Number Of Merozoites Research Articles

Mathematical model and analysis for within-host dynamics of the malaria parasite infection with optimal control strategies

This study develops and analyzes the within-host dynamics of malaria parasite infection with optimal control strategies. We considered both the blood and liver stages of the malaria infection, taking into account the immunological response against the infection. The basic reproduction number, which indicates the potential spread of the parasite is evaluated. The parasite-free equilibrium point is locally and globally asymptotically stable when the basic reproduction is less than a unit. Sensitivity analysis reveals that the invention rate of red blood cells and the average number of merozoites per rupture infected red blood cell are the most influential parameters of the model. Furthermore, to investigate the most effective measurement of malaria parasite infection, we performed the optimal control strategies. The pre-erythrocytic vaccine, blood-stage vaccine, primary tissue schizontocides, blood schizontocides, and gametocytocidal drugs are incorporated as control measures. The controls are implemented to minimize the infected hepatocytes, infected red blood cells, gametocytes, and merozoites in the human host, as well as the associated costs. Pontryagin’s Minimum Principle is applied to establish optimal control strategies against infected red blood cells, infected hepatocytes, and malaria parasites. Several simulation situations are conducted to assess the analytical results and determine the effective control intervention measures. The results indicate that administering all four controls simultaneously would eradicate the prevalence of malaria infection in the human host.

A Mathematical Analysis of Competitive Dynamics and Aggressive Treatment in the Evolution of Drug Resistance in Malaria Parasites

Experimental evidence supports the counterintuitive notion that rapid eradication of pathogens within a host, infected with both drug-sensitive and -resistant malaria parasites, can actually accelerate the evolution of drug-resistant pathogens. This study aims to analyze the competitive dynamics between these two strains through a mathematical model and evaluate the impact of aggressive treatment on the spread of drug resistance. We conducted equilibrium, uncertainty, and sensitivity analyses to assess the model, identifying and measuring the influence of key factors on the outcome variable (the population of drug-resistant parasites). Both equilibrium and local sensitivity analyses concurred that the density of drug-resistant parasites is notably affected by genetic instability, the production rate of red blood cells, the number of merozoites, and competition factors. Conversely, there is a negative relationship between genetic instability and one of the competition coefficients. Global sensitivity analysis offers a comprehensive examination of the impact of each input parameter on the temporal propagation of drug resistance, effectively accounting for the interplay among parameters. Both local and global sensitivity analyses underscore the continuous impact of drug treatment on the progression of drug resistance over time. This paper anticipates exploring the underlying mechanisms of drug resistance and providing theoretical support for developing more effective drug treatment strategies.

A cell-level dynamical model for malaria parasite infection with antimalarial drug treatment

Malaria is an infectious disease caused by intracellular parasites of the genus Plasmodium. It is a major health problem around the world. In this study, a cell-level mathematical model of malaria parasites with antimalarial drug treatments is formulated and analyzed. The model consists of seven compartments for cell populations. We analyzed the qualitative behavior of the model using various techniques. The stability analysis of the parasite-free equilibrium is obtained, whereas it is locally and globally stable if the basic reproduction number R0<1. The parasite persistence equilibrium point exists, and it is locally asymptotically stable if R0>1. The sensitivity analysis of the basic reproduction number is computed, and the results show that the infection rate of the erythrocyte by merozoites, the average number of merozoites per ruptured infected erythrocyte cells, the natural death rate of merozoites, and the requirement rate of the uninfected erythrocyte are the most influential parameters within-host dynamics of malaria infection. Different numerical simulations are performed to supplement our analytical findings. The effect of primary tissue schizontocides, blood schizontocides, and gametocytocides on infected hepatocytes, infected erythrocytes, and gametocytes have been investigated, respectively. Finally, some counterplots are presented in order to investigate the impact of parameters on the basic reproduction number. The in-host basic reproduction number decreases as the antimalarial treatment administration increases. Therefore, increasing antimalarial treatment administration is the best way to mitigate the in-host malaria infection.

