an epidemiological model, we show that plant sugar source can be a significant driver of malaria transmission dynamics, with some plant species exhibiting either transmission-reducing or -enhancing activities.

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<![CDATA[Open Source Drug Discovery with the Malaria Box Compound Collection for Neglected Diseases and Beyond]]> https://www.researchpad.co/article/5989da7aab0ee8fa60b980fe

A major cause of the paucity of new starting points for drug discovery is the lack of interaction between academia and industry. Much of the global resource in biology is present in universities, whereas the focus of medicinal chemistry is still largely within industry. Open source drug discovery, with sharing of information, is clearly a first step towards overcoming this gap. But the interface could especially be bridged through a scale-up of open sharing of physical compounds, which would accelerate the finding of new starting points for drug discovery. The Medicines for Malaria Venture Malaria Box is a collection of over 400 compounds representing families of structures identified in phenotypic screens of pharmaceutical and academic libraries against the Plasmodium falciparum malaria parasite. The set has now been distributed to almost 200 research groups globally in the last two years, with the only stipulation that information from the screens is deposited in the public domain. This paper reports for the first time on 236 screens that have been carried out against the Malaria Box and compares these results with 55 assays that were previously published, in a format that allows a meta-analysis of the combined dataset. The combined biochemical and cellular assays presented here suggest mechanisms of action for 135 (34%) of the compounds active in killing multiple life-cycle stages of the malaria parasite, including asexual blood, liver, gametocyte, gametes and insect ookinete stages. In addition, many compounds demonstrated activity against other pathogens, showing hits in assays with 16 protozoa, 7 helminths, 9 bacterial and mycobacterial species, the dengue fever mosquito vector, and the NCI60 human cancer cell line panel of 60 human tumor cell lines. Toxicological, pharmacokinetic and metabolic properties were collected on all the compounds, assisting in the selection of the most promising candidates for murine proof-of-concept experiments and medicinal chemistry programs. The data for all of these assays are presented and analyzed to show how outstanding leads for many indications can be selected. These results reveal the immense potential for translating the dispersed expertise in biological assays involving human pathogens into drug discovery starting points, by providing open access to new families of molecules, and emphasize how a small additional investment made to help acquire and distribute compounds, and sharing the data, can catalyze drug discovery for dozens of different indications. Another lesson is that when multiple screens from different groups are run on the same library, results can be integrated quickly to select the most valuable starting points for subsequent medicinal chemistry efforts.

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<![CDATA[Marked Rise in the Prevalence of Asymptomatic Plasmodium falciparum Infection in Rural Gabon]]> https://www.researchpad.co/article/5989da44ab0ee8fa60b8aef2

Control strategies implemented a decade ago led to a marked reduction in the prevalence of malaria in many countries. In Dienga, southeastern Gabon, the prevalence of microscopic P. falciparum infection was 7% in 2003, close to the pre-elimination threshold of 5%. The aim of this work was to determine the prevalence of P. falciparum infection in the same community a decade later. A cohort of 370 individuals aged from 3 to 85 years living in Dienga was investigated for P. falciparum infection; during six passages (P) in 15-month period. Demographic data were collected, along with behaviors and attitudes towards malaria. Plasmodium infection was diagnosed by microscopy (ME), followed by PCR to detect submicroscopic infection. The prevalence of P. falciparum infection in P1, P2, P3, P4, P5 and P6 was respectively 43.5% (25.1% ME+, 18.4% PCR+); 40.9% (27.0% ME+, 13.9% PCR+), 52.7% (26.1% ME+, 26.6% PCR+); 34.1% (14.1% ME+, 20% PCR+), 57.7% (25.4.% ME+, 32.3% PCR+); and 46.2% (21.4% ME+, 24.8% PCR+) with an overall average of 45.9% (95%CI [37.0–54.7], 23.2% ME+ and 22.7% PCR+). P4 and P5 prevalences were statically different throughout the six passages. Microscopic prevalence was significantly higher than that observed ten years ago (23% [n = 370] vs 7% [n = 323], p < 0.001). Asymptomatic infections were the most frequent (96%). Gametocytes were detected in levels ranging from 5.9% to 13.9%. Insecticide-treated nets, indoor residual insecticides, and self-medication were used by respectively 33.2% (95%CI [29.0–37.4]), 17.7% (95%CI [15.5–19.9]) and 12.1% (95%CI [10.6–13.6]) of the study population. A near-threefold increase in P. falciparum infection has been observed in a rural area of southeastern Gabon during a 10-year period. Most infections were asymptomatic, but these subjects likely represent a parasite reservoir. These findings call for urgent reinforcement of preventive measures.

