Chronic malaria exposure is associated with inhibitory markers on T cells that correlate with atypical memory and marginal zone-like B cells.

Chronic immune activation from persistent malaria infections can induce immunophenotypic changes associated with T-cell exhaustion. However, associations between T and B cells during chronic exposure remain undefined. We analyzed peripheral blood mononuclear cells from malaria-exposed pregnant women from Papua New Guinea and Spanish malaria-naïve individuals using flow cytometry to profile T-cell exhaustion markers phenotypically. T-cell lineage (CD3, CD4, and CD8), inhibitory (PD1, TIM3, LAG3, CTLA4, and 2B4), and senescence (CD28-) markers were assessed. Dimensionality reduction methods revealed increased PD1, TIM3, and LAG3 expression in malaria-exposed individuals. Manual gating confirmed significantly higher frequencies of PD1+CD4+ and CD4+, CD8+, and double-negative (DN) T cells expressing TIM3 in malaria-exposed individuals. Increased frequencies of T cells co-expressing multiple markers were also found in malaria-exposed individuals. T-cell data were analyzed with B-cell populations from a previous study where we reported an alteration of B-cell subsets, including increased frequencies of atypical memory B cells (aMBC) and reduction in marginal zone (MZ-like) B cells during malaria exposure. Frequencies of aMBC subsets and MZ-like B cells expressing CD95+ had significant positive correlations with CD28+PD1+TIM3+CD4+ and DN T cells and CD28+TIM3+2B4+CD8+ T cells. Frequencies of aMBC, known to associate with malaria anemia, were inversely correlated with hemoglobin levels in malaria-exposed women. Similarly, inverse correlations with hemoglobin levels were found for TIM3+CD8+ and CD28+PD1+TIM3+CD4+ T cells. Our findings provide further insights into the effects of chronic malaria exposure on circulating B- and T-cell populations, which could impact immunity and responses to vaccination.

Efficacy and safety of hydroxychloroquine for treatment of mild SARS-CoV-2 infection and prevention of COVID-19 severity in pregnant and postpartum women: A randomized, double-blind, placebo-controlled trial.

INTRODUCTION: Pregnant women have an increased risk of severe COVID-19. Evaluation of drugs with a safety reproductive toxicity profile is a priority. At the beginning of the pandemic, hydroxychloroquine (HCQ) was recommended for COVID-19 treatment. MATERIAL AND METHODS: A randomized, double-blind, placebo-controlled clinical trial was conducted in eight teaching hospitals in Spain to evaluate the safety and efficacy of HCQ in reducing viral shedding and preventing COVID-19 progression. Pregnant and postpartum women with a positive SARS-CoV-2 PCR (with or without mild COVID-19 signs/symptoms) and a normal electrocardiogram were randomized to receive either HCQ orally (400 mg/day for 3 days and 200 mg/day for 11 days) or placebo. PCR and electrocardiogram were repeated at day 21 after treatment start. Enrollment was stopped before reaching the target sample due to low recruitment rate. Trial registration EudraCT #: 2020-001587-29, on April 2, 2020. CLINICAL TRIALS: gov # NCT04410562, registered on June 1, 2020. RESULTS: A total of 116 women (75 pregnant and 41 post-partum) were enrolled from May 2020 to June 2021. The proportion of women with a positive SARS-CoV-2 PCR at day 21 was lower in the HCQ group (21.8%, 12/55) than in the placebo group (31.6%, 18/57), although the difference was not statistically significant (P = 0.499). No differences were observed in COVID-19 progression, adverse events, median change in QTc, hospital admissions, preeclampsia or poor pregnancy and perinatal outcomes between groups. CONCLUSIONS: HCQ was found to be safe in pregnant and postpartum women with asymptomatic or mild SARS-CoV-2 infection. Although the prevalence of infection was decreased in the HCQ group, the statistical power was insufficient to confirm the potential beneficial effect of HCQ for COVID-19 treatment.

Immunisation coverage and factors associated with incomplete immunisation in children under two during the COVID-19 pandemic in Sierra Leone.

