Targeted newborn metabolomics: prediction of gestational age from cord blood.

OBJECTIVE: Our study sought to determine whether metabolites from a retrospective collection of banked cord blood specimens could accurately estimate gestational age and to validate these findings in cord blood samples from Busia, Uganda. STUDY DESIGN: Forty-seven metabolites were measured by tandem mass spectrometry or enzymatic assays from 942 banked cord blood samples. Multiple linear regression was performed, and the best model was used to predict gestational age, in weeks, for 150 newborns from Busia, Uganda. RESULTS: The model including metabolites and birthweight, predicted the gestational ages within 2 weeks for 76.7% of the Ugandan cohort. Importantly, this model estimated the prevalence of preterm birth <34 weeks closer to the actual prevalence (4.67% and 4.00%, respectively) than a model with only birthweight which overestimates the prevalence by 283%. CONCLUSION: Models that include cord blood metabolites and birth weight appear to offer improvement in gestational age estimation over birth weight alone.

Assessment of Plasmodium antigens and CRP in dried blood spots with multiplex malaria array.

Dried blood spots (DBS) typically prepared on filter papers are an ideal sample type for malaria surveillance by offering easy and cost-effective methods in terms of sample collection, storage, and transport. The objective of this study was to evaluate the applicability of DBS with a commercial multiplex malaria assay, developed to concurrently measure Plasmodium antigens, histidine-rich protein 2 (HRP2), Plasmodium lactate dehydrogenase (pLDH), and a host inflammatory biomarker, C-reactive protein (CRP), in whole blood. The assay conditions were optimized for DBS, and thermal stability for measurement of Plasmodium antigens and CRP in dried blood were determined. Performance of the multiplex assay on matched DBS and whole blood pellet samples was also evaluated using the clinical samples. The results indicate the acceptable performance in multiplex antigen detection using DBS samples. At cutoff levels for DBS, with a diagnostic specificity with a lower 95% confidence bound > 92%, diagnostic sensitivities against polymerase chain reaction (PCR)-confirmed malaria for HRP2, Pf LDH, Pv LDH, and Pan LDH were 93.5%, 80.4%, 21.3%, and 55.6%, respectively. The half-life of pLDH was significantly less than that of HRP2 in thermal stability studies. Results with DBS samples collected from Peru indicate that the uncontrolled storage conditions of DBS can result in inaccurate reporting for infection with P. falciparum parasites with hrp2/3 deletions. With careful consideration that minimizing the unfavorable DBS storage environment is essential for ensuring integrity of heat-labile Plasmodium antigens, DBS samples can be used as an alternative to liquid whole blood to detect P. falciparum with hrp2/3 deletions in malaria surveillance. SUPPLEMENTARY INFORMATION: The online version of this article (10.1007/s12639-020-01325-2) contains supplementary material, which is available to authorized users.

Age-Related Changes in Malaria Clinical Phenotypes During Infancy Are Modified by Sickle Cell Trait.

BACKGROUND: Infants are protected against Plasmodium falciparum malaria. Mechanisms that drive this protection remain unclear due to a poor understanding of malaria clinical phenotypes during infancy. METHODS: We enrolled a birth cohort of 678 infants in Busia, Uganda, an area of high malaria transmission. We followed infants through 12 months of age and quantified protection against parasitemia and clinical disease. RESULTS: Symptomatic malaria incidence increased from 1.2 to 2.6 episodes per person-year between 0 and <6 months and between 6 and 12 months of age, while the monthly probability of asymptomatic parasitemia given infection decreased from 32% to 21%. Sickle cell trait (HbAS) was protective against symptomatic malaria (incidence rate ratio  = 0.57 comparing HbAS vs hemoglobin AA (HbAA); 95% confidence interval, 0.44-0.74; P < .001), but age modified this relationship (Pint = <0.001), with nonlinear protection that waned between 0 and 9 months of age before increasing. Increasing age was associated with higher parasite densities at the time of infection and, in infants with HbAS, a reduced ability to tolerate high parasite densities without fever. CONCLUSIONS: Age-dependent changes in HbAS protective efficacy in infancy were accompanied by differential loss of antiparasite and antidisease protection among HbAS and HbAA infants. This provides a framework for investigating the mechanisms that underlie infant protection against malaria. CLINICAL TRIALS REGISTRATION: NCT02793622.

Multiplex Human Malaria Array: Quantifying Antigens for Malaria Rapid Diagnostics.