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<![CDATA[Single dose primaquine to reduce gametocyte carriage and Plasmodium falciparum transmission in Cambodia: An open-label randomized trial]]> https://www.researchpad.co/article/5989db5dab0ee8fa60be03fd

Background

Single low dose primaquine (SLD PQ, 0.25mg/kg) is recommended in combination with artemisinin-based combination therapy (ACT) as a gametocytocide to prevent Plasmodium falciparum transmission in areas threatened by artemisinin resistance. To date, no randomized controlled trials have measured primaquine’s effect on infectiousness to Anopheline mosquitoes in Southeast Asia.

Methods

Cambodian adults with uncomplicated falciparum malaria were randomized to receive a single 45mg dose of primaquine (equivalent to three SLD PQ) or no primaquine after the third dose of dihydroartemisin-piperaquine (DHP) therapy. A membrane-feeding assay measured infectiousness to Anopheles dirus on days 0, 3, 7, and 14 of blood-stage therapy. Gametocytemia was evaluated by microscopy and reverse-transcriptase PCR.

Results

Prior to trial halt for poor DHP treatment efficacy, 101 participants were randomized and 50 received primaquine. Overall microscopic gametocyte prevalence was low (9%), but gametocytemic subjects given primaquine were gametocyte-free by day 14, and significantly less likely to harbor gametocytes by day 7 compared to those treated with DHP-alone, who remained gametocytemic for a median of two weeks. Only one infectious subject was randomized to the primaquine group, precluding assessment of transmission-blocking efficacy. However, he showed a two-fold reduction in oocyst density of infected mosquitoes less than 24 hours after primaquine dosing. In the DHP-alone group, four subjects remained infectious through day 14, infecting roughly the same number of mosquitoes pre and post-treatment. Overall, microscopic gametocytemia was an excellent predictor of infectiousness, and performed better than submicroscopic gametocytemia post-treatment, with none of 474 mosquitoes infected post-treatment arising from submicroscopic gametocytes.

Conclusions

In a setting of established ACT resistance, a single dose of 45mg primaquine added to DHP rapidly and significantly reduced gametocytemia, while DHP-alone failed to reduce gametocytemia and prevent malaria transmission to mosquitoes. Continued efforts to make single dose primaquine widely available are needed to help achieve malaria elimination.

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<![CDATA[Stage-Specific Changes in Plasmodium Metabolism Required for Differentiation and Adaptation to Different Host and Vector Environments]]> https://www.researchpad.co/article/5989da9eab0ee8fa60ba4c28

Malaria parasites (Plasmodium spp.) encounter markedly different (nutritional) environments during their complex life cycles in the mosquito and human hosts. Adaptation to these different host niches is associated with a dramatic rewiring of metabolism, from a highly glycolytic metabolism in the asexual blood stages to increased dependence on tricarboxylic acid (TCA) metabolism in mosquito stages. Here we have used stable isotope labelling, targeted metabolomics and reverse genetics to map stage-specific changes in Plasmodium berghei carbon metabolism and determine the functional significance of these changes on parasite survival in the blood and mosquito stages. We show that glutamine serves as the predominant input into TCA metabolism in both asexual and sexual blood stages and is important for complete male gametogenesis. Glutamine catabolism, as well as key reactions in intermediary metabolism and CoA synthesis are also essential for ookinete to oocyst transition in the mosquito. These data extend our knowledge of Plasmodium metabolism and point towards possible targets for transmission-blocking intervention strategies. Furthermore, they highlight significant metabolic differences between Plasmodium species which are not easily anticipated based on genomics or transcriptomics studies and underline the importance of integration of metabolomics data with other platforms in order to better inform drug discovery and design.

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