BACKGROUND: Routine childhood immunisation is one of the most important life-saving public health interventions. However, many children still have inadequate access to these vaccines and millions remain (partially) unvaccinated globally. As the COVID-19 pandemic disrupted health systems worldwide, its effects on immunisation have become apparent. This study aimed to estimate routine immunisation coverage among children under two in Sierra Leone and to identify factors associated with incomplete immunisation during the COVID-19 pandemic. METHODS: A cross-sectional household survey was conducted in three districts in Sierra Leone: Bombali, Tonkolili and Port Loko. A three-stage cluster sampling method was followed to enrol children aged 10-23 months. Information regarding immunisation status was based on vaccination cards or caretaker's recall. Using WHO's definition, a fully immunised child received one BCG dose, three oral polio vaccine doses, three pentavalent vaccine doses and one measles-containing vaccine dose. Following the national schedule, full immunisation status can be achieved at 9 months of age. Data were weighted to reflect the survey's sampling design. Associations between incomplete immunisation and sociodemographic characteristics were assessed through multivariable logistic regression. RESULTS: A total of 720 children were enrolled between November and December 2021. Full vaccination coverage was estimated at 65.8% (95% CI 60.3%-71.0%). Coverage estimates were highest for vaccines administered at birth and decreased with doses administered subsequently. Adjusting for age, the lowest estimated coverage was 40.7% (95% CI 34.5%-47.2%) for the second dose of the measles-containing vaccine. Factors found to be associated with incomplete immunisation status were: living in Port Loko district (aOR = 3.47, 95% CI = 2.00-6.06; p-value < 0.001), the interviewed caretaker being Muslim (aOR = 1.94, 95% CI = 1.25-3.02; p-value = 0.015) and the interviewed caretaker being male (aOR = 1.93, 95% CI = 1.03-3.59, p-value = 0.039). CONCLUSION: Though full immunisation coverage at district level improved compared with pre-pandemic district estimates from 2019, around one in three surveyed children had missed at least one basic routine vaccination and over half of eligible children had not received the recommended two doses of a measles-containing vaccine. These findings highlight the need to strengthen health systems to improve vaccination uptake in Sierra Leone, and to further explore barriers that may jeopardise equitable access to these life-saving interventions.

Prevalence and risk factors associated with malaria infection in children under two years of age in southern Togo prior to perennial malaria chemoprevention implementation.

BACKGROUND: Malaria remains the leading cause of mortality and morbidity in young children in sub-Saharan Africa. To prevent malaria in children living in moderate-to-high malaria transmission areas, the World Health Organization has recommended perennial malaria chemoprevention (PMC). Prior to piloting PMC implementation in southern Togo, a household survey was conducted to estimate malaria infection prevalence in children under 2 years of age (U2). METHODS: A cross-sectional community-based household survey was conducted in the Haho district in the Togo Plateaux region. A three-stage random sampling method was used to select study participants aged 10-23 months whose caretakers gave informed consent. The prevalence of Plasmodium infection, defined as a positive rapid diagnostic test (RDT), was estimated with 95% confidence interval (CI). Clinical malaria was defined as having a positive RDT plus fever (≥ 37.5 °C) or history of fever in the last 24 h. Mixed-effects logistic regression models were used to assess the child's, caretaker's, and household's factors associated with malaria infection. RESULTS: A total of 685 children were included in the survey conducted January-February in 2022 (dry season). Median age was 17 months (interquartile range: 13-21). About 80% of the children slept under a bed net the night before the interview. Malaria infection prevalence was 32.1% (95% CI 27.7-37.0) with significant area variation (cluster range: 0.0-73.3). Prevalence of clinical malaria was 15.4% (95% CI 12.2-19.2). Children whose caretakers were animist (aOR: 1.71, 95% CI 1.19-2.46) and those living in mother-headed households (aOR: 2.39, 95% CI 1.43-3.99) were more likely to have a positive RDT. Living more than 5 km away from the nearest health facility (aOR: 1.60, 95% CI 1.04-2.44) and presence of two or more under-5-years children in the household (aOR: 1.44, 95% CI 1.01-2.07) were also associated with increased risk of infection. CONCLUSION: One-third of the children U2 who participated in this survey had malaria infection, thus PMC could be a promising strategy to reduce malaria burden in young children in Plateaux region. Reinforcement of outreach services and targeting the poorest households should be prioritized to reduce the inequity in malaria prevention in children exposed to the infection.

Coverage of intermittent preventive treatment of malaria in infants after four years of implementation in Sierra Leone.

BACKGROUND: Intermittent Preventive Treatment of malaria in infants (IPTi) is a malaria control strategy consisting of the administration of an anti-malarial drug alongside routine immunizations. So far, this is being implemented nationwide in Sierra Leone only. IPTi has been renamed as Perennial Malaria Chemoprevention -PMC-, accounting for its recently recommended expansion into the second year of life. Before starting a pilot implementation on PMC, the currently implemented strategy and malaria prevalence were assessed in young children in selected areas of Sierra Leone. METHODS: A cross-sectional, community-based, multi-stage cluster household survey was conducted from November to December 2021 in selected districts of the Northern and northwestern provinces of Sierra Leone among 10-23 months old children, whose caretakers gave written informed consent to participate in the survey. Coverage of IPTi and malaria prevalence-assessed with rapid diagnostic tests-were calculated using percentages and 95% confidence intervals weighted for the sampling design and adjusted for non-response within clusters. Factors associated with RDT + and iPTi coverage were also assessed. RESULTS: A total of 720 children were recruited. Coverage of three IPTi doses was 50.57% (368/707; 95% CI 45.38-55.75), while prevalence of malaria infection was 28.19% (95% CI 24.81-31.84). Most children had received IPTi1 (80.26%, 574/707; 95% CI 75.30-84.44), and IPTi2 (80.09%, 577/707; 95% CI 76.30-83.40) and over half of the children also received IPTi3 (57.72%, 420/707; 95% CI 53.20-62.11). The uptake of each IPTi dose was lower than that of the vaccines administered at the same timepoint at all contacts. CONCLUSION: In Sierra Leone, half of the children received the three recommended doses of IPTi indicating an increase in its uptake compared to previous data of just a third of children receiving the intervention. However, efforts need to be made in improving IPTi coverage, especially in the planned expansion of the strategy into the second year of life following recent WHO guidelines.

