Malaria Journal

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Online ISSN: 1475-2875
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Article
We read with interest the publication on malaria treatment by Obonyo et al. (Malaria J 21:30, 2022). This commentary questions the methodology, especially the chosen time points of treatment outcome measures.
 
Article
The World Malaria Report, released in December 2021, reflects the unique challenges currently facing the global malaria community. The report showed the devastating toll of malaria, with an estimated 627,000 people losing their lives to the disease in 2020. The improved methodological approach used for calculating cause of death for young children revealed a systematic underestimation of disease burden over the past two decades; and that Africa has an even greater malaria crisis than previously known. While countries were able to prevent the worst-case scenarios, the disruptions due to the COVID-19 pandemic revealed how weak health systems and inadequate financing can limit the capacity of the continent to address the malaria challenge. African countries also face a convergence of biological threats that could redefine malaria control, notably widespread pyrethroid resistance and emerging resistance to artemisinin. Despite these challenges, there is cause for optimism in lessons learned from the COVID-19 pandemic, recent acceleration of cutting edge research and development, and new partnerships that encourage leadership from and ownership by affected countries. This article presents key insights from the 2021 World Malaria Report and reflections on the future trajectories: it was informed by an in-depth discussion with leading malaria experts from the World Health Organization (WHO), the Bill & Melinda Gates Foundation, and the U.S. President’s Malaria Initiative (PMI). The discussion took place during the 34th edition of the Ifakara Master Classes, held virtually on December 15th, 2021.
 
Article
Building on an exercise that identified potential harms from simulated investigational releases of a population suppression gene drive for malaria vector control, a series of online workshops identified nine recommendations to advance future environmental risk assessment of gene drive applications.
 
Study districts in Amhara, Tigray, Oromia and Gambella regions, Ethopia
Article
Background Malaria rapid diagnostic tests (RDTs) have expanded diagnostic service to remote endemic communities in Ethiopia, where 70% of malaria services per annum are reliant on them. However, diagnostic strategies are threatened by Plasmodium falciparum parasites with deletions of the histidine-rich protein 2 and/or 3 ( pfhrp2/3 ) genes. Studies have reported pfhrp2/3 gene deletion prevalence in Ethiopia that exceeds the WHO recommended threshold to switch to non-HRP2 targeted RDTs for detection of P. falciparum . Therefore, RDTs that target alternative antigens, such as P. falcipar um lactate dehydrogenase (PfLDH) are increasingly in programmatic use. Methods Malaria suspected patients visiting health facilities of Amhara, Tigray, Gambella, and Oromia regions of Ethiopia were screened by community health workers using Carestart Pf/Pv ( HRP2/Pv-LDH ) and SD-Bioline Pf ( HRP2 for Pf/LDH for Pf ) RDTs. Dried blood spot (DBS) samples were collected from selected patients for molecular and serological analysis. The clinical data and RDT results were recorded on standard forms, entered into EpiInfo, and analysed using STATA. The Pf-LDH detecting RDT results were compared with real-time PCR and bead-based immunoassay to determine their diagnostic performance. Results The 13,172 (56% male and 44% female, median age of 19 years ranging from 1 to 99 year) study participants were enrolled and tested with PfHRP2 and PfLDH detection RDTs; 20.6% (95% CI: 19.6 to 21.6) were P. falciparum RDT positive. A subset of samples (n = 820) were previously tested using P. falciparum lactate dehydrogenase ( pfldh ) quantitative real-time PCR, and 456 of these further characterized using bead-based immunoassay. The proportion of samples positive for P. falciparum by the PfHRP2 Carestart and SD-Bioline RDTs were 66% (539/820) and 59% (481/820), respectively; 68% (561/820) were positive for the PfLDH band on the SD-Bioline RDT. The sensitivity and specificity of the PfLDH RDT band were 69% and 38%, respectively, versus pfldh qPCR; and 72% and 36%, respectively, versus PfLDH detection by immunoassay. Among samples with results for RDT, qPCR, and immunoassay, higher proportions of P. falciparum were recorded by pfldh qPCR (90%, 411/456) and PfLDH immunoassay (88%, 363/413) compared to the PfLDH band on the SD-Bioline RDT (74.6%, 340/456). Conclusion and recommendation Both PfHRP2 RDTs detected fewer P. falciparum cases than PfLDH, and fewer cases than qPCR or immunoassay. The poor sensitivity and specificity of the PfLDH RDT compared to qPCR and to immunoassay in this study raises concern. Continuous operator training and RDTs quality assurance programme to ensure quality diagnostic services are recommended.
 
Geographical location of the mosquito collection sites in Western Kenya
Distribution of TEP1 genotypes and alleles circulating in An. gambiae and An. arabiensis in Bungoma, Kakamega, Homa Bay, and Kisumu Counties in western Kenya
The evolutionary history was inferred by using the Maximum Likelihood method and Kimura 2-parameter model [1]. The tree with the highest log likelihood (-1872.22) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches. Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances estimated using the Maximum Composite Likelihood (MCL) approach and then selecting the topology with a superior log-likelihood value. A discrete Gamma distribution was used to model evolutionary rate differences among sites (5 categories (+G, parameter = 0.7700). The rate variation model allowed for some sites to be evolutionarily invariable ([+I], 43.64% sites). The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. This analysis involved 15 nucleotide sequences. There were a total of 873 positions in the final dataset. Evolutionary analyses were conducted in MEGA X [2]. Red and green dots indicate haplotypes identified in this study; squares with different colors represent reference haplotypes extracted from GenBank
Article
Abstract Background Evolutionary pressures lead to the selection of efficient malaria vectors either resistant or susceptible to Plasmodium parasites. These forces may favour the introduction of species genotypes that adapt to new breeding habitats, potentially having an impact on malaria transmission. Thioester-containing protein 1 (TEP1) of Anopheles gambiae complex plays an important role in innate immune defenses against parasites. This study aims to characterize the distribution pattern of TEP1 polymorphisms among populations of An. gambiae sensu lato (s.l.) in western Kenya. Methods Anopheles gambiae adult and larvae were collected using pyrethrum spray catches (PSC) and plastic dippers respectively from Homa Bay, Kakamega, Bungoma, and Kisumu counties between 2017 and 2020. Collected adults and larvae reared to the adult stage were morphologically identified and then identified to sibling species by PCR. TEP1 alleles were determined in 627 anopheles mosquitoes using restriction fragment length polymorphisms-polymerase chain reaction (RFLP-PCR) and to validate the TEP1 genotyping results, a representative sample of the alleles was sequenced. Results Two TEP1 alleles (TEP1*S1 and TEP1*R2) and three corresponding genotypes (*S1/S1, *R2/S1, and *R2/R2) were identified. TEP1*S1 and TEP1*R2 with their corresponding genotypes, homozygous *S1/S1 and heterozygous *R2/S1 were widely distributed across all sites with allele frequencies of approximately 80% and 20%, respectively both in Anopheles gambiae and Anopheles arabiensis. There was no significant difference detected among the populations and between the two mosquito species in TEP1 allele frequency and genotype frequency. The overall low levels in population structure (F ST = 0.019) across all sites corresponded to an effective migration index (Nm = 12.571) and low Nei’s genetic distance values (
 
Map of the study site
Types of houses in Makenene. Four types were found consisting of A cement blocks, B Mud and plank, C earth brick and D plank
Home management and episodes of malaria cases in households in Makenene from January to August 2021
Article
Background To contribute to the mission of the National Malaria Control Programme (NMCP) and guide future interventions in Cameroon in general, and in Makenene in particular, this study assessed the knowledge, attitudes and practices of the population of Makenene towards the fight against malaria. Methods Using a semi-structured questionnaire, a descriptive cross-sectional household community survey was carried out in randomly selected households in Makenene, a locality situated between forest and savannah ecotypes. Results Out of the 413 households surveyed, all (100%) claimed to have heard of malaria with over 94% (n = 391) associating disease transmission with mosquito bites. The main mosquito control tools used in the area were mosquito nets (92.25%). The majority of participants had good knowledge (55.93%; n = 231), good practices (71.67%, n = 296) but moderate attitudes (47.94%; n = 198) towards malaria control and fight. Good knowledge and practices were recorded mostly in educated persons including public servants and students. Good attitudes were adopted mostly by public servants and students of secondary and higher levels of education. Conclusion In Makenene, the population exhibits good knowledge and practices towards malaria and its control. However, despite high LLINs ownership and use, people still complain about malaria in the area. Control tools should be monitored, repaired or replaced when necessary to support the achievement of the NMCP mission.
 