Ponazuril: Clinical efficacy, ultrastructure, and histopathology studies of in vivo anticoccidial action against Eimeria tenella

Ponazuril, a novel antiprotozoal drug in the class of triazine, has shown a promising application on apicomplexan infections in poultry and livestock. However, the effect and mechanism of action of ponazuril against Eimeria tenella (E. tenella) are unclear. The efficacy against E. tenella was initially studied by administering different doses of ponazuril in drinking water. The treated stage and site of ponazuril on E. tenella were observed through ultrastructural and histopathological analyses. Chicks were orally treated with a dose of 15 mg/kg body weight of ponazuril at different endogenous stages of E. tenella post-infection. According to the clinical study, the values of anticoccidial indices (ACI) were 157.0, 162.3, 196.9, 194.5, and 190.9, respectively, when the ponazuril was administered in drinking water at doses of 5, 10, 20, 40, and 50 mg/L for two consecutive days after infection. Among them, the 20 mg/L ponazuril group showed the best anticoccidial effect, which was superior to that of the toltrazuril treatment group, with an ACI value of 191.7. Histological analysis indicated that ponazuril effectively relieved cecal lesions, and decreased the number of merozoites. Transmission electron micrographs (TEM) observed that merozoites became irregular in shape, and some apparent protrusions of the outer membrane were presented especially the second-generation merozoites. Additionally, abnormalities in the development of WFBI and WFBII in the macrogametocyte were observed, which may affect the formation of the ovule wall. Moreover, merozoites in the treated group showed uneven and marginalized chromatin and mitochondrial swelling. These results suggested ponazuril is a potential anticoccidial drug, providing information on the mechanism of anticoccidial effects.

Differential Gene Expression of Malaria Parasite in Response to Red Blood Cell-Specific Glycolytic Intermediate 2,3-Diphosphoglycerate (2,3-DPG)

Innovative strategies to control malaria are urgently needed. Exploring the interplay between Plasmodium sp. parasites and host red blood cells (RBCs) offers opportunities for novel antimalarial interventions. Pyruvate kinase deficiency (PKD), characterized by heightened 2,3-diphosphoglycerate (2,3-DPG) concentration, has been associated with protection against malaria. Elevated levels of 2,3-DPG, a specific mammalian metabolite, may hinder glycolysis, prompting us to hypothesize its potential contribution to PKD-mediated protection. We investigated the impact of the extracellular supplementation of 2,3-DPG on the Plasmodium falciparum intraerythrocytic developmental cycle in vitro. The results showed an inhibition of parasite growth, resulting from significantly fewer progeny from 2,3-DPG-treated parasites. We analyzed differential gene expression and the transcriptomic profile of P. falciparum trophozoites, from in vitro cultures subjected or not subjected to the action of 2,3-DPG, using Nanopore Sequencing Technology. The presence of 2,3-DPG in the culture medium was associated with the significant differential expression of 71 genes, mostly associated with the GO terms nucleic acid binding, transcription or monoatomic anion channel. Further, several genes related to cell cycle control were downregulated in treated parasites. These findings suggest that the presence of this RBC-specific glycolytic metabolite impacts the expression of genes transcribed during the parasite trophozoite stage and the number of merozoites released from individual schizonts, which supports the potential role of 2,3-DPG in the mechanism of protection against malaria by PKD.

What exactly does the PfK13 C580Y mutation in Plasmodium falciparum influence?

BackgroundThe emergence and spread of artemisinin resistance threaten global malaria control and elimination goals, and encourage research on the mechanisms of drug resistance in malaria parasites. Mutations in Plasmodium falciparum Kelch 13 (PfK13) protein are associated with artemisinin resistance, but the unique or common mechanism which results in this resistance is unclear.MethodsWe analyzed the effects of the PfK13 mutation on the transcriptome and proteome of P. falciparum at different developmental stages. Additionally, the number of merozoites, hemozoin amount, and growth of P. falciparum 3D7C580Y and P. falciparum 3D7WT were compared. The impact of iron supplementation on the number of merozoites of P. falciparum 3D7C580Y was also examined.ResultsWe found that the PfK13 mutation did not significantly change glycolysis, TCA, pentose phosphate pathway, or oxidative phosphorylation, but did reduce the expression of reproduction- and DNA synthesis-related genes. The reduced number of merozoites, decreased level of hemozoin, and slowed growth of P. falciparum 3D7C580Y were consistent with these changes. Furthermore, adding iron supply could increase the number of the merozoites of P. falciparum 3D7C580Y.ConclusionsThese results revealed that the PfK13 mutation reduced hemoglobin ingestion, leading to artemisinin resistance, likely by decreasing the parasites' requirement for haem and iron. This study helps elucidate the mechanism of artemisinin resistance due to PfK13 mutations.Graphical

Extraordinary high level of propagation of Babesia divergens in severe human babesiosis.