Burden of malaria in pregnancy among adolescent girls compared to adult women in 5 sub-Saharan African countries: A secondary individual participant data meta-analysis of 2 clinical trials.

BACKGROUND: Malaria is among the top causes of death in adolescent girls (10 to 19 years) globally. Adolescent motherhood is associated with increased risk of adverse maternal and neonatal outcomes. The interaction of malaria, adolescence, and pregnancy is especially relevant in malaria endemic areas, where rates of adolescent pregnancy are high. However, data on burden of malaria among adolescent girls are limited. This study aimed at investigating whether adolescent girls were at a greater risk of experiencing malaria-related outcomes in pregnancy-parasitaemia and clinical disease-than adult women. METHODS AND FINDINGS: An individual secondary participant-level meta-analysis was conducted using data from 5,804 pregnant women participating in 2 malaria prevention clinical trials in Benin, Gabon, Kenya, Mozambique, and Tanzania between 2009 and 2014. Of the sample, 1,201 participants were adolescent girls with a mean age of 17.5 years (standard deviation (SD) 1.3) and 886 (73.8%) of them primigravidae. Among the 4,603 adult women with mean age of 27.0 years (SD 5.4), 595 (12.9%) were primigravidae. Mean gestational age at enrolment was 20.2 weeks (SD 5.2) and 1,069 (18.4%) participants were HIV-infected. Women were followed monthly until the postpartum visit (1 month to 6 weeks after delivery). This study considered outcomes including clinical episodes during pregnancy, peripheral parasitaemia at delivery, and placental malaria. A 2-stage meta-analysis approach was followed by pooling single multivariable regression results into standard DerSimonian-Laird random-effects models. Adolescent girls were more likely than adult women to present with clinical malaria during pregnancy (incidence risk ratio (IRR) 1.70, 95% confidence interval (CI) 1.20; 2.39, p-value = 0.003, I2 = 0.0%, N = 4,092), peripheral parasitaemia at delivery (odds ratio (OR) 2.28, 95% CI 1.46; 3.55, p-value < 0.001, I2 = 0.0%, N = 3,977), and placental infection (OR 1.97, 95% CI 1.31; 2.98, p-value = 0.001, I2 = 1.4%, N = 4,797). Similar associations were observed among the subgroup of HIV-uninfected participants: IRR 1.72 (95% CI 1.22; 2.45, p-value = 0.002, I2 = 0.0%, N = 3,531) for clinical malaria episodes, OR 2.39 (95% CI 1.49; 3.86, p-value < 0.001, I2 = 0.0%, N = 3,053) for peripheral parasitaemia, and OR 1.88 (95% CI 1.06 to 3.33, p-value = 0.03, I2 = 34.9%, N = 3,847) for placental malaria. Among HIV-infected subgroups statistically significant associations were not observed. Similar associations were found in the subgroup analysis by gravidity. The small sample size and outcome prevalence in specific countries limited the inclusion of some countries in the meta-analysis. Furthermore, peripheral parasitaemia and placental malaria presented a considerable level of missing data-12.6% and 18.2% of participants had missing data on those outcomes, respectively. Given the original scope of the clinical trials, asymptomatic malaria infection was only assessed at the end of pregnancy through peripheral and placental parasitaemia. CONCLUSIONS: In this study, we observed that adolescent girls in sub-Saharan Africa (SSA) are more prone to experience clinical malaria episodes during pregnancy and have peripheral malaria and placental infection at delivery than adult women. Moreover, to the best of our knowledge, for the first time this study disaggregates figures and stratifies analyses by HIV infection. Similar associations were found for both HIV-infected and uninfected women, although those for HIV-infected participants were not statistically significant. Our finding suggests that adolescent girls may benefit from targeted malaria prevention strategies even before they become pregnant.

Eotaxin-2 and eotaxin-3 in malaria exposure and pregnancy.

