Adjunctive rosiglitazone treatment for severe pediatric malaria: A randomized placebo-controlled trial in Mozambican children.

OBJECTIVES: We tested the hypothesis that adjunctive rosiglitazone treatment would reduce levels of circulating angiopoietin-2 (Angpt-2) and improve outcomes of Mozambican children with severe malaria. METHODS: A randomized, double-blind, placebo-controlled trial of rosiglitazone vs placebo as adjunctive treatment to artesunate in children with severe malaria was conducted. A 0.045 mg/kg/dose of rosiglitazone or matching placebo were administered, in addition to standard of malaria care, twice a day for 4 days. The primary endpoint was the rate of decline of Angpt-2 over 96 hours. Secondary outcomes included the longitudinal dynamics of angiopoietin-1 (Angpt-1) and the Angpt-2/Angpt-1 ratio over 96 hours, parasite clearance kinetics, clinical outcomes, and safety metrics. RESULTS: Overall, 180 children were enrolled; 91 were assigned to rosiglitazone and 89 to placebo. Children who received rosiglitazone had a steeper rate of decline of Angpt-2 over the first 96 hours of hospitalization compared to children who received placebo; however, the trend was not significant (P = 0.28). A similar non-significant trend was observed for Angpt-1 (P = 0.65) and the Angpt-2/Angpt-1 ratio (P = 0.34). All other secondary and safety outcomes were similar between groups (P >0.05). CONCLUSION: Adjunctive rosiglitazone at this dosage was safe and well tolerated but did not significantly affect the longitudinal kinetics of circulating Angpt-2.

Evaluation of the Performance of a 3D-Printed Smartphone-Based Retinal Imaging Device as a Screening Tool for Retinal Pathology in Mozambique.

Low-income countries carry approximately 90% of the global burden of visual impairment, and up to 80% of this could be prevented or cured. However, there are only a few studies on the prevalence of retinal disease in these countries. Easier access to retinal information would allow differential diagnosis and promote strategies to improve eye health, which are currently scarce. This pilot study aims to evaluate the functionality and usability of a tele-retinography system for the detection of retinal pathology, based on a low-cost portable retinal scanner, manufactured with 3D printing and controlled by a mobile phone with an application designed ad hoc. The study was conducted at the Manhiça Rural Hospital in Mozambique. General practitioners, with no specific knowledge of ophthalmology or previous use of retinography, performed digital retinographies on 104 hospitalized patients. The retinographies were acquired in video format, uploaded to a web platform, and reviewed centrally by two ophthalmologists, analyzing the image quality and the presence of retinal lesions. In our sample there was a high proportion of exudates and hemorrhages-8% and 4%, respectively. In addition, the presence of lesions was studied in patients with known underlying risk factors for retinal disease, such as HIV, diabetes, and/or hypertension. Our tele-retinography system based on a smartphone coupled with a simple and low-cost 3D printed device is easy to use by healthcare personnel without specialized ophthalmological knowledge and could be applied for the screening and initial diagnosis of retinal pathology.

Endothelial transcriptomic analysis identifies biomarkers of severe and cerebral malaria.

Malaria can quickly progress from an uncomplicated infection into a life-threatening severe disease. However, the unspecificity of early symptoms often makes it difficult to identify patients at high risk of developing severe disease. Additionally, one of the most feared malaria complications - cerebral malaria - is challenging to diagnose, often resulting in treatment delays that can lead to adverse outcomes. To identify candidate biomarkers for the prognosis and/or diagnosis of severe and cerebral malaria, we have analyzed the transcriptomic response of human brain microvascular endothelial cells to erythrocytes infected with Plasmodium falciparum. Candidates were validated in plasma samples from a cohort of pediatric patients with malaria from Mozambique, resulting in the identification of several markers with capacity to distinguish uncomplicated from severe malaria, the most potent being the metallopeptidase ADAMTS18. Two other biomarkers, Angiopoietin-like-4 and Inhibin-ßE were able to differentiate children with cerebral malaria within the severe malaria group, showing increased sensitivity after combination in a biomarker signature. The validation of the predicted candidate biomarkers in plasma of children with severe and cerebral malaria underscores the power of this transcriptomic approach and indicates that a specific endothelial response to P. falciparum-infected erythrocytes is linked to the pathophysiology of severe malaria.

Provider adherence to clinical care recommendations for infants and children who died in seven low- and middle-income countries in the Child Health and Mortality Prevention Surveillance (CHAMPS) network.