Babesias are obligate apicomplexan parasites that affect the red blood cells (RBCs) of animals. Humans can serve as accidental hosts for them. Asexual reproduction of a parasite occurs in a vertebrate host through asynchronous binary fission, yielding a complex pleomorphic population of intraerythrocytic forms. In natural hosts (Bos taurus), paired pyriforms (‘figure 8’) of Babesia divergens are usual, but tetrads (‘Maltese Cross’) are very rare (only in 0.02% infected erythrocytes); in humans, however, up to 5% of infected erythrocytes show tetrads. The current study shows that B. divergens proliferating in an accidental human host can promote extraordinarily high level of fission. This phenomenon is expressed as the simultaneous division of the parasite into 6 and possibly a greater number of merozoites, forming a ‘daisy head’ (vs the usual 2, less often 4 merozoites). Reproduction is possible without egressing merozoites from the erythrocyte, which results in multi-occupancy of an RBC (≥5 parasites per RBC). An unusually high polyparasitism – up to 14 parasites developed in the affected erythrocytes – was observed. This phenomenon is rare in natural hosts (usually ≤5), but when B. divergens is cultured in vitro it can be 10–12.

Overexpression of Plasmodium falciparum M1 Aminopeptidase Promotes an Increase in Intracellular Proteolysis and Modifies the Asexual Erythrocytic Cycle Development.

Plasmodium falciparum, the most virulent of the human malaria parasite, is responsible for high mortality rates worldwide. We studied the M1 alanyl-aminopeptidase of this protozoan (PfA-M1), which is involved in the final stages of hemoglobin cleavage, an essential process for parasite survival. Aiming to help in the rational development of drugs against this target, we developed a new strain of P. falciparum overexpressing PfA-M1 without the signal peptide (overPfA-M1). The overPfA-M1 parasites showed a 2.5-fold increase in proteolytic activity toward the fluorogenic substrate alanyl-7-amido-4-methylcoumarin, in relation to the wild-type group. Inhibition studies showed that overPfA-M1 presented a lower sensitivity against the metalloaminopeptidase inhibitor bestatin and to other recombinant PfA-M1 inhibitors, in comparison with the wild-type strain, indicating that PfA-M1 is a target for the in vitro antimalarial activity of these compounds. Moreover, overPfA-M1 parasites present a decreased in vitro growth, showing a reduced number of merozoites per schizont, and also a decrease in the iRBC area occupied by the parasite in trophozoite and schizont forms when compared to the controls. Interestingly, the transgenic parasite displays an increase in the aminopeptidase activity toward Met-, Ala-, Leu- and Arg-7-amido-4-methylcoumarin. We also investigated the potential role of calmodulin and cysteine proteases in PfA-M1 activity. Taken together, our data show that the overexpression of PfA-M1 in the parasite cytosol can be a suitable tool for the screening of antimalarials in specific high-throughput assays and may be used for the identification of intracellular molecular partners that modulate their activity in P. falciparum.

Lessons Learned for Pathogenesis, Immunology, and Disease of Erythrocytic Parasites: Plasmodium and Babesia.

Malaria caused by Plasmodium species and transmitted by Anopheles mosquitoes affects large human populations, while Ixodes ticks transmit Babesia species and cause babesiosis. Babesiosis in animals has been known as an economic drain, and human disease has also emerged as a serious healthcare problem in the last 20–30 years. There is limited literature available regarding pathogenesis, immunity, and disease caused by Babesia spp. with their genomes sequenced only in the last decade. Therefore, using previous studies on Plasmodium as the foundation, we have compared similarities and differences in the pathogenesis of Babesia and host immune responses. Sexual life cycles of these two hemoparasites in their respective vectors are quite similar. An adult Anopheles female can take blood meal several times in its life such that it can both acquire and transmit Plasmodia to hosts. Since each tick stage takes blood meal only once, transstadial horizontal transmission from larva to nymph or nymph to adult is essential for the release of Babesia into the host. The initiation of the asexual cycle of these parasites is different because Plasmodium sporozoites need to infect hepatocytes before egressed merozoites can infect erythrocytes, while Babesia sporozoites are known to enter the erythrocytic cycle directly. Plasmodium metabolism, as determined by its two- to threefold larger genome than different Babesia, is more complex. Plasmodium replication occurs in parasitophorous vacuole (PV) within the host cells, and a relatively large number of merozoites are released from each infected RBC after schizogony. The Babesia erythrocytic cycle lacks both PV and schizogony. Cytoadherence that allows the sequestration of Plasmodia, primarily P. falciparum in different organs facilitated by prominent adhesins, has not been documented for Babesia yet. Inflammatory immune responses contribute to the severity of malaria and babesiosis. Antibodies appear to play only a minor role in the resolution of these diseases; however, cellular and innate immunity are critical for the clearance of both pathogens. Inflammatory immune responses affect the severity of both diseases. Macrophages facilitate the resolution of both infections and also offer cross-protection against related protozoa. Although the immunosuppression of adaptive immune responses by these parasites does not seem to affect their own clearance, it significantly exacerbates diseases caused by coinfecting bacteria during coinfections.

