RNAscope in situ hybridization reveals microvascular sequestration of Plasmodium relictum pSGS1 blood stages but absence of exo-erythrocytic dormant stages during latent infection of Serinus canaria.

BACKGROUND: Birds chronically infected with avian malaria parasites often show relapses of parasitaemia after latent stages marked by absence of parasites in the peripheral circulation. These relapses are assumed to result from the activation of dormant exo-erythrocytic stages produced during secondary (post-erythrocytic) merogony of avian Plasmodium spp. Yet, there is no morphological proof of persistent or dormant tissue stages in the avian host during latent infections. This study investigated persistence of Plasmodium relictum pSGS1 in birds with latent infections during winter, with the goal to detect presumed persisting tissue stages using a highly sensitive RNAscope® in situ hybridization technology. METHODS: Fourteen domestic canaries were infected with P. relictum pSGS1 by blood-inoculation in spring, and blood films examined during the first 4 months post infection, and during winter and spring of the following year. After parasitaemia was no longer detectable, half of the birds were dissected, and tissue samples investigated for persisting tissue stages using RNAscope ISH and histology. The remaining birds were blood-checked and dissected after re-appearance of parasitaemia, and their tissues equally examined. RESULTS: Systematic examination of tissues showed no exo-erythrocytic stages in birds exhibiting latent infections by blood-film microscopy, indicating absence of dormant tissue stages in P. relictum pSGS1-infected canaries. Instead, RNAscope ISH revealed rare P. relictum blood stages in capillaries of various tissues and organs, demonstrating persistence of the parasites in the microvasculature. Birds examined after re-appearance of parasitemia showed higher numbers of P. relictum blood stages in both capillaries and larger blood vessels, indicating replication during early spring and re-appearance in the peripheral circulation. CONCLUSIONS: The findings suggest that persistence of P. relictum pSGS1 during latent infection is mediated by continuous low-level erythrocytic merogony and possibly tissue sequestration of infected blood cells. Re-appearance of parasitaemia in spring seems to result from increased erythrocytic merogony, therefore representing recrudescence and not relapse in blood-inoculated canaries. Further, the study highlights strengths and limitations of the RNAscope ISH technology for the detection of rare parasite stages in tissues, providing directions for future research on persistence and tissue sequestration of avian malaria and related haemosporidian parasites.

Variability in white blood cell count during uncomplicated malaria and implications for parasite density estimation: a WorldWide Antimalarial Resistance Network individual patient data meta-analysis.

BACKGROUND: The World Health Organization (WHO) recommends that when peripheral malarial parasitaemia is quantified by thick film microscopy, an actual white blood cell (WBC) count from a concurrently collected blood sample is used in calculations. However, in resource-limited settings an assumed WBC count is often used instead. The aim of this study was to describe the variability in WBC count during acute uncomplicated malaria, and estimate the impact of using an assumed value of WBC on estimates of parasite density and clearance. METHODS: Uncomplicated malaria drug efficacy studies that measured WBC count were selected from the WorldWide Antimalarial Resistance Network data repository for an individual patient data meta-analysis of WBC counts. Regression models with random intercepts for study-site were used to assess WBC count variability at presentation and during follow-up. Inflation factors for parasitaemia density, and clearance estimates were calculated for methods using assumed WBC counts (8000 cells/µL and age-stratified values) using estimates derived from the measured WBC value as reference. RESULTS: Eighty-four studies enrolling 27,656 patients with clinically uncomplicated malaria were included. Geometric mean WBC counts (× 1000 cells/µL) in age groups < 1, 1-4, 5-14 and ≥ 15 years were 10.5, 8.3, 7.1, 5.7 and 7.5, 7.0, 6.5, 6.0 for individuals with falciparum (n = 24,978) and vivax (n = 2678) malaria, respectively. At presentation, higher WBC counts were seen among patients with higher parasitaemia, severe anaemia and, for individuals with vivax malaria, in regions with shorter regional relapse periodicity. Among falciparum malaria patients, using an assumed WBC count of 8000 cells/µL resulted in parasite density underestimation by a median (IQR) of 26% (4-41%) in infants < 1 year old but an overestimation by 50% (16-91%) in adults aged ≥ 15 years. Use of age-stratified assumed WBC values removed systematic bias but did not improve precision of parasitaemia estimation. Imprecision of parasite clearance estimates was only affected by the within-patient WBC variability over time, and remained < 10% for 79% of patients. CONCLUSIONS: Using an assumed WBC value for parasite density estimation from a thick smear may lead to underdiagnosis of hyperparasitaemia and could adversely affect clinical management; but does not result in clinically consequential inaccuracies in the estimation of the prevalence of prolonged parasite clearance and artemisinin resistance.

Unravelling factors associated with malaria parasitaemia among children 6-24 months to inform malaria interventions in Nigeria: evidence from 2021 Malaria Indicator Survey.