BACKGROUND: Eotaxin-1 concentrations in plasma have been inversely associated with malaria exposure, malaria infection and pregnancy, but the effect of these conditions on the levels of the related chemokines eotaxin-2 and eotaxin-3 remains unknown. METHODS: Eotaxin-2 and -3 concentrations were measured in 310 peripheral or placental plasma samples from pregnant and non-pregnant individuals from Papua New Guinea (malaria-endemic country) and Spain (malaria-naïve individuals) with previous data on eotaxin-1 concentrations. Correlations between eotaxin concentrations were examined with the Spearman's test. Differences in eotaxin concentrations among groups were evaluated with the Kruskal-Wallis or Mann Whitney tests. The pairwise Wilcoxon test was performed to compare eotaxin-2 concentration between peripheral and placental matched plasmas. Univariable and multivariable linear regression models were estimated to assess the association between eotaxins and Plasmodium infection or gestational age. RESULTS: Eotaxin-2 concentrations in plasma showed a weak positive correlation with eotaxin-3 (rho = 0.35, p < 0.05) concentrations. Eotaxin-2 concentrations in the malaria-exposed non-pregnant group were significantly lower than the in the malaria-naive non-pregnant and the malaria-exposed pregnant groups. Eotaxin-3 plasma concentrations were lower in malaria-exposed than in non-exposed groups (p < 0.05), but no differences were found associated to pregnancy. Eotaxin-2 and eotaxin-3 plasma concentrations were negatively correlated with anti-Plasmodium IgG levels: PfDBL5ε-IgG (rhoEo2 = - 0.35, p = 0.005; rhoEo3 =- 0.37, p = 0.011), and eotaxin-3 was negatively correlated with PfDBL3x-IgG levels (rhoEo3 =- 0.36; p = 0.011). Negative correlations of eotaxin-2 and 3 in plasma were also observed with atypical memory B cells (rhoEo2 = - 0.37, p < 0.001; rhoEo3= - 0.28, p = 0.006), a B cell subset expanded in malaria-exposed individuals. In addition, a borderline negative association was observed between eotaxin-3 concentrations and Plasmodium infection (adjusted effect estimate, ß = - 0.279, 95% CI - 0.605; 0.047, p = 0.091). Moreover, eotaxin-2 placental concentrations were significantly increased compared to peripheral concentrations in the malaria-exposed pregnant group whereas the contrary was observed in the non-exposed pregnant group (p < 0.005). CONCLUSION: Although a clear epidemiological negative association is observed between eotaxins concentrations and malaria exposure and/or infection, pregnancy may alter this association for eotaxin-2. Further research is required to understand the role of these chemokines in this disease and in combination with pregnancy.

Folic acid supplementation and malaria susceptibility and severity among people taking antifolate antimalarial drugs in endemic areas.

BACKGROUND: Description of the condition Malaria, an infectious disease transmitted by the bite of female mosquitoes from several Anopheles species, occurs in 87 countries with ongoing transmission (WHO 2020). The World Health Organization (WHO) estimated that, in 2019, approximately 229 million cases of malaria occurred worldwide, with 94% occurring in the WHO's African region (WHO 2020). Of these malaria cases, an estimated 409,000 deaths occurred globally, with 67% occurring in children under five years of age (WHO 2020). Malaria also negatively impacts the health of women during pregnancy, childbirth, and the postnatal period (WHO 2020). Sulfadoxine/pyrimethamine (SP), an antifolate antimalarial, has been widely used across sub-Saharan Africa as the first-line treatment for uncomplicated malaria sTo examine the effects of folic acid supplementation, at various doses, on malaria susceptibility (risk of infection) and severity among people living in areas with various degrees of malaria endemicity. We will examine the interaction between folic acid supplements and antifolate antimalarial drugs. Specifically, we will aim to answer the following. Among uninfected people living in malaria endemic areas, who are taking or not taking antifolate antimalarials for malaria prophylaxis, does taking a folic acid-containing supplement increase susceptibility to or severity of malaria infection? Among people with malaria infection who are being treated with antifolate antimalarials, does folic acid supplementation increase the risk of treatment failure?Criteria for considering studies for this review Types of studies Inclusion criteria Randomized controlled trials (RCTs) Quasi-RCTs with randomization at the individual or cluster level conducted in malaria-endemic areas (areas with ongoing, local malaria transmission, including areas approaching elimination, as listed in the World Malaria Report 2020) (WHO 2020) Exclusion criteria Ecological studies Observational studies In vivo/in vitro studies Economic studies Systematic literature reviews and meta-analyses (relevant systematic literature reviews and meta-analyses will be excluded but flagged for grey literature screening) Types of participants Inclusion criteria Individuals of any age or gender, living in a malaria endemic area, who are taking antifolate antimalarial medications (inclu

Burden, clinical presentation and risk factors of advanced HIV disease in pregnant Mozambican women.

BACKGROUND: Information on the frequency and clinical features of advanced HIV disease (AHD) in pregnancy and its effects on maternal and perinatal outcomes is limited. The objective of this study was to describe the prevalence and clinical presentation of AHD in pregnancy, and to assess the impact of AHD in maternal and perinatal outcomes in Mozambican pregnant women. METHODS: This is a prospective and retrospective cohort study including HIV-infected pregnant women who attended the antenatal care (ANC) clinic at the Manhiça District Hospital between 2015 and 2020. Women were followed up for 36 months. Levels of CD4 + cell count were determined to assess AHD immune-suppressive changes. Risk factors for AHD were analyzed and the immune-suppressive changes over time and the effect of AHD on pregnancy outcomes were assessed. RESULTS: A total of 2458 HIV-infected pregnant women were enrolled. The prevalence of AHD at first ANC visit was 14.2% (349/2458). Among women with AHD at enrolment, 76.2% (260/341) were on antiretroviral therapy (ART). The proportion of women with AHD increased with age reaching 20.5% in those older than 35 years of age (p < 0.001). Tuberculosis was the only opportunistic infection diagnosed in women with AHD [4.9% (17/349)]. There was a trend for increased CD4 + cell count in women without AHD during the follow up period; however, in women with AHD the CD4 + cell count remained below 200 cells/mm3 (p < 0.001). Forty-two out of 2458 (1.7%) of the women were severely immunosuppressed (CD4 + cell count < 50 cells/mm3). No significant differences were detected between women with and without AHD in the frequency of maternal mortality, preterm birth, low birth weight and neonatal HIV infection. CONCLUSIONS: After more than two decades of roll out of ART in Mozambique, over 14% and nearly 2% of HIV-infected pregnant women present at first ANC clinic visit with AHD and severe immunosuppression, respectively. Prompt HIV diagnosis in women of childbearing age, effective linkage to HIV care with an optimal ART regimen and close monitoring after ART initiation may contribute to reduce this burden and improve maternal and child survival.

