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In this study, a determination of Troponin I and creatine kinase activity in whole-blood samples in a cohort of 100 small infants in the age of 2-5 years from Uganda with complicated Plasmodium falciparum malaria suggests the prevalence of cardiac symptoms in comparison to non-infected, healthy patients. Troponin I and creatine kinase activity increased during infection. Different reports showed that complicated malaria coincides with hypoxia in children. The obtained clinical data prompted us to further elucidate the underlying regulatory mechanisms of cardiac involvement in human cardiac ventricular myocytes. Complicated malaria is the most common clinical presentation and might induce cardiac impairment by hypoxia. Eukaryotic initiation factor 5A (eIF-5A) is involved in hypoxia induced factor (HIF-1α) expression. EIF-5A is a protein posttranslationally modified by hypusination involving catalysis of the two enzymes deoxyhypusine synthase (DHS) and deoxyhypusine hydroxylase. Treatment of human cardiomyocytes with GC7, an inhibitor of DHS, catalyzing the first step in hypusine biosynthesis led to a decrease in proinflammatory and proapoptotic myocardial caspase-1 activity in comparison to untreated cardiomyocytes. This effect was even more pronounced after co-administration of GC7 and GPI from P. falciparum simulating the pathology of severe malaria. Moreover, in comparison to untreated and GC7-treated cardiomyocytes, co-administration of GC7 and GPI significantly decreased the release of cytochrome C and lactate from damaged mitochondria. In sum, coadministration of GC7 prevented cardiac damage driven by hypoxia in vitro. Our approach demonstrates the potential of the pharmacological inhibitor GC7 to ameliorate apoptosis in cardiomyocytes in an in vitro model simulating severe malaria. This regulatory mechanism is based on blocking EIF-5A hypusination.

Antimalarial drugs are vital for treating malaria and controlling transmission. Measuring drug efficacy in the field requires large clinical trials and thus we have identified proxy measures of drug efficacy such as the parasite clearance curve. This is often assumed to measure the rate of drug activity against parasites and is used to predict optimal treatment regimens required to completely clear a blood-stage infection. We discuss evidence that the clearance curve is not measuring the rate of drug killing. This has major implications for how we assess optimal treatment regimens, as well as how we prioritise new drugs in the drug development pipeline., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Plasmodium falciparum resistance against artemisinin has not emerged in Africa; however, there are reports of the presence of polymerase chain reaction-determined residual submicroscopic parasitaemia detected on day 3 after artemisinin-based combination therapy (ACT). These residual submicroscopic parasites are thought to represent tolerant/resistant parasites against artemisinin, the fast-acting component of the combination. This review focused on residual submicroscopic parasitaemia, what it represents, and its significance on the emergence and spread of artemisinin resistance in Africa. Presence of residual submicroscopic parasitemia on day 3 after treatment initiation leaves question on whether successful treatment is attained with ACT. Thus there is a need to determine the potential public health implication of the PCR-determined residual submicroscopic parasitaemia observed on day 3 after ACT. Robust techniques, such as in vitro cultivation, should be used to evaluate if the residual submicroscopic parasites detected on day 3 after ACT are viable asexual parasites, or gametocytes, or the DNA of the dead parasites waiting to be cleared from the circulation. Such techniques would also evaluate the transmissibility of these residual parasites.

Malaria and lymphatic filariasis (LF) are two leading and common mosquito-borne parasitic diseases worldwide. These two diseases are co-endemic in many tropical and sub-tropical regions and are known to share vectors. The interactions between malaria and filarial parasites are poorly understood. Thus, this study aimed at establishing the interactions that occur between Brugia pahangi and Plasmodium berghei ANKA (PbA) co-infection in gerbils. Briefly, the gerbils were matched according to age, sex, and weight and grouped into filarial-only infection, PbA-only infection, co-infection, and control group. The parasitemia, survival and clinical assessment of the gerbils were monitored for a period of 30 days post Plasmodium infection. The immune responses of gerbils to both mono and co-infection were monitored. Findings show that co-infected gerbils have higher survival rate than PbA-infected gerbils. Food and water consumption were significantly reduced in both PbA-infected and co-infected gerbils, although loss of body weight, hypothermia, and anemia were less severe in co-infected gerbils. Plasmodium-infected gerbils also suffered hypoglycemia, which was not observed in co-infected gerbils. Furthermore, gerbil cytokine responses to co-infection were significantly higher than PbA-only-infected gerbils, which is being suggested as a factor for their increased longevity. Co-infected gerbils had significantly elicited interleukin-4, interferon-gamma, and tumor necrotic factor at early stage of infection than PbA-infected gerbils. Findings from this study suggest that B. pahangi infection protect against severe anemia and hypoglycemia, which are manifestations of PbA infection.

Cytauxzoon felis is a pathogen responsible for cytauxzoonosis, a highly fatal disease in domestic cats. Although most studies of C. felis have focused on this parasite in domestic cats, bobcats are the reservoir host. In stark contrast, there is little information relative to the progression of C. felis infections in bobcats. We studied bobcats in southern Illinois during 2014-2017 to evaluate which environmental factors (i.e., ambient temperature; number of daylight hours; trapping year, month, and day) influenced C. felis parasitemia levels. Mean ambient temperature at 1 wk and 2 wk prior to sampling was associated with increased parasitemia levels. Vector activity intensifies with higher temperatures, suggesting that increased parasitemia levels are an adaptation to facilitate transmission., (© American Society of Parasitologists 2020.)

Background: Owing to the large amount of host DNA in clinical samples, generation of high-quality Plasmodium falciparum whole genome sequencing (WGS) data requires enrichment for parasite DNA. Enrichment is often achieved by leukocyte depletion of infected blood prior to storage. However, leukocyte depletion is difficult in low-resource settings and limits analysis to prospectively-collected samples. As a result, approaches such as selective whole genome amplification (sWGA) are being used to enrich for parasite DNA. However, sWGA has had limited success in generating reliable sequencing data from low parasitaemia samples. In this study, enzymatic digestion with MspJI prior to sWGA and whole genome sequencing was evaluated to determine whether this approach improved genome coverage compared to sWGA alone. The potential of sWGA to cause amplification bias in polyclonal infections was also examined., Methods: DNA extracted from laboratory-created dried blood spots was treated with a modification-dependent restriction endonuclease, MspJI, and filtered via vacuum filtration. Samples were then selectively amplified using a previously reported sWGA protocol and subjected to WGS. Genome coverage statistics were compared between the optimized sWGA approach and the previously reported sWGA approach performed in parallel. Differential amplification by sWGA was assessed by comparing WGS data generated from lab-created mixtures of parasite isolates, from the same geographical region, generated with or without sWGA., Results: MspJI digestion did not enrich for parasite DNA. Samples that underwent vacuum filtration (without MspJI digestion) prior to sWGA had the highest parasite DNA concentration and displayed greater genome coverage compared to MspJI + sWGA and sWGA alone, particularly for low parasitaemia samples. The optimized sWGA (filtration + sWGA) approach was successfully used to generate WGS data from 218 non-leukocyte depleted field samples from Malawi. Sequences from lab-created mixtures of parasites did not show evidence of differential amplification of parasite strains compared to directly sequenced samples., Conclusion: This optimized sWGA approach is a reliable method to obtain WGS data from non-leukocyte depleted, low parasitaemia samples. The absence of amplification bias in data generated from mixtures of isolates from the same geographic region suggests that this approach can be appropriately used for molecular epidemiological studies.

Background: Growth rate of malaria parasites in the blood of infected subjects is an important measure of efficacy of drugs and vaccines., Methods: We used log-linear and sine-wave models to estimate the parasite growth rate of the 3D7 strain of Plasmodium falciparum using data from 177 subjects from 14 induced blood stage malaria (IBSM) studies conducted at QIMR Berghofer. We estimated parasite multiplication rate per 48 hours (PMR48), PMR per life-cycle (PMRLC), and parasite life-cycle duration. We compared these parameters to those from studies conducted elsewhere with infections induced by IBSM (n = 66), sporozoites via mosquito bite (n = 336), or injection (n = 51)., Results: The parasite growth rate of 3D7 in QIMR Berghofer studies was 0.75/day (95% confidence interval [CI], .73-.77/day), PMR48 was 31.9 (95% CI, 28.7-35.4), PMRLC was 16.4 (95% CI, 15.1-17.8), and parasite life-cycle was 38.8 hours (95% CI, 38.3-39.2 hours). These parameters were similar to estimates from IBSM studies elsewhere (0.71/day, 95% CI, .67-.75/day; PMR48 26.6, 95% CI, 22.2-31.8) but significantly higher (P < .001) than in sporozoite studies (0.47/day, 95% CI, .43-.50/day; PMR48 8.6, 95% CI, 7.3-10.1)., Conclusions: Parasite growth rates were similar across different IBSM studies and higher than infections induced by sporozoite., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)

Background: Plasmodium malariae is considered a minor malaria parasite, although its global disease burden is underappreciated. The aim of this study was to develop an induced blood-stage malaria (IBSM) model of P. malariae to study parasite biology, diagnostic assays, and treatment., Methods: This clinical trial involved 2 healthy subjects who were intravenously inoculated with cryopreserved P. malariae-infected erythrocytes. Subjects were treated with artemether-lumefantrine after development of clinical symptoms. Prior to antimalarial therapy, mosquito-feeding assays were performed to investigate transmission, and blood samples were collected for rapid diagnostic testing and parasite transcription profiling. Serial blood samples were collected for biomarker analysis., Results: Both subjects experienced symptoms and signs typical of early malaria. Parasitemia was detected 7 days after inoculation, and parasite concentrations increased until antimalarial treatment was initiated 25 and 21 days after inoculation for subjects 1 and 2 respectively (peak parasitemia levels, 174 182 and 50 291 parasites/mL, respectively). The parasite clearance half-life following artemether-lumefantrine treatment was 6.7 hours. Mosquito transmission was observed for 1 subject, while in vivo parasite transcription and biomarkers were successfully profiled., Conclusions: An IBSM model of P. malariae has been successfully developed and may be used to study the biology of, diagnostic testing for, and treatment of this neglected malaria species., Clinical Trials Registration: ACTRN12617000048381., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)