Parasitaemia and PfHRP2 concentration over study period for three anti-malarial treatments in IBSM studies. Relationship between mean parasitaemia (Grey solid line) and mean PfHRP2 concentration (Black dashed line) over the study period used for model development for each of the three anti-malarial treatments in the IBSM study. Bars at each time-point represent minimum and maximum
PfHRP2 concentration over time post administration of anti-malarial treatment for estimation of PfHRP2 elimination half-life. PfHRP2 concentration over time post administration of anti-malarial drug for the 10 IBSM individuals with maximum PfHRP2 concentrations exceeding 300 pg/mL that were used to estimate the PfHRP2 elimination half-life. The red dashed line is the reported limit of detection (LOD) for Q-plex ELISA 2-Plex (LOD 5 pg/mL)
Example fits of the Base Model and the Final Model for two IBSM individuals. The observed parasitaemia over the course of infection is represented by black solid line, observed PfHRP2 concentration is represented by circles (pre-treatment in solid circles and post-treatment in open circles), and the predicted minimum and maximum PfHRP2 concentration from the model is shown as blue and red dashed lines, respectively
Fits of the final model with elimination half-life of 4.5 days to the Namibia study. Fits of the final PfHRP2 model with elimination half-life of 4.5 days to the 6 individuals from the Namibia study, with observed PfHRP2 concentration represented by closed circles, and minimum and maximum predicted PfHRP2 concentration represented by the blue and red dashed lines, respectively. The dashed grey horizontal line represents the threshold of 800 pg/mL and the solid grey horizontal line represents the threshold of 80 pg/mL which correspond to the positivity threshold of an RDT and usRDT, respectively
Article
Background Rapid diagnostic tests (RDTs) that rely on the detection of Plasmodium falciparum histidine-rich protein 2 ( Pf HRP2) have become key tools for diagnosing P. falciparum infection. The utility of RDTs can be limited by Pf HRP2 persistence, however it can be a potential benefit in low transmission settings where detection of persistent Pf HRP2 using newer ultra-sensitive Pf HRP2 based RDTs can serve as a surveillance tool to identify recent exposure. Better understanding of the dynamics of Pf HRP2 over the course of a malaria infection can inform optimal use of RDTs. Methods A previously published mathematical model was refined to mimic the production and decay of Pf HRP2 during a malaria infection. Data from 15 individuals from volunteer infection studies were used to update the original model and estimate key model parameters. The refined model was applied to a cohort of patients from Namibia who received treatment for clinical malaria infection for whom longitudinal Pf HRP2 concentrations were measured. Results The refinement of the Pf HRP2 dynamic model indicated that in malaria naïve hosts, P. falciparum parasites of the 3D7 strain produce 33.6 × 10 ⁻¹⁵ g (95% CI 25.0–42.1 × 10 ⁻¹⁵ g) of Pf HRP2 in vivo per parasite replication cycle, with an elimination half-life of 1.67 days (95% CI 1.11–3.40 days). The refined model included these updated parameters and incorporated individualized body fluid volume calculations, which improved predictive accuracy when compared to the original model. The performance of the model in predicting clearance of Pf HRP2 post treatment in clinical samples from six adults with P. falciparum infection in Namibia improved when using a longer elimination half-life of 4.5 days, with 14% to 67% of observations for each individual within the predicted range. Conclusions The updated mathematical model can predict the growth and clearance of Pf HRP2 during the production and decay of a mono-infection with P. falciparum , increasing the understanding of Pf HRP2 antigen dynamics. This model can guide the optimal use of Pf HRP2-based RDTs for reliable diagnosis of P. falciparum infection and re-infection in endemic settings, but also for malaria surveillance and elimination programmes in low transmission areas.
 
The annual malaria incidence in Sri Lanka 1962–2012 showing two very similar epidemic cycles in which the declining arms (area shaded in gray) were shown to be dynamically similar. Financial expenditure was compared in the 9 and 8 years (range of years shaded in yellow) which followed two pivotal years in this period (marked by arrows), in which the incidence was similar
Malaria expenditure and the number of cases during the periods of malaria control (1980–1989) and malaria elimination (2002–2010)
Expenditure per case of malaria in the two periods of malaria control (left) and elimination (right). Note that the vertical axes scales are different for the two time periods
AMC expenditure (proportions) on budget categories in 1989 (during malaria control) and 2010 (during malaria elimination)
Article
Background Malaria was endemic in Sri Lanka for centuries and was eliminated in 2012. It is widely assumed that the costs of elimination are generally greater than that of control. The costs of malaria elimination in Sri Lanka with that of malaria control in the past using periods in which starting transmission dynamics were similar were compared. Methods The expenditure of the Anti-Malaria Campaign (AMC), total and by budget category, during 2002–2010 is compared with that of malaria control during the period 1980–1989, using regression analyses and the Mann Whitney U statistic. Results The expenditure on malaria control and malaria elimination was similar ranging from 21 to 45 million USD per year when adjusted for inflation. In both periods, external funding for the malaria progamme constituted around 24% of the total budget; during the control phase in the 1980s, external funds came from bilateral agencies and were disbursed in accordance with government budget guidelines. In the elimination phase in the 2000s, most of external funding was from the Global Fund and had flexibility of disbursement. In the 1980s, most funds were expended on commodities—insecticides, diagnostics and medicines and their delivery; in the elimination phase, they were spent on programme management, human resources, technical assistance and monitoring and evaluation; monitoring and evaluation was not a budget line in the 1980s. Although the cost per case of malaria was considerably higher during the elimination phase than in the control phase, expenditure was not on individual cases but on general systems strengthening. Conclusion Malaria elimination in Southeast Asia may not require more funding than malaria control. But sustained funding for an agile programme with flexibility in fund utilization and improved efficiencies in programme management with stringent monitoring and evaluation appears to be critically important.
 
Article
Background Data integration and visualisation techniques have been widely used in scientific research to allow the exploitation of large volumes of data and support highly complex or long-lasting research questions. Integration allows data from different sources to be aggregated into a single database comprising variables of interest for different types of studies. Visualisation allows large and complex data sets to be manipulated and interpreted in a more intuitive way. Methods Integration and visualisation techniques were applied in a malaria surveillance ecosystem to build an integrated database comprising notifications, deaths, vector control and climate data. This database is accessed through Malaria-VisAnalytics, a visual mining platform for descriptive and predictive analysis supporting decision and policy-making by governmental and health agents. Results Experimental and validation results have proved that the visual exploration and interaction mechanisms allow effective surveillance for rapid action in suspected outbreaks, as well as support a set of different research questions over integrated malaria electronic health records. Conclusion The integrated database and the visual mining platform (Malaria-VisAnalytics) allow different types of users to explore malaria-related data in a user-friendly interface. Summary data and key insights can be obtained through different techniques and dimensions. The case study on Manaus can serve as a reference for future replication in other municipalities. Finally, both the database and the visual mining platform can be extended with new data sources and functionalities to accommodate more complex scenarios (such as real-time data capture and analysis).
 
Map showing the geographical distribution of study sites in East Shewa, Ethiopia
Distribution (violin plot) of P. vivax asexual parasitemia by Duffy genotypes detected in patients in East Shewa, Ethiopia
Article
Abstract Background The increase in detections of Plasmodium vivax infection in Duffy-negative individuals in Africa has challenged the dogma establishing the unique P. vivax Duffy Binding Protein-Duffy antigen receptor for chemokines (PvDBP-DARC) pathway used by P. vivax merozoites to invade reticulocytes. Information on the impact of Duffy antigen polymorphisms on the epidemiology of P. vivax malaria remains elusive. The objective of this study was to determine the distribution of asexual parasitaemia of P. vivax according to the Duffy antigen polymorphisms in Ethiopia. Methods DNA was extracted from dried blood spots (DBS) collected from prospectively recruited 138 P. vivax-infected patients from health centres. The identification and estimation of P. vivax asexual parasitaemia were performed by microscopic examination and quantitative real-time polymerase chain reaction (PCR). Duffy genotyping was conducted by DNA sequencing in a total of 138 P.vivax infected samples. Results The proportion of Duffy-negatives (FY*BES/FY*BES) in P. vivax infected patients was 2.9% (4/138). Duffy genotype FY*B/FY*BES (48.6%) was the most common, followed by FY*A/FY*BES genotype (25.4%). In one patient, the FY*02 W.01/FY*02 N.01 genotype conferring a weak expression of the Fyb antigen was observed. All P.vivax infected Duffy-negative patients showed low asexual parasitaemia (≤ 110 parasites/µL). The median P. vivax parasitaemia in Duffy-negative patients (53 parasites/µL) was significantly lower than those found in homozygous and heterozygous individuals (P
 