How Many Is Enough? - Challenges of Multinucleated Cell Division in Malaria Parasites.

Regulating the number of progeny generated by replicative cell cycles is critical for any organism to best adapt to its environment. Classically, the decision whether to divide further is made after cell division is completed by cytokinesis and can be triggered by intrinsic or extrinsic factors. Contrarily, cell cycles of some species, such as the malaria-causing parasites, go through multinucleated cell stages. Hence, their number of progeny is determined prior to the completion of cell division. This should fundamentally affect how the process is regulated and raises questions about advantages and challenges of multinucleation in eukaryotes. Throughout their life cycle Plasmodium spp. parasites undergo four phases of extensive proliferation, which differ over three orders of magnitude in the amount of daughter cells that are produced by a single progenitor. Even during the asexual blood stage proliferation parasites can produce very variable numbers of progeny within one replicative cycle. Here, we review the few factors that have been shown to affect those numbers. We further provide a comparative quantification of merozoite numbers in several P. knowlesi and P. falciparum parasite strains, and we discuss the general processes that may regulate progeny number in the context of host-parasite interactions. Finally, we provide a perspective of the critical knowledge gaps hindering our understanding of the molecular mechanisms underlying this exciting and atypical mode of parasite multiplication.

A Plasmodium berghei putative serine-threonine kinase 2 (PBANKA_0311400) is required for late liver stage development and timely initiation of blood stage infection.

ABSTRACTIn Plasmodium, protein kinases govern key biological processes of the parasite life cycle involved in the establishment of infection, dissemination and sexual reproduction. The rodent malaria model Plasmodium berghei encodes for 66 putative eukaryotic protein kinases (ePKs) as identified through modelling domain signatures and are highly conserved in Plasmodium falciparum. We report here the functional characterisation of a putative serine-threonine kinase PBANKA_0311400 identified in this kinome analysis and designate it as Pbstk2. To elucidate its role, we knocked out Pbstk2 locus and performed a detailed phenotypic analysis at different life cycle stages. The Pbstk2 knockout (KO) was not compromised in asexual blood stage propagation, transmission and development in the mosquito vector. The Pbstk2 KO produced viable salivary gland sporozoites that successfully transformed into exo-erythrocytic forms (EEFs) and were morphologically indistinguishable from wild-type GFP (WT GFP) with regard to size and shape until 48 h. An intravenous dose of 1×103 Pbstk2 KO sporozoites in C57BL/6 mice failed to establish blood stage infection and a higher dose of 5X103 showed a 2–3 day delay in prepatency as compared to WT GFP parasites. Consistent with such an observation, analysis of in vitro EEF development at 62 h revealed that the hepatic merozoite numbers were reduced to nearly 40% as compared to WT GFP and showed meagre expression of MSP1. Our studies provide evidence for the role of PbSTK2 in late liver stage development and for the successful establishment of a timely blood stage infection.