Minimally Invasive Tissue Sampling as an Alternative to Complete Diagnostic Autopsies in the Context of Epidemic Outbreaks and Pandemics: The Example of Coronavirus Disease 2019 (COVID-19).

BACKGROUND: Infectious diseases' outbreak investigation requires, by definition, conducting a thorough epidemiological assessment while simultaneously obtaining biological samples for an adequate screening of potential responsible pathogens. Complete autopsies remain the gold-standard approach for cause-of-death evaluation and characterization of emerging diseases. However, for highly transmissible infections with a significant associated lethality, such as COVID-19, complete autopsies are seldom performed due to biosafety challenges, especially in low-resource settings. Minimally invasive tissue sampling (MITS) is a validated new approach based on obtaining postmortem samples from key organs and body fluids, a procedure that does not require advanced biosafety measures or a special autopsy room. METHODS: We aimed to review the use of MITS or similar procedures for outbreak investigation up to 27 March 2021 and their performance for evaluating COVID-19 deaths. RESULTS: After a literature review, we analyzed in detail the results of 20 studies conducted at international sites, whereby 216 COVID-19-related deaths were investigated. MITS provided a general and more granular understanding of the pathophysiological changes secondary to the infection and high-quality samples where the extent and degree of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-related damage could be evaluated. CONCLUSIONS: MITS is a useful addition in the investigation and surveillance of infections occurring in outbreaks or epidemics. Its less invasive nature makes the tool more acceptable and feasible and reduces the risk of procedure-associated contagion, using basic biosafety measures. Standardized approaches protocolizing which samples should be collected-and under which exact biosafety measures-are necessary to facilitate and expand its use globally.

Minimally Invasive Tissue Sampling: A Tool to Guide Efforts to Reduce AIDS-Related Mortality in Resource-Limited Settings.

BACKGROUND: Available information on the causes of death among people living with human immunodeficiency virus (PLHIV) in low- and middle-income countries (LMICs) remains scarce. We aimed to provide data on causes of death in PLHIV from two LMICs, Brazil and Mozambique, to assess the impact of clinical misdiagnosis on mortality rates and to evaluate the accuracy of minimally invasive tissue sampling (MITS) in determining the cause of death in PLHIV. METHODS: We performed coupled MITS and complete autopsy on 164 deceased PLHIV (18 children, 36 maternal deaths, and 110 adults). HIV antibody levels and HIV RNA viral loads were determined from postmortem serum samples. RESULTS: Tuberculosis (22.7%), toxoplasmosis (13.9%), bacterial infections (13.9%), and cryptococcosis (10.9%) were the leading causes of death in adults. In maternal deaths, tuberculosis (13.9%), bacterial infections (13.9%), cryptococcosis (11.1%), and cerebral malaria (8.3%) were the most frequent infections, whereas viral infections, particularly cytomegalovirus (38.9%), bacterial infections (27.8%), pneumocystosis (11.1%), and HIV-associated malignant neoplasms (11.1%) were the leading cause among children. Agreement between the MITS and the complete autopsy was 100% in children, 91% in adults, and 78% in maternal deaths. The MITS correctly identified the microorganism causing death in 89% of cases. CONCLUSIONS: Postmortem studies provide highly granular data on the causes of death in PLHIV. The inaccuracy of clinical diagnosis may play a significant role in the high mortality rates observed among PLHIV in LMICs. MITS might be helpful in monitoring the causes of death in PLHIV and in highlighting the gaps in the management of the infections.

Minimally Invasive Tissue Sampling Findings in 12 Patients With Coronavirus Disease 2019.

BACKGROUND: Minimally invasive tissue sampling (MITS), a postmortem procedure that uses core needle biopsy samples and does not require opening the body, may be a valid alternative to complete autopsy (CA) in highly infectious diseases such as coronavirus disease-19 (COVID-19). This study aimed to (1) compare the performance of MITS and CA in a series of COVID-19 deaths and (2) evaluate the safety of the procedure. METHODS: From October 2020 to February 2021, MITS was conducted in 12 adults who tested positive before death for COVID-19, in a standard, well-ventilated autopsy room, where personnel used reinforced personal protective equipment. In 9 cases, a CA was performed after MITS. A thorough histological evaluation was conducted, and the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was evaluated by real-time reverse-transcription polymerase chain reaction (RT-PCR) and immunohistochemistry. RESULTS: The diagnoses provided by MITS and CA matched almost perfectly. In 9 patients, COVID-19 was in the chain of events leading to death, being responsible for diffuse alveolar damage and mononuclear T-cell inflammatory response in the lungs. No specific COVID-19 features were identified. Three deaths were not related to COVID-19. All personnel involved in MITS repeatedly tested negative for COVID-19. SARS-CoV-2 was identified by RT-PCR and immunohistochemistry in the MITS samples, particularly in the lungs. CONCLUSIONS: MITS is useful for evaluating COVID-19-related deaths in settings where a CA is not feasible. The results of this simplified and safer technique are comparable to those of CA.