Article
Background The automated haematology analyzer XN-31 prototype (XN-31p) is a new flow cytometry-based device developed to measure the number and the ratio of malaria-infected red blood cells (MI-RBC) with a complete blood count (CBC). The XN-31p can provide results in about one minute and also can simultaneously provide information on the malaria parasite ( Plasmodium ) species. In this study, clinical testing of the XN-31p was performed using blood samples from patients with imported malaria in Japan. Methods Blood samples were collected from 80 patients who visited the hospital of the National Center for Global Health and Medicine, Tokyo, Japan, for malaria diagnosis from January 2017 to January 2019. The test results by the XN-31p were compared with those by other standard methods, such as microscopic observation, rapid diagnostic tests and the nested PCR. Results Thirty-three patients were diagnosed by the nested PCR as being malaria positive (28 Plasmodium falciparum , 2 Plasmodium vivax , 1 Plasmodium knowlesi , 1 mixed infection of P. falciparum and Plasmodium malariae , and 1 mixed infection of P. falciparum and Plasmodium ovale ), and the other 47 were negative. The XN-31p detected 32 patients as “MI-RBC positive”, which almost matched the results by the nested PCR and, in fact, completely matched with the microscopic observations. The ratio of RBCs infected with malaria parasites as determined by the XN-31p showed a high correlation coefficient of more than 0.99 with the parasitaemia counted under microscopic observation. The XN-31p can analyse the size and nucleic acid contents of each cell, and the results were visualized on a two-dimensional cytogram termed the “M scattergram”. Information on species and developmental stages of the parasites could also be predicted from the patterns visualized in the M scattergrams. The XN-31p showed a positive coincidence rate of 0.848 with the nested PCR in discriminating P. falciparum from the other species. Conclusions The XN-31p could rapidly provide instructive information on the ratio of MI-RBC and the infecting Plasmodium species. It was regarded to be of great help for the clinical diagnosis of malaria.
 
Article
Background The use of long-lasting insecticide-treated nets (LLINs) is one of the main malaria prevention method promoted by the World Health Organization (WHO) in Côte d'Ivoire. LLIN-coverage has reached 95% since 2015 and nearly 16 million LLINs were distributed in 2017. Despite these efforts, malaria incidence at the national level remains high (120‰ in 2012 to 164‰ in 2017) although this could be partly explained by increased screening efforts. This study aimed at determining what preventative measures were used against mosquito bites, as well as LLIN maintenance practices used by the inhabitants of the city of Bouaké, capital city of the Gbêkê region with a malaria incidence of 257‰ in 2017. Methods A descriptive qualitative investigation took place in Bouaké, in four neighbourhoods that were selected through purposive sampling based on their social composition. Data were collected using an interview guide based on convenience sampling. Results The results of the study reveal that LLINs are the most reported used malaria prevention measure (66.4%). Environmental health (28.8%) came second in their declarations, smoke coils (23.5%) third and aerosol cans (18.8%) last. The percentage of respondents who answered that they had slept under an LLIN the previous night was 53%. 57.7% reported that they wash their LLINs, 12.1% that they do not wash them, and 4% that they replace dirty LLINs with new ones. The LLINs washing methods described by the respondents did not comply with the WHO recommendations and there was no mention of LLINs repairs. Conclusion Despite mass distributions of LLINs in Côte d'Ivoire, this key malaria control tool remains under-used by the population. Regarding LLIN maintenance, more than half of the population reports that they wash their nets while not complying with recommended practices or repairing them.
 
Article
Background Gene drives are a genetic engineering method where a suite of genes is inherited at higher than Mendelian rates and has been proposed as a promising new vector control strategy to reinvigorate the fight against malaria in sub-Saharan Africa. Methods Using an agent-based model of malaria transmission with vector genetics, the impacts of releasing population-replacement gene drive mosquitoes on malaria transmission are examined and the population replacement gene drive system parameters required to achieve local elimination within a spatially-resolved, seasonal Sahelian setting are quantified. The performance of two different gene drive systems—“classic” and “integral”—are evaluated. Various transmission regimes (low, moderate, and high—corresponding to annual entomological inoculation rates of 10, 30, and 80 infectious bites per person) and other simultaneous interventions, including deployment of insecticide-treated nets (ITNs) and passive healthcare-seeking, are also simulated. Results Local elimination probabilities decreased with pre-existing population target site resistance frequency, increased with transmission-blocking effectiveness of the introduced antiparasitic gene and drive efficiency, and were context dependent with respect to fitness costs associated with the introduced gene. Of the four parameters, transmission-blocking effectiveness may be the most important to focus on for improvements to future gene drive strains because a single release of classic gene drive mosquitoes is likely to locally eliminate malaria in low to moderate transmission settings only when transmission-blocking effectiveness is very high (above ~ 80–90%). However, simultaneously deploying ITNs and releasing integral rather than classic gene drive mosquitoes significantly boosts elimination probabilities, such that elimination remains highly likely in low to moderate transmission regimes down to transmission-blocking effectiveness values as low as ~ 50% and in high transmission regimes with transmission-blocking effectiveness values above ~ 80–90%. Conclusion A single release of currently achievable population replacement gene drive mosquitoes, in combination with traditional forms of vector control, can likely locally eliminate malaria in low to moderate transmission regimes within the Sahel. In a high transmission regime, higher levels of transmission-blocking effectiveness than are currently available may be required.
 
Geographical situation of Ankazomborona and Matanga (Source: BD 500 and FTM)
Fragment sampling of RDT strip. A Distal part, B central part, C proximal part, D all parts
PCR product from DNA extracted by the two methods: Instagena matrix (1 to 8) and simple elution in water (9 to 16). Expected size 206 bp for P. falciparum (A) and 120 bp for P. vivax (B), PM: 100 bp DNA ladder marker, 1–16: samples
Polymorphism of P. falciparumglurp gene
Article
Background Rapid diagnostic tests (RDT) are widely used for malaria diagnosis in Madagascar, where Plasmodium falciparum is the predominant species. Molecular diagnosis is essential for malaria surveillance, but requires additional blood samples for DNA extraction. Used RDTs is an attractive alternative that can be used as a source of DNA. Plasmodium falciparum genetic diversity and multiplicity of infection, usually determined by the genotyping of polymorphic regions of merozoite surface proteins 1 and 2 genes (msp1, msp2), and the repeated region RII of the glutamate-rich protein gene (glurp) have been associated with malaria transmission levels and subsequently with the impact of the deployed control strategies. Thus, the study aims to use RDT as DNA source to detect Plasmodium species, to characterize Plasmodium falciparum genetic diversity and determine the multiplicity of infection. Methods A pilot study was conducted in two sites with different epidemiological patterns: Ankazomborona (low transmission area) and Matanga (high transmission area). On May 2018, used RDT (SD BIOLINE Malaria Ag P.f/Pan, 05FK63) were collected as DNA source. Plasmodium DNA was extracted by simple elution with nuclease free water. Nested-PCR were performed to confirm Plasmodium species and to analyse P. falciparum msp1, msp2 and glurp genes polymorphisms. Results Amongst the 170 obtained samples (N = 74 from Ankazomborona and N = 96 from Matanga), Plasmodium positivity rate was 23.5% (40/170) [95% CI 17.5–30.8%] by nested-PCR with 92.2% (37/40) positive to P. falciparum, 5% (2/40) to Plasmodium vivax and 2.5% (1/40) to P. falciparum/P. vivax mixed infection. Results showed high polymorphisms in P. falciparum msp1, msp2 and glurp genes. Multiple infection rate was 28.6% [95% CI 12.2–52.3%]. The mean of MOI was 1.79 ± 0.74. Conclusion This pilot study highlighted that malaria diagnosis and molecular analysis are possible by using used malaria RDT. A large-scale study needs to be conducted to assess more comprehensively malaria parasites transmission levels and provide new data for guiding the implementation of local strategies for malaria control and elimination. Trial registration Retrospectively registered
 
Ethical principles informing Target Malaria's engagement strategy [32]
The evolution of Target Malaria’s engagement strategy: an iterative process
Definitions
Article
Gene drive mosquitoes are increasingly considered a potential transformational tool for vector control of malaria mosquitoes. As part of efforts to promote responsible research in this field, a number of guidance documents have been published by the World Health Organization, National Academies and expert groups. While virtually all recent guidance documents on gene drive research stress the importance of stakeholder engagement activities, no specific guidelines on implementing them have been established. Target Malaria, a not-for-profit research consortium developing a vector-control gene drive approach to eliminate malaria, has reflected on how its stakeholder engagement strategy translates engagement guidance documents into practice. The project analysed and addressed the tension between the context specificities and the international recommendations. The engagement strategy combines published recommendations for responsible gene drive research, information collected from the local context where the project operates and a set of principles guiding the choices made. This strategy was first developed during the early phases of the project’s research, years ahead of any activities with gene drive mosquitoes in those countries of operations. These earlier activities, and their related engagement, allow the project to develop and adapt an engagement strategy appropriate for potential gene drive research in its field site countries. This paper offers a description of a stakeholder engagement strategy operationalization based on (1) adaptation to stakeholder preferences, (2) inclusiveness and (3) empowerment and accountability. The authors hope to offer concrete examples to support other projects with the development and implementation of their engagement strategies with particular attention to the co-development principle.
 