Plasticity and genetic variation in traits underpinning asexual replication of the rodent malaria parasite, Plasmodium chabaudi

BackgroundThe ability of malaria (Plasmodium) parasites to adjust investment into sexual transmission stages versus asexually replicating stages is well known, but plasticity in other traits underpinning the replication rate of asexual stages in the blood has received less attention. Such traits include burst size (the number of merozoites produced per schizont), the duration of the asexual cycle, and invasion preference for different ages of red blood cell (RBC).MethodsHere, plasticity [environment (E) effects] and genetic variation [genotype (G) effects] in traits relating to asexual replication rate are examined for 4 genotypes of the rodent malaria parasite Plasmodium chabaudi. An experiment tested whether asexual dynamics differ between parasites infecting control versus anaemic hosts, and whether variation in replication rate can be explained by differences in burst size, asexual cycle, and invasion rates.ResultsThe within-host environment affected each trait to different extents but generally had similar impacts across genotypes. The dynamics of asexual densities exhibited a genotype by environment effect (G×E), in which one of the genotypes increased replication rate more than the others in anaemic hosts. Burst size and cycle duration varied between the genotypes (G), while burst size increased and cycle duration became longer in anaemic hosts (E). Variation in invasion rates of differently aged RBCs was not explained by environmental or genetic effects. Plasticity in burst size and genotype are the only traits making significant contributions to the increase in asexual densities observed in anaemic hosts, together explaining 46.4% of the variation in replication rate.ConclusionsThat host anaemia induces several species of malaria parasites to alter conversion rate is well documented. Here, previously unknown plasticity in other traits underpinning asexual replication is revealed. These findings contribute to mounting evidence that malaria parasites deploy a suite of sophisticated strategies to maximize fitness by coping with, or exploiting the opportunities provided by, the variable within-host conditions experienced during infections. That genetic variation and genotype by environment interactions also shape these traits highlights their evolutionary potential. Asexual replication rate is a major determinant of virulence and so, understanding the evolution of virulence requires knowledge of the ecological (within-host environment) and genetic drivers of variation among parasites.

Kinetics of the invasion and egress processes of Babesia divergens, observed by time-lapse video microscopy

Based on confocal fluorescence and bright field video microscopy, we present detailed observations on the processes of invasion and egress of erythrocytes by the apicomplexan parasite Babesia divergens. Time-lapse images reveal numerous unexpected findings associated with the dynamics of B. divergens and its ability to manipulate the erythrocyte during both processes in its asexual cycle under in vitro conditions. Despite the speed at which these processes occur and the small size of the parasite, we capture infective merozoites moving vigorously and causing striking deformations in the erythrocyte’s plasma membrane during an active invasion. We also observed intraerythrocytic dynamic stages as paired pyriforms, double paired pyriforms, tetrads, unattached pyriform sister cells and multiple parasite stages resulting in the release of large numbers of merozoites over a short period. Of considerable interest is that time-lapse images reveal a novel mechanism of egress used by B. divergens to exit the human erythrocyte. The release occurs when B. divergens parasites establish contacts with the plasma membrane of the erythrocyte from within, before exiting the cell. Visualization and analysis of the images enabled us to obtain useful information and broaden our knowledge of complex and crucial events involved with parasitisation of human erythrocytes by B. divergens.

Effect of dihydroartemisinin at low concentration on intervention of Plasmodium falciparum 3D7 strain

Malaria is still the most severe strain of the human malaria parasites, and malaria disease is life-threatening which can result in severe anemia and cerebral malaria, especially in children in tropical Africa. Previous studies have shown that artemisinin and its derivatives could selectively kill erythrocytic stage of malaria and have a greater impact on the ring period. In recent years, there have been new findings of its mechanism continually. However, the concentration of artemisinin and its derivatives used in these studies can reach 50 to 80 times the half-inhibitory concentration in vitro. In this study, the international standard strain 3D7 of Plasmodium falciparum was used to culture in vitro. After half-inhibitory concentration of dihydroartemisinin was treated, the morphological changes of P. falciparum intraerythrocytic stage were observed, and then the 3D7 life cycle and effects of different developmental stages after dosing was explored. The 3D7 strain of P. falciparum was continuously synchronised more than 3 times. And dihydroartemisinin (DHA) at half maximal inhibitory concentration (10 nmol·L⁻¹) was administered for 6 hours after the last synchronization, and 3 life cycles were continuously observed (132 h). The results showed that compared with the parasites untreated by DHA, there was a noticeable delay in the life cycle of at least 36 h, indicating that the growth of 3D7 was significantly inhibited by DHA (P<0.001), and the rate of ring formation was significantly reduced (P<0.05). The trophozoites were abnormal in shape, such as shrink in size, and the number of merozoites in schizonts was significantly decreased (P<0.05). These results suggested that non-killing concentrations of DHA (meaning parasites can be inhibited but not killed) can significantly inhibit the growth of P. falciparum, which may not only affect the ring stage, but also have an impact on other stages of the P. falciparum.