Background: Most childhood deaths globally are considered preventable through high-quality clinical care, which includes adherence to clinical care recommendations. Our objective was to describe adherence to World Health Organization recommendations for the management of leading causes of death among children. Methods: We conducted a retrospective, descriptive study examining clinical data for children aged 1-59 months who were hospitalized and died in a Child Health and Mortality Prevention Surveillance (CHAMPS) catchment, December 2016-June 2021. Catchment areas included: Baliakandi and Faridpur, Bangladesh; Kersa, Haramaya, and Harar, Ethiopia; Kisumu and Siaya, Kenya; Bamako, Mali; Manhiça and Quelimane, Mozambique; Makeni, Sierra Leone; Soweto, South Africa. We reviewed medical records of those who died from lower respiratory tract infections, sepsis, malnutrition, malaria, and diarrheal diseases to determine the proportion who received recommended treatments and compared adherence by hospitalization duration. Findings: CHAMPS enrolled 460 hospitalized children who died from the leading causes (median age 12 months, 53.0% male). Median hospital admission was 31 h. There were 51.0% (n = 127/249) of children who died from lower respiratory tract infections received supplemental oxygen. Administration of intravenous fluids for sepsis (15.9%, n = 36/226) and supplemental feeds for malnutrition (14.0%, n = 18/129) were uncommon. There were 51.4% (n = 55/107) of those who died from malaria received antimalarials. Of the 80 children who died from diarrheal diseases, 76.2% received intravenous fluids. Those admitted for ≥24 h more commonly received antibiotics for lower respiratory tract infections and sepsis, supplemental feeds for malnutrition, and intravenous fluids for sepsis than those admitted <24 h. Interpretation: Provision of recommended clinical care for leading causes of death among young children was suboptimal. Further studies are needed to understand the reasons for deficits in clinical care recommendation adherence. Funding: Bill & Melinda Gates Foundation.

Causes of Death Among Infants and Children in the Child Health and Mortality Prevention Surveillance (CHAMPS) Network.

Importance: The number of deaths of children younger than 5 years has been steadily decreasing worldwide, from more than 17 million annual deaths in the 1970s to an estimated 5.3 million in 2019 (with 2.8 million deaths occurring in those aged 1-59 months [53% of all deaths in children aged <5 years]). More detailed characterization of childhood deaths could inform interventions to improve child survival. Objective: To describe causes of postneonatal child deaths across 7 mortality surveillance sentinel sites in Africa and Asia. Design, Setting, and Participants: The Child Health and Mortality Prevention Surveillance (CHAMPS) Network conducts childhood mortality surveillance in sub-Saharan Africa and South Asia using innovative postmortem minimally invasive tissue sampling (MITS). In this cross-sectional study, MITS was conducted in deceased children aged 1 to 59 months at 7 sites in sub-Saharan Africa and South Asia from December 3, 2016, to December 3, 2020. Data analysis was conducted between October and November 2021. Main Outcomes and Measures: The expert panel attributed underlying, intermediate, and immediate conditions in the chain of events leading to death, based on histopathologic analysis, microbiological diagnostics, clinical data, and verbal autopsies. Results: In this study, MITS was performed in 632 deceased children (mean [SD] age at death, 1.3 [0.3] years; 342 [54.1%] male). The 6 most common underlying causes of death were malnutrition (104 [16.5%]), HIV (75 [11.9%]), malaria (71 [11.2%]), congenital birth defects (64 [10.1%]), lower respiratory tract infections (LRTIs; 53 [8.4%]), and diarrheal diseases (46 [7.2%]). When considering immediate causes only, sepsis (191 [36.7%]) and LRTI (129 [24.8%]) were the 2 dominant causes. An infection was present in the causal chain in 549 of 632 deaths (86.9%); pathogens most frequently contributing to infectious deaths included Klebsiella pneumoniae (155 of 549 infectious deaths [28.2%]; 127 [81.9%] considered nosocomial), Plasmodium falciparum (122 of 549 [22.2%]), and Streptococcus pneumoniae (109 of 549 [19.9%]). Other organisms, such as cytomegalovirus (57 [10.4%]) and Acinetobacter baumannii (39 [7.1%]; 35 of 39 [89.7%] considered nosocomial), also played important roles. For the top underlying causes of death, the median number of conditions in the chain of events leading to death was 3 for malnutrition, 3 for HIV, 1 for malaria, 3 for congenital birth defects, and 1 for LRTI. Expert panels considered 494 of 632 deaths (78.2%) preventable and 26 of 632 deaths (4.1%) preventable under certain conditions. Conclusions and Relevance: In this cross-sectional study investigating causes of child mortality in the CHAMPS Network, results indicate that, in these high-mortality settings, infectious diseases continue to cause most deaths in infants and children, often in conjunction with malnutrition. These results also highlight opportunities for action to prevent deaths and reveal common interaction of various causes in the path toward death.

Experiences of parents and caretakers going through the consent process to perform minimally invasive tissue sampling (MITS) on their deceased children in Quelimane, Mozambique: A qualitative study.