RDT stock-outs and monthly rainfall in Kondoa over a 12-month inventory period
Article
Background Despite the large-scale rollout of malaria rapid diagnostic tests (RDTs) in Tanzania, many healthcare providers (HCPs) continue using blood film microscopy (BFM) and clinical examination to diagnose malaria, which can increase the risk of mal-diagnosis and over-prescribing of anti-malarials. Patients disregarding negative test results and self-treating exacerbate the problem. This study explored the knowledge, attitudes and practices of HCPs and healthcare-seekers regarding RDTs in comparison to BFM testing. Methods A situational analysis was, therefore, conducted in Kondoa District, Dodoma Region, Tanzania. A multi-methodological approach was adopted including (i) a health facility inventory and screening of logbooks from May 2013 to April 2014 with 77,126 patient entries from 33 health facilities; (ii) a survey of 40 HCPs offering malaria services; and iii) a survey of 309 randomly selected household members from the facilities’ catchment area. Surveys took place in April and May 2014. Results Health facility records revealed that out of 77,126 patient entries, 22% (n = 17,235) obtained a malaria diagnosis. Of those, 45% were made with BFM, 33% with RDT and 22% with clinical diagnosis. A higher rate of positive diagnoses was observed with BFM compared with RDT (71% vs 14%). In the HCP survey, 48% preferred using BFM for malaria testing, while 52% preferred RDT. Faced with a negative RDT result for a patient presenting with symptoms typical for malaria, 25% of HCPs stated they would confirm the result with a microscopy test, 70% would advise or perform a clinical diagnosis and 18% would prescribe anti-malarials. Interviews with household members revealed a preference for microscopy testing (58%) over RDT (23%), if presented with malaria symptoms. For participants familiar with both tests, a second opinion was desired in 45% after a negative microscopy result and in 90% after an RDT. Conclusions Non-adherence to negative diagnostics by HCPs and patients continues to be a concern. Frequent training and supportive supervision for HCPs diagnosing and treating malaria and non-malaria febrile illnesses is essential to offer quality services that can instil confidence in HCPs and patients alike. The introduction of new diagnostic devices should be paired with context-specific behaviour change interventions targeting healthcare-seekers and healthcare providers.
 
Article
Effective control of infectious diseases is facilitated by informed decisions that require accurate and timely diagnosis of disease. For malaria, improved access to malaria diagnostics has revolutionized malaria control and elimination programmes. However, for COVID-19, diagnosis currently remains largely centralized and puts many low- and middle-income countries (LMICs) at a disadvantage. Malaria and COVID-19 are infectious diseases that share overlapping symptoms. While the strategic responses to disease control for malaria and COVID-19 are dependent on the disease ecologies of each disease, the fundamental need for accurate and timely testing remains paramount to inform accurate responses. This review highlights how the roll-out of rapid diagnostic tests has been fundamental in the fight against malaria, primarily within the Asia Pacific and along the Greater Mekong Subregion. By learning from the successful elements of malaria control programmes, it is clear that improving access to point-of-care testing strategies for COVID-19 will provide a suitable framework for COVID-19 diagnosis in not only the Asia Pacific, but all malarious countries. In malaria-endemic countries, an integrated approach to point-of-care testing for COVID-19 and malaria would provide bi-directional benefits for COVID-19 and malaria control, particularly due to their paralleled likeness of symptoms, infection control strategies and at-risk individuals. This is especially important, as previous disease pandemics have disrupted malaria control infrastructure, resulting in malaria re-emergence and halting elimination progress. Understanding and combining strategies may help to both limit disruptions to malaria control and support COVID-19 control.
 
Article
Background Thailand’s strong malaria elimination programme relies on effective implementation of its 1-3-7 surveillance strategy, which was endorsed and implemented nationwide in 2016. For each confirmed malaria patient, the Ministry of Public Health’s Division of Vector Borne Diseases (DVBD) ensures completion of case notification within 1 day, case investigation within 3 days, and foci investigation within 7 days. To date, there has not been a comprehensive assessment of the performance and achievements of the 1-3-7 surveillance strategy although such results could help Thailand’s future malaria elimination strategic planning. Methods This study examined adherence to the 1-3-7 protocols, tracked progress against set targets, and examined geographic variations in implementation of the 1-3-7 strategy in the programme’s initial 5 years. An auto-regressive integrated moving average (ARIMA) time series analysis with seasonal decomposition assessed the plausible implementation effect of the 1-3-7 strategy on malaria incidence in the programme’s initial 5 years. The quantitative analysis included all confirmed malaria cases from public health and non-governmental community facilities from October 2014 to September 2021 (fiscal year [FY] 2015 to FY 2021) (n = 77,405). The spatial analysis included active foci with known geocoordinates that reported more than five cases from FY 2018 to FY 2021. Results From FY 2017 to FY 2021, on-time case notification improved from 24.4% to 89.3%, case investigations from 58.0% to 96.5%, and foci investigations from 37.9% to 87.2%. Adherence to timeliness protocols did not show statistically significant variation by area risk classification. However, adherence to 1-3-7 protocols showed a marked spatial heterogeneity among active foci, and the ARIMA model showed a statistically significant acceleration in the reduction of malaria incidence. The 1-3-7 strategy national indicators and targets in Thailand have shown progressive success, and most targets were achieved for FY 2021. Conclusion The results of Thailand’s 1-3-7 surveillance strategy are associated with a decreased incidence in the period following the adoption of the strategy although there is notable geographic variation. The DVBD will continue to implement and adapt the 1-3-7 strategy to accelerate progress toward malaria elimination. This assessment may be useful for domestic strategic planning and to other countries considering more intensive case and foci investigation and response strategies.
 
Relationship between saliva and plasma levels of angiopoietin-1, angiopoietin-2 and CXCL10 among malaria patients. A Correlation between saliva and plasma levels of CXCL10 among malaria patients. B Correlation between saliva and plasma levels of angiopoietin-1 among malaria patients. C Correlation between saliva and plasma levels of angiopoietin-2 among malaria patients
Saliva and plasma levels of angiopoietin-1 protein among the study participants. A Association of saliva angiopoietin-1 protein levels with malaria infection. Malaria patients have significantly lower saliva angiopoietin-1 levels than non-malaria subjects (p = 0.009). B Association of plasma angiopoietin-1 protein levels with malaria infection. Malaria patients have significantly lower plasma angiopoietin-1 levels than non- malaria subjects (p < 0.001). Box plot represent medians with 25th and 75th percentiles, bars for 10th and 90th percentiles. Significant differences of angiopoietin-1 median levels between the groups were determined by Mann–Whitney U test or Kruskal Wallis tests
Saliva and plasma levels of angiopoietin-2 protein among the study participants. A Association of saliva angiopoietin-2 protein levels with malaria infection. Malaria patients have significantly higher saliva angiopoietin-2 levels than non-malaria subjects (p = 0.001). B Association of plasma angiopoietin-2 protein levels with malaria infection. Malaria patients have significantly higher plasma angiopoietin-2 levels than non-malaria subjects (p < 0.001). Box plot represent medians with 25th and 75th percentiles, bars for 10th and 90th percentiles. Significant differences of angiopoietin-2 median levels between the groups were determined by Mann–Whitney U test or Kruskal Wallis tests
Saliva and plasma levels of CXCL10 protein among the study participants. A Association of saliva CXCL10 protein levels with malaria infection. Malaria patients have significantly higher saliva CXCL10 levels than non-malaria subjects (p = 0.004). B Association of plasma CXCL10 protein levels with malaria infection. Malaria patients have significantly higher plasma CXCL10 levels than non-malaria subjects (p < 0.001). Box plot represent medians with 25th and 75th percentiles, bars for 10th and 90th percentiles. Significant differences of CXCL10 median levels between the groups were determined by Mann–Whitney U test or Kruskal Wallis tests
Article
Background: Malaria related mortality is associated with significant deregulation of host inflammatory factors such as interferon-inducible protein 10, a member of the CXC or α-subfamily (CXCL10), and host angiogenic factors such as angiopoietin 1 (Ang-1) and angiopoietin 2 (Ang-2). However, detection of these factors in malaria patients requires the drawing of blood, which is invasive and increases the risk of accidental blood-borne infections. There has been an increased interest in the use of saliva as the body fluid of choice for the diagnosis of many infectious diseases including malaria. Here, saliva levels of CXCL10, Ang-1, and Ang-2 previously shown to be predictive of severe malaria in malaria patients in Ghana were assessed in malaria patients. Methods: This study was conducted in the Shai-Osudoku District Hospital in Dodowa, Accra, Ghana and the study population comprised 119 malaria patients and 94 non-malaria subjects. The non-malaria subjects are healthy community participants with no malaria infection. Plasma and saliva levels of CXCL10, Ang-1 and Ang-2 of the study participants were measured using an enzyme-linked immunoassay. Complete blood counts of each participant were measured with a haematology autoanalyzer. Pearson correlation was used to evaluate the correlation between plasma and saliva levels of each biomarker in malaria patients. A p-value of < 0.05 was considered significant. Box plots of median biomarker concentrations were plotted. SPSS version 14.2 software was used for statistical analysis. Results: The non-malaria subjects had a median age of 29 years compared to 23 years for malaria patients (p = 0.001). Among the malaria patients, there was a strong significant relationship between CXCL10 (R2 = 0.7, p < 0.0001) and Ang-1 (R2 = 0.7, p < 0.0001). Malaria patients had lower saliva levels of Ang-1 (p = 0.009) and higher saliva levels of CXCL10 (p = 0.004) and Ang-2 (p = 0.001) compared to non-malaria subjects. Conclusions: This study provides the first evidence of elevated levels of CXCL10 and Ang-2 in the saliva of malaria patients. Detection of CXCL10, Ang-1 and Ang-2 in saliva may have a potential application for non-invasive malaria diagnosis.
 