Background: This study determined the rate of mother-to-child transmission (MTCT) of HIV among HIV positive women with placenta malaria and factors associated with placenta malaria., Methods: This was a prospective observational study of booked HIV positive pregnant women in labour. A smear for malaria parasite was made from blood taken from the placental tissue post-delivery. The baby HIV testing was done with DNA polymerase chain reaction at 6 weeks postpartum. Data on age, parity, gestational age, religion, address, highest educational attainment and knowledge about malaria prevention in pregnancy was obtained with questionnaires and analysed using SPSS version 20. The P-value was set at 0.05 providing a confidence interval of 95%., Results: A total of 174 booked HIV women participated in this study. The placental malaria parasitaemia prevalence was 44.8%. Overall rate of MTCT of HIV infection was 17.2%. Number of infants with HIV infection among women with maternal placental malarial parasitaemia was 30/78 (38.5%), while it was 0/96 (0%) for women without placenta malaria. There was significant relationship between placenta malaria density and infant HIV status (P-value = 0.001). The relative risk for MTCT of HIV for women with placenta malaria Density > 5000 was 25% with 95% confidence interval of 11.41-54.76%., Conclusion: The mother-to-child transmission rate of HIV was high among HIV positive women with placental malaria parasitaemia. There is the need to review the malarial treatment and prophylactic measures at least in this group of women and to establish the nature of relationship between placenta malaria and MTCT of HIV infection.

Background: Human blood cells (erythrocytes) serve as hosts for the malaria parasite Plasmodium falciparum during its 48-h intraerythrocytic developmental cycle (IDC). Established in vitro protocols allow for the study of host-parasite interactions during this phase and, in particular, high-resolution metabolomics can provide a window into host-parasite interactions that support parasite development., Methods: Uninfected and parasite-infected erythrocyte cultures were maintained at 2% haematocrit for the duration of the IDC, while parasitaemia was maintained at 7% in the infected cultures. The parasite-infected cultures were synchronized to obtain stage-dependent information of parasite development during the IDC. Samples were collected in quadruplicate at six time points from the uninfected and parasite-infected cultures and global metabolomics was used to analyse cell fractions of these cultures., Results: In uninfected and parasite-infected cultures during the IDC, 501 intracellular metabolites, including 223 lipid metabolites, were successfully quantified. Of these, 19 distinct metabolites were present only in the parasite-infected culture, 10 of which increased to twofold in abundance during the IDC. This work quantified approximately five times the metabolites measured in previous studies of similar research scope, which allowed for more detailed analyses. Enrichment in lipid metabolism pathways exhibited a time-dependent association with different classes of lipids during the IDC. Specifically, enrichment occurred in sphingolipids at the earlier stages, and subsequently in lysophospholipid and phospholipid metabolites at the intermediate and end stages of the IDC, respectively. In addition, there was an accumulation of 18-, 20-, and 22-carbon polyunsaturated fatty acids, which produce eicosanoids and promote gametocytogenesis in infected erythrocyte cultures., Conclusions: The current study revealed a number of heretofore unidentified metabolic components of the host-parasite system, which the parasite may exploit in a time-dependent manner to grow over the course of its development in the blood stage. Notably, the analyses identified components, such as precursors of immunomodulatory molecules, stage-dependent lipid dynamics, and metabolites, unique to parasite-infected cultures. These conclusions are reinforced by the metabolic alterations that were characterized during the IDC, which were in close agreement with those known from previous studies of blood-stage infection.

MHC class II (MHCII) molecules are cell surface glycoproteins that play an important role to develop adaptive immune responses. MHCII-disease association is not restricted to structural variation alone but also may extend to genetic variations, which may modulate gene expression. The observed variations in class II gene expression make it possible that the association of MHCII polymorphism with diseases may relate to the level of gene expression in addition to the restriction of response to Ag. Understanding the extent of, and the mechanisms underlying, transcription factor DNA binding variation is therefore key to elucidate the molecular determinants of complex phenotypes. In this study, we investigated whether single nucleotide polymorphisms in MHCII-DRB regulatory gene may be associated with clinical outcomes of malaria in Plasmodium -infected individuals. To this end, we conducted a case-control study to compare patients who had mild malaria with those patients who had asymptomatic Plasmodium infection. It demonstrates that GTAT haplotype exerts an increased DRB transcriptional activity, resulting in higher DRB expression and subsequently perturbed Ag presentation and T cell activation, higher TLR-mediated innate immune gene expression, and Ag clearance, so low parasitemia in comparison with haplotypes other than GTAT (GTAC, GGGT). Hence, we hypothesized that DRB gene promoter polymorphism might lead to altered DRB gene expression, which could possibly affect the TLR-triggered innate immune responses in malaria patients. These genetic findings may contribute to the understanding of the pathogenesis of malaria and will facilitate the rational vaccine design for malaria., (Copyright © 2020 by The American Association of Immunologists, Inc.)

Apicomplexan parasites contain rhoptries, which are specialized secretory organelles that coordinate host cell invasion. During the process of invasion, rhoptries secrete their contents to facilitate interaction with, and entry into, the host cell. Here, we report the crystal structure of the rhoptry protein Armadillo Repeats-Only (ARO) from the human malaria parasite, Plasmodium falciparum (PfARO). The structure of PfARO comprises five tandem Armadillo-like (ARM) repeats, with adjacent ARM repeats stacked in a head-to-tail orientation resulting in PfARO adopting an elongated curved shape. Interestingly, the concave face of PfARO contains two distinct patches of highly conserved residues that appear to play an important role in protein-protein interaction. We functionally characterized the P. falciparum homolog of ARO interacting protein (PfAIP) and demonstrate that it localizes to the rhoptries. We show that conditional mislocalization of PfAIP leads to deficient red blood cell invasion. Guided by the structure, we identified mutations of PfARO that lead to mislocalization of PfAIP. Using proximity-based biotinylation we probe into PfAIP interacting proteins., (Copyright © 2019. Published by Elsevier Ltd.)

Background: Naphthoquine (NQ) is a suitable partner anti-malarial for the artemisinin-based combination therapy (ACT), which is recommended to be taken orally as a single-dose regimen. The metabolism of NQ was mainly mediated by CYP2D6, which is well-known to show gender-specific differences in its expression. In spite of its clinical use, there is limited information on the pharmacokinetics of NQ, and no data are available for females. In this study, the effect of gender on the pharmacokinetics and antiplasmodial efficacy of NQ in rodents was evaluated. The underlying factors leading to the potential gender difference, i.e., plasma protein binding and metabolic clearance, were also evaluated., Methods: The pharmacokinetic profiles of NQ were investigated in healthy male or female rats after a single oral administration of NQ. The antiplasmodial efficacy of NQ was studied in male or female mice infected with Plasmodium yoelii. The recrudescence and survival time of infected mice were also recorded after drug treatment. Plasma protein binding of NQ was determined in pooled plasma collected from male or female mice, rat or human. In vitro metabolism experiments were performed in the liver microsomes of male or female mice, rat or human., Results: The results showed that the gender of rats did not affect NQ exposure (AUC 0-t and C max ) significantly (P > 0.05). However, a significant (P < 0.05) longer t 1/2 was found for NQ in male rats (192.1 ± 47.7), compared with female rats (143.9 ± 27.1). Slightly higher but not significant (P > 0.05) antiplasmodial activity was found for NQ in male mice (ED 90 , 1.10 mg/kg) infected with P. yoelii, compared with female mice (ED 90 , 1.67 mg/kg). The binding rates of NQ to plasma protein were similar in males and females. There was no metabolic difference for NQ in male and female mice, rat or human liver microsomes., Conclusions: These results indicated that the pharmacokinetic profiles of NQ were similar between male and female rats, except for a longer t 1/2 in male rats. The difference was not associated with plasma protein binding or hepatic metabolic clearance. Equivalent antiplasmodial activity was found for NQ in male and female mice infected with P. yoelii. This study will be helpful for the rational design of clinical trials for NQ.

Background: Day 7 plasma lumefantrine concentration is suggested as a predictor for malaria treatment outcomes and a cut-off of ≥ 200 ng/ml is associated with day 28 cure rate in the general population. However, day 7 lumefantrine plasma concentration can be affected by age, the extent of fever, baseline parasitaemia, and bodyweight. Therefore, this study assessed the usefulness of day 7 lumefantrine plasma concentration as a predictor of malaria treatment outcome in under-fives children treated with generic or innovator drug-containing artemether-lumefantrine (ALu) in Tanzania., Methods: This study was nested in an equivalence prospective study that aimed at determining the effectiveness of a generic ALu (Artefan ® ) in comparison with the innovator's product (Coartem ® ). Children with uncomplicated malaria aged 6-59 months were recruited and randomized to receive either generic or innovator's product. Children were treated with ALu as per World Health Organization recommendations. The clinical and parasitological outcomes were assessed after 28 days of follow up. PCR was performed to distinguish recrudescence and re-infections among children with recurrent malaria. Analysis of day 7 lumefantrine plasma concentration was carried out using a high-performance liquid chromatographic method with UV detection., Results: The PCR corrected cure rates were 98.7% for children treated with generic and 98.6% for those treated with the innovator product (p = 1.00). The geometric mean (± SD) of day 7 plasma lumefantrine concentration was 159.3 (± 2.4) ng/ml for the generic and 164 (± 2.5) ng/ml for the innovator groups, p = 0.87. Geometric mean (± SD) day 7 lumefantrine plasma concentration between cured and recurrent malaria was not statistically different in both treatment arms [158.5 (± 2.4) vs 100.0 (± 1.5) ng/ml, (p = 0.28) for generic arm and 158.5 (± 2.3) vs 251.2 (± 4.2) ng/ml, (p = 0.24) for innovator arm]. Nutritional status was found to be a determinant of recurrent malaria (adjusted hazardous ratio (95% confidence interval) = 3(1.1-8.2), p = 0.029., Conclusion: Using the recommended cut-off point of ≥ 200 ng/ml, day 7 plasma lumefantrine concentration failed to predict malaria treatment outcome in children treated with ALu in Tanzania. Further studies are recommended to establish the day 7 plasma lumefantrine concentration cut-off point to predict malaria treatment outcome in children.