BACKGROUND: In Mozambique, the Countrywide Mortality Surveillance for Action (COMSA) Program implemented a child mortality surveillance to strengthen vital events registration (pregnancies, births, and deaths) and investigate causes of death using verbal autopsies. In Quelimane district, in addition to the abovementioned cause of death determination approaches, minimally invasive tissue sampling (MITS) was performed on deceased children <5years of age. This study focused on understanding deceased children parents' and caretakers' experiences of the consent process to perform MITS in order to contribute to the improvement of approaches to cause of death investigation and inform efforts to maximize acceptability of mortality surveillance activities. METHODS: A qualitative study was conducted in six urban and semi-urban communities in Quelimane district. A total of 40 semi-structured interviews with family members of deceased children and 50 non-participant observations of the consent process were conducted to explore their experience with informed consent request to perform MITS on their child. Data analysis of the interviews and observations was thematic, being initially deductive (predetermined codes) followed by the generation of new codes according to the data (inductive).The Consolidated criteria for reporting qualitative research (COREQ) guidelines for reporting qualitative studies were performed. FINDINGS: Although most participants consented to the performance of MITS on their deceased child, some stated they had not fully understood the MITS procedure despite the informed consent process due to unclear information and their state of mind after their loss. Consenting to MITS and doing so with family members disagreeing were also identified as stress-enhancing factors. Participants also described dissatisfaction of family members, resulting from the condition of the body delivered after tissue collection. In addition, the waiting time to receive the body and resulting delays for the funeral were considered additional factors that may increase stress and compromise the acceptability of MITS. CONCLUSION: Family experiences were influenced by operational and logistical issues linked to the procedure itself and by it being in tension with social and cultural issues, which caused stress and discontentment on parents and caretakers of deceased children. The main factors that contributed to the experience of going through the MITS process were the state of mind after the death, complex decision making processes within the family, washing of the body for purification after MITS and seepage, and limited understanding of consent for MITS. When requesting consent for MITS, emphasis should be placed on transmitting clear and understandable information about MITS procedures to participants.

Tracking SARS-CoV-2 introductions in Mozambique using pandemic-scale phylogenies: a retrospective observational study

From the start of the SARS-CoV-2 outbreak, global sequencing efforts have generated an unprecedented amount of genomic data. Nonetheless, unequal sampling between high-income and low-income countries hinders the implementation of genomic surveillance systems at the global and local level. Filling the knowledge gaps of genomic information and understanding pandemic dynamics in low-income countries is essential for public health decision making and to prepare for future pandemics. In this context, we aimed to discover the timing and origin of SARS-CoV-2 variant introductions in Mozambique, taking advantage of pandemic-scale phylogenies. Methods: We did a retrospective, observational study in southern Mozambique. Patients from Manhiça presenting with respiratory symptoms were recruited, and those enrolled in clinical trials were excluded. Data were included from three sources: (1) a prospective hospital-based surveillance study (MozCOVID), recruiting patients living in Manhiça, attending the Manhiça district hospital, and fulfilling the criteria of suspected COVID-19 case according to WHO; (2) symptomatic and asymptomatic individuals with SARS-CoV-2 infection recruited by the National Surveillance system; and (3) sequences from SARS-CoV-2-infected Mozambican cases deposited on the Global Initiative on Sharing Avian Influenza Data database. Positive samples amenable for sequencing were analysed. We used Ultrafast Sample placement on Existing tRees to understand the dynamics of beta and delta waves, using available genomic data. This tool can reconstruct a phylogeny with millions of sequences by efficient sample placement in a tree. We reconstructed a phylogeny (~7·6 million sequences) adding new and publicly available beta and delta sequences. Findings: A total of 5793 patients were recruited between Nov 1, 2020, and Aug 31, 2021. During this time, 133 328 COVID-19 cases were reported in Mozambique. 280 good quality new SARS-CoV-2 sequences were obtained after the inclusion criteria were applied and an additional 652 beta (B.1.351) and delta (B.1.617.2) public sequences were included from Mozambique. We evaluated 373 beta and 559 delta sequences. We identified 187 beta introductions (including 295 sequences), divided in 42 transmission groups and 145 unique introductions, mostly from South Africa, between August, 2020 and July, 2021. For delta, we identified 220 introductions (including 494 sequences), with 49 transmission groups and 171 unique introductions, mostly from the UK, India, and South Africa, between April and November, 2021. Interpretation: The timing and origin of introductions suggests that movement restrictions effectively avoided introductions from non-African countries, but not from surrounding countries. Our results raise questions about the imbalance between the consequences of restrictions and health benefits. This new understanding of pandemic dynamics in Mozambique can be used to inform public health interventions to control the spread of new variants. Funding: European and Developing Countries Clinical Trials, European Research Council, Bill & Melinda Gates Foundation, and Agència de Gestió d'Ajuts Universitaris i de Recerca.

Tracking SARS-CoV-2 introductions in Mozambique using pandemic-scale phylogenies: a retrospective observational study.