Article
Background Many Plasmodium infections in endemic regions exist at densities below the limit of detection of standard diagnostic tools. These infections threaten control efforts and may impact vaccine and therapeutic drug studies. Simple, cost-effective methods are needed to study the natural history of asymptomatic submicroscopic parasitaemia. Self-collected dried blood spots (DBS) analysed using pooled and individual quantitative reverse transcription polymerase chain reaction (qRT-PCR) provide such a solution. Here, the feasibility and acceptability of daily at-home DBS collections for qRT-PCR was studied to better understand low-density infections. Methods Rapid diagnostic test (RDT)-negative individuals in Katakwi District, northeastern Uganda, were recruited between April and May 2021. Venous blood samples and clinic-collected DBS were taken at enrollment and at four weekly clinic visits. Participants were trained in DBS collection and asked to collect six DBS weekly between clinic visits. Opinions about the collection process were solicited using daily Diary Cards and a Likert scale survey at the final study visit. Venous blood and DBS were analysed by Plasmodium 18S rRNA qRT-PCR. The number of participants completing the study, total DBS collected, and opinions of the process were analysed to determine compliance and acceptability. The human internal control mRNA and Plasmodium 18S rRNA were evaluated for at-home vs. clinic-collected DBS and venous blood to assess quality and accuracy of at-home collected samples. Results One-hundred two adults and 29 children were enrolled, and 95 and 26 completed the study, respectively. Three individuals withdrew due to pain or inconvenience of procedures. Overall, 96% of participants collected ≥ 16 of 24 at-home DBS, and 87% of DBS contained ≥ 40 µL of blood. The procedure was well tolerated and viewed favourably by participants. At-home collected DBS were acceptable for qRT-PCR and showed less than a one qRT-PCR cycle threshold shift in the human control mRNA compared to clinic-collected DBS. Correlation between Plasmodium falciparum 18S rRNA from paired whole blood and DBS was high (R = 0.93). Conclusions At-home DBS collection is a feasible, acceptable, and robust method to obtain blood to evaluate the natural history of low-density Plasmodium infections by qRT-PCR.
 
The schematic diagram illustrating the experimental design of the study
Chemical structure of the major constituents of Pelargonium roseum (citronellol, geraniol, linalool, L-menthone, and citronellyl formate) and Juniperus virginiana (sabinene, β-myrcene, dl-limonene, terpinen-4-ol, and bornyl acetate) essential oils along with their total ion chromatograms of GC-MS analysis
Probit regression line of larvae of Anopheles gambiae s.s. exposed to different concentrations of JVO, PRO, and the main components of PRO essential oils in laboratory and semi-field environments at times of 24–72 h
Adult percentage mortality post exposure in different oils (Juniperus virginiana, Pelargonium roseum, and the main constituents of P. roseum) for laboratory-reared and field populations of Anopheles gambiae complex. PER permethrin, JVO Juniperus virginiana, PRO Pelargonium roseum, CO citronellol, GO geraniol, LO linalool, MO L-menthone MIX mixture of all four ingredients
Article
Background More than 90% of malaria cases occur in Africa where the disease is transmitted by Anopheles gambiae and Anopheles arabiensis . This study evaluated the anti-mosquito properties of Juniperus virginiana (JVO) and Pelargonium roseum (PRO) essential oils (EOs) against larvae and adults of An. gambiae sensu lato (s.l. ) from East Africa in laboratory and semi-field conditions. Methods EOs was extracted from the aerial green parts of Asian herbs by hydrodistillation. Their constituents were characterized by gas chromatography-mass spectrometry (GC-MS). Larvicidal activities of JVO, PRO, and PRO components [citronellol (CO), linalool (LO), and geraniol (GO)] were investigated against An. gambiae sensu stricto ( s.s .). The percentage of knockdown effects and mortality rates of all oils were also evaluated in the adults of susceptible An. gambiae s.s. and permethrin-resistant An. arabiensis . Results GC-MS analyses identified major constituents of JVO (sabinene, dl-limonene, β-myrcene, bornyl acetate, and terpinen-4-ol) and PRO (citronellol, citronellyl formate, L-menthone, linalool, and geraniol). Oils showed higher larvicidal activity in the laboratory than semi-field trials. The LC 50 values for JVO/PRO were computed as 10.82–2.89/7.13–0.9 ppm and 10.75–9.06/13.63–8.98 ppm in laboratory and semi-field environments, respectively at exposure time of 24–72 h. The percentage of knockdown effects of the oils were also greater in An. gambiae s.s. than in An. arabiensis . Filter papers impregnated with JVO (100 ppm) and PRO (25 ppm) displayed 100% mortality rates for An. gambiae s.s. and 3.75% and 90% mortality rates, for An. arabiensis populations, respectively. Each component of CO, LO, and GO exhibited 98.13%, 97.81%, and 87.5%, respectively, and a mixture of the PRO components indicated 94.69% adult mortality to permethrin-resistant An. arabiensis . Conclusions The findings of this study show that PRO and its main constituents, compared to JVO, have higher anti-mosquito properties in terms of larvicidal, knockdown, and mortality when applied against susceptible laboratory and resistant wild populations of An. gambiae s.l. Consequently, these oils have the potential for the development of new, efficient, safe, and affordable agents for mosquito control.
 
A Location of Bandarban District in Bangladesh. B Villages sampled for this study (24 of 28). Villages in which Plasmodium spp. samples were detected are labelled by colour. Villages in which no infections were detected are marked with a (+). Not pictured are Faruk Para, Hebron Para, Hati Bhanga Para, Paschim Antaha Para, and Purbo Antaha Para for which GPS location was not recorded. C Distribution of Plasmodium spp. infections detected by qPCR. Plasmodium falciparum and P. vivax were both found throughout the study area. D The majority of infections were clustered among the Khumi and Mro
Structure plots showed no structure among P. falciparum or P. vivax populations. Plots were generated using k = 2–5, and all plots looked very similar
Relatedness networks based on pairwise IBS  ≥ 0.5. In the top row, samples are colored according to ethnicity; in the bottom row, samples are colored by village. Among P. falciparum there is a small network of related infections from the Khumi and Mro, plus one infection from the Tripura. Among P. vivax there are two smaller networks of related infections from the Mro plus one infection from the Tripura. There are very few linkages for either parasite species
Principal components analysis based on principal components 1 and 2. Plasmodium vivax haplotypes showed some separation by ethnicity, with the Khumi and Mro forming loose clusters. P. falciparum samples showed very little separation, with samples appearing closely around the origin
Age distribution of participants with Plasmodium spp. Infections
Article
Background Malaria remains endemic in Bangladesh, with the majority of cases occurring in forested, mountainous region in the Chittagong Hill Tracts (CHT). This area is home to Bengali and diverse groups of indigenous people (Pahari) residing largely in mono-ethnic villages. Methods 1002 individuals of the 9 most prominent Pahari and the Bengali population were randomly selected and screened by RDT and qPCR. Parasites were genotyped by msp2 and deep sequencing of 5 amplicons ( ama1-D3, cpmp, cpp, csp , and msp7 ) for Plasmodium falciparum (n = 20), and by microsatellite (MS) typing of ten loci and amplicon sequencing of msp1 for Plasmodium vivax (n = 21). Population structure was analysed using STRUCTURE software. Identity-by-state (IBS) was calculated as a measure of parasite relatedness and used to generate relatedness networks. Results The prevalence of P. falciparum and P. vivax infection was 0.7% by RDT ( P. falciparum 6/1002; P. vivax 0/1002, mixed: 1/1002) and 4% by qPCR ( P. falciparum 21/1002; P. vivax 16/1002, mixed: 5/1002). Infections were highly clustered, with 64% (27/42) of infections occurring in only two Pahari groups, the Khumi and Mro. Diversity was high; expected heterozygosity was 0.93 for P. falciparum and 0.81 for P. vivax . 85.7% (18/21) of P. vivax and 25% (5/20) of P. falciparum infections were polyclonal. No population structure was evident for either species, suggesting high transmission and gene flow among Pahari groups. Conclusions High subclinical infection prevalence and genetic diversity mirror ongoing transmission. Control activities should be specifically directed to Pahari groups at greatest risk.
 