BACKGROUND: As an additional two million malaria cases were reported in 2021 compared to the previous year, concerted efforts toward achieving a steady decline in malaria cases are needed to achieve malaria elimination goals. This work aimed at determining the factors associated with malaria parasitaemia among children 6-24 months for better targeting of malaria interventions. METHODS: A cross-sectional study analysed 2021 Nigeria Malaria Indicator Survey dataset. Data from 3058 children 6-24 months were analyzed. The outcome variable was children 6-24 months whose parasitaemia was determined using a rapid diagnostic test (RDT). Independent variables include child age in months, mothers' age, mothers' education, region, place of residence, household ownership and child use of insecticide-treated net (ITN), exposure to malaria messages and knowledge of ways to prevent malaria. Logistic regression analysis was conducted to examine possible factors associated with malaria parasitaemia in children 6-24 months. RESULTS: Findings revealed that 28.7% of the 3058 children aged 6-24 months tested positive for malaria by RDT. About 63% of children 12-17 months (aOR = 1.63, 95% CI 1.31-2.03) and 91% of children 18 to 24 months (aOR = 1.91, 95% CI 1.51-2.42) were more likely to have a positive malaria test result. Positive malaria test result was also more likely in rural areas (aOR = 1.79, 95% CI 2.02-24.46), northeast (aOR = 1.54, 95% CI 1.02-2.31) and northwest (aOR = 1.63, 95% CI 1.10-2.40) region. In addition, about 39% of children who slept under ITN had a positive malaria test result (aOR = 1.39 95% CI 1.01-1.90). While children of mothers with secondary (aOR = 0.40, 95% CI 0.29-0.56) and higher (aOR = 0.26, 95% CI 0.16-0.43) levels of education and mothers who were aware of ways of avoiding malaria (aOR = 0.69, 95% CI 0.53-0.90) were less likely to have a malaria positive test result. CONCLUSION: As older children 12 to 24 months, children residing in the rural, northeast, and northwest region are more likely to have malaria, additional intervention should target them in an effort to end malaria.

Sub-microscopic Plasmodium falciparum parasitaemia, dihydropteroate synthase (dhps) resistance mutations to sulfadoxine-pyrimethamine, transmission intensity and risk of malaria infection in pregnancy in Mount Cameroon Region.

BACKGROUND: Plasmodium falciparum resistance to intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) continues to spread throughout sub-Saharan Africa. This study assessed the occurrence of microscopic and sub-microscopic P. falciparum parasitaemia, dihydropteroate synthase mutations associated with resistance to SP and maternal anaemia in the Mount Cameroon area. METHODS: Consenting pregnant women living in semi-rural and semi-urban/urbanized settings were enrolled in this cross-sectional study. Socio-demographic, antenatal and clinical data were documented. Microscopic and sub-microscopic parasitaemia were diagnosed using peripheral blood microscopy and nested polymerase chain reaction (PCR) respectively. The dhps mutations were genotyped by restriction fragment length polymorphism analysis. The presence of A437G, K540E, and A581G was considered a marker for high-level resistance. Haemoglobin levels and anaemia status were determined. RESULTS: Among the women, the prevalence of microscopic and sub-microscopic P. falciparum infection were 7.7% (67/874) and 18.6% (93/500) respectively. Predictors of microscopic infection were younger age (< 21 years) (AOR = 2.89; 95% CI 1.29-6.46) and semi-rural settings (AOR = 2.27; 95% CI 1.31-3.96). Determinants of sub-microscopic infection were the rainy season (AOR, 3.01; 95% CI 1.77-5.13), primigravidity (AOR = 0.45; 95% CI 0.21-0.94) and regular ITN usage (AOR = 0.49; 95% CI 0.27-0.90). Of the145 P. falciparum isolates genotyped, 66.9% (97) carried mutations associated with resistance to SP; 33.8% (49), 0%, 52.4% (76) and 19.3% (28) for A437G, K540E, A581G and A437G + A581G respectively. The A581G mutation was associated with ≥ 3 SP doses evident only among sub-microscopic parasitaemia (P = 0.027) and multigravidae (P = 0.009). Women with microscopic infection were more likely from semi-rural settings (AOR = 7.09; 95% CI 2.59-19.42), to report history of fever (AOR = 2.6; 95% CI 1.07-6.31), to harbour parasites with double resistant mutations (AOR = 6.65; 95% CI 1.85-23.96) and were less likely to have received 2 SP doses (AOR = 0.29; 95% CI 1.07-6.31). Microscopic infection decreased Hb levels more than sub-microscopic infection. CONCLUSION: The occurrence of sub-microscopic P. falciparum parasites resistant to SP and intense malaria transmission poses persistent risk of malaria infection during pregnancy in the area. ITN usage and monitoring spread of resistance are critical.

Performance of a novel melting curve-based qPCR assay for malaria parasites in routine clinical practice in non-endemic setting.

BACKGROUND: High-quality malaria diagnosis is essential for effective treatment and clinical disease management. Microscopy and rapid diagnostic tests are the conventional methods performed as first-line malaria diagnostics in non-endemic countries. However, these methods lack the characteristic to detect very low parasitaemia, and accurate identification of the Plasmodium species can be difficult. This study evaluated the performance of the MC004 melting curve-based qPCR for the diagnosis of malaria in routine clinical practice in non-endemic setting. METHODS AND RESULTS: Whole blood samples were collected from 304 patients with clinical suspicion of malaria and analysed by both the MC004 assay and conventional diagnostics. Two discrepancies were found between the MC004 assay and microscopy. Repeated microscopic analysis confirmed the qPCR results. Comparison of the parasitaemia of nineteen Plasmodium falciparum samples determined by both microscopy and qPCR showed the potential of the MC004 assay to estimate the parasite load of P. falciparum. Eight Plasmodium infected patients were followed after anti-malarial treatment by the MC004 assay and microscopy. The MC004 assay still detected Plasmodium DNA although no parasites were seen with microscopy in post-treatment samples. The rapid decline in Plasmodium DNA showed the potential for therapy-monitoring. CONCLUSION: Implementation of the MC004 assay in non-endemic clinical setting improved the diagnosis of malaria. The MC004 assay demonstrated superior Plasmodium species identification, the ability to indicate the Plasmodium parasite load, and can potentially detect submicroscopic Plasmodium infections.

Haematological parameters and their correlation with the degree of malaria parasitaemia among outpatients attending a polyclinic.