BACKGROUND: From the start of the SARS-CoV-2 outbreak, global sequencing efforts have generated an unprecedented amount of genomic data. Nonetheless, unequal sampling between high-income and low-income countries hinders the implementation of genomic surveillance systems at the global and local level. Filling the knowledge gaps of genomic information and understanding pandemic dynamics in low-income countries is essential for public health decision making and to prepare for future pandemics. In this context, we aimed to discover the timing and origin of SARS-CoV-2 variant introductions in Mozambique, taking advantage of pandemic-scale phylogenies. METHODS: We did a retrospective, observational study in southern Mozambique. Patients from Manhiça presenting with respiratory symptoms were recruited, and those enrolled in clinical trials were excluded. Data were included from three sources: (1) a prospective hospital-based surveillance study (MozCOVID), recruiting patients living in Manhiça, attending the Manhiça district hospital, and fulfilling the criteria of suspected COVID-19 case according to WHO; (2) symptomatic and asymptomatic individuals with SARS-CoV-2 infection recruited by the National Surveillance system; and (3) sequences from SARS-CoV-2-infected Mozambican cases deposited on the Global Initiative on Sharing Avian Influenza Data database. Positive samples amenable for sequencing were analysed. We used Ultrafast Sample placement on Existing tRees to understand the dynamics of beta and delta waves, using available genomic data. This tool can reconstruct a phylogeny with millions of sequences by efficient sample placement in a tree. We reconstructed a phylogeny (~7·6 million sequences) adding new and publicly available beta and delta sequences. FINDINGS: A total of 5793 patients were recruited between Nov 1, 2020, and Aug 31, 2021. During this time, 133 328 COVID-19 cases were reported in Mozambique. 280 good quality new SARS-CoV-2 sequences were obtained after the inclusion criteria were applied and an additional 652 beta (B.1.351) and delta (B.1.617.2) public sequences were included from Mozambique. We evaluated 373 beta and 559 delta sequences. We identified 187 beta introductions (including 295 sequences), divided in 42 transmission groups and 145 unique introductions, mostly from South Africa, between August, 2020 and July, 2021. For delta, we identified 220 introductions (including 494 sequences), with 49 transmission groups and 171 unique introductions, mostly from the UK, India, and South Africa, between April and November, 2021. INTERPRETATION: The timing and origin of introductions suggests that movement restrictions effectively avoided introductions from non-African countries, but not from surrounding countries. Our results raise questions about the imbalance between the consequences of restrictions and health benefits. This new understanding of pandemic dynamics in Mozambique can be used to inform public health interventions to control the spread of new variants. FUNDING: European and Developing Countries Clinical Trials, European Research Council, Bill & Melinda Gates Foundation, and Agència de Gestió d'Ajuts Universitaris i de Recerca.

A Qualitative Assessment of Community Acceptability and Its Determinants in the Implementation of Minimally Invasive Tissue Sampling in Children in Quelimane City, Mozambique.

The Countrywide Mortality Surveillance for Action project aims to implement a child mortality surveillance program through strengthening vital registration event reporting (pregnancy, birth, and death) and investigating causes of death (CODs) based on verbal autopsies. In Quelimane (central Mozambique), Minimally Invasive Tissue Sampling (MITS) procedures were added to fine-tune the COD approaches. Before the implementation of MITS, an evaluation of the acceptability and ethical considerations of child mortality surveillance was considered fundamental. A socio-anthropological study was conducted in Quelimane, using observations, informal conversations, semi-structured interviews, and focus group discussions with healthcare providers, nharrubes (traditional authorities who handle bodies before the funeral), community and religious leaders, and traditional birth attendants to understand the locally relevant potential facilitators and barriers to the acceptability of MITS. Audio materials were transcribed, systematically coded, and analyzed using NVIVO12®. The desire to know the COD, intention to discharge the elders from accusations of witchcraft, involvement of leaders in disseminating project information, and provision of transport for bodies back to the community constitute potential facilitators for the acceptability of MITS implementation. In contrast, poor community mobilization, disagreement with Islamic religious practices, and local traditional beliefs were identified as potential barriers. MITS was considered a positive innovation to determine the COD, although community members remain skeptical about the procedure due to tensions with religion and tradition. Therefore, the implementation of MITS in Quelimane should prioritize the involvement of a variety of influential community and religious leaders.

Causes of death identified in neonates enrolled through Child Health and Mortality Prevention Surveillance (CHAMPS), December 2016 -December 2021.

Each year, 2.4 million children die within their first month of life. Child Health and Mortality Prevention Surveillance (CHAMPS) established in 7 countries aims to generate accurate data on why such deaths occur and inform prevention strategies. Neonatal deaths that occurred between December 2016 and December 2021 were investigated with MITS within 24-72 hours of death. Testing included blood, cerebrospinal fluid and lung cultures, multi-pathogen PCR on blood, CSF, nasopharyngeal swabs and lung tissue, and histopathology examination of lung, liver and brain. Data collection included clinical record review and family interview using standardized verbal autopsy. The full set of data was reviewed by local experts using a standardized process (Determination of Cause of Death) to identify all relevant conditions leading to death (causal chain), per WHO recommendations. For analysis we stratified neonatal death into 24-hours of birth, early (1-<7 days) and late (7-<28 days) neonatal deaths. We analyzed 1458 deaths, 41% occurring within 24-hours, 41% early and 18% late neonatal deaths. Leading underlying causes of death were complications of intrapartum events (31%), complications of prematurity (28%), infections (17%), respiratory disorders (11%), and congenital malformations (8%). In addition to the underlying cause, 62% of deaths had additional conditions and 14% had ≥3 other conditions in the causal chain. The most common causes considering the whole causal chain were infection (40%), prematurity (32%) and respiratory distress syndrome (28%). Common maternal conditions linked to neonatal death were maternal hypertension (10%), labour and delivery complications (8%), multiple gestation (7%), placental complications (6%) obstructed labour and chorioamnionitis (5%, each). CHAMPS' findings showing the full causal chain of events that lead to death, in addition to maternal factors, highlights the complexities involved in each death along with the multiple opportunities for prevention. Highlighting improvements to prenatal and obstetric care and infection prevention are urgently needed in high-mortality settings.