Ethnopharmacological Relevance: Coccinia barteri (Hook. F.) is traditional used in Southeast of Nigeria in management of fever. This study aimed to evaluate the antimalarial activities of hydro-methanol crude extract and solvent fractions of Coccinia barteri leaf., Materials and Methods: Two animal models employed for the study were, 4-day suppressive and curative assays against chloroquine sensitive Plasmodium berghei NK65. Level of parasitaemia, mean survival time (MST), anal temperature and weight loss were measured to assess antimalarial efficacy of the extract/fractions. Chloroquine (10 mg kg -1 ) was used as positive control. Chemo-profile of extract was evaluated using GC-MS, HPLC techniques and standard phytochemical analysis. Preliminary toxicity test was done using modified Lorke's method., Results: The crude extract (100-400 mg kg- 1 ) and solvent fractions (20-80 mg kg- 1 ) demonstrated antimalarial activity in both models compared to controls. Semi purified fractions of the extract produced stronger percentage chemosuppression and inhibition of parasite. The % inhibition of the fractions, hexane, chloroform, ethyl acetate and aqueous at 80 mg kg -1 were 96.0 0, 95.29, 89.86 and 96.00% respectively on day 8 (D 8 ). While on D 14 , 100% parasite clearance, indicating cure was obtained for hexane, chloroform and aqueous fraction treatment groups, no death occurred in these groups. Ethyl acetate fraction treated groups lived longer but were not fully protected. Some marker compounds were identified., Conclusions: These results support the use of C. barteri as malaria remedy and potential source of antimalarial templates. Long acting parasitaemia reduction effect indicates its possible combination potential in poly-herbal combination therapy., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Malaria is a worldwide problem that affects millions of people yearly. In rural areas where anti-malarial drugs are not easily accessible, many people use herbal treatments, such as Moringa oleifera, to treat a variety of diseases and ailments including malaria. While Moringa is reported to possess potent and curative anti-malarial properties, previous studies have mostly been restricted to assessment of parasitaemia. In this study, the effect of Moringa on malaria immunity in a murine model was investigated., Methods: Using a high dose (60 mg/mouse) for a short time (7 days) or low dose Moringa (30 mg/mouse) for a longer time (3 weeks), cytokine production, and Tbet expression by effector CD4 + T cells (Teff) were determined. Mice were also treated with Moringa after infection (curatively) or before infection (prophylactically) to determine the effect of the plant extract on parasitaemia and immunity. Given that Moringa also possess many nutritional benefits, the contribution of Moringa on malnourished malaria infected mice was determined. Malnutrition was induced by limiting access to food to only 4 h a day for 4 weeks, while control mice had unlimited access to mouse laboratory chow. All data was collected by flow cytometry and analysed using one-Way ANOVA or two tailed Student's t test., Results: Moringa-treated mice had increased numbers of effector CD4 + T cells accompanied by an increase in Tbet expression compared to control untreated mice. Mice that were treated with Moringa curatively also exhibited increased effector CD4 + T cell numbers, IFN-gamma and TNF secretion. Interestingly, the mice that were treated prophylactically had significantly higher Tbet expression. In the absence of adaptive immunity, high parasitaemia was observed in the RAG1 knockout mice. The food limited mice (malnourished) had reduced numbers of CD4 + T cells, TNF proportions, and significantly greater Tbet expression compared to the control group. Supplementation with Moringa in the limited group slightly restored CD4 + T cell activation, IL-2, and IL-10 production., Conclusions: Taken together, these data suggest that Moringa treatment leads to increased CD4 + T cell activation, Th1 differentiation and production of pro-inflammatory cytokines after malaria infection. Thus, Moringa may be immunologically useful in the treatment of malaria and malnutrition. Further investigations are required to identify the active components in Moringa.

Background: Multiple red blood cell (RBC) variants appear to offer protection against the most severe forms of Plasmodium falciparum malaria. Associations between these variants and uncomplicated malaria are less clear., Methods: Data from a longitudinal cohort study conducted in 3 sub-counties in Uganda was used to quantify associations between three red blood cell variants Hb [AA, AS, S (rs334)], alpha thalassaemia 3.7 kb deletion, and glucose-6-phosphate dehydrogenase deficiency A-(G6PD 202A genotype) and malaria incidence, parasite prevalence, parasite density (a measure of anti-parasite immunity) and body temperature adjusted for parasite density (a measure of anti-disease immunity). All analyses were adjusted for age, average household entomological inoculation rate, and study site. Results for all variants were compared to those for wild type genotypes., Results: In children, HbAS was associated, compared to wild type, with a lower incidence of malaria (IRR = 0.78, 95% CI 0.66-0.92, p = 0.003), lower parasite density upon infection (PR = 0.66, 95% CI 0.51-0.85, p = 0.001), and lower body temperature for any given parasite density (- 0.13 ℃, 95% CI - 0.21, - 0.05, p = 0.002). In children, HbSS was associated with a lower incidence of malaria (IRR = 0.17, 95% CI 0.04-0.71, p = 0.02) and lower parasite density upon infection (PR = 0.31, 95% CI 0.18-0.54, p < 0.001). α-/αα thalassaemia, was associated with higher parasite prevalence in both children and adults (RR = 1.23, 95% CI 1.06-1.43, p = 0.008 and RR = 1.52, 95% CI 1.04-2.23, p = 0.03, respectively). G6PD deficiency was associated with lower body temperature for any given parasite density only among male hemizygote children (- 0.19 ℃, 95% CI - 0.31, - 0.06, p = 0.003)., Conclusion: RBC variants were associated with non-severe malaria outcomes. Elucidation of the mechanisms by which they confer protection will improve understanding of genetic protection against malaria.

Hepatozoon pyramidumi sp. n. is described from the blood of the Egyptian saw-scaled viper, Echis pyramidum, captured from Saudi Arabia. Five out of ten viper specimens examined (50%) were found infected with Hepatozoon pyramidumi sp. n. with parasitaemia level ranged from 20-30%. The infection was restricted only to the erythrocytes. Two morphologically different forms of intraerythrocytic stages were observed; small and mature gamonts. The small ganomt with average size of 10.7 × 3.5 μm. Mature gamont was sausage-shaped with recurved poles measuring 16.3 × 4.2 μm in average size. Infected erythrocytes were hypertrophied; their nuclei were deformed and sometimes displaced from their central position in the normal uninfected cell. Merogonic stages were observed in the lung endothelial cell and the liver parenchyma cells. Mature meront was 17.8 × 13.6 µm and contained banana-shaped merozoites with average size of ~15 × 2 µm. Phylogenetic analysis based on the SSU rDNA sequence clustered Hepatozoon pyramidumi sp. n with previously sequenced Hepatozoon spp., most of them infected reptilian hosts without geographic consideration. The morphological and molecular comparison with closely related species proved the taxonomic uniqueness and novelty of the present form.

The aim of this study was to investigate the antimalarial activities and toxicity of Pogostemon cablin extracts. In vitro activities against the chloroquine-resistant Plasmodium falciparum K1 strain were assessed by using the Plasmodium lactate dehydrogenase enzyme (pLDH) assay, while in vivo activity against the Plasmodium berghei ANKA strain in mice was investigated using a 4-day suppressive test. The in vitro and in vivo toxicity were determined in Vero cells and mice, respectively. The ethanolic extract possessed antimalarial activity with an IC 50 of 24.49 ± 0.01 µg/ml, whereas the aqueous extract showed an IC 50 of 549.30 ± 0.07 µg/ml. Cytotoxic analyses of the ethanolic and aqueous extracts revealed a nontoxic effect on Vero cells at a concentration of 80 µg/ml. Based on a preliminary study of in vitro antimalarial activity, the ethanolic extract was chosen as a potential agent for further in vivo antimalarial activity analysis in mice. The ethanolic extract, which showed no toxic effect on mice at a dose of 2000 mg/kg body weight, significantly suppressed parasitemia in mice by 38.41%, 45.12% and 89.00% at doses of 200, 400 and 600 mg/kg body weight, respectively. In conclusion, this study shows that the ethanolic P. cablin extract possesses in vitro and in vivo antimalarial activity without toxic effects.

Artefenomel and DSM265 are two new compounds that have been shown to be well tolerated and effective when administered as monotherapy malaria treatment. This study aimed to determine the safety, pharmacokinetics, and pharmacodynamics of artefenomel and DSM265 administered in combination to healthy subjects in a volunteer infection study using the Plasmodium falciparum -induced blood-stage malaria model. Thirteen subjects were inoculated with parasite-infected erythrocytes on day 0 and received a single oral dose of artefenomel and DSM265 on day 7. Cohort 1 ( n  = 8) received 200 mg artefenomel plus 100 mg DSM265, and cohort 2 ( n  = 5) received 200 mg artefenomel plus 50 mg DSM265. Blood samples were collected to measure parasitemia, gametocytemia, and artefenomel-DSM265 plasma concentrations. There were no treatment-related adverse events. The pharmacokinetic profiles of artefenomel and DSM265 were similar to those of the compounds when administered as monotherapy, suggesting no pharmacokinetic interactions. A reduction in parasitemia occurred in all subjects following treatment (log 10 parasite reduction ratios over 48 h [PRR 48 ] of 2.80 for cohort 1 and 2.71 for cohort 2; parasite clearance half-lives of 5.17 h for cohort 1 and 5.33 h for cohort 2). Recrudescence occurred in 5/8 subjects in cohort 1 between days 19 and 28 and in 5/5 subjects in cohort 2 between days 15 and 22. Low-level gametocytemia (1 to 330 female gametocytes/ml) was detected in all subjects from day 14. The results of this single-dosing combination study support the further clinical development of the use of artefenomel and DSM265 in combination as a treatment for falciparum malaria. (This study has been registered at ClinicalTrials.gov under identifier NCT02389348.)., (Copyright © 2019 McCarthy et al.)