BACKGROUND: Malaria is a parasitic disease caused by various species of the blood parasite Plasmodium; of all the parasitic diseases, malaria has the highest prevalence and mortality with an estimated 247 million cases and 619,000 deaths recorded worldwide as of 2021. Malaria causes febrile illness with several changes in blood cell parameters. Some of these changes include leucopenia, thrombocytopenia, and anaemia. If these changes could be correlated with the degree of parasitaemia, it can serve as a guide to physicians when treating malaria. This study was therefore aimed at correlating haematological parameters with levels of parasitaemia during malaria infection. METHODS: The study was a cross-sectional study involving 89 malaria positive patients. About 5 ml of blood was collected from each participant who gave his or her informed consent to partake in the study. A full blood count was performed on their samples to determine their haematological parameters using a haematology auto-analyzer. A parasite count was also performed via microscopy to determine the degree of parasitaemia. The data obtained from the study was entered into a database and statistically analysed using Statistical Package for Social Sciences (SPSS) version 23 and Microsoft Excel 2016. RESULTS: The study comprised of 89 participants out of which 35 were males and 54 were females with the mean age of 26.15 years. Secondary education participants were the highest with quaternary education the lowest. The highest parasite count recorded was 398,174 parasites/µl of blood, lowest count was 101 with the average being 32,942.32584. There was also a significant positive Pearson's correlation between total WBC and parasitaemia and with the WBC differentials, neutrophils, lymphocytes and monocytes had positive correlations while eosinophils and basophils had negative correlations. Furthermore, platelets, total RBC's, haemoglobin, MCH, MCHC and Hct all showed negative correlations. Linear regression also showed a linear relationship between parasite density and the various haematological parameters. CONCLUSION: The linear relationship (correlation) between WBC and MCH were the only significant ones at 95% and 99% confidence interval, respectively based on a two-tail t-test. Also, based on the regression analysis, the changes caused by WBC and PLT were the only significant changes at 95% confidence level in a two-tailed t-test.

PET-PCR reveals low parasitaemia and submicroscopic malarial infections in Honduran Moskitia.

BACKGROUND: Malaria remains a main parasitic disease of humans. Although the largest number of cases is reported in the African region, there are still endemic foci in the Americas. Central America reported 36,000 malaria cases in 2020, which represents 5.5% of cases in the Americas and 0.015% of cases globally. Most malaria infections in Central America are reported in La Moskitia, shared by Honduras and Nicaragua. In the Honduran Moskitia, less than 800 cases were registered in 2020, considering it an area of low endemicity. In low endemicity settings, the number of submicroscopic and asymptomatic infections tends to increase, leaving many cases undetected and untreated. These reservoirs challenge national malaria elimination programmes. This study aimed to assess the diagnostic performance of Light Microscopy (LM), a nested PCR test and a photoinduced electron transfer polymerase chain reaction (PET-PCR) in a population of febrile patients from La Moskitia. METHODS: A total of 309 febrile participants were recruited using a passive surveillance approach at the Puerto Lempira hospital. Blood samples were analysed by LM, nested PCR, and PET-PCR. Diagnostic performance including sensitivity, specificity, negative and positive predictive values, kappa index, accuracy, and ROC analysis was evaluated. The parasitaemia of the positive samples was quantified by both LM and PET-PCR. RESULTS: The overall prevalence of malaria was 19.1% by LM, 27.8% by nPCR, and 31.1% by PET-PCR. The sensitivity of LM was 67.4% compared to nPCR, and the sensitivity of LM and nPCR was 59.6% and 80.8%, respectively, compared to PET-PCR. LM showed a kappa index of 0.67, with a moderate level of agreement. Forty positive cases by PET-PCR were not detected by LM. CONCLUSIONS: This study demonstrated that LM is unable to detect parasitaemia at low levels and that there is a high degree of submicroscopic infections in the Honduran Moskitia.

Vivax malaria in Duffy-negative patients shows invariably low asexual parasitaemia: implication towards malaria control in Ethiopia.

BACKGROUND: The increase in detections of Plasmodium vivax infection in Duffy-negative individuals in Africa has challenged the dogma establishing the unique P. vivax Duffy Binding Protein-Duffy antigen receptor for chemokines (PvDBP-DARC) pathway used by P. vivax merozoites to invade reticulocytes. Information on the impact of Duffy antigen polymorphisms on the epidemiology of P. vivax malaria remains elusive. The objective of this study was to determine the distribution of asexual parasitaemia of P. vivax according to the Duffy antigen polymorphisms in Ethiopia. METHODS: DNA was extracted from dried blood spots (DBS) collected from prospectively recruited 138 P. vivax-infected patients from health centres. The identification and estimation of P. vivax asexual parasitaemia were performed by microscopic examination and quantitative real-time polymerase chain reaction (PCR). Duffy genotyping was conducted by DNA sequencing in a total of 138 P.vivax infected samples. RESULTS: The proportion of Duffy-negatives (FY*BES/FY*BES) in P. vivax infected patients was 2.9% (4/138). Duffy genotype FY*B/FY*BES (48.6%) was the most common, followed by FY*A/FY*BES genotype (25.4%). In one patient, the FY*02 W.01/FY*02 N.01 genotype conferring a weak expression of the Fyb antigen was observed. All P.vivax infected Duffy-negative patients showed low asexual parasitaemia (≤ 110 parasites/µL). The median P. vivax parasitaemia in Duffy-negative patients (53 parasites/µL) was significantly lower than those found in homozygous and heterozygous individuals (P < 0.0001). CONCLUSION: Plasmodium vivax in Duffy-negative patients shows invariably low asexual parasitaemia. This finding suggests that the pathway used by P. vivax to invade Duffy-negative reticulocytes is much less efficient than that used in Duffy-positives. Moreover, the low asexual parasitaemia observed in Duffy-negative individuals could constitute an 'undetected silent reservoir', thus likely delaying the elimination of vivax malaria in Ethiopia.