Prognostic accuracy of biomarkers of immune and endothelial activation in Mozambican children hospitalized with pneumonia.

Pneumonia is a leading cause of child mortality. However, currently we lack simple, objective, and accurate risk-stratification tools for pediatric pneumonia. Here we test the hypothesis that measuring biomarkers of immune and endothelial activation in children with pneumonia may facilitate the identification of those at risk of death. We recruited children <10 years old fulfilling WHO criteria for pneumonia and admitted to the Manhiça District Hospital (Mozambique) from 2010 to 2014. We measured plasma levels of IL-6, IL-8, Angpt-2, sTREM-1, sFlt-1, sTNFR1, PCT, and CRP at admission, and assessed their prognostic accuracy for in-hospital, 28-day, and 90-day mortality. Healthy community controls, within same age strata and location, were also assessed. All biomarkers were significantly elevated in 472 pneumonia cases versus 80 controls (p<0.001). IL-8, sFlt-1, and sTREM-1 were associated with in-hospital mortality (p<0.001) and showed the best discrimination with AUROCs of 0.877 (95% CI: 0.782 to 0.972), 0.832 (95% CI: 0.729 to 0.935) and 0.822 (95% CI: 0.735 to 0.908), respectively. Their performance was superior to CRP, PCT, oxygen saturation, and clinical severity scores. IL-8, sFlt-1, and sTREM-1 remained good predictors of 28-day and 90-day mortality. These findings suggest that measuring IL-8, sFlt-1, or sTREM-1 at hospital presentation can guide risk-stratification of children with pneumonia, which could enable prioritized care to improve survival and resource allocation.

Prioritizing Health Care Strategies to Reduce Childhood Mortality.

Importance: Although child mortality trends have decreased worldwide, deaths among children younger than 5 years of age remain high and disproportionately circumscribed to sub-Saharan Africa and Southern Asia. Tailored and innovative approaches are needed to increase access, coverage, and quality of child health care services to reduce mortality, but an understanding of health system deficiencies that may have the greatest impact on mortality among children younger than 5 years is lacking. Objective: To investigate which health care and public health improvements could have prevented the most stillbirths and deaths in children younger than 5 years using data from the Child Health and Mortality Prevention Surveillance (CHAMPS) network. Design, Setting, and Participants: This cross-sectional study used longitudinal, population-based, and mortality surveillance data collected by CHAMPS to understand preventable causes of death. Overall, 3390 eligible deaths across all 7 CHAMPS sites (Bangladesh, Ethiopia, Kenya, Mali, Mozambique, Sierra Leone, and South Africa) between December 9, 2016, and December 31, 2021 (1190 stillbirths, 1340 neonatal deaths, 860 infant and child deaths), were included. Deaths were investigated using minimally invasive tissue sampling (MITS), a postmortem approach using biopsy needles for sampling key organs and fluids. Main Outcomes and Measures: For each death, an expert multidisciplinary panel reviewed case data to determine the plausible pathway and causes of death. If the death was deemed preventable, the panel identified which of 10 predetermined health system gaps could have prevented the death. The health system improvements that could have prevented the most deaths were evaluated for each age group: stillbirths, neonatal deaths (aged <28 days), and infant and child deaths (aged 1 month to <5 years). Results: Of 3390 deaths, 1505 (44.4%) were female and 1880 (55.5%) were male; sex was not recorded for 5 deaths. Of all deaths, 3045 (89.8%) occurred in a healthcare facility and 344 (11.9%) in the community. Overall, 2607 (76.9%) were deemed potentially preventable: 883 of 1190 stillbirths (74.2%), 1010 of 1340 neonatal deaths (75.4%), and 714 of 860 infant and child deaths (83.0%). Recommended measures to prevent deaths were improvements in antenatal and obstetric care (recommended for 588 of 1190 stillbirths [49.4%], 496 of 1340 neonatal deaths [37.0%]), clinical management and quality of care (stillbirths, 280 [23.5%]; neonates, 498 [37.2%]; infants and children, 393 of 860 [45.7%]), health-seeking behavior (infants and children, 237 [27.6%]), and health education (infants and children, 262 [30.5%]). Conclusions and Relevance: In this cross-sectional study, interventions prioritizing antenatal, intrapartum, and postnatal care could have prevented the most deaths among children younger than 5 years because 75% of deaths among children younger than 5 were stillbirths and neonatal deaths. Measures to reduce mortality in this population should prioritize improving existing systems, such as better access to antenatal care, implementation of standardized clinical protocols, and public education campaigns.