Background: Global efforts to scale-up malaria control interventions are gaining steam. These include the use of Long-Lasting Insecticide Nets, Indoor Residual Spraying, Intermittent Preventive Treatment and Test, Treat and Track. Despite these, the drive for malaria elimination is far from being realistic in endemic communities in Africa. This is partly due to the fact that asymptomatic parasite carriage, not specifically targeted by most interventions, remains the bedrock that fuels transmission. This has led to mass testing, treatment and tracking (MTTT) as an alternative strategy to target asymptomatic individuals. We report the impact of MTTT on the prevalence of asymptomatic malaria parasitaemia over a one-year period in Ghana, hypothesizing that implementing MTTT could reduce the rate of asymptomatic parasitaemia., Methods: A population of about 5000 individuals in seven communities in the Pakro sub-district of Ghana participated in this study. A register was developed for each community following a census. MTTT engaged trained community-based health volunteers who conducted house-to-house testing using RDTs every 4 months and treated positive cases with Artemisinin-based Combination Therapy. Between interventions, community-based management of malaria was implemented for symptomatic cases., Results: MTTT Coverage was 98.8% in July 2017 and 79.3% in July 2018. Of those tested, asymptomatic infection with malaria parasites reduced from 36.3% (1795/4941) in July 2017 to 32.9% (1303/3966) in July 2018 (p = 0.001). Prevalence of asymptomatic parasitaemia among children under 15 years declined from 52.6% (1043/1984) in July 2017 to 47.5% (820/1728) in July 2018 (p = 0.002). Implementing MTTT significantly reduced asymptomatic parasitaemia by 24% from July 2017 to July 2018 after adjusting for age, ITN use and axillary temperature (OR = 0.76, CI = 0.67, 0.85 p ≤ 0.001)., Conclusion: This study has demonstrated that implementing MTTT is feasible and could reduce the prevalence of asymptomatic malaria parasitaemia in children under 15 years of age. Furthermore, the use of community-based health volunteers could ensure high coverage at lower cost of implementation., Trial Registration: NCT04167566, Date 14/11/2019. Retrospective registration.

Loiasis is a vector-borne parasitic disease caused by the filarial Loa loa (L. loa). Definitive diagnosis can be done by identifying and counting microfilariae in the peripheral blood by microscopy and with L.loa-specific PCR. An additional diagnostic method is the detection of L.loa-specific antibodies. Accurate methods are needed to automate quantification of microfilaria (mf) in peripheral blood. Indeed, the treatment procedure depends on the microfilarial L. loa load in blood. We report the first documented use of flow cytometry as a new method to count microfilaraemia in peripheral blood from a patient with L. loa infection. The diagnosis of loiasis was strongly suspected based on clinical presentation and rapidly confirmed by identifying typical features of L. loa in the peripheral blood. This diagnosis was achieved by flow cytometry using a specific fluorescence pattern for microfilaraemia count. The current report highlights the potential of flow cytometry to assess microfilarial L. loa load from a patient with loiasis infection., (Copyright © 2019 Elsevier Inc. All rights reserved.)

We investigated the immunomodulatory, antiparasitic and cardioprotective effects of a sesquiterpene lactone (SL) administered alone or combined with benznidazole (Bz), in a murine model of Chagas' disease by in vitro and in vivo assays. Antiparasitic and cytotoxic potential of tagitinin C (SL) and Bz were tested in vitro against T. cruzi epimastigotes and cardiomyocytes. Swiss mice challenged with T. cruzi were also treated for 20 days with tagitinin C (10 mg/kg) alone and combined with Bz (100 mg/kg). Tagitinin C exhibited a higher antiparasitic (IC 50 : 1.15 µM) and cytotoxic (CC 50 at 6.54 µM) potential than Bz (IC 50 : 35.81 µM and CC 50: 713.5 µM, respectively). When combined, these drugs presented an addictive interaction, determining complete suppression of parasitemia and parasitological cure in all infected mice (100%) compared to those receiving Bz alone (70%). Anti-T. cruzi immunoglobulin G, and pro-inflammatory cytokines IFN-γ and TNF-α levels were reduced in animals treated with tagitinin C combined with Bz, while IL-10 production was unaffected. Heart inflammation was undetectable in 90% of the animals receiving this combination, while only 50% of the animals receiving Bz alone showed no evidence of myocarditis. Together, our findings indicated that the combination of tagitinin C and Bz exerts potent antiparasitic, immunomodulatory and cardioprotective effects. Due to the remarkable suppression of parasitemia and high parasitological cure, this combination was superior to Bz monotherapy, indicating a high potential for the treatment of Chagas's disease., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Although CRIg was originally identified as a macrophage receptor for binding complement C3b/iC3b in vitro, recent studies reveal that CRIg functions as a pattern recognition receptor in vivo for Kupffer cells (KCs) to directly bind bacterial pathogens in a complement-independent manner. This raises the critical question of whether CRIg captures circulating pathogens through interactions with complement in vivo under flow conditions. Furthermore, the role of CRIg during parasitic infection is unknown. Taking advantage of intravital microscopy and using African trypanosomes as a model, we studied the role of CRIg in intravascular clearance of bloodborne parasites. Complement C3 is required for intravascular clearance of African trypanosomes by KCs, preventing the early mortality of infected mice. Moreover, antibodies are essential for complement-mediated capture of circulating parasites by KCs. Interestingly, reduced antibody production was observed in the absence of complement C3 during infection. We further demonstrate that CRIg but not CR3 is critically involved in KC-mediated capture of circulating parasites, accounting for parasitemia control and host survival. Of note, CRIg cannot directly catch circulating parasites and antibody-induced complement activation is indispensable for CRIg-mediated parasite capture. Thus, we provide evidence that CRIg, by interacting with complement in vivo, plays an essential role in intravascular clearance of bloodborne parasites. Targeting CRIg may be considered as a therapeutic strategy., Competing Interests: The authors declare no competing interest.

Background: Malaria remains a public health issue, particularly in sub-Saharan Africa with special features of seriousness in young children and pregnant women. Adolescents and adults are reported to have acquired a semi-immune status and, therefore, present with low parasitaemia. Children are understood to present with a much higher parasitaemia and severe malaria. It is a concern that effective malaria control programmes targeting young children may lead to a delay in the acquisition of acquired immunity and, therefore, causing a shift in the epidemiology of malaria. Prevalence and parasitaemia were explored in adolescents and adults with Plasmodium falciparum infections compared to young children in the area of Lambaréné, Gabon as an indicator for semi-immunity., Methods: A cross-sectional study was conducted at the Centre de Recherches Médicales de Lambaréné (CERMEL) during a 6-month period in 2018. Symptomatic patients, of all ages were screened for malaria at health facilities in Lambaréné and Fougamou and their respective surrounding villages in the central region of Gabon. Plasmodium falciparum infections were determined either by rapid diagnostic test (RDT) or by microscopy. Descriptive analysis of data on parasite densities, anaemia, and fever are presented., Results: 1589 individuals screened were included in this analysis, including 731 (46%) adolescents and adults. Out of 1377 assessed, the proportion of P. falciparum positive RDTs was high among adolescents (68%) and adults (44%), compared to young children (55%) and school children (72%). Out of 274 participants assessed for malaria by microscopy, 45 (16%) had a parasite count above 10,000/µl of which 9 (20%) were adults., Conclusion: This study shows a high rate of P. falciparum infections in adolescents and adults associated with high-level parasitaemia similar to that of young children. Adolescents and adults seem to be an at-risk population, suggesting that malaria programmes should consider adolescents and adults during the implementation of malaria prevention and case management programmes with continuous care, since they also act as reservoirs for P. falciparum.

A model is developed and used to study within-human malaria parasite dynamics. The model integrates actors involved in the development-progression of parasitemia, gametocytogenesis and mechanisms for immune response activation. Model analyses under immune suppression reveal different dynamical behaviours for different healthy red blood cell (HRBC) generation functions. Existence of a threshold parameter determines conditions for HRBCs depletion. Oscillatory dynamics reminiscent of malaria parasitemia are obtained. A dependence exists on the type of recruitment function used to generate HRBCs, with complexities observed for a more nonlinear function. An upper bound that delimits the size of feasible parasitized steady-state solution exists for a logistic function but not a constant function. The upper bound is completely characterized and is affected by parameters associated with HRBCs recruitment, parasitized red blood cells generation and the release and time-to-release of free merozoites. A stable density size for mature gametocytes, the bridge to invertebrate hosts, is derived.