Trends of malaria infection in pregnancy in Ghana over the past two decades: a review.

BACKGROUND: There has been a global decline in malaria transmission over the past decade. However, not much is known of the impact of this observation on the burden of malaria infection in pregnancy in endemic regions including Ghana. A narrative review was undertaken to help describe trends in malaria infection in pregnancy in Ghana. Among others, such information is important in showing any progress made in malaria in pregnancy control. METHODS: To describe trends in pregnancy-associated malaria infection in Ghana, a search and review of literature reporting data on the prevalence of asymptomatic Plasmodium falciparum infection in pregnancy was conducted. RESULTS: Thirty-six (36) studies, conducted over 1994-2019, were included in the review. In the northern savannah zone with largely seasonal malaria transmission, prevalence appeared to reduce from about 50-60% in 1994-2010 to 13-26% by 2019. In the middle transitional/forest zone, where transmission is perennial with peaks in the rainy season, prevalence apparently reduced from 60% in the late 1990 s to about 5-20% by 2018. In the coastal savannah area, there was apparent reduction from 28 to 35% in 2003-2010 to 5-11% by 2018-2019. The burden of malaria infection in pregnancy continues to be highest among teenagers and younger-aged pregnant women and paucigravidae. CONCLUSIONS: There appears to be a decline in asymptomatic parasite prevalence in pregnancy in Ghana though this has not been uniform across the different transmission zones. The greatest declines were noticeably in urban settings. Submicroscopic parasitaemia remains a challenge for control efforts. Further studies are needed to evaluate the impact of the reduced parasite prevalence on maternal anaemia and low birthweight and to assess the local burden of submicroscopic parasitaemia in relation to pregnancy outcomes.

Heavy Malaria Parasitaemia in Young Nigerian Infants: Prevalence, Determinants and Implication for the Health System.

BACKGROUND: Infants who are aged six months and below are often protected from malaria and usually present with light parasitaemia when infected. However, complications following heavy malaria parasitaemia in this age group are being increasingly reported. This study set out to determine the prevalence, determinants and the public health implications of heavy malaria parasitaemia in young infants (aged one to six months) at the Wesley Guild Hospital, Ilesa (a unit of the Obafemi Awolowo University Teaching Hospitals Complex). METHODS: Ill infants aged one to six months in out-patient and in-patient care were recruited over an 11-month period. Clinical examinations and blood film for malaria parasite were done for all the study participants. Heavy parasitaemia was defined as > 5000 parasites/µl. Clinical predictors of heavy parasitaemia were determined. RESULTS: Heavy parasitaemia was observed in 16(23.9%) of the sixty-seven participants with malaria infection. Presence of fever at presentation (p=0.007), excessive crying (p=0.003) and pallor (p=0.001) were associated with heavy malaria parasitaemia. However, pallor (OR = 20.653; 95%CI 2.091-203.958; p=0.010) was the only independent predictor of heavy parasitaemia among the young infants. CONCLUSION: About one-in-four ill young infants with malaria had heavy parasitaemia, which was predicted by pallor. Hence, the presence of pallor and factors related to low parental socio-economic status should increase the suspicion of heavy malaria parasitaemia in ill young infants in malaria endemic settings.

CONTEXTE: Nourrissons âgés de six mois et moins sont souvent protégés du paludisme et généralement présents avec de la lumière parasitémie lorsqu'il est infecté. Cependant, les complications qui suivent une parasitémie palustre lourde dans ce groupe d'âge est en cours de plus en plus signalés. Cette étude visait à déterminer la prévalence, les déterminants et les répercussions de l'action sur la santé publique parasitémie palustre sévère chez les jeunes nourrissons (âgés de un à six ans)mois) à l'hôpital Wesley Guild, Ilesa (une unité de l'Obafemi Complexe des hôpitaux universitaires d'Awolowo). MÉTHODES: Nourrissons malades âgés de un à six mois en ambulatoire et les soins aux patients hospitalisés ont été recrutés sur une période de 11 mois. Les examens cliniques et le film sanguin pour le parasite du paludisme ont été fait pour tous les participants à l'étude. La parasitémie lourde était défini comme > 5000 parasites/µl. Prédicteurs cliniques de lourd la parasitémie a été déterminée. RÉSULTATS: Une parasitémie sévère a été observée chez 16 (23,9%) des soixante-sept participants atteints d'une infection palustre. Présence de fièvre à la présentation (p = 0,007), pleurs excessifs (p = 0,003) et la pâleur (p = 0,001) était associée à un paludisme lourdparasitémie. Cependant, pâleur (OR = 20,653; IC à 95 % 2,091-203.958; p=0,010) était le seul prédicteur indépendant de parasitémie chez les jeunes nourrissons. CONCLUSION: Environ un jeune nourrisson malade sur quatre atteint de paludisme avait une parasitémie lourde, qui était prédite par pâleur. D'où la présence de pâleur et de facteurs liés à un faible niveau parental le statut socio-économique devrait accroître la suspicion de lourd parasitémie palustre chez les jeunes nourrissons malades dans le paludisme endémique Paramètres. Mots-clés: Jeunes nourrissons, Parasitémie palustre, Parasite lourddensité, prévalence.

Evaluation of Asymptomatic Malaria Antigenaemia and Parasitaemia in Human Immunodeficiency Virus-Infected Children in Benin City, Nigeria.