Malaria antigen detection through rapid diagnostic tests (RDTs) is widely used to diagnose malaria and estimate prevalence. To support more sensitive next-generation RDT development and screen asymptomatic malaria, we developed and evaluated the Q-Plex™ Human Malaria Array (Quansys Biosciences, Logan, UT), which quantifies the antigens commonly used in RDTs-Plasmodium falciparum-specific histidine-rich protein 2 (HRP2), P. falciparum-specific lactate dehydrogenase (Pf LDH), Plasmodium vivax -specific LDH (Pv LDH), and Pan malaria lactate dehydrogenase (Pan LDH), and human C-reactive protein (CRP), a biomarker of severity in malaria. At threshold levels yielding 99.5% or more diagnostic specificity, diagnostic sensitivities against polymerase chain reaction-confirmed malaria for HRP2, Pf LDH, Pv LDH, and Pan LDH were 92.7%, 71.5%, 46.1%, and 83.8%, respectively. P. falciparum culture strains and samples from Peru indicated that HRP2 and Pf LDH combined improves detection of P. falciparum parasites with hrp2 and hrp3 deletions. This array can be used for antigen-based malaria screening and detecting hrp2/3 deletion mutants of P. falciparum.

Mind the gap: scaling up the utilization of insecticide treated mosquito nets using a knowledge translation model in Isingiro district, rural south western Uganda.

Background: The phenomenon of Knowledge Translation (KT) is a key intervention towards bridging the 'know-do' gap. We conducted a KT initiative in Isingiro district to positively change attitude and improve on the uptake of Insecticide Treated Mosquito Nets (ITNs) as a malaria prevention strategy. Methods: This was a community based interactive initiative that was carried out within the seventeen administrative units of Isingiro district using varied dissemination activities, namely: health talks; drama activities, and the sharing of ITNs success stories. Results: We reached out to 34 dissemination groups, comprising communal gathering, religious crusades, open markets, secondary schools, and district administration. In addition, we spot-visited 46 households to ascertain the physical presence of ITNs, and their appropriate use. The major intervention was improved knowledge base of malaria causation and prevention strategies. The indicators for improved knowledge were hinged on the five-interventions, namely: (a) communal sensitization on malaria to provide, (b) monitoring and support of selected households, (c) emphasis of ITN use as a malaria prevention strategy, (d) promotion of care for ITNs, and (e) promotion of ITN use. In all, the major output was improved knowledge base of malaria causation and prevention strategies by providing accurate information to redress the myths and misconceptions related to malaria and ITNs use. Conclusion: This undertaking describes a consolidated community intervention to promote ITN utilization. It is plausible that this intervention positively enhances and promotes uptake and utilization of ITNs.

Simultaneous Quantification of Plasmodium Antigens and Host Factor C-Reactive Protein in Asymptomatic Individuals with Confirmed Malaria by Use of a Novel Multiplex Immunoassay.

Malaria rapid diagnostic tests (RDTs) primarily detect Plasmodium falciparum antigen histidine-rich protein 2 (HRP2) and the malaria-conserved antigen lactate dehydrogenase (LDH) for P. vivax and other malaria species. The performance of RDTs and their utility is dependent on circulating antigen concentration distributions in infected individuals in a population in which malaria is endemic and on the limit of detection of the RDT for the antigens. A multiplexed immunoassay for the quantification of HRP2, P. vivax LDH, and all-malaria LDH (pan LDH) was developed to accurately measure circulating antigen concentration and antigen distribution in a population with endemic malaria. The assay also measures C-reactive protein (CRP) levels as an indicator of inflammation. Validation was conducted with clinical specimens from 397 asymptomatic donors from Myanmar and Uganda, confirmed by PCR for infection, and from participants in induced blood-stage malaria challenge studies. The assay lower limits of detection for HRP2, pan LDH, P. vivax LDH, and CRP were 0.2 pg/ml, 9.3 pg/ml, 1.5 pg/ml, and 26.6 ng/ml, respectively. At thresholds for HRP2, pan LDH, and P. vivax LDH of 2.3 pg/ml, 47.8 pg/ml, and 75.1 pg/ml, respectively, and a specificity ≥98.5%, the sensitivities for ultrasensitive PCR-confirmed infections were 93.4%, 84.9%, and 48.9%, respectively. Plasmodium LDH (pLDH) concentration, in contrast to that of HRP2, correlated closely with parasite density. CRP levels were moderately higher in P. falciparum infections with confirmed antigenemia versus those in clinical specimens with no antigen. The 4-plex array is a sensitive tool for quantifying diagnostic antigens in malaria infections and supporting the evaluation of new ultrasensitive RDTs.

Knowledge, attitude and behaviour towards the use of insecticide treated mosquito nets among pregnant women and children in rural Southwestern Uganda.