Characterization of complete life cycles of haemoparasites requires the maintenance of suitable susceptible vertebrate hosts and vectors for long periods in captivity, in order to follow the complete parasitic cycle in definitive and intermediate hosts. Currently, there are few host-parasite models established in avian haemosporidian research, and those have been developed mainly for species of Passeriformes and their parasites. This study aimed to develop an experimental methodology to access the complete life cycle of Haemoproteus columbae (cytb lineage HAECOL1), which parasitizes the Rock Pigeon (Columba livia) and louse fly (Pseudolynchia canariensis). A colony of louse flies, which are the natural vectors of this parasite, was established. Thirty newly emerged insects were exposed to H. columbae infection and used to infect naïve Rock Pigeons. The peak of parasitaemia (acute stage) was seen between 27 and 32 days p.i. when up to 70.8% of red blood cells were infected. The crisis occurred approximately 1 week after the peak, and the long-lasting chronic parasitaemia stage followed. Exo-erythrocytic meronts were seen mainly in the lungs where extensive tissue damage was reported, but also in the kidneys and spleen. In the vector, the sporogonic cycle of H. columbae was completed between 13 and 16 days p.i., at an average temperature ranging between 12 and 15 °C. This host-parasite model is tractable for maintenance in captivity. It is recommended for use in studies aiming for detailed characterization of host-parasite relationships in areas such as physiology, pathology, immunobiology, genetics, as well as for evaluative treatments and to follow the infection in any stage of parasite development both in the vertebrate or invertebrate host., (Copyright © 2019 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Sepsis can be caused by malaria infection, but little is known about the utility of the quick Sequential (Sepsis-Related) Organ Failure Assessment (qSOFA) and SOFA score in malaria. We conducted a prospective observational study from March 2013 to February 2017 to examine adults admitted with community-acquired infection in a tertiary-care hospital in Ubon Ratchathani, Northeast Thailand (Ubon-sepsis). Subjects were classified as having sepsis if they had a modified SOFA score ≥2 within 24 hours of admission. Serum was stored and later tested for malaria parasites using a nested PCR assay. Presence of severe malaria was defined using modified World Health Organization criteria. Of 4,989 patients enrolled, 153 patients (3%) were PCR positive for either Plasmodium falciparum (74 [48%]), P. vivax (69 [45%]), or both organisms (10 [7%]). Of 153 malaria patients, 80 were severe malaria patients presenting with sepsis, 70 were non-severe malaria patients presenting with sepsis, and three were non-severe malaria patients presenting without sepsis. The modified SOFA score (median 5; IQR 4-6; range 1-18) was strongly correlated with malaria severity determined by the number of World Health Organization severity criteria satisfied by the patient (Spearman's rho = 0.61, p<0.001). Of 80 severe malaria patients, 2 (2.5%), 11 (14%), 62 (77.5%) and 5 (6%), presented with qSOFA scores of 0, 1, 2 and 3, respectively. Twenty eight-day mortality was 1.3% (2/153). In conclusion, qSOFA and SOFA can serve as markers of disease severity in adults with malarial sepsis. Patients presenting with a qSOFA score of 1 may also require careful evaluation for sepsis; including diagnosis of cause of infection, initiation of medical intervention, and consideration for referral as appropriate., Competing Interests: The authors have declared that no competing interests exist.

Ethnopharmacological Relevance: An aqueous decoction of root bark of Alstonia congensis Engl. (Apocynaceae) is used in several African countries to treat various ailments including malaria., Materials and Methods: Extracts of different polarity and isolated constituents were tested in vitro for their antiplasmodial activity against the chloroquine-resistant strain Plasmodium falciparum K1 and the chloroquine-sensitive strain P. falciparum NF54A19A, as well as for their cytotoxic effects againt MRC-5 cells (human lung fibroblasts). Extracts and fractions were evaluated in vivo against the chloroquine-resistant strain P. yoelii N67 and the chloroquine-sensitive strain P. berghei berghei ANKA., Results: The aqueous extract, the 80% methanol extract and the alkaloid-enriched extract exhibited strong antiplasmodial activity against P. falciparum K1 with IC 50 values < 10 µg/ml and against P. falciparum NF54 A19A with IC 50 values < 0.02 µg/ml. In vivo against P. yoelii N67, at the highest oral dose of 400 mg/kg body weight, all extracts produced 70-73% chemosuppression, while against P. berghei berghei, more than 75% chemosuppression was observed. With regard to the isolated constituents, boonein was inactive in vitro against P. falciparum K-1 (IC 50 > 64 µM), while echitamine, 6,7-seco-angustilobine B and β-amyrin exhibited moderate activity (IC 50 < 30 µM). Against P. falciparum NF54 A19A, boonein was inactive (IC 50 > 64 µM), while echitamine, 6,7-secoangustilobine and β-amyrin showed moderate IC 50 values of 11.07, 21.26 and 40.70 µM, respectively., Conclusion: These results demonstrated that all extracts from A. congensis root bark possessed antiplasmodial activity in vitro and in vivo. They can be used as raw materials for the preparation of ameliorated remedies for the treatment of uncomplicated malaria. The observed antiplasmodial activity may be due in part to the presence of indole alkaloids., (Copyright © 2019. Published by Elsevier B.V.)

C57BL/6 mice infected with Plasmodium berghei ANKA (PbA) develop neurological symptoms and die 6--7-day post-inoculation in the absence of high parasitemia. The effects of chronic intake of a high-fat diet on this process are largely unknown. In this study, we assessed the effect of a high-fat diet on the host-parasite response to malarial infection. Mice were fed ad libitum with either standard or a high-fat diet for 8 weeks and afterwards were infected with PbA. PbA-infected mice feeding a standard diet presented blood parasitemia, hepatic and cerebral histopathological alterations, and hepatic injury with increased hemozoin deposition in the liver. By contrast, these changes were not observed in the malaria high-fat diet group. In addition, mice fed a high-fat diet did not develop the expected neurological symptoms of cerebral malaria and were resistant to death. Taken together, our results indicate that chronic ingestion of high-fat diet prevents the development of experimental malaria induced by PbA injection, suggesting a relationship between a high-fat diet and malaria, which is an interesting subject for further study in humans.

Based on morphology and morphometry of gametocytes in blood and molecular phylogenetic analysis, we described a new species of hemoparasite from the genus Haemogregarina isolated from Lepidosiren paradoxa in the eastern Amazon region. Haemogregarina daviesensis sp. nov. is characterized by monomorphic gametocytes of varying maturity stage and their dimensions were 16 ± 0.12 μm (range 13-18) in length and 6 ± 0.97 μm (range 5-8) in width. The morphological and morphometric data were not identical with other haemogregarine species from fish. All specimens of L. paradoxa analyzed were infected by H. daviesensis sp. nov. and the parasitemia level was moderate (1-28/2000 blood erythrocytes). Two sequences were obtained from L. paradoxa, and these constituted a monophyletic sister clade to the Haemogregarina species. In addition, H. daviesensis sp. nov. detected here grouped with Haemogregarina sp. sequences isolated from chelonian Macrochelys temminckii, with 99% bootstrap support. This study provides the first data on the molecular phylogeny of an intraerythrocytic haemogregarine of freshwater fish and highlights the importance of obtaining additional information on aspects of the general biology of these hemoparasites in fish populations, in order to achieve correct taxonomic classification.

Emerging malaria parasite sulfadoxine-pyrimethamine (SP) resistance has prompted assessment of alternatives for intermittent preventive treatment in pregnancy (IPTp). The objective was to evaluate the tolerability and prophylactic efficacy of azithromycin (AZ) plus piperaquine (PQ) in pregnant women in Papua New Guinea. The study was an open-label, randomized, parallel-group trial. A total of 122 women (median gestation, 26 weeks [range, 14 to 32 weeks]) were randomized 1:1 to three daily doses of 1 g AZ plus 960 mg PQ tetraphosphate or single-dose SP (4,500 mg sulfadoxine plus 225 mg pyrimethamine), based on computer-generated block randomization. Tolerability was assessed to day 7, and efficacy was assessed to day 42 (when participants were returned to usual care) and at delivery. Data for 119 participants (AZ-PQ, n  = 61; SP, n  = 58) were analyzed. Both regimens were well tolerated, but AZ-PQ was associated with more gastrointestinal side effects (31%) and dizziness (21%). Eight women (6.7%) were parasitemic at recruitment but all were aparasitemic by 72 h. There were no differences in blood smear positivity rates between AZ-PQ and SP up to day 42 (0% versus 5.2%; relative risk [RR], 0.14 [95% confidence interval [CI], 0.01 to 2.58] [ P =  0.18]; absolute risk reduction [ARR], 5.2% [95% CI, -1.3 to 11.6%]) and at the time of delivery (0% versus 8.7%; RR, 0.11 [95% CI, 0.01 to 2.01] [ P =  0.14]; ARR, 8.7% [95% CI, -0.2 to 17.6%]). Of 92 women who were monitored to parturition, 89 (97%) delivered healthy babies; there were 3 stillbirths (SP, n  = 1; AZ-PQ, n  = 2 [twins]). There was a higher live birth weight (mean ± standard deviation) in the AZ-PQ group (3.13 ± 0.42 versus 2.88 ± 0.55 kg [ P =  0.016]; mean difference, 0.25 kg [95% CI, 0.02 to 0.48 kg]). AZ-PQ is a promising candidate for IPTp., (Copyright © 2019 American Society for Microbiology.)

Plasmodium relapses are attributed to the activation of dormant liver-stage parasites and are responsible for a significant number of recurring malaria blood-stage infections. While characteristic of human infections caused by P. vivax and P. ovale, their relative contribution to malaria disease burden and transmission remains poorly understood. This is largely because it is difficult to identify 'bona fide' relapse infections due to ongoing transmission in most endemic areas. Here, we use the P. cynomolgi-rhesus macaque model of relapsing malaria to demonstrate that clinical immunity can form after a single sporozoite-initiated blood-stage infection and prevent illness during relapses and homologous reinfections. By integrating data from whole blood RNA-sequencing, flow cytometry, P. cynomolgi-specific ELISAs, and opsonic phagocytosis assays, we demonstrate that this immunity is associated with a rapid recall response by memory B cells that expand and produce anti-parasite IgG1 that can mediate parasite clearance of relapsing parasites. The reduction in parasitemia during relapses was mirrored by a reduction in the total number of circulating gametocytes, but importantly, the cumulative proportion of gametocytes increased during relapses. Overall, this study reveals that P. cynomolgi relapse infections can be clinically silent in macaques due to rapid memory B cell responses that help to clear asexual-stage parasites but still carry gametocytes., Competing Interests: The authors have declared that no competing interests exist.