Sensitivity analysis for a within-human-host immuno-pathogenesis dynamics of Plasmodium falciparum parasites

Sensitivity analysis has become increasingly useful in many fields of engineering and sciences. Researchers use sensitivity and uncertainty analysis in the mathematical modelling of biological phenomena because of its value in identifying essential parameters for model's output. Moreover, it can help in the process of experimental analysis, model order reduction, parameter estimation, decision making or development of recommendations for decision makers. Here, we demonstrate the use of local sensitivity analysis to understand the influence of different parameters on a threshold parameter, R_0^I, resulting from the analysis of a within human-host model for the dynamics of malaria parasites. %We highlight the different methods used in sensitivity analysis.Our results reveal that the obtained R_0^I is most sensitive to the infection rate of healthy red blood cells (RBCs) by merozoites, the average number of merozoites released per bursting parasitized RBCs, the proportion of parasitized RBCs that continue asexual reproduction and the per capita natural death rate of merozoites.

Critical role of Erythrocyte Binding-Like protein of the rodent malaria parasite Plasmodium yoelii to establish an irreversible connection with the erythrocyte during invasion

Plasmodium malaria parasites multiply within erythrocytes and possess a repertoire of proteins whose function is to recognize and invade these vertebrate host cells. One such protein involved in erythrocyte invasion is the micronemal protein, Erythrocyte Binding-Like (EBL), which has been studied as a potential target of vaccine development in Plasmodium vivax (PvDBP) and Plasmodium falciparum (EBA-175). In the rodent malaria parasite model Plasmodium yoelii, specific substitutions in the EBL regions responsible for intracellular trafficking (17XL parasite line) or receptor recognition (17X1.1pp. parasite line), paradoxically increase invasion ability and virulence rather than abolish EBL function. Attempts to disrupt the ebl gene locus in the 17XL and 17XNL lines were unsuccessful, suggesting EBL essentiality. To understand the mechanisms behind these potentially conflicting outcomes, we generated 17XL-based transfectants in which ebl expression is suppressed with anhydrotetracycline (ATc) and investigated merozoite behavior during erythrocyte invasion. In the absence of ATc, EBL was secreted to the merozoite surface, whereas following ATc administration parasitemia was negligible in vivo. Merozoites lacking EBL were unable to invade erythrocytes in vitro, indicating that EBL has a critical role for erythrocyte invasion. Quantitative time-lapse imaging revealed that with ATc administration a significant number of merozoites were detached from the erythrocyte after the erythrocyte deformation event and no echinocytosis was observed, indicating that EBL is required for merozoites to establish an irreversible connection with erythrocytes during invasion.

Mathematical Model for Hepatocytic-Erythrocytic Dynamics of Malaria

Human malaria remains a major killer disease worldwide, with nearly half (3.2 billion) of the world’s population at risk of malaria infection. The infectious protozoan disease is endemic in tropical and subtropical regions, with an estimated 212 million new cases and 429,000 malaria-related deaths in 2015. An in-host mathematical model ofPlasmodium falciparummalaria that describes the dynamics and interactions of malaria parasites with the host’s liver cells (hepatocytic stage), the red blood cells (erythrocytic stage), and macrophages is reformulated. By a theoretical analysis, an in-host basic reproduction numberR0is derived. The disease-free equilibrium is shown to be locally and globally asymptotically stable. Sensitivity analysis reveals that the erythrocyte invasion rateβr, the average number of merozoites released per bursting infected erythrocyteK, and the proportion of merozoites that cause secondary invasions at the blood phaseζare the most influential parameters in determining the malaria infection outcomes. Numerical results show that macrophages have a considerable impact in clearing infected red blood cells through phagocytosis. Moreover, the density of infected erythrocytes and hence the severity of malaria are shown to increase with increasing density of merozoites in the blood. Concurrent use of antimalarial drugs and a potential erythrocyte invasion-avoidance vaccine would minimize the density of infected erythrocytes and hence malaria disease severity.