Article
Background Interleukin (IL)-4 had been linked to malaria severity, but the findings are controversial, and the evidence is inconsistent and imprecise. In the current investigation, data on IL-4 levels in patients with severe and uncomplicated malaria were compiled. Methods The systematic review was registered at PROSPERO (CRD42022323387). Searches for relevant articles on IL-4 levels in patients with severe malaria and studies that examined IL-4 levels in both uncomplicated malaria and healthy controls were performed in PubMed, Embase, and Scopus using the search strategy without limitation to publication years or language. The quality of all included studies was evaluated using The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) Statement: standards for reporting observational studies . Qualitative and quantitative data syntheses were performed. The random-effects model, which weights each study according to its between- and within-study variance, was used to pool the mean difference (MD) of individual studies. The degree of heterogeneity was determined using Cochran's Q and I ² statistics. Additionally, meta-regression and subgroup analyses were perfomed to investigate possible sources of heterogeneity. The outliers were identified using the leave-one-out method and assessed publication bias using funnel plots, Egger’s test, and a contour-enhanced funnel plot . Results A total of 2300 studies were identified through database searches, and 36 were included for analyses. The meta-analysis results showed lower mean IL-4 levels in severe malaria (434 cases) than in uncomplicated malaria (611 cases) ( P = 0.01, pooled MD: −3.36 pg/mL, 95% confidence intervals CI −5.55 to −1.16 pg/mL, I 2 : 98.15%, 11 studies). The meta-analysis results showed no difference in mean IL-4 levels between cerebral malaria (96 cases) and noncerebral severe malaria (108 cases) ( P = 0.71, pooled MD: 0.86 pg/mL, 95% CI −3.60 to 5.32 pg/mL, I 2 92.13%, four studies). Finally, no difference was found in mean IL-4 levels between uncomplicated malaria (635 cases) and healthy controls (674 cases) ( P = 0.57, pooled MD: 0.79 pg/mL, 95% CI −1.92 to 3.50 pg/mL, I 2 : 99.89%, 11 studies). Conclusion The meta-analysis revealed lower IL-4 levels in patients with severe malaria than in those with uncomplicated malaria, though a trend toward comparable IL-4 levels between both groups was more likely because several sources of heterogeneities were observed. Based on the limited number of studies included in the meta-analysis, until additional investigations have been conducted, IL-4 consideration as an alternative prognostic factor for malaria severity is not warranted.
 
Partial purification of native 5′ to 3′ DNA helicase from P. falciparum crude extract. A Parasite crude extract was purified on the Resource Q column with 0–100% KCl linear gradient. B Resource Q pooled fraction containing helicase activity was purified on the Mono S column with 0–100% KCl linear gradient. C Mono S pooled active fraction was further purified on the ssDNA column with 0–100% KCl linear gradient. Each fraction was measured for the amount of proteins by using the Bradford reagent and assayed for unwinding activity using the M13-17-mer substrate
Unwinding activity of partially purified P. falciparum 3′ to 5′ DNA helicase eluted from ssDNA column (Fraction D). The standard displacement assay with M13-17-mer substrate and 2 ml of the enzyme from each fraction was done. Numbers above the figure are the fraction eluted from the column
Directionality of P. falciparum 5′ to 3′ DNA helicase activity. The 43-mer linear partial duplex was used as a substrate and then digested with SmaI. Lane 1 corresponds to a heat-denatured substrate, lane 2 is negative control (reaction without enzyme), and lane 3 is a reaction with 0.2 mg of partially purified P. falciparum 5′–3′ DNA helicase
Unwinding activity of P. falciparum 5′ to 3′ DNA helicase with different substrates. A the M13-17-mer partial duplex. B the 41-mer blunt end. C the M13-44-mer with 3′ hanging tail. D the M13-45-mer with 5' hanging tail. E the 5′ to 3′ hanging tails on the same M13 template. Asterisks denote the ³²P-labelled end. Lane 1 is the control without enzyme, Lane 2 is a reaction with partially purified enzyme, and Lane 3 is the heat-denatured substrate. UD: unwound DNA
Inhibitory effects of anthracyclines on P. falciparum 5′–3′ DNA helicase. Lane 1 is the heat-denatured substrate. Lane 2 is negative control and lanes 3 and 20 are positive controls. Lanes 4–7 are nogalamycin at concentrations of 10, 5, 1, 0.1 μM, respectively. Lanes 8–19 are aclarubicin, daunorubicin, and doxorubicin at the same series of drug concentrations at 10, 1, 0.1, 0.01 μM, respectively. Lane 21 is an enzyme with 0.2% (v/v) DMSO
Article
Background Plasmodium falciparum has been becoming resistant to the currently used anti-malarial drugs. Searching for new drug targets is urgently needed for anti-malarial development. DNA helicases separating double-stranded DNA into single-stranded DNA intermediates are essential in nearly all DNA metabolic transactions, thus they may act as a candidate for new drug targets against malarial parasites. Methods In this study, a P. falciparum 5′ to 3′ DNA helicase (PfDH-B) was partially purified from the crude extract of chloroquine- and pyrimethamine-resistant P. falciparum strain K1, by ammonium sulfate precipitation and three chromatographic procedures. DNA helicase activity of partially purified PfDH-B was examined by measuring its ability to unwind ³² P-labelled partial duplex DNA. The directionality of PfDH-B was determined, and substrate preference was tested by using various substrates. Inhibitory effects of DNA intercalators such as anthracycline antibiotics on PfDH-B unwinding activity and parasite growth were investigated. Results The native PfDH-B was partially purified with a specific activity of 4150 units/mg. The PfDH-B could unwind M13-17-mer, M13-31-mer with hanging tail at 3′ or 5′ end and a linear substrate with 3′ end hanging tail but not blunt-ended duplex DNA, and did not need a fork-like substrate. Anthracyclines including aclarubicin, daunorubicin, doxorubicin, and nogalamycin inhibited the unwinding activity of PfDH-B with an IC 50 value of 4.0, 7.5, 3.6, and 3.1 µM, respectively. Nogalamycin was the most effective inhibitor on PfDH-B unwinding activity and parasite growth (IC 50 = 0.1 ± 0.002 µM). Conclusion Partial purification and characterization of 5′–3′ DNA helicase of P. falciparum was successfully performed. The partially purified PfDH-B does not need a fork-like substrate structure found in P. falciparum 3′ to 5′ DNA helicase (PfDH-A). Interestingly, nogalamycin was the most potent anthracycline inhibitor for PfDH-B helicase activity and parasite growth in culture. Further studies are needed to search for more potent but less cytotoxic inhibitors targeting P. falciparum DNA helicase in the future.
 