Plasmodium falciparum sexual conversion rates can be affected by artemisinin-based treatment in naturally infected malaria patients.

BACKGROUND: Artemisinins (ART) are the key component of the frontline antimalarial treatment, but their impact on Plasmodium falciparum sexual conversion rates in natural malaria infections remains unknown. This is an important knowledge gap because sexual conversion rates determine the relative parasite investment between maintaining infection in the same human host and transmission to mosquitoes. METHODS: The primary outcome of this study was to assess the impact of ART-based treatment on sexual conversion rates by comparing the relative transcript levels of pfap2-g and other sexual ring biomarkers (SRBs) before and after treatment. We analysed samples from previously existing cohorts in Vietnam, Burkina Faso and Mozambique (in total, n=109) collected before treatment and at 12 h intervals after treatment. As a secondary objective, we investigated factors that may influence the effect of treatment on sexual conversion rates. FINDINGS: In the majority of infections from the African cohorts, but not from Vietnam, we observed increased expression of pfap2-g and other SRBs after treatment. Estimated parasite age at the time of treatment was negatively correlated with the increase in pfap2-g transcript levels, suggesting that younger parasites are less susceptible to stimulation of sexual conversion. INTERPRETATION: We observed enhanced expression of SRBs after ART-based treatment in many patients, which suggests that in natural malaria infections sexual conversion rates can be altered by treatment. ART-based treatment reduces the potential of a treated individual to transmit the disease, but we hypothesise that under some circumstances this reduction may be attenuated by ART-enhanced sexual conversion. FUNDING: Spanish Agencia Estatal de Investigación (AEI), European Regional Development Fund (ERDF, European Union), Belgium Development Cooperation (DGD), Canadian University Health Network (UHN), TransGlobalHealth-Erasmus Mundus (European Union).

Postmortem investigations and identification of multiple causes of child deaths: An analysis of findings from the Child Health and Mortality Prevention Surveillance (CHAMPS) network.

BACKGROUND: The current burden of >5 million deaths yearly is the focus of the Sustainable Development Goal (SDG) to end preventable deaths of newborns and children under 5 years old by 2030. To accelerate progression toward this goal, data are needed that accurately quantify the leading causes of death, so that interventions can target the common causes. By adding postmortem pathology and microbiology studies to other available data, the Child Health and Mortality Prevention Surveillance (CHAMPS) network provides comprehensive evaluations of conditions leading to death, in contrast to standard methods that rely on data from medical records and verbal autopsy and report only a single underlying condition. We analyzed CHAMPS data to characterize the value of considering multiple causes of death. METHODS AND FINDINGS: We examined deaths identified from December 2016 through November 2020 from 7 CHAMPS sites (in Bangladesh, Ethiopia, Kenya, Mali, Mozambique, Sierra Leone, and South Africa), including 741 neonatal, 278 infant, and 241 child <5 years deaths for which results from Determination of Cause of Death (DeCoDe) panels were complete. DeCoDe panelists included all conditions in the causal chain according to the ICD-10 guidelines and assessed if prevention or effective management of the condition would have prevented the death. We analyzed the distribution of all conditions listed as causal, including underlying, antecedent, and immediate causes of death. Among 1,232 deaths with an underlying condition determined, we found a range of 0 to 6 (mean 1.5, IQR 0 to 2) additional conditions in the causal chain leading to death. While pathology provides very helpful clues, we cannot always be certain that conditions identified led to death or occurred in an agonal stage of death. For neonates, preterm birth complications (most commonly respiratory distress syndrome) were the most common underlying condition (n = 282, 38%); among those with preterm birth complications, 256 (91%) had additional conditions in causal chains, including 184 (65%) with a different preterm birth complication, 128 (45%) with neonatal sepsis, 69 (24%) with lower respiratory infection (LRI), 60 (21%) with meningitis, and 25 (9%) with perinatal asphyxia/hypoxia. Of the 278 infant deaths, 212 (79%) had ≥1 additional cause of death (CoD) beyond the underlying cause. The 2 most common underlying conditions in infants were malnutrition and congenital birth defects; LRI and sepsis were the most common additional conditions in causal chains, each accounting for approximately half of deaths with either underlying condition. Of the 241 child deaths, 178 (75%) had ≥1 additional condition. Among 46 child deaths with malnutrition as the underlying condition, all had ≥1 other condition in the causal chain, most commonly sepsis, followed by LRI, malaria, and diarrheal disease. Including all positions in the causal chain for neonatal deaths resulted in 19-fold and 11-fold increases in attributable roles for meningitis and LRI, respectively. For infant deaths, the proportion caused by meningitis and sepsis increased by 16-fold and 11-fold, respectively; for child deaths, sepsis and LRI are increased 12-fold and 10-fold, respectively. While comprehensive CoD determinations were done for a substantial number of deaths, there is potential for bias regarding which deaths in surveillance areas underwent minimally invasive tissue sampling (MITS), potentially reducing representativeness of findings. CONCLUSIONS: Including conditions that appear anywhere in the causal chain, rather than considering underlying condition alone, markedly changed the proportion of deaths attributed to various diagnoses, especially LRI, sepsis, and meningitis. While CHAMPS methods cannot determine when 2 conditions cause death independently or may be synergistic, our findings suggest that considering the chain of events leading to death can better guide research and prevention priorities aimed at reducing child deaths.