BACKGROUND: Malaria and HIV/AIDS are rampant in subSaharan Africa with prevalence of one reinforcing the other and control of one impactful on control of the other. Malaria parasitaemia (MP) prevalence is increased in HIV-infected individuals while certain drugs used in latter cause decline in MP but it is uncertain how they affect malaria antigenaemia (MA). How certain bio-social and disease characteristics affect MA and MP in this cohort is unknown. OBJECTIVES: To determine prevalence of asymptomatic MP and MA and their clinical and social determinants in HIV infected. METHODS: In a prospective cross-sectional study carried out at the University of Benin Teaching Hospital (April to June 2016), 221 HIV-infected children (aged 1-17years) asymptomatic for malaria and 221 apparently healthy HIV-negative controls were studied. MA was assessed using rapid diagnostic test while MP was evaluated using microscopy. Standard method was used to determine parasite count. RESULTS: Prevalence of asymptomatic MP was 24.4% in subjects and 17.6% in controls while MA prevalence in subjects and controls were comparable (20.8% vs 18.1%). Malaria parasitaemia rate (MPr) of 24.4% was higher than malaria antigenaemia rate (MAr) (20.8%). MP and MA rates were independent of socioeconomic status, access to anti-retroviral drugs, their duration of use and clinical disease stage. CONCLUSION: MA occurred frequently enough to warrant its use as malaria case definition surrogate in asymptomatic children with HIV/AIDS receiving trimethoprim-sulfamethoxazole prophylaxis and protease inhibitors.

CONTEXTE: Le paludisme et le VIH / SIDA sont endémiques en Afrique subsaharienne, la prévalence de l'un renforçant l'autre et le contrôle de l'un ayant un impact sur le contrôle de l'autre. La prévalence de la parasitémie du paludisme (MP) est augmentée chez les personnes infectées par le VIH, tandis que certains médicaments utilisés dans ces derniers entraînent une baisse de la MP, mais on ignore comment ils affectent l'antigénémie du paludisme (AM). On ne sait pas comment certaines caractéristiques biosociales et pathologiques affectent l'AMM et la MP dans cette cohorte. OBJECTIFS: Déterminer la prévalence de la MP et de l'AMM et leurs déterminants cliniques-sociaux chez les enfants infectés par le VIH. METHODES: Dans une étude transversale prospective menée à l'hôpital universitaire de l'Université du Bénin (avril et juin 2016), 221 enfants infectés par le VIH (âgés de 1 à 17 ans) asymptomatiques pour le paludisme et 221 témoins séronégatifs apparemment en bonne santé ont été étudiés. La MA a été évaluée à l'aide d'un test de diagnostic rapide tandis que la MP a été évaluée à l'aide de la microscopie à coloration de Giemsa. La méthode standard a été utilisée pour déterminer le nombre de parasites. RESULTATS: La prévalence de la MP asymptomatique était de 24,4% chez les sujets et de 17,6% chez les témoins, tandis que la prévalence de l'AM chez les sujets et les témoins était comparable (20,8% vs 18,1%). Le taux de parasitémie du paludisme (MPr) de 24,4% était plus élevé que le taux d'antigénémie du paludisme (MAr) (20,8%). Les taux de MP et d'AM étaient indépendants du statut socio-économique, de l'accès aux médicaments antirétroviraux, de leur durée d'utilisation et du stade clinique de la maladie. CONCLUSION: l'AMM est survenue suffisamment fréquemment pour justifier son utilisation comme substitut de la définition de cas dans la prise en charge de ces sujets, étant donné l'implication de la parasitémie dans la physiopathologie et la virulence du VIH. MOTS CLÉS: Paludisme, Antigénémie, Parasitémie, Enfants infectés par le VIH, Benin City.

Artemisinin Cocrystals for Bioavailability Enhancement. Part 2: In Vivo Bioavailability and Physiologically Based Pharmacokinetic Modeling.

We report the evaluation and prediction of the pharmacokinetic (PK) performance of artemisinin (ART) cocrystal formulations, that is, 1:1 artemisinin/orcinol (ART-ORC) and 2:1 artemisinin/resorcinol (ART2-RES), using in vivo murine animal and physiologically based pharmacokinetic (PBPK) models. The efficacy of the ART cocrystal formulations along with the parent drug ART was tested in mice infected with Plasmodium berghei. When given at the same dose, the ART cocrystal formulation showed a significant reduction in parasitaemia at day 4 after infection compared to ART alone. PK parameters including Cmax (maximum plasma concentration), Tmax (time to Cmax), and AUC (area under the curve) were obtained by determining drug concentrations in the plasma using liquid chromatography-high-resolution mass spectrometry (LC-HRMS), showing enhanced ART levels after dosage with the cocrystal formulations. The dose-response tests revealed that a significantly lower dose of the ART cocrystals in the formulation was required to achieve a similar therapeutic effect as ART alone. A PBPK model was developed using a PBPK mouse simulator to accurately predict the in vivo behavior of the cocrystal formulations by combining in vitro dissolution profiles with the properties of the parent drug ART. The study illustrated that information from classical in vitro and in vivo experimental investigations of the parent drug of ART formulations can be coupled with PBPK modeling to predict the PK parameters of an ART cocrystal formulation in an efficient manner. Therefore, the proposed modeling strategy could be used to establish in vitro and in vivo correlations for different cocrystals intended to improve dissolution properties and to support clinical candidate selection, contributing to the assessment of cocrystal developability and formulation development.

Prevalence of asymptomatic malaria, submicroscopic parasitaemia and anaemia in Korogwe District, north-eastern Tanzania.