Background: Primaquine is the only widely used drug that prevents Plasmodium vivax malaria relapses, but adherence to the standard 14-day regimen is poor. We aimed to assess the efficacy of a shorter course (7 days) of primaquine for radical cure of vivax malaria., Methods: We did a randomised, double-blind, placebo-controlled, non-inferiority trial in eight health-care clinics (two each in Afghanistan, Ethiopia, Indonesia, and Vietnam). Patients (aged ≥6 months) with normal glucose-6-phosphate dehydrogenase (G6PD) and presenting with uncomplicated vivax malaria were enrolled. Patients were given standard blood schizontocidal treatment and randomly assigned (2:2:1) to receive 7 days of supervised primaquine (1·0 mg/kg per day), 14 days of supervised primaquine (0·5 mg/kg per day), or placebo. The primary endpoint was the incidence rate of symptomatic P vivax parasitaemia during the 12-month follow-up period, assessed in the intention-to-treat population. A margin of 0·07 recurrences per person-year was used to establish non-inferiority of the 7-day regimen compared with the 14-day regimen. This trial is registered at ClinicalTrials.gov (NCT01814683)., Findings: Between July 20, 2014, and Nov 25, 2017, 2336 patients were enrolled. The incidence rate of symptomatic recurrent P vivax malaria was 0·18 (95% CI 0·15 to 0·21) recurrences per person-year for 935 patients in the 7-day primaquine group and 0·16 (0·13 to 0·18) for 937 patients in the 14-day primaquine group, a difference of 0·02 (-0·02 to 0·05, p=0·3405). The incidence rate for 464 patients in the placebo group was 0·96 (95% CI 0·83 to 1·08) recurrences per person-year. Potentially drug-related serious adverse events within 42 days of starting treatment were reported in nine (1·0%) of 935 patients in the 7-day group, one (0·1%) of 937 in the 14-day group and none of 464 in the control arm. Four of the serious adverse events were significant haemolysis (three in the 7-day group and one in the 14-day group)., Interpretation: In patients with normal G6PD, 7-day primaquine was well tolerated and non-inferior to 14-day primaquine. The short-course regimen might improve adherence and therefore the effectiveness of primaquine for radical cure of P vivax malaria., Funding: UK Department for International Development, UK Medical Research Council, UK National Institute for Health Research, and the Wellcome Trust through the Joint Global Health Trials Scheme (MR/K007424/1) and the Bill & Melinda Gates Foundation (OPP1054404)., (Copyright © 2019 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: The aim was to assess factors affecting disease severity in imported P. falciparum and non-falciparum malaria., Methods: We reviewed medical records from 2793/3260 (85.7%) of all episodes notified in Sweden between 1995 and 2015 and performed multivariable logistic regression., Results: Severe malaria according to WHO 2015 criteria was found in P. falciparum (9.4%), P. vivax (7.7%), P. ovale (5.3%), P. malariae (3.3%), and mixed P. falciparum episodes (21.1%). Factors associated with severe P. falciparum malaria were age <5 years and >40 years, origin in nonendemic country, pregnancy, HIV, region of diagnosis, and health care delay. Moreover, oral treatment of P. falciparum episodes with parasitemia ≥2% without severe signs at presentation was associated with progress to severe malaria with selected criteria. In non-falciparum, age >60 years, health care delay and endemic origin were identified as risk factors for severe disease. Among patients originating in endemic countries, a higher risk for severe malaria, both P. falciparum and non-falciparum, was observed among newly arrived migrants., Conclusions: Severe malaria was observed in P. falciparum and non-falciparum episodes. Current WHO criteria for severe malaria may need optimization to better guide the management of malaria of different species in travelers and migrants in nonendemic areas., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)

Malaria and babesiosis are bloodborne protozoan infections for which the emergence of drug-resistant strains poses a threat. Our previous phage display cDNA screens established the essentiality of Plasmodium falciparum signal peptide peptidase (SPP) in asexual development at the blood stage of malaria infection. Given the structural similarities between SPP inhibitors and HIV protease inhibitors, we screened ten HIV protease inhibitors and selected Lopinavir and Atazanavir for their ability to inhibit PfSPP activity. Using a transcription-based assay, we observed that Lopinavir inhibits both parasite-and host-derived SPP activities whereas Atazanavir inhibited only parasite derived SPP activity. Consistent with their inhibitory effect on Plasmodium growth, both Lopinavir and Atazanavir strongly inhibited intraerythrocytic Babesia microti growth ex vivo. Moreover, Lopinavir prevented the steep rise in Babesia microti parasitemia typically observed in rag1-deficient mice. Our data provide first evidence that inhibition of parasite-derived SPPs by HIV protease inhibitors offers a promising therapeutic avenue for the treatment of severe babesiosis and infections caused by other Apicomplexa parasites., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Studies of the association between malaria in pregnancy (MiP) and malaria during infancy have provided mixed results. A systematic review was conducted to evaluate available evidence on the impact of Plasmodium falciparum malaria infection during pregnancy, and intermittent preventive treatment of malaria during pregnancy (IPTp), on the risk of clinical malaria or parasitaemia during infancy., Methods: MEDLINE, EMBASE, Global Health, and Malaria in Pregnancy Library databases were searched from inception to 22 May 2018 for articles published in English that reported on associations between MiP and malaria risk in infancy. Search terms included malaria, Plasmodium falciparum, pregnancy, placenta, maternal, prenatal, foetal, newborn, infant, child or offspring or preschool. Randomized controlled trials and prospective cohort studies, which followed infants for at least 6 months, were included if any of the following outcomes were reported: incidence of clinical malaria, prevalence of parasitaemia, and time to first episode of parasitaemia or clinical malaria. Substantial heterogeneity between studies precluded meta-analysis. Thus, a narrative synthesis of included studies was conducted., Results: The search yielded 14 published studies, 10 prospective cohort studies and four randomized trials; all were conducted in sub-Saharan Africa. Infants born to mothers with parasitaemia during pregnancy were at higher risk of malaria in three of four studies that assessed this association. Placental malaria detected by microscopy or histology was associated with a higher risk of malaria during infancy in nine of 12 studies, but only one study adjusted for malaria transmission intensity. No statistically significant associations between the use of IPTp or different IPTp regimens and the risk of malaria during infancy were identified., Conclusion: Evidence of an association between MiP and IPTp and risk of malaria in infancy is limited and of variable quality. Most studies did not adequately adjust for malaria transmission intensity shared by mothers and their infants. Further research is needed to confirm or exclude an association between MiP and malaria in infancy. Randomized trials evaluating highly effective interventions aimed at preventing MiP, such as IPTp with dihydroartemisinin-piperaquine, may help to establish a causal association between MiP and malaria in infancy.

Background: Intermittent preventive treatment (IPT) of malaria with dihydroartemisinin-piperaquine is a promising strategy for malaria prevention in young African children. However, the optimal dosing strategy is unclear and conflicting evidence exists regarding the risk of malaria after cessation of chemoprevention. We aimed to compare two dosing strategies of IPT with dihydroartemisinin-piperaquine in young Ugandan children, and to evaluate the risk of malaria after cessation of IPT., Methods: In this double-blind, randomised controlled phase 2 trial, women and their unborn children were recruited at Tororo District Hospital (Tororo, Uganda). Eligible participants were HIV-negative women aged 16 years or older with a viable pregnancy (gestational age 12-20 weeks). Women and their unborn children were randomly assigned (1:1:1:1) to one of four treatment groups, all receiving dihydroartemisinin-piperaquine, on the basis of the IPT intervention received by the woman during pregnancy: women every 8 weeks, children every 4 weeks; women every 4 weeks, children every 4 weeks; women every 8 weeks, children every 12 weeks; and women every 4 weeks, children every 12 weeks. Block randomisation was done by an independent investigator using a computer-generated randomisation list (permuted block sizes of six and 12). We analysed children on the basis of their random assignment to receive dihydroartemisinin-piperaquine (20 mg/160 mg tablets) once daily for 3 consecutive days every 4 weeks or 12 weeks. Children received study drugs from age 8 weeks to 24 months and were followed-up to age 36 months. Participants and investigators were masked to treatment allocation. The primary outcome was the incidence of symptomatic malaria during the intervention and following cessation of the intervention, adjusted for potential confounders. The primary outcome and safety were assessed in the modified intention-to-treat population, which included all children who reached 8 weeks of age and received at least one dose of study drug. This trial is registered with ClinicalTrials.gov, number NCT02163447., Findings: Between Oct 21, 2014, and May 18, 2015, 191 children were born, of whom 183 reached 8 weeks of age and received at least one dose of study drug and thus were included in the primary analysis (96 children in the 4-week group and 87 in the 12-week group). During the intervention, the incidence of symptomatic malaria was significantly lower among children treated every 4 weeks than children treated every 12 weeks; three episodes occurred among children treated every 4 weeks (incidence 0·018 episodes per person-year) compared with 61 episodes among children treated every 12 weeks (incidence 0·39 episodes per person-year; adjusted incidence rate ratio [aIRR] 0·041, 95% CI 0·012-0·150, p<0·0001). After cessation of IPT, children who had previously received dihydroartemisinin-piperaquine every 4 weeks had a lower incidence of symptomatic malaria than children who were treated every 12 weeks; 62 episodes occurred among children previously treated every 4 weeks (incidence 0·73 episodes per person-year) compared with 83 episodes among children treated every 12 weeks (incidence 1·1 episodes per person-year; aIRR 0·62, 0·40-0·95, p=0·028). In the 4-week group, 94 (98%) of 96 children had adverse events versus 87 (100%) of 87 children in the 12-week group. The most commonly reported adverse event was cough in both treatment groups (94 [98%] in the 4-week group vs 87 [100%] in the 12-week group). 16 children had severe adverse events (seven [7%] children in the 4-week group vs nine [10%] children in the 12-week group). No severe adverse events were thought to be related to study drug administration. One death occurred during the intervention (age 8 weeks to 24 months), which was due to respiratory failure unrelated to malaria., Interpretation: IPT with dihydroartemisinin-piperaquine given every 4 weeks was superior to treatment every 12 weeks for the prevention of malaria during childhood, and this protection was extended for up to 1 year after cessation of IPT., Funding: Eunice Kennedy Shriver National Institute of Child Health and Human Development., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Avian malaria is a widespread infection caused by parasites from the Order Haemosporida. Indeed, Neotropical swifts are interesting models for host-parasite coevolution studies due to their unique life history that may allow them to escape parasitism. Considering this, we evaluated haemosporidians from Neotropical swifts. We collected blood and prepared smears from 277 individuals from waterfalls in Brazil. Despite low parasitemia or absence of parasites detected by microscopy haemosporidian infection was detected in 44 swifts (15.8%) using a screening PCR that amplifies a 154-nucleotide segment of ribosomal RNA coding sequence within the mitochondrial DNA (mtDNA) of Plasmodium and Haemoproteus. Although previous studies reported absence of these parasites in European and North American swifts our data suggest that Neotropical swifts are susceptible to haemosporidians. Further studies will add information about evolutionary and ecological aspects of avian haemosporidia in Neotropical Cypseloidinae swifts., (Copyright © 2019 Elsevier B.V. All rights reserved.)