Deaths Attributed to Respiratory Syncytial Virus in Young Children in High-Mortality Rate Settings: Report from Child Health and Mortality Prevention Surveillance (CHAMPS).

BACKGROUND: Lower respiratory tract infections are a leading cause of death in young children, but few studies have collected the specimens needed to define the role of specific causes. The Child Health and Mortality Prevention Surveillance (CHAMPS) platform aims to investigate causes of death in children aged <5 years in high-mortality rate settings, using postmortem minimally invasive tissue sampling and other advanced diagnostic techniques. We examined findings for deaths identified in CHAMPS sites in 7 countries in sub-Saharan Africa and south Asia to evaluate the role of respiratory syncytial virus (RSV). METHODS: We included deaths that occurred between December 2016 and December 2019. Panels determined causes of deaths by reviewing all available data including pathological results from minimally invasive tissue sampling, polymerase chain reaction screening for multiple infectious pathogens in lung tissue, nasopharyngeal swab, blood, and cerebrospinal fluid samples, clinical information from medical records, and verbal autopsies. RESULTS: We evaluated 1213 deaths, including 695 in neonates (aged <28 days), 283 in infants (28 days to <12 months), and 235 in children (12-59 months). RSV was detected in postmortem specimens in 67 of 1213 deaths (5.5%); in 24 deaths (2.0% of total), RSV was determined to be a cause of death, and it contributed to 5 other deaths. Younger infants (28 days to <6 months of age) accounted for half of all deaths attributed to RSV; 6.5% of all deaths in younger infants were attributed to RSV. RSV was the underlying and only cause in 4 deaths; the remainder (n = 20) had a median of 2 (range, 1-5) other conditions in the causal chain. Birth defects (n = 8) and infections with other pathogens (n = 17) were common comorbid conditions. CONCLUSIONS: RSV is an important cause of child deaths, particularly in young infants. These findings add to the substantial body of literature calling for better treatment and prevention options for RSV in high-mortality rate settings.

Klebsiella spp. cause severe and fatal disease in Mozambican children: antimicrobial resistance profile and molecular characterization.

BACKGROUND: Klebsiella spp. are important pathogens associated with bacteremia among admitted children and is among the leading cause of death in children < 5 years in postmortem studies, supporting a larger role than previously considered in childhood mortality. Herein, we compared the antimicrobial susceptibility, mechanisms of resistance, and the virulence profile of Klebsiella spp. from admitted and postmortem children. METHODS: Antimicrobial susceptibility and virulence factors of Klebsiella spp. recovered from blood samples collected upon admission to the hospital (n = 88) and postmortem blood (n = 23) from children < 5 years were assessed by disk diffusion and multiplex PCR. RESULTS: Klebsiella isolates from postmortem blood were likely to be ceftriaxone resistant (69.6%, 16/23 vs. 48.9%, 43/88, p = 0.045) or extended-spectrum ß-lactamase (ESBL) producers (60.9%, 14/23 vs. 25%, 22/88, p = 0.001) compared to those from admitted children. blaCTX-M-15 was the most frequent ESBL gene: 65.3%, 9/14 in postmortem isolates and 22.7% (5/22) from admitted children. We found higher frequency of genes associated with hypermucoviscosity phenotype and invasin in postmortem isolates than those from admitted children: rmpA (30.4%; 7/23 vs. 9.1%, 8/88, p = 0.011), wzi-K1 (34.7%; 8/23 vs. 8%; 7/88, p = 0.002) and traT (60.8%; 14/23 vs. 10.2%; 9/88, p < 0.0001), respectively. Additionally, serine protease auto-transporters of Enterobacteriaceae were detected from 1.8% (pic) to 12.6% (pet) among all isolates. Klebsiella case fatality rate was 30.7% (23/75). CONCLUSION: Multidrug resistant Klebsiella spp. harboring genes associated with hypermucoviscosity phenotype has emerged in Mozambique causing invasive fatal disease in children; highlighting the urgent need for prompt diagnosis, appropriate treatment and effective preventive measures for infection control.

Plasma MicroRNA Profiling of Plasmodium falciparum Biomass and Association with Severity of Malaria Disease.