BACKGROUND: Asymptomatic malaria infections largely remain undetected and act as a reservoir for continuous transmission. The study assessed the prevalence of submicroscopic asymptomatic malaria infections and anaemia in two rural low (300 m above sea level) and highland (700 m asl) settings of Korogwe District north-eastern Tanzania. METHODS: A cross-sectional malariometric survey involving individuals aged 0-19 years was conducted in June 2018 in the two rural villages. Venous blood was collected from eligible study participants for estimation of haemoglobin level, detection of malaria by rapid diagnostic test (RDT), quantification of malaria parasitaemia by microscopy, as well as dried blood spot (DBS) for determining submicroscopic infections by PCR targeting the small subunit of the ribosomal ribonucleic acid (ssrRNA) of human Plasmodium. RESULTS: Out of 565 individuals tested, 211 (37.3%) were malaria positive based on RDT, whereas only 81 (14.3%) were positive by microscopy. There was no significant difference in the prevalence between the highland and the lowland village, p = 0.19 and p = 0.78 microscopy and RDT, respectively. Three out of 206 (1.5%) RDT/microscopy negative samples were P. falciparum positive by PCR. Of the 211 RDT and 81 microscopy positive, 130 (61.6%) and 33 (40.7%), respectively, were defined as being asymptomatic. Of the 565 individuals, 135 (23.9%) were anaemic (haemoglobin < 11 g/dL) out of which 5.2% were severely anaemic. The risk of being anaemic was significantly higher among individuals with asymptomatic malaria as compared to those without malaria as confirmed by RDT (AOR = 2.06 (95% CI 1.32-3.20) while based on microscopic results there was no significant differences observed (AOR = 2.09, 95% CI 0.98-4.47). Age and altitude had no effect on the risk of anaemia even after adjusting for asymptomatic malaria. CONCLUSIONS: Asymptomatic malaria is associated with an increased risk of having anaemia in the study communities. The findings highlight the need for targeted interventions focusing on asymptomatic infections which is an important risks factor for anaemia in the community and act as a source of continued transmission of malaria in the study area.

Plasmodium falciparum outbreak in native communities of Condorcanqui, Amazonas, Perú.

BACKGROUND: Malaria remains a serious health threat in the Amazonas Region of Peru and approximately 95% of the cases, mainly Plasmodium vivax, are found in native communities of The Rio Santiago District, Condorcanqui Province. In 2019, more than one thousand malaria cases were reported, with an unusual number of Plasmodium falciparum autochthonous cases. The present study aims to report this P. falciparum outbreak while describing the epidemiology of malaria and the risk factors associated in the native communities of Amazonas, Peru. METHODS: The DIRESA-Amazonas in collaboration with the Condorcanqui Health Network and the Institute of Tropical Diseases of the UNTRM carried out a malaria Active Case Detection (ACD III) between January 31st and February 10th of 2020. A total of 2718 (47.4%) individuals from 21 native communities grouped in eight sanitary districts, were screened for malaria infections. Each participant was screened for malaria using microscopy. Follow-up surveys were conducted for all malaria positive individuals to collect socio-demographic data. Spatial clustering of infection risk was calculated using a generalized linear model (GLM). Analysis of risk considered factors such as gender, age, type of infection, symptomatology, and parasitaemia. RESULTS: The study suggests that the P. falciparum index case was imported from Loreto and later spread to other communities of Rio Santiago during 2019. The ACD III reported 220 (8.1%) malaria cases, 46 P. falciparum, 168 P. vivax and 6 mixed infections. SaTScan analysis detected a cluster of high infection risk in Middle Rio Santiago and a particular high P. falciparum infection risk cluster in Upper Rio Santiago. Interestingly, the evaluation of different risk factors showed significant associations between low parasitaemia and P. falciparum asymptomatic cases. CONCLUSION: This is the first report of a P. falciparum outbreak in native communities of Condorcanqui, Amazonas. Timely identification and treatment of symptomatic and asymptomatic cases are critical to achieve malaria control and possible elimination in this area. However, the current malaria situation in Condorcanqui is uncertain, given that malaria ACD activities have been postponed due to COVID-19.

Malaria parasitaemia and mRDT diagnostic performances among symptomatic individuals in selected health care facilities across Ghana.

BACKGROUND: Parasitological diagnosis generates data to assist malaria-endemic countries determine their status within the malaria elimination continuum and also inform the deployment of proven interventions to yield maximum impact. This study determined prevalence of malaria parasitaemia and mRDT performances among febrile patients in selected health care facilities across Ghana. METHODS: This study was a cross-sectional survey conducted in the previously 10 regions of Ghana from May to August 2018. Each patient suspected to have uncomplicated malaria was tested using microscopy and two malaria rapid diagnostic tests (mRDTs): routinely used CareStart™ Malaria HRP2 (Pf) and SD Bioline Malaria Ag Pf (HRP2/pLDH). Main outcome variables were malaria slide and CareStart™ Malaria HRP2 (Pf) positivity rates; and diagnostic accuracy of CareStart™ Malaria HRP2 (Pf) and SD Bioline Malaria Ag Pf (HRP2/pLDH) using microscopy as "gold standard". RESULTS: Overall parasite positivity rates were 32.3% (6266/19402) by mRDT and 16.0% (2984/18616) by microscopy, with Plasmodium falciparum mono-infection accounting for 98.0% of all infections. The odds of parasitaemia by microscopy was significantly lower among female patients compared with males (OR = 0.78; 95% CI: 0.66-0.91), and among patients with history of previous antimalarial intake compared with those with no such history (OR = 0.72; 95% CI: 0.54-0.95). Overall sensitivity of CareStart™ Malaria HRP2 (Pf) was statistically similar to that of the HRP2 band of SD Bioline Malaria Ag Pf (HRP2/pLDH) combo kit (95.4%; 95% CI: 94.6-96.1 vs 94.3%; 95% CI: 93.4-95.1; p = 0.065) but significantly higher than the pLDH band (89.3%; 95% CI: 88.1-90.4; p < 0.001). The same pattern was observed for negative predictive value. CONCLUSIONS: Malaria control interventions should be targeted at the general population, and history of antimalarial intake considered a key predictor of malaria slide negativity. Furthermore, HRP2-based mRDTs remain effective diagnostic tool in the management of suspected uncomplicated malaria in the country.