To date, there is no clear standard to monitor drug treatment for canine Chagas disease. We used 2 real-time PCR (rtPCR) assays targeting Trypanosoma cruzi kinetoplast DNA (kDNA) and nuclear satellite DNA (nDNA) to detect T. cruzi in canine whole blood. Samples were collected randomly from 131 untreated dogs with unknown T. cruzi infection status in Texas. The kDNA-based rtPCR was slightly more sensitive (diagnostic sensitivity of kDNA = 49% vs. nDNA = 44%; p = 0.5732) but slightly less specific (diagnostic specificity of kDNA = 96% vs. nDNA = 97%; p > 0.9999) than the nDNA-based rtPCR. However, the differences in sensitivity and specificity between the nDNA- and kDNA-based rtPCR assays were not statistically significant. Using the nDNA- and kDNA-based qualitative rtPCR assays to monitor parasitemia from 137 itraconazole- and amiodarone-treated cases with nDNA- and kDNA-based PCR-positive baselines showed that the PCR positive rate decreased to 0% in 30 d. Using kDNA-based quantitative rtPCR to monitor normalized T. cruzi DNA copies in 4 representative dogs demonstrated that drug treatment could reduce parasite loads within 7-30 d. The kDNA-based qualitative rtPCR may be used for routine parasitemia screening of drug-treated Chagas-positive dogs, whereas nDNA-based qualitative rtPCR may be used for confirmation.

Developing ultrasensitive methods capable of detecting submicroscopic parasitemia-a challenge that persists in low transmission areas, asymptomatic carriers, and patients showing recrudescence-is vital to achieving malaria eradication. Nucleic acid amplification techniques offer improved analytical sensitivity but are limited by the number of copies of the amplification targets. Herein, we perform a novel genome mining approach to identify a pair of identical multirepeat sequences (IMRSs) that constitute 170 and 123 copies in the Plasmodium falciparum genome and explore their potential as primers for PCR. Real-time quantitative PCR analyses have shown the ability of P. falciparum IMRSs to amplify as low as 2.54 fg of P. falciparum genomic DNA (approximately 0.1 parasite), with a striking 100-fold increase in detection limit when compared with P. falciparum 18S rRNA (251.4 fg; approximately 10 parasites). Validation with clinical samples from malaria-endemic regions has shown 6.70 ± 1.66 cycle better detection threshold in terms of Ct value for P. falciparum IMRSs, with approximately 100% sensitivity and specificity. Plasmodium falciparum IMRS assays are also capable of detecting submicroscopic infections in asymptomatic samples. To summarize, this approach of initiating amplification at multiple loci across the genome and generating more products with increased analytical sensitivity is different from classic approaches amplifying multicopy genes or tandem repeats. This can serve as a platform technology to develop advanced diagnostics for various pathogens., (Copyright © 2019 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

Background: Plasmodium elongatum (cytochrome b lineage pGRW6) is a widespread avian malaria parasite, often causing severe disease in non-adapted hosts. This parasite lineage is of global distribution however, its virulence remains insufficiently understood, particularly in wild birds. Surprisingly, this infection has never been reported in Common starlings Sturnus vulgaris and Common crossbills Loxia curvirostra, common European songbirds which were extensively sampled across Europe. A hypothesis was proposed that these birds might be resistant to the pGRW6 infection. The aim of this study was to test this hypothesis., Methods: Lineage pGRW6 was isolated from a naturally infected Eurasian reed warbler, multiplied in vivo and inoculated in Common starlings and Common crossbills. Experimental and control groups (8 birds in each) were maintained in controlled conditions and examined microscopically every 4 days. Haematocrit value and body mass were monitored in parallel. At the end of the experiment (44 days post exposure), samples of internal organs were collected and examined using histological methods for possible presence of phanerozoites., Results: All control birds remained uninfected. Experimental starlings were resistant. All exposed crossbills were susceptible and survived until the end of this study. Prepatent period was 12-16 days post exposure. Light parasitaemia (< 0.7%) developed in all birds, and only few phanerozoites were seen in bone marrow cells of 5 of 8 experimentally infected crossbills. Significant changes were reported only in haematocrit value but not body mass in the exposed crossbills compared to controls., Conclusion: Plasmodium elongatum (pGRW6) is of low virulence in Common crossbills and is unable to develop in Common starlings, indicating innate resistance of the later bird species. Low virulence in Common crossbills is likely due to the inability or low ability of this parasite lineage to develop phanerozoites resulting in light (if at all) damage of stem bone marrow cells. This study suggests that susceptibility of different bird species to the lineage pGRW6 is markedly variable. The global distribution of this parasite might be due to low virulence in wild adapted avian hosts, which survive this infection and serve as reservoirs host for non-adapted birds in whom this infection is often lethal.

When administered alone, preinfection exercise training and benznidazole-based chemotherapy induce cardioprotection in Chagas disease. However, the effect of concomitant exercise and benznidazole treatment is unknown. We investigated whether exercise and specific chemotherapy could interact to modulate parasitemia, inflammation, redox status and heart damage in a murine model of T. cruzi infection. Wistar rats were randomized into an uninfected control group (CNT) and four groups infected with T. cruzi: sedentary untreated (SUN) and treated (STR), and trained untreated (TUN) and treated (TTR). Running training was administered 5 days/week for 4 weeks. Treated animals concomitantly received 100 mg/kg/day benznidazole. Heart inflammation and reactive damage were not detected in CNT animals. Compared to SUN, TUN animals presented increased levels of parasitemia, myocarditis, nitric oxide, hydrogen peroxide, protein carbonyl, malondialdehyde, cytokines (IFN-γ, TNF-α, IL-4, IL-6, IL-10 and IL-17), catalase, superoxide dismutase and glutathione reductase activity, as well as reduced heart non-protein antioxidant levels (P < 0.05). TTR animals exhibited higher levels of parasitemia, myocarditis, hydrogen peroxide, malondialdehyde, IFN-γ, TNF-α and IL-6 than STR animals (P < 0.05), which showed the lowest levels of all analyzed parameters compared to the other groups (P < 0.05). Our findings indicate that exercise aggravates acute infection. When concomitantly administered with benznidazole, exercise training impaired parasitic control and chemotherapy-induced cardioprotection in T. cruzi-infected rats. Considering that exercise training and T. cruzi infection constitute independent metabolic challenges, the negative effects of concomitant treatment are potentially related to the overlapping oxidative and immunoinflammatory demands of exercise and the infection itself., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: The Mindray BC-6800 haematology analyzer (BC-6800) provides a dedicated flag 'Infected RBC' (InR) and the number of InR (InR#)/the permillage of InR (InR‰) in routine blood testing as a screening tool for malaria in endemic areas. This study sought to evaluate the effectiveness of the BC-6800 flag parameter for aiding the diagnosis of malaria., Methods: A total of 181 samples were tested using the Mindray BC-6800 haematology analyzer, including 117 malaria-infected samples collected from Yunnan, China, and 64 samples from healthy controls. Microscopy examination was conducted as reference when stained thick blood film revealed the presence of malaria parasites identified as Plasmodium vivax and Plasmodium falciparum. The receiver operating characteristic (ROC) curve analysis was developed using Analyse-it v4.92.3. The Kappa value was determined to evaluate the agreement between BC-6800 and light microscopy., Results: The sensitivity of InR‰ generated by BC-6800 for P. vivax and P. falciparum was 88.3 and 24.1%, respectively; specificity of InR‰ for malaria parasites was 84.3 and 84.3%, respectively; positive predictive value and negative predictive value was 89.4 and 82.7% for P. vivax, and 52.8 and 60.3% for P. falciparum. There was a strong correlation between ΔWBC and InR‰ (R 2  = 0.9731 for P. vivax and R 2  = 0.9757 for P. falciparum). There was also a significant correlation between parasitaemia and InR# in P. vivax-infected samples (R 2  = 0.734). InR# was evaluated using ROC curve analysis, the area under the ROC curve is 0.95 with a 95% confidence interval of 0.926 to 0.974, and the cut-off value is 0.01 × 10 9 /L for P. vivax. However, the ring stage and the early trophozoite stage of Plasmodium cannot be detected easily on BC-6800, possibly because of the small size and low nucleic acid content of these stages., Conclusions: The findings suggest that the flag 'InR' and the parameters 'InR#/InR‰' provided by the BC-6800 haematology analyzer could be used to screen for malaria in a clinical setting.