Severe malaria (SM) is a major public health problem in malaria-endemic countries. Sequestration of Plasmodium falciparum-infected erythrocytes in vital organs and the associated inflammation leads to organ dysfunction. MicroRNAs (miRNAs), which are rapidly released from damaged tissues into the host fluids, constitute a promising biomarker for the prognosis of SM. We applied next-generation sequencing to evaluate the differential expression of miRNAs in SM and in uncomplicated malaria (UM) in children in Mozambique. Six miRNAs were associated with in vitro P. falciparum cytoadhesion, severity in children, and P. falciparum biomass. Relative expression of hsa-miR-4497 quantified by TaqMan-quantitative reverse transcription PCR was higher in plasma of children with SM than those with UM (p<0.048) and again correlated with P. falciparum biomass (p = 0.033). These findings suggest that different physiopathological processes in SM and UM lead to differential expression of miRNAs and suggest a pathway for assessing their prognostic value malaria.

The impact of delayed treatment of uncomplicated P. falciparum malaria on progression to severe malaria: A systematic review and a pooled multicentre individual-patient meta-analysis.

BACKGROUND: Delay in receiving treatment for uncomplicated malaria (UM) is often reported to increase the risk of developing severe malaria (SM), but access to treatment remains low in most high-burden areas. Understanding the contribution of treatment delay on progression to severe disease is critical to determine how quickly patients need to receive treatment and to quantify the impact of widely implemented treatment interventions, such as 'test-and-treat' policies administered by community health workers (CHWs). We conducted a pooled individual-participant meta-analysis to estimate the association between treatment delay and presenting with SM. METHODS AND FINDINGS: A search using Ovid MEDLINE and Embase was initially conducted to identify studies on severe Plasmodium falciparum malaria that included information on treatment delay, such as fever duration (inception to 22nd September 2017). Studies identified included 5 case-control and 8 other observational clinical studies of SM and UM cases. Risk of bias was assessed using the Newcastle-Ottawa scale, and all studies were ranked as 'Good', scoring ≥7/10. Individual-patient data (IPD) were pooled from 13 studies of 3,989 (94.1% aged <15 years) SM patients and 5,780 (79.6% aged <15 years) UM cases in Benin, Malaysia, Mozambique, Tanzania, The Gambia, Uganda, Yemen, and Zambia. Definitions of SM were standardised across studies to compare treatment delay in patients with UM and different SM phenotypes using age-adjusted mixed-effects regression. The odds of any SM phenotype were significantly higher in children with longer delays between initial symptoms and arrival at the health facility (odds ratio [OR] = 1.33, 95% CI: 1.07-1.64 for a delay of >24 hours versus ≤24 hours; p = 0.009). Reported illness duration was a strong predictor of presenting with severe malarial anaemia (SMA) in children, with an OR of 2.79 (95% CI:1.92-4.06; p < 0.001) for a delay of 2-3 days and 5.46 (95% CI: 3.49-8.53; p < 0.001) for a delay of >7 days, compared with receiving treatment within 24 hours from symptom onset. We estimate that 42.8% of childhood SMA cases and 48.5% of adult SMA cases in the study areas would have been averted if all individuals were able to access treatment within the first day of symptom onset, if the association is fully causal. In studies specifically recording onset of nonsevere symptoms, long treatment delay was moderately associated with other SM phenotypes (OR [95% CI] >3 to ≤4 days versus ≤24 hours: cerebral malaria [CM] = 2.42 [1.24-4.72], p = 0.01; respiratory distress syndrome [RDS] = 4.09 [1.70-9.82], p = 0.002). In addition to unmeasured confounding, which is commonly present in observational studies, a key limitation is that many severe cases and deaths occur outside healthcare facilities in endemic countries, where the effect of delayed or no treatment is difficult to quantify. CONCLUSIONS: Our results quantify the relationship between rapid access to treatment and reduced risk of severe disease, which was particularly strong for SMA. There was some evidence to suggest that progression to other severe phenotypes may also be prevented by prompt treatment, though the association was not as strong, which may be explained by potential selection bias, sample size issues, or a difference in underlying pathology. These findings may help assess the impact of interventions that improve access to treatment.

Post-malarial anemia in Mozambican children treated with quinine or artesunate: A retrospective observational study.

OBJECTIVES: This retrospective analysis performed in Manhiça, Southern Mozambique, aimed to describe the frequency of post-malarial anemia (measured as a decrease of hematocrit ≥10%) and the need for blood transfusions in children with severe malaria treated with intravenous quinine or parenteral artesunate. METHODS: All children <15 years admitted with a parasitologically-confirmed diagnosis of malaria from 1st January 2003 to 31st December 2017, alive at hospital discharge, and with at least one measurement of hematocrit within 28 days after hospital discharge, detected by passive case detection, were included. RESULTS: The overall prevalence of post-malarial anemia observed in the study was 23.13%, with an estimated incidence rate of 288.84 episodes/1,000 children-month at risk in the follow-up period (28 days after discharge). There were no differences between treatment groups, although the study showed a higher association between blood transfusions and artesunate treatment. CONCLUSIONS: In this setting, children with severe malaria frequently present a meaningful decrease of hematocrit (>=10%) in the first weeks after their episode, sometimes requiring blood transfusions. Because of the high underlying prevalence of anemia in malaria-endemic settings, all children with severe malaria need to be actively followed up, irrespective of the treatment received.