Circulating Trypanosoma cruzi load and major cardiovascular outcomes in patients with chronic Chagas cardiomyopathy: a prospective cohort study.

OBJECTIVES: To analyse the effect of parasite load assessed by quantitative reverse transcription PCR (RT-qPCR) in serum on the prognosis of patients with chronic Chagas cardiomyopathy (CCM) after a 2-year follow-up. METHODS: Prospective cohort study conducted between 2015 and 2017. One hundred patients with CCM were included. Basal parasitaemia levels of Trypanosoma cruzi (T. cruzi) were measured using a quantitative polymerase chain reaction (qPCR) test. The primary composite outcome (CO) was all-cause mortality, cardiac transplantation and implantation of a left ventricular assist device. Secondary outcomes were the baseline levels of serum biomarkers and echocardiographic variables. RESULTS: After a 2 years of follow-up, the primary CO rate was 16%. A positive qPCR was not associated with a higher risk of the CO. However, when parasitaemia was evaluated by comparing tertiles (tertile 1: undetectable parasitaemia, tertile 2: low parasitaemia and tertile 3: high parasitaemia), a higher risk of the CO (HR 3.66; 95% CI 1.11-12.21) was evidenced in tertile 2. Moreover, patients in tertile 2 had significantly higher levels of high-sensitivity troponin T and cystatin C and more frequently exhibited an ejection fraction <50%. CONCLUSION: Low parasitaemia was associated with severity markers of myocardial injury and a higher risk of the composite outcome when compared with undetectable parasitaemia. This finding could be hypothetically explained by a more vigorous immune response in patients with low parasitaemia that could decrease T. cruzi load more efficiently, but be associated with increased myocardial damage. Additional studies with a larger number of patients and cytokine measurement are required to support this hypothesis.

OBJECTIFS: Analyser l'effet de la charge parasitaire évaluée par PCR quantitative de transcription inverse (RT-qPCR) dans le sérum sur le pronostic des patients atteints de cardiomyopathie chronique de Chagas (CCM) après un suivi de deux ans. MÉTHODES: Etude de cohorte prospective menée entre 2015 et 2017. Une centaine de patients atteints de CCM ont été inclus. Les niveaux de parasitémie basale de Trypanosoma cruzi (T. cruzi) ont été mesurés en utilisant un test de réaction en chaîne de la polymérase quantitative (qPCR). Le principal résultat composite (RC) était la mortalité toutes causes, la transplantation cardiaque et l'implantation d'un dispositif d'assistance ventriculaire gauche. Les critères secondaires étaient les niveaux de base des biomarqueurs sériques et des variables échocardiographiques. RÉSULTATS: Après 2 ans de suivi, le taux de RC primaire était de 16%. Une qPCR positive n'était pas associée à un risque plus élevé de RC. Cependant, lorsque la parasitémie était évaluée en comparant les tertiles (tertile 1: parasitémie indétectable, tertile 2: parasitémie faible et tertile 3: parasitémie élevée), un risque plus élevé de RC (HR: 3,66; IC95%: 1,11-12,21) a été mis en évidence dans le tertile 2. De plus, les patients du tertile 2 avaient des niveaux significativement plus élevés de troponine T et de cystatine-C à haute sensibilité et présentaient plus fréquemment une fraction d'éjection <50%. CONCLUSION: Une faible parasitémie était associée à des marqueurs de sévérité des lésions myocardiques et à un risque plus élevé de résultat composite par rapport à une parasitémie indétectable. Cette découverte pourrait être hypothétiquement expliquée par une réponse immunitaire plus vigoureuse chez les patients présentant une faible parasitémie qui pourrait diminuer la charge de T. cruzi plus efficacement mais être associée à une augmentation des lésions myocardiques. Des études supplémentaires avec un plus grand nombre de patients et une mesure des cytokines sont nécessaires pour étayer cette hypothèse.

Performance of IRS on malaria prevalence and incidence using pirimiphos-methyl in the context of pyrethroid resistance in Koulikoro region, Mali.

BACKGROUND: Koulikoro Health District is one of three districts of Mali where the indoor residual spray (IRS) has been implemented from 2008 to 2016. With widespread of resistance to pyrethroid, IRS was shifted from pyrethroid to pirimiphos-methyl from 2014 to 2016. The objective of this study was to assess the added value of IRS to LLINs on the prevalence of parasitaemia and malaria incidence among children under 10 years old. METHODS: A comparative study was carried out to assess the effects of pirimiphos-methyl based IRS on malaria prevalence and incidence among children from 6 months to 10 years old in selected pyrethroid resistance villages of two health districts in Mali: one where IRS was implemented in combination with LLINs (intervention area) and one with LLINs-only (control area). Two cross-sectional surveys were carried out at the beginning (June) and end of the rainy season (October) to assess seasonal changes in malaria parasitaemia by microscopy. A passive detection case (PCD) was set-up in each study village for 9 months to estimate the incidence of malaria using RDT. RESULTS: There was an increase of 220% in malaria prevalence from June to October in the control area (14% to 42%) versus only 53% in the IRS area (9.2% to 13.2%). Thus, the proportional rise in malaria prevalence from the dry to the rainy season in 2016 was 4-times greater in the control area compared to the IRS area. The overall malaria incidence rate was 2.7 per 100 person-months in the IRS area compared with 6.8 per 100 person-month in the control areas. The Log-rank test of Kaplan-Meier survival analysis showed that children living in IRS area remain much longer free from malaria (Hazard ratio (HR) = 0.45, CI 95% 0.37-0.54) than children of the control area (P < 0.0001). CONCLUSIONS: IRS using pirimiphos-methyl has been successful in reducing substantially both the prevalence and the incidence of malaria in children under 10 years old in the area of pyrethroid resistance of Koulikoro, Mali. Pirimiphos-methyl is a better alternative than pyrethroids for IRS in areas with widespread of pyrethroid resistance.