Map showing study sites in Gaza and Inhambane provinces, southern Mozambique
Various mosquito collection tools used in the study. A Human-baited Tent Trap outdoors; B Human-baited Tent Trap indoors, C window exit trap, D Pyrethreum Spray Catch, a fieldworker spraying the eaves (from the outside)
Anopheles biting, resting, and exiting behaviors in southern Mozambique during the rainy and dry season of 2018. A-C Gaza Province (A: Bilene, B: Chokwe, cidade de Xai Xai), D-F Inhambane Province (D: cidade de Inhambane, E: Jangamo, F: Massinga). All indicators are expressed as the mean number of mosquitoes per person
Detection of Plasmodium falciparum in Anopheles species that were collected in Gaza and Inhambane province (southern Mozambique) in 2018
Article
Background To eliminate malaria in southern Mozambique, the National Malaria Control Programme and its partners are scaling up indoor residual spraying (IRS) activities in two provinces, Gaza and Inhambane. An entomological surveillance planning tool (ESPT) was used to answer the programmatic question of whether IRS would be effective in target geographies, given limited information on local vector bionomics. Methods Entomological intelligence was collected in six sentinel sites at the end of the rainy season (April–May 2018) and the beginning of the dry season (June–July 2018). The primary objective was to provide an ‘entomological snapshot’ by collecting question-based, timely and high-quality data within one single week in each location. Host-seeking behaviour (both indoors and outdoors) was monitored by human-baited tent traps. Indoor resting behaviour was quantified by pyrethrum spray catches and window exit traps. Results Five different species or species groups were identified: Anopheles funestus sensu lato (s.l.) (66.0%), Anopheles gambiae s.l. (14.0%), Anopheles pharoensis (1.4%), Anopheles tenebrosus (14.1%) and Anopheles ziemanni (4.5%). Anopheles funestus sensu stricto (s.s.) was the major vector among its sibling species, and 1.9% were positive for Plasmodium falciparum infections. Anopheles arabiensis was the most abundant vector species within the An. gambiae complex, but none tested positive for P. falciparum infections. Some An. tenebrosus were positive for P. falciparum (1.3%). When evaluating behaviours that impact IRS efficacy, i.e. endophily, the known primary vector An. funestus s.s. , was found to rest indoors—demonstrating at least part of its population will be impacted by the intervention if insecticides are selected to which this vector is susceptible. However, other vector species, including An. gambiae s.l ., An. tenebrosus , An. pharoensis and An. ziemanni , showed exophilic and exophagic behaviours in several of the districts surveilled. Conclusion The targeted approach to entomological surveillance was successful in collecting question-based entomological intelligence to inform decision-making about the use of IRS in specific districts. Endophilic An. funestus s.s. was documented as being the most prevalent and primary malaria vector suggesting that IRS can reduce malaria transmission, but the presence of other vector species both indoors and outdoors suggests that alternative vector control interventions that target these gaps in protection may increase the impact of vector control in southern Mozambique.
 
Study Commune Boundaries, Care-Seeking Behavior Study, Madagascar, 2018
Article
Background Prompt diagnosis and treatment of malaria contributes to reduced morbidity, particularly among children and pregnant women; however, in Madagascar, care-seeking for febrile illness is often delayed. To describe factors influencing decisions for prompt care-seeking among caregivers of children aged < 15 years and pregnant women, a mixed-methods assessment was conducted with providers (HP), community health volunteers (CHV) and community members. Methods One health district from each of eight malaria-endemic zones of Madagascar were purposefully selected based on reported higher malaria transmission. Within districts, one urban and one rural community were randomly selected for participation. In-depth interviews (IDI) and focus group discussions (FGD) were conducted with caregivers, pregnant women, CHVs and HPs in these 16 communities to describe practices and, for HPs, system characteristics that support or inhibit care-seeking. Knowledge tests on malaria case management guidelines were administered to HPs, and logistics management systems were reviewed. Results Participants from eight rural and eight urban communities included 31 HPs from 10 public and 8 private Health Facilities (HF), five CHVs, 102 caregivers and 90 pregnant women. All participants in FGDs and IDIs reported that care-seeking for fever is frequently delayed until the ill person does not respond to home treatment or symptoms become more severe. Key care-seeking determinants for caregivers and pregnant women included cost, travel time and distance, and perception that the quality of care in HFs was poor. HPs felt that lack of commodities and heavy workloads hindered their ability to provide quality malaria care services. Malaria commodities were generally more available in public versus private HFs. CHVs were generally not consulted for malaria care and had limited commodities. Conclusions Reducing cost and travel time to care and improving the quality of care may increase prompt care-seeking among vulnerable populations experiencing febrile illness. For patients, perceptions and quality of care could be improved with more reliable supplies, extended HF operating hours and staffing, supportive demeanors of HPs and seeking care with CHVs. For providers, malaria services could be improved by increasing the reliability of supply chains and providing additional staffing. CHVs may be an under-utilized resource for sick children.
 
Trends in Thailand’s malaria incidence, 2000–2020
Active malaria transmission foci in Thailand, 2020
Health-related public administration structure in Thailand
Timeline of key activities in LAO engagement, 2018–2021
“LAO Collaboration” Tableau dashboard in Malaria Online
Article
Thailand’s National Malaria Elimination Strategy 2017–2026 seeks to increase domestic support and financing for malaria elimination. During 2018–2020, through a series of training sessions, public health officials in Thailand utilized foci-level malaria data to engage subdistrict-level government units known as Local Administrative Organizations (LAOs) with the aim of increasing their understanding of their local malaria situation, collaboration with public health networks, and advocacy for financial support of targeted interventions in villages within their jurisdictions. As a result of these efforts, total LAO funding support for malaria nearly doubled from the 2017 baseline to 2020. In 2021, a novel “LAO collaboration” feature was added to Thailand’s national malaria information system that enables tracking and visualization of LAO financial support of malaria in areas with transmission, by year, down to the subdistrict level. This case study describes Thailand’s experience implementing the LAO engagement strategy, quantifying and monitoring the financial support mobilized from LAOs, and results from a qualitative study in five high-performance provinces examining factors and approaches that foster successful local collaboration between LAOs, public health networks, and communities for malaria prevention and response. Results from the study showed that significant malaria endemicity or local outbreaks helped spur collaboration in multiple provinces. Increases in LAO support and involvement were attributable to four approaches employed by public health officials: (a) strengthening malaria literacy and response capacity of LAOs, (b) organizational leadership in response to outbreaks, (c) utilization of structural incentives, and (d) multisectoral involvement in malaria response. In two provinces, capacity building of LAOs in malaria vector control, following a precedent set by Thailand’s dengue programme, enabled LAO personnel to play both funding and implementation roles in local malaria response. Wider replication of the LAO engagement strategy across Thailand may sustain gains and yield efficiencies in the fight against malaria as the vector-borne disease workforce declines. Lessons from Thailand’s experience may be useful for malaria programmes in other geographies to support the goals and sustainability of elimination and prevention of re-establishment by improving financing through local collaboration between the health system and elected officials.
 
Article
Background Quality assurance (QA) of insecticide-treated nets (ITNs) delivered to malaria-endemic countries is conducted by measuring physiochemical parameters, but not bioefficacy against malaria mosquitoes. This study explored utility of cone bioassays for pre-delivery QA of pyrethroid ITNs to test the assumption that cone bioassays are consistent across locations, mosquito strains, and laboratories. Methods Double-blinded bioassays were conducted on twenty unused pyrethroid ITNs of 4 brands (100 nets, 5 subsamples per net) that had been delivered for mass distribution in Papua New Guinea (PNG) having passed predelivery inspections. Cone bioassays were performed on the same net pieces following World Health Organization (WHO) guidelines at the PNG Institute of Medical Research (PNGIMR) using pyrethroid susceptible Anopheles farauti sensu stricto ( s.s .) and at Ifakara Health Institute (IHI), Tanzania using pyrethroid susceptible Anopheles gambiae s.s . Additionally, WHO tunnel tests were conducted at IHI on ITNs that did not meet cone bioefficacy thresholds. Results from IHI and PNGIMR were compared using Spearman’s Rank correlation, Bland–Altman (BA) analysis and analysis of agreement. Literature review on the use of cone bioassays for unused pyrethroid ITNs testing was conducted. Results In cone bioassays, 13/20 nets (65%) at IHI and 8/20 (40%) at PNGIMR met WHO bioefficacy criteria. All nets met WHO bioefficacy criteria on combined cone/tunnel tests at IHI. Results from IHI and PNGIMR correlated on 60-min knockdown (KD60) (r s = 0.6, p = 0.002,n = 20) and 24-h mortality (M24) (r s = 0.9, p < 0.0001,n = 20) but BA showed systematic bias between the results. Of the 5 nets with discrepant result between IHI and PNGIMR, three had confidence intervals overlapping the 80% mortality threshold, with averages within 1–3% of the threshold. Including these as a pass, the agreement between the results to predict ITN failure was good with kappa = 0.79 (0.53–1.00) and 90% accuracy. Conclusions Based on these study findings, the WHO cone bioassay is a reproducible bioassay for ITNs with > 80% M24, and for all ITNs provided inherent stochastic variation and systematic bias are accounted for. The literature review confirms that WHO cone bioassay bioefficacy criteria have been previously achieved by all pyrethroid ITNs (unwashed), without the need for additional tunnel tests. The 80% M24 threshold remains the most reliable indicator of pyrethroid ITN quality using pyrethroid susceptible mosquitoes. In the absence of alternative tests, cone bioassays could be used as part of pre-delivery QA.
 