Modulation of cholesterol-related sterols during Eimeria bovis macromeront formation and impact of selected oxysterols on parasite development

Obligate intracellular apicomplexan parasites are considered as deficient in cholesterol biosynthesis and scavenge cholesterol from their host cell in a parasite-specific manner. Compared to fast proliferating apicomplexan species producing low numbers of merozoites per host cell, (e. g. Toxoplasma gondii), the macromeront-forming protozoa Eimeria bovis is in extraordinary need for cholesterol for offspring production (≥ 170,000 merozoites I/macromeront). Interestingly, optimized in vitro E. bovis merozoite I production occurs under low foetal calf serum (FCS, 1.2%) supplementation. To analyze the impact of extensive E. bovis proliferation on host cellular sterol metabolism we here compared the sterol profiles of E. bovis-infected primary endothelial host cells grown under optimized (1.2% FCS) and non-optimized (10% FCS) cell culture conditions. Therefore, several sterols indicating endogenous de novo cholesterol synthesis, cholesterol conversion and sterol uptake (phytosterols) were analyzed via GC–MS-based approaches. Overall, significantly enhanced levels of phytosterols were detected in both FCS conditions indicating infection-triggered sterol uptake from extracellular sources as a major pathway of sterol acquisition. Interestingly, a simultaneous induction of endogenous cholesterol synthesis based on increased levels of distinct cholesterol precursors was only observed in case of optimized parasite proliferation indicating a parasite proliferation-dependent effect. Considering side-chain oxysterols, 25 hydroxycholesterol levels were selectively found increased in E. bovis-infected host cells, while 24 hydroxycholesterol and 27 hydroxycholesterol contents were not significantly altered by infection. Exogenous treatments with 25 hydroxycholesterol, 27 hydroxycholesterol, and 7 ketocholesterol revealed significant adverse effects on E. bovis intracellular development. Thus, the number and size of developing macromeronts and merozoite I production was significantly reduced indicating that these oxysterols bear direct or indirect antiparasitic properties. Overall, the current data indicate parasite-driven changes in the host cellular sterol profile reflecting the huge demand of E. bovis for cholesterol during macromeront formation and its versatility in the acquisition of cholesterol sources.

Intestinal Life Cycle of Eimeria caliginosa (Apicomplexa: Eimeriidae) From the Dusky Rice Rat, Melanomys caliginosus (Rodentia: Cricetidae: Sigmodontinae), In Costa Rica

The endogenous life cycle of Eimeria caliginosa was studied in experimentally infected dusky rice rats, Melanomys caliginosus. All endogenous stages were located in the epithelial cells of villi in the small intestine. Both Giemsa-stained mucosal scrapings and histological sections were studied for the diagnosis of all the life-cycle stages. Eimeria caliginosa has 3 generations of meronts (M) that differ by size, shape, and number of merozoites (m), which also differ in their size, shape, and location of their nuclei within the cytoplasm of the meronts. The 3 meront types, M1-M3, respectively, had 20-33 (m1), 5-9 (m2), and 13-16 (m3) merozoites. Macrogametocytes and microgametocytes, as well as macrogametes and microgametes, completed the sexual cycle, which concludes with the formation of unsporulated oocysts. This parasite's endogenous development produced severe intestinal lesions in the experimentally infected dusky rice rats. In our ongoing work to understand the biodiversity present in plants and animals of the protected Reserva Biológica Alberto Manuel Brenes (ReBAMB) field station in Costa Rica, we now have discovered 3 new Eimeria species, and this is the second complete life cycle in which we document both the asexual and sexual stages.

Mathematical Model for the In-Host Malaria Dynamics Subject to Malaria Vaccines

Despite the success of the existing malaria control strategies, reported malaria cases are still quite high. In 2016, the WHO reported about 216 million malaria cases; 90% of which occurred in the WHO African Region. In this paper, a mathematical model for the in-host Plasmodium falciparum malaria subject to malaria vaccines is formulated and analysed. An efficacious pre-erythrocytic vaccine is shown to greatly reduce the severity of clinical malaria. Based on the normalized forward sensitivity index technique, the average number of merozoites released per bursting blood schizont is shown to be the most sensitive parameter in the model. Numerical simulation results further suggest that an efficacious blood stage vaccine has the potential to reduce the burst size of the blood schizonts and maximize the rate of activation of CD8+ T cells during malaria infection. Moreover, vaccine combinations that are efficacious might help in achieving a malaria free population by the year 2030. This paper provides useful insights in malaria vaccine control and a unique opportunity to intensify support and funding for malaria vaccine development.