Counter-Selection of Antimalarial Resistance Polymorphisms by Intermittent Preventive Treatment in Pregnancy.

BACKGROUND: Innovative approaches are needed to limit antimalarial resistance evolution. Understanding the role of intermittent preventive treatment in pregnancy (IPTp) on the selection for resistance and the impact such selection has on pregnancy outcomes can guide future interventions. METHODS: Plasmodium falciparum isolates (n = 914) from 2 randomized clinical trials were screened for pfmdr1 copy number variation and pfcrt, pfmdr1, pfdhfr, and pfdhps resistance markers. The trials were conducted between 2010 and 2013 in Benin, Gabon, Kenya, and Mozambique to establish the efficacy of IPTp-mefloquine (MQ) compared with IPTp-sulphadoxine-pyrimethamine (SP) in human immunodeficiency virus (HIV)-uninfected and to IPTp-placebo in HIV-infected women. RESULTS: In HIV-uninfected women, the prevalence of pfcrt mutants, pfdhfr/pfdhps quintuple mutants, and pfmdr1 copy number was similar between women receiving IPT-SP and IPTp-MQ. However, prevalence of pfmdr1 polymorphism 86Y was lower in the IPTp-MQ group than in the IPTp-SP group, and within the IPTp-MQ group it was lower at delivery compared with recruitment. No effect of IPTp-MQ on resistance markers was observed among HIV-infected women. The carriage of resistance markers was not associated with pregnancy outcomes. CONCLUSIONS: Selection of wild-type pfmdr1 polymorphism N86 by IPTp-MQ highlights the strong selective pressure IPTp can exert and the opportunity for using negative cross-resistance in drug choice for clinical treatment and IPTp.

A multiphase program for malaria elimination in southern Mozambique (the Magude project): A before-after study.

BACKGROUND: Malaria eradication remains the long-term vision of the World Health Organization (WHO). However, whether malaria elimination is feasible in areas of stable transmission in sub-Saharan Africa with currently available tools remains a subject of debate. This study aimed to evaluate a multiphased malaria elimination project to interrupt Plasmodium falciparum malaria transmission in a rural district of southern Mozambique. METHODS AND FINDINGS: A before-after study was conducted between 2015 and 2018 in the district of Magude, with 48,448 residents living in 10,965 households. Building on an enhanced surveillance system, two rounds of mass drug administrations (MDAs) per year over two years (phase I, August 2015-2017), followed by one year of reactive focal mass drug administrations (rfMDAs) (phase II, September 2017-June 2018) were deployed with annual indoor residual spraying (IRS), programmatically distributed long-lasting insecticidal nets (LLINs), and standard case management. The four MDA rounds covered 58%-72% of the population, and annual IRS reported coverage was >70%. Yearly parasite surveys and routine surveillance data were used to monitor the primary outcomes of the study-malaria prevalence and incidence-at baseline and annually since the onset of the project. Parasite prevalence by rapid diagnostic test (RDT) declined from 9.1% (95% confidence interval [CI] 7.0-11.8) in May 2015 to 2.6% (95% CI 2.0-3.4), representing a 71.3% (95% CI 71.1-71.4, p < 0.001) reduction after phase I, and to 1.4% (95% CI 0.9-2.2) after phase II. This represented an 84.7% (95% CI 81.4-87.4, p < 0.001) overall reduction in all-age prevalence. Case incidence fell from 195 to 75 cases per 1,000 during phase I (61.5% reduction) and to 67 per 1,000 during phase II (65.6% overall reduction). Interrupted time series (ITS) analysis was used to estimate the level and trend change in malaria cases associated with the set of project interventions and the number of cases averted. Phase I interventions were associated with a significant immediate reduction in cases of 69.1% (95% CI 57.5-77.6, p < 0.001). Phase II interventions were not associated with a level or trend change. An estimated 76.7% of expected cases were averted throughout the project (38,369 cases averted of 50,005 expected). One malaria-associated inpatient death was observed during the study period. There were 277 mild adverse events (AEs) recorded through the passive pharmacovigilance system during the four MDA rounds. One serious adverse event (SAE) that resulted in death was potentially related to the drug. The study was limited by the incomplete coverage of interventions, the quality of the routine and cross-sectional data collected, and the restricted accuracy of ITS analysis with a short pre-intervention period. CONCLUSION: In this study, we observed that the interventions deployed during the Magude project fell short of interrupting P. falciparum transmission with the coverages achieved. While new tools and strategies may be required to eventually achieve malaria elimination in stable transmission areas of sub-Saharan Africa, this project showed that innovative mixes of interventions can achieve large reductions in disease burden, a necessary step in the pathway towards elimination. TRIAL REGISTRATION: ClinicalTrials.gov NCT02914145.