Background: More than 200 medicinal plants including Euphorbia abyssinica are utilized for treatment of malaria in Ethiopian traditional medical practices. However, the safety, efficacy and quality of these medicinal plants are largely unknown. Pharmacological and toxicological investigations of these plants are among the prioritized issues in every country. The aim of this study was, therefore, to evaluate the anti-malarial activity of Euphorbia abyssinica root extract against Plasmodium berghei infection in mice., Methods: The fresh roots of Euphorbia abyssinica were identified and collected. They were dried and extracted by 80% methanol using maceration. Acute toxicity of the extract was done using female Swiss albino mice. Anti-malarial activity of the extract was done by a standard 4-day suppressive test using chloroquine-sensitive Plasmodium berghei. Twenty-five male Swiss albino mice were randomly grouped into 5 groups of 5 mice each. Group I was treated with distilled water (10 ml/kg), group II, III, and IV were treated with 200, 400, and 600 mg/kg of extract, respectively and group V was treated with chloroquine (25 mg/kg). The level of parasitaemia, survival time, and variation in weight were utilized to determine the anti-malarial activity of the extract. Data was analysed using ANOVA followed by Tukey test., Results: The plant extract did not show any sign of toxicity and mortality at 2000 mg/kg. The 4-day chemosuppressive anti-malarial activities produced by the crude extract were 66.87% (P < 0.001), 84.94% (P < 0.001) and 93.69% (P < 0.001) at 200, 400 and 600 mg/kg extract, respectively, compared to distilled water treated group. Mice treated with 400 mg/kg (P < 0.01), and 600 mg/kg extract (P < 0.001) showed significant chemosuppressive anti-malarial activity variations as compared to mice treated with 200 mg/kg extract. Mice treated with 600 mg/kg extract significantly (P < 0.001) lived longer than distilled water treated mice. However, the crude extract did not cause any significant change on body weights of mice., Conclusions: From this study, it can be concluded that the root of Euphorbia abyssinica showed very good 4-day chemosuppressive anti-malarial activity. The plant might contain biologically active compounds which are relevant for treatment of malaria. Further phytochemical, toxicological and pharmacological investigations are, therefore, required to evaluate its anti-malarial potential.

Post-transcriptional regulons coordinate the expression of groups of genes in eukaryotic cells, yet relatively few have been characterized. Parasitic trypanosomatids are particularly good models for studies on such mechanisms because they exhibit almost exclusive polycistronic, and unregulated, transcription. Here, we identify the Trypanosoma brucei ZC3H39/40 RNA-binding proteins as regulators of the respiratome; the mitochondrial electron transport chain (complexes I-IV) and the FoF1-ATP synthase (complex V). A high-throughput RNAi screen initially implicated both ZC3H proteins in variant surface glycoprotein (VSG) gene silencing. This link was confirmed and both proteins were shown to form a cytoplasmic ZC3H39/40 complex. Transcriptome and mRNA-interactome analyses indicated that the impact on VSG silencing was indirect, while the ZC3H39/40 complex specifically bound and stabilized transcripts encoding respiratome-complexes. Quantitative proteomic analyses revealed specific positive control of >20 components from complexes I, II and V. Our findings establish a link between the mitochondrial respiratome and VSG gene silencing in bloodstream form T. brucei. They also reveal a major respiratome regulon controlled by the conserved trypanosomatid ZC3H39/40 RNA-binding proteins., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Agro-ecological systems have been associated with increased malaria intensity. This study determined association between different agro-ecological systems, prevalence of malaria parasitaemia and anaemia in Mvomero district, Tanzania., Methods: The study was carried out in three agro-ecosystems namely, savannah, rice-irrigation, and sugarcane. Malaria and anaemia prevalence were measured in four seasons of a year. Villages were categorized according to environmental characteristics, proportion of water-shaded areas and agro-ecosystems. Mixed-effects logistic regression analysis was used to determine factors associated with malaria infection., Results: A total of 7888 individuals were involved with the overall malaria prevalence of 34.4%. Plasmodium falciparum was the dominant (99.52%) malaria species. Malaria prevalence was highest (42.9%) in children of 10-15 years of age, and significantly low during dry and hot season. Of the infected individuals, 78.1% were from rice-irrigation, 18.7% savannah and 3.2% sugarcane ecosystem. Individuals living in villages with high levels of water-shaded areas had highest malaria risk. Over three-quarters (78.9%) of the individuals slept under a mosquito net, with the highest (88.5%) coverage among individuals in sugarcane ecosystem. On average 47.1% of the children were anaemic. Anaemia was more prevalent (60.5%) among individuals in the savannah than in the rice-irrigation (48.2%) or sugarcane communities (23%). Analysis indicated that ecosystems and levels of water-shaded area were highly correlated, and altered levels of malaria infection. Gender, age, mosquito net-use, and season were other significant determinants of P. falciparum infection. Males had higher odds than females (OR = 1.16, 95% CI 1.05, 1.29). The risk for children 6-9 years and older children (10-15 years) was over 50% and 24%, respectively, higher compared to young ones (0-5 years). Use of mosquito net reduced malaria risk by 26%. The risk of infection was higher during dry and cool season (OR = 1.92, 95 %CI 1.66, 2.23) compared to other seasons. Living in villages with high level of water-shaded areas increased the chances of getting malaria up to 15 times than living in drier areas. Similarly, infection odds increased when living in savannah and rice-irrigation ecosystems than in the sugarcane ecosystem., Conclusions: Findings show significant variations in malaria prevalence between communities living in different agro-ecosystems within the same district. Local malaria control strategies should consider these variations and liaise with agricultural experts while designing interventions to maximize effectiveness.

In epidemiological surveys and surveillance the application of molecular tools is essential in detecting submicroscopic malaria. A genus-specific conventional cytochrome b (cytb) PCR has shown high sensitivity in field studies, detecting 70% submicroscopic malaria. The main objective of this study was to assess the conversion from conventional to real-time PCR testing both SYBR and probe protocols, and including quantitative (q) PCR. The protocols were assessed applying well-defined clinical patient material consisting of 33 positive and 80 negative samples. Sequencing of positive PCR products was performed. In addition, a sensitivity comparison of real-time PCR methods was done by including five relevant assays investigating the effect of amplification target and platform. Sensitivity was further examined using field material consisting of 111 P.falciparum positive samples from Tanzanian children (< 5 years), as well as using related patient data to assess the application of q-PCR with focus on low-level parasitaemia. Both the cytb SYBR and probe PCR protocols showed as high sensitivity and specificity as their conventional counterpart, except missing one P. malariae sample. The SYBR protocol was more sensitive and specific than using probe. Overall, choice of amplification target applied is relevant for achieving ultra-sensitivity, and using intercalating fluorescence dye rather than labelled hydrolysis probes is favourable. Application of q-PCR analysis in field projects is important for the awareness and understanding of low-level parasitaemia. For use in clinical diagnosis and epidemiological studies the highly sensitive and user-friendly cytb SYBR q-PCR method is a relevant tool. The genus-specific method has the advantage that species identification by sequencing can be performed as an alternative to species-specific PCR., Competing Interests: The authors have declared that no competing interests exist.

Background: Placental malaria is a major cause of adverse birth outcomes. However, data are limited on the relationships between longitudinal measures of parasitemia during pregnancy and placental malaria., Methods: Data came from 637 women enrolled in a randomized controlled trial of intermittent preventive treatment of malaria in pregnancy (IPTp) from Uganda. Plasmodium falciparum parasitemia was assessed using microscopy and ultrasensitive quantitative PCR at intervals of 28 days from 12 to 20 weeks gestation through delivery. Multivariate analysis was used to measure associations between characteristics of parasitemia during pregnancy and the risk of placental malaria based on histopathology., Results: Overall risk of placental malaria was 44.6%. None of the 34 women without parasitemia detected during pregnancy had evidence of placental malaria. Increasing proportion of interval assessments with parasitemia and higher parasite densities were independently associated with an increased risk of placental malaria. Higher gravidity and more effective IPTp were associated with a decreased risk of placental malaria. Women with parasitemia only detected before the third trimester still had an increased risk of placental malaria., Conclusions: The frequency, density, and timing of parasitemia are all important risk factors for placental malaria. Interventions should target the prevention of all levels of parasitemia throughout pregnancy., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: Tafenoquine (TQ) was recently approved by the US Food and Drug Administration for prophylaxis of malaria and, in addition, for eradication of the hepatic phase of the relevant Plasmodium species. In this study, we evaluated the efficacy of TQ for treatment of Babesia microti infection in mice with severe combined immunodeficiency (SCID)., Methods: SCID mice were infected with 1.1-1.5 × 108 B. microti-infected red blood cells by intraperitoneal injection. On day 3 or 4 postinfection, when parasitemia levels typically exceeded 10%, mice were treated with TQ vs vehicle alone, both administered by oral gavage., Results: A single dose of TQ completely eliminated detectable parasites, with a >90% reduction in the level of parasitemia within just 4 days. Before elimination, a conspicuous phenotypic change in the parasite was observed. Although parasitologic cure was not achieved, there was no evidence for the development of drug resistance., Conclusions: This study suggests that TQ may be a highly useful drug to treat B. microti infection in patients. If further animal studies establish that a marked reduction in B. microti parasitemia can be reliably achieved with peak blood levels of TQ known to be well tolerated in humans, a clinical trial in patients should be considered., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)