BACKGROUND: The burden of malaria in Uganda remains unacceptably high, especially among children and pregnant women. To prevent malaria related complications, household possession and use of Insecticide Treated mosquito Nets (ITNs) has become a common practice in the country. Despite the availability of ITNs, malaria remains a foremost public health concern in Uganda. We sought to explore knowledge, attitude, and behaviour towards the use of ITNs as a nightly malaria prevention strategy among pregnant women and children under five years of age in Isingiro district, Southwestern Uganda. MATERIALS AND METHODS: This was a community based, descriptive cross-sectional study, in which households with children under 5 years, and/or pregnant women were enrolled. We used a structured questionnaire to collect data on participants' understanding of the causes, signs and symptoms of malaria; use of ITNs to prevent malaria; attitudes and behaviours towards the use of ITNs. We also conducted key informant interviews (KIIs) to get in-depth understanding of responses from the participants. We analysed quantitative data using STATA version 12.Qualitative findings from the KIIs were transcribed and translated, and manually analysed using thematic content analysis. RESULTS: Of the 369 households enrolled, 98.4% (N = 363) households had children under five. Most participants (41.2%, N = 152) were in the 21-30 age category (mean age; 32.2 years). 98.1% (N = 362) of the respondents considered ITNs a key malaria prevention strategy. The ITN possession rate was 84.0% (N = 310), of these, 66.1% (N = 205) consistently used them. 39% of the respondents did not have a positive attitude towards ITNs. CONCLUSIONS: Although 84.0% of the respondents possessed ITNs, many were not consistently using them. To this, there is need to engage all stakeholders (including cultural leaders, community health workers, religious leaders and the government) in the malaria prevention campaigns using ITNs through: a) government's concerted effort to ensure universal access of right fit ITNs, b) end-user directed health education to emphasize positive attributes of ITN use, c) telling the ITN success stories to improve on the usage.

Performance of a High-Sensitivity Rapid Diagnostic Test for Plasmodium falciparum Malaria in Asymptomatic Individuals from Uganda and Myanmar and Naive Human Challenge Infections.

Sensitive field-deployable diagnostic tests can assist malaria programs in achieving elimination. The performance of a new Alere™ Malaria Ag P.f Ultra Sensitive rapid diagnostic test (uRDT) was compared with the currently available SD Bioline Malaria Ag P.f RDT in blood specimens from asymptomatic individuals in Nagongera, Uganda, and in a Karen Village, Myanmar, representative of high- and low-transmission areas, respectively, as well as in pretreatment specimens from study participants from four Plasmodium falciparum-induced blood-stage malaria (IBSM) studies. A quantitative reverse transcription PCR (qRT-PCR) and a highly sensitive enzyme-linked immunosorbent assay (ELISA) test for histidine-rich protein II (HRP2) were used as reference assays. The uRDT showed a greater than 10-fold lower limit of detection for HRP2 compared with the RDT. The sensitivity of the uRDT was 84% and 44% against qRT-PCR in Uganda and Myanmar, respectively, and that of the RDT was 62% and 0% for the same two sites. The specificities of the uRDT were 92% and 99.8% against qRT-PCR for Uganda and Myanmar, respectively, and 99% and 99.8% against the HRP2 reference ELISA. The RDT had specificities of 95% and 100% against qRT-PCR for Uganda and Myanmar, respectively, and 96% and 100% against the HRP2 reference ELISA. The uRDT detected new infections in IBSM study participants 1.5 days sooner than the RDT. The uRDT has the same workflow as currently available RDTs, but improved performance characteristics to identify asymptomatic malaria infections. The uRDT may be a useful tool for malaria elimination strategies.

Sociocultural and structural factors contributing to delays in treatment for children with severe malaria: a qualitative study in southwestern Uganda.

Malaria is a leading cause of pediatric mortality, and Uganda has among the highest incidences in the world. Increased morbidity and mortality are associated with delays to care. This qualitative study sought to characterize barriers to prompt allopathic care for children hospitalized with severe malaria in the endemic region of southwestern Uganda. Minimally structured, qualitative interviews were conducted with guardians of children admitted to a regional hospital with severe malaria. Using an inductive and content analytic approach, transcripts were analyzed to identify and define categories that explain delayed care. These categories represented two broad themes: sociocultural and structural factors. Sociocultural factors were 1) interviewee's distinctions of "traditional" versus "hospital" illnesses, which were mutually exclusive and 2) generational conflict, where deference to one's elders, who recommended traditional medicine, was expected. Structural factors were 1) inadequate distribution of health-care resources, 2) impoverishment limiting escalation of care, and 3) financial impact of illness on household economies. These factors perpetuate a cycle of illness, debt, and poverty consistent with a model of structural violence. Our findings inform a number of potential interventions that could alleviate the burden of this preventable, but often fatal, illness. Such interventions could be beneficial in similarly endemic, low-resource settings.