The duration of chemoprophylaxis against malaria after treatment with artesunate-amodiaquine and artemether-lumefantrine and the effects of pfmdr1 86Y and pfcrt 76T: a meta-analysis of individual patient data.

BACKGROUND: The majority of Plasmodium falciparum malaria cases in Africa are treated with the artemisinin combination therapies artemether-lumefantrine (AL) and artesunate-amodiaquine (AS-AQ), with amodiaquine being also widely used as part of seasonal malaria chemoprevention programs combined with sulfadoxine-pyrimethamine. While artemisinin derivatives have a short half-life, lumefantrine and amodiaquine may give rise to differing durations of post-treatment prophylaxis, an important additional benefit to patients in higher transmission areas. METHODS: We analyzed individual patient data from 8 clinical trials of AL versus AS-AQ in 12 sites in Africa (n = 4214 individuals). The time to PCR-confirmed reinfection after treatment was used to estimate the duration of post-treatment protection, accounting for variation in transmission intensity between settings using hidden semi-Markov models. Accelerated failure-time models were used to identify potential effects of covariates on the time to reinfection. The estimated duration of chemoprophylaxis was then used in a mathematical model of malaria transmission to determine the potential public health impact of each drug when used for first-line treatment. RESULTS: We estimated a mean duration of post-treatment protection of 13.0 days (95% CI 10.7-15.7) for AL and 15.2 days (95% CI 12.8-18.4) for AS-AQ overall. However, the duration varied significantly between trial sites, from 8.7-18.6 days for AL and 10.2-18.7 days for AS-AQ. Significant predictors of time to reinfection in multivariable models were transmission intensity, age, drug, and parasite genotype. Where wild type pfmdr1 and pfcrt parasite genotypes predominated (<=20% 86Y and 76T mutants, respectively), AS-AQ provided ~ 2-fold longer protection than AL. Conversely, at a higher prevalence of 86Y and 76T mutant parasites (> 80%), AL provided up to 1.5-fold longer protection than AS-AQ. Our simulations found that these differences in the duration of protection could alter population-level clinical incidence of malaria by up to 14% in under-5-year-old children when the drugs were used as first-line treatments in areas with high, seasonal transmission. CONCLUSION: Choosing a first-line treatment which provides optimal post-treatment prophylaxis given the local prevalence of resistance-associated markers could make a significant contribution to reducing malaria morbidity.

Blood cytokine, chemokine and growth factor profiling in a cohort of pregnant women from tropical countries.

The immune status of women changes during and after pregnancy, differs between blood compartments at delivery and is affected by environmental factors particularly in tropical areas endemic for multiple infections. We quantified the plasma concentration of a set of thirty-one TH1, TH2, TH17 and regulatory cytokines, pro-inflammatory and anti-inflammatory cytokines and chemokines, and growth factors (altogether biomarkers), in a cohort of 540 pregnant women from five malaria-endemic tropical countries. Samples were collected at recruitment (first antenatal visit), delivery (periphery, cord and placenta) and postpartum, allowing a longitudinal analysis. We found the lowest concentration of biomarkers at recruitment and the highest at postpartum, with few exceptions. Among them, IL-6, HGF and TGF-ß had the highest levels at delivery, and even higher concentrations in the placenta compared to peripheral blood. Placental concentrations were generally higher than peripheral, except for eotaxin that was lower. We also compared plasma biomarker concentrations between the tropical cohort and a control group from Spain at delivery, presenting overall higher biomarker levels the tropical cohort, particularly pro-inflammatory cytokines and growth factors. Only IL-6 presented lower levels in the tropical group. Moreover, a principal component analysis of biomarker concentrations at delivery showed that women from Spain grouped more homogenously, and that IL-6 and IL-8 clustered together in the tropical cohort but not in the Spanish one. Plasma cytokine concentrations correlated with Plasmodium antibody levels at postpartum but not during pregnancy. This basal profiling of immune mediators over gestation and in different compartments at delivery is important to subsequently understand response to infections and clinical outcomes in mothers and infants in tropical areas.

Systematic review of artemisinin embryotoxicity in animals: Implications for malaria control in human pregnancy.

Pregnant women are one of the most susceptible and vulnerable groups to malaria, the most important parasitic disease worldwide. Artemisinin-based combination therapies (ACTs) are recommended for the treatment of uncomplicated malaria in all population groups including pregnant women. However, due to the embryotoxicity observed in animal studies, ACTs have long been contraindicated during the first trimester in pregnant women. Despite the safety concerns raised in pre-clinical studies, recent findings on ACTs's use in pregnant women appear to be reassuring regarding safety and have prompted a revision of malaria treatment guidelines for first trimester of pregnancy. To contribute to the risk-benefit assessment of ACTs, we conducted a systematic literature review of animal studies published between 2007 and 2019, which evaluated the embryotoxic effects of artemisinin and its derivatives among pregnant mammals. Eighteen experimental studies fitted the inclusion criteria. These studies confirmed and further characterized the severe embryolethal and embryotoxic dose-dependent effects of artemisinin and its derivatives when administered during the organogenesis period in rats, rabbits and monkeys. Timing of administration and dosage of the drug were found to be key factors in the appearance of embryo damage. Overall, the translation of the findings of artemisinin derivatives use in animal studies to pregnant women remains disturbing. Thus, a policy change in the use of ACTs during the first trimester in pregnant women for the treatment of uncomplicated malaria does not seem pertinent and if implemented, it should be accompanied by solid pharmacovigilance systems, which are challenging to establish in malaria endemic countries.