Epidemiology of malaria among pregnant women during their first antenatal clinic visit in the middle belt of Ghana: a cross sectional study.

BACKGROUND: Malaria during pregnancy may result in unfavourable outcomes in both mothers and their foetuses. This study sought to document the current burden and factors associated with malaria and anaemia among pregnant women attending their first antenatal clinic visit in an area of Ghana with perennial malaria transmission. METHODS: A total of 1655 pregnant women aged 18 years and above with a gestational age of 13-22 weeks, who attended an antenatal care (ANC) clinic for the first time, were consented and enrolled into the study. A structured questionnaire was used to collect socio-demographic and obstetric data and information on use of malaria preventive measures. Venous blood (2 mL) was collected before sulfadoxine-pyrimethamine administration. Malaria parasitaemia and haemoglobin concentration were determined using microscopy and an automated haematology analyser, respectively. Data analysis was carried out using Stata 14. RESULTS: Mean age (SD) and gestational age (SD) of women at enrolment were 27.4 (6.2) years and 16.7 (4.3) weeks, respectively. Overall malaria parasite prevalence was 20.4% (95% CI 18.5-22.4%). Geometric mean parasite density was 442 parasites/µL (95% CI 380-515). Among women with parasitaemia, the proportion of very low (1-199 parasites/µL), low (200-999 parasites/µL), medium (1000-9999 parasites/µL) and high (≥ 10,000 parasites/µL) parasite density were 31.1, 47.0, 18.9, and 3.0%, respectively. Age ≥ 25 years (OR 0.57, 95% CI 0.41-0.79), multigravid (OR 0.50, 95% CI 0.33-0.74), educated to high school level or above (OR 0.53, 95% CI 0.33-0.83) and in household with higher socio-economic status (OR 0.34, 95% CI 0.21-0.54) were associated with a lower risk of malaria parasitaemia. The prevalence of anaemia (< 11.0 g/dL) was 56.0%, and the mean haemoglobin concentration in women with or without parasitaemia was 9.9 g/dL or 10.9 g/dL, respectively. CONCLUSION: One out of five pregnant women attending their first ANC clinic visit in an area of perennial malaria transmission in the middle belt of Ghana had Plasmodium falciparum infection. Majority of the infections were below 1000 parasites/µL and with associated anaemia. There is a need to strengthen existing malaria prevention strategies to prevent unfavourable maternal and fetal birth outcomes in this population.

Field performance of ultrasensitive and conventional malaria rapid diagnostic tests in southern Mozambique.

BACKGROUND: An ultrasensitive malaria rapid diagnostic test (RDT) was recently developed for the improved detection of low-density Plasmodium falciparum infections. This study aimed to compare the diagnostic performance of the PfHRP2-based Abbott Malaria Ag P. falciparum ultrasensitive RDT (uRDT) to that of the conventional SD-Bioline Malaria Ag P. falciparum RDT (cRDT) when performed under field conditions. METHODS: Finger-prick blood samples were collected from adults and children in two cross-sectional surveys in May of 2017 in southern Mozambique. Using real-time quantitative PCR (RT-qPCR) as the reference method, the age-specific diagnostic performance indicators of the cRDT and uRDT were compared. The presence of histidine-rich protein 2 (HRP2) and Plasmodium lactate dehydrogenase (pLDH) antigens was evaluated in a subset from dried blood spots by a quantitative antigen assay. pfhrp2 and pfhrp3 gene deletions were assessed in samples positive by RT-qPCR and negative by both RDTs. RESULTS: Among the 4,396 participants with complete test results, the sensitivity of uRDTs (68.2; 95% CI 60.8 to 74.9) was marginally better than that of cRDTs (61.5; 95% CI 53.9 to 68.6) (p-value = 0.004), while the specificities were similar (uRDT: 99.0 [95% CI 98.6 to 99.2], cRDT: 99.2 [95% CI 98.9 to 99.4], p-value = 0.02). While the performance of both RDTs was lowest in ≥ 15-year-olds, driven by the higher prevalence of low parasite density infections in this group, the sensitivity of uRDTs was significantly higher in this age group (54.9, 95% CI 40.3 to 68.9) compared to the sensitivity of cRDTs (39.2, 95% CI 25.8 to 53.9) (p-value = 0.008). Both RDTs detected P. falciparum infections at similar geometric mean parasite densities (112.9  parasites/µL for uRDTs and 145.5 parasites/µL for cRDTs). The presence of HRP2 antigen was similar among false positive (FP) samples of both tests (80.5% among uRDT-FPs and 84.4% among cRDT-FPs). Only one false negative sample was detected with a partial pfhrp2 deletion. CONCLUSION: This study showed that the uRDTs developed by Abbott do not substantially outperform SD-Bioline Pf malaria RDTs in the community and are still not comparable to molecular methods to detect P. falciparum infections in this study setting.

Description of Plasmodium falciparum infections in central Gabon demonstrating high parasite densities among symptomatic adolescents and adults.

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.