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma.

BACKGROUND: Malaria diagnostics by rapid diagnostic test (RDT) relies primarily on the qualitative detection of Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and Plasmodium spp lactate dehydrogenase (pLDH). As novel RDTs with increased sensitivity are being developed and implemented as point of care diagnostics, highly sensitive laboratory-based assays are needed for evaluating RDT performance. Here, a quantitative suspension array technology (qSAT) was developed, validated and applied for the simultaneous detection of PfHRP2 and pLDH in a variety of biological samples (whole blood, plasma and dried blood spots) from individuals living in different endemic countries. RESULTS: The qSAT was specific for the target antigens, with analytical ranges of 6.8 to 762.8 pg/ml for PfHRP2 and 78.1 to 17076.6 pg/ml for P. falciparum LDH (Pf-LDH). The assay detected Plasmodium vivax LDH (Pv-LDH) at a lower sensitivity than Pf-LDH (analytical range of 1093.20 to 187288.5 pg/ml). Both PfHRP2 and pLDH levels determined using the qSAT showed to positively correlate with parasite densities determined by quantitative PCR (Spearman r = 0.59 and 0.75, respectively) as well as microscopy (Spearman r = 0.40 and 0.75, respectively), suggesting the assay to be a good predictor of parasite density. CONCLUSION: This immunoassay can be used as a reference test for the detection and quantification of PfHRP2 and pLDH, and could serve for external validation of RDT performance, to determine antigen persistence after parasite clearance, as well as a complementary tool to assess malaria burden in endemic settings.

Lay knowledge of cervical cancer in Manhiça district, Mozambique: a qualitative study.

BACKGROUND: Mozambique has one of the highest cervical cancer incidence rates in the world. Health interventions are still being conceived solely from a non-communicable disease standpoint despite that it is also a sexual and reproductive health problem. The objective of this study was to assess the extent to which lay perceptions of cervical cancer align with biomedical knowledge from the standpoint of sexual and reproductive health. METHODS: 10 focus group discussions were carried out with 10 target groups in Manhiça. The target groups were diverse in terms of age, sex, educational level and occupation. There were a total of 116 participants. The focus groups discussions were applied to obtain verbal information and trigger debates around beliefs and attitudes about cervical cancer as well as to explore notions of transmission and aetiology of the disease. The discussions were recorded for later transcription and analysis, following a combination of content and thematic analysis. RESULTS: Participants were familiar with the biomedical term 'cervical cancer' but knowledge of its aetiology and transmission was limited. Cervical cancer was readily associated to sexual transmission and sexually transmitted infections, and conceived as a 'wound that does not heal'. The term 'cancer' caused confusion, as it was perceived to happen only in limbs, understood as hereditary, not transmissible and as an illness of the West. CONCLUSIONS: Lay perceptions of cervical cancer do, to a large extent, align with biomedical ones, thus, there is common ground to frame future health interventions from a sexual and reproductive health standpoint. Some misperceptions were identified which could be reduced through social behaviour change communication initiatives.

Standardization of Minimally Invasive Tissue Sampling Specimen Collection and Pathology Training for the Child Health and Mortality Prevention Surveillance Network.

BACKGROUND: Minimally invasive tissue sampling (MITS) is a simplified postmortem examination technique that has shown to be an adequate approach for cause of death investigation in low-resource settings. It requires relatively low level of infrastructures and can be performed by health professionals with no background in pathology. A training program has been developed for the Child Health and Mortality Prevention Surveillance (CHAMPS) network to guarantee standardization of specimen collection techniques, procedures, and laboratory methods. METHODS: The training program has included assessment of the site capacities and training on a standardized protocol of MITS sampling and histological processing. The project has also introduced a program of training for trainers for the personnel from Mozambique. To guarantee the adequacy of the procedure in each site, a trainer accompanied the local teams when the activities started. Training outcomes were assessed by evaluating the quality of the samples obtained and the quality of the slides produced locally. RESULTS: Between June 2016 and October 2018, the laboratories of 7 sites (Bangladesh, Ethiopia, Kenya, Mali, Mozambique, Sierra Leone, and South Africa) have been evaluated and upgraded. Training has been delivered to 63 staff members from all sites. More than 600 MITS procedures have been performed. The quantity of tissue obtained in the MITS by the local teams was sufficient or abundant in 73%, and 87% of the slides were considered as technically acceptable or excellent. CONCLUSIONS: Satisfactory standardization of MITS and histology procedures has been achieved across all CHAMPS sites through organized capacity-building plans.