Top panels indicate the geographic location of the study site. Yellow circles in the bottom panel represent the total counts of An. gibbinsi captured at each household throughout the study period. Households with no An. gibbinsi are represented by red triangles
Article
Background Nchelenge District in northern Zambia suffers from holoendemic malaria transmission despite a decade of yearly indoor residual spraying (IRS) and insecticide-treated net (ITN) distributions. One hypothesis for this lack of impact is that some vectors in the area may forage in the early evening or outdoors. Anopheles gibbinsi specimens were identified in early evening mosquito collections performed in this study area, and further insight was gleaned into this taxon, including characterizing its genetic identity, feeding preferences, and potential role as a malaria vector. Methods Mosquitoes were collected in July and August 2019 by CDC light traps in Nchelenge District in indoor sitting rooms, outdoor gathering spaces, and animal pens from 16:00–22:00. Host detection by PCR, COI and ITS2 PCR, and circumsporozoite (CSP) ELISA were performed on all samples morphologically identified as An. gibbinsi, and a subset of specimens were selected for COI and ITS2 sequencing. To determine risk factors for increased abundance of An. gibbinsi, a negative binomial generalized linear mixed-effects model was performed with household-level variables of interest. Results Comparison of COI and ITS2 An. gibbinsi reference sequences to the NCBI database revealed > 99% identity to “Anopheles sp. 6” from Kenya. More than 97% of specimens were morphologically and molecularly consistent with An. gibbinsi. Specimens were primarily collected in animal pen traps (59.2%), followed by traps outdoors near where humans gather (24.3%), and traps set indoors (16.5%). Host DNA detection revealed a high propensity for goats, but 5% of specimens with detected host DNA had fed on humans. No specimens were positive for Plasmodium falciparum sporozoites. Animal pens and inland households > 3 km from Lake Mweru were both associated with increased An. gibbinsi abundance. Conclusions This is the first report of An. gibbinsi in Nchelenge District, Zambia. This study provided a species identity for unknown “An. sp. 6” in the NCBI database, which has been implicated in malaria transmission in Kenya. Composite data suggest that this species is largely zoophilic and exophilic, but comes into contact with humans and the malaria parasites they carry. This species should continue to be monitored in Zambia and neighbouring countries as a potential malaria vector.
 
Selected sites to receive Intensification Plan interventions and number of mobile malaria workers (MMWs) assigned based on identified malaria hotspots
Number of malaria tests performed in intensification plan (IP) health facility catchment areas before and during IP, stratified by health cadre. Total: All tests (cadre disaggregation not available for pre-IP data); HC: health center; MMW (Not IP): mobile malaria workers in IP sites but not managed directly through IP, MMW: mobile malaria workers managed through IP; VMW: volunteer malaria worker. Pre-IP includes IP Phase 1 HFCAs only. IP Phase 1 (30 HFCAs), IP Phase 2 (36 HFCAs)
Number of Plasmodium falciparum/mix malaria cases (grey bars) and test positivity rate (TPR—red solid line) in intensification plan (IP) health facility catchment areas (HFCAs) over the course of the IP. Left of black dotted line: IP Phase 1 (30 HFCAs); right of black dotted line: IP Phase 2 (36 HFCAs)
Number of Plasmodium falciparum/mix malaria cases (red dots) in intensification plan (IP) health facility catchment areas (HFCAs). Left) Beginning of IP (IP1 HFCAs)—October 2018 and Right) End of IP (IP2 HFCAs)—October 2020. Blue polygons: IP operational districts; green polygons: non-IP operational districts
Cumulative number of P. falciparum/mix cases in all health facility catchment areas (HFCAs) in Cambodia. Black dashed line: rollout date of IP. Red: IP HFCAs (n = 45); Blue: Non-IP HFCAs (n = 897). Red dashed line: extrapolated counterfactual from pre-IP model. HFCAs are classified as an IP HFCA if they were included in either/both of Phase 1 or Phase 2
Article
Background Malaria in Cambodia has decreased by 90.8% between 2010 and 2020, driven by the commitment of the National Center for Parasitology, Entomology and Malaria (CNM) and the achievements of the roll-out of a village malaria worker programme. However, in the first seven months of 2018, CNM identified a 207% increase (11,969 to 36,778) in confirmed malaria cases compared to the same months in the previous year. To address this increase, CNM developed the “Intensification Plan” (IP), implemented between October 2018 and December 2020. Methods The structure of the IP was summarized, including the selection of sites, the interventions implemented in the selected health facility catchment areas (HFCAs) and the monitoring and evaluation process. Data on IP interventions were collected by CNM and civil society organisations. Data on malaria cases and tests from all HFCAs in Cambodia from January 2018 to December 2020 were sourced from the Cambodia Malaria Information System (MIS) and WHO Malaria Elimination Database. Malaria data from IP HFCAs and non-IP HFCAs was analysed and compared to present the changes in malaria testing and confirmed cases before and during implementation of the IP. Results Between October 2018 and December 2020, through the IP 16,902 forest packs and 293,090 long-lasting insecticide treated nets were distributed. In the 45 HFCAs included in the IP, 431,143 malaria tests were performed and 29,819 malaria cases were diagnosed, 5364 (18%) of which were Plasmodium falciparum /mixed cases. During the intervention period, over all HFCAs included in IP, P. falciparum /mixed cases declined from 1029 to 39, a 96.2% decrease, and from 25.4 P. falciparum /mixed cases per HFCA to 0.9. HFCAs not included in IP declined from 468 to 43 cases, a 90.8% decrease, showing that routine malaria activities in Cambodia were also playing an important contribution to malaria control. Conclusions Over the course of IP implementation there was a substantial increase in malaria testing and both overall malaria cases and P. falciparum /mixed cases decreased month on month. The initiative yields lessons learned for Cambodia to reach the final stage of elimination as well as for other countries aiming to accelerate their malaria control programmes.
 
Localities where LLINs were distributed and durability monitored
Retained proportions not removed (red), retention after sampling (blue) and cumulative number of nets removed by type and semester after 3 years
Categories of holes by type and semester after 3 years. Size 1, size 2, size 3
Insecticide concentration in mg/m² according to the type LLINs and semester
Correlation of chemical and bioassay results by type of LLIN and semester
Article
Background The preventive and curative strategies of malaria are based on promoting the use of long-lasting insecticidal nets (LLINs) and treating confirmed cases with artemisinin-based combination therapy. These strategies have led to a sharp decline in the burden of malaria, which remains a significant public health problem in sub-Saharan countries. The objective of this study was to determine and compare the residual efficacy of LLINs recommended by the World Health Organization. Methods The study was conducted in six villages in two sites in Senegal located in the Sahelo-Sudanian area of the Thiès region, 70 km from Dakar and in Mbagame, a semi-urban zone in the Senegal River Valley. A census was conducted of all sleeping places in each household to be covered by LLINs. Five brands of LLIN were distributed, and every six months, retention rates, net use, maintenance, physical integrity, insecticide chemical content, and biological efficacy were examined for each type of LLIN. Results A total of 3012 LLINs were distributed in 1249 households in both sites, with an average coverage rate of 94% (95% CI 92.68–95.3). After 36 months, the average retention rate was 12.5% and this rate was respectively 20.5%, 15.1%, 10%, 7%, and 3% for Olyset Net ® , Dawa Plus ® 2.0, PermaNet ® 2.0, NetProtect ® and Life Net ® , respectively. The proportion of LLINs with holes and the average number of holes per mosquito net increased significantly during each follow-up, with a large predominance of size 1 (small) holes for all types of LLINs distributed. During the three-year follow-up, bioassay mortality rates of a susceptible strain of insectary reared Anopheles coluzzii decreased in the following net types: in Dawa Plus ® 2.0 (100% to 51.7%), PermaNet ® 2.0 (96.6% to 83%), and Olyset Net ® (96.6% to 33.3%). Mortality rates remained at 100% in Life Net ® over the same time period. After 36 months, the average insecticide content per brand of LLIN decreased by 40.9% for Dawa Plus ® 2.0, 31% for PermaNet ® 2.0, 39.6% for NetProtect ® and 51.9% for Olyset Net ® and 40.1% for Life Net. Conclusions Although some net types retained sufficient insecticidal activity, based on all durability parameters measured, none of the net types survived longer than 2 years.
 
VCT apparatus and design. A bespoke stand made from fused PLA plastic filament (A) supports the Perspex cone board at 45°. Openings in the side of the stand allows the host arm to be placed horizontally behind the net. The smartphone is clamped to the board and positioned in front of the cone. The recording field (B) comprises of the entire volume of the cone and net. The presence of a gap between the lower rim and upper opening of the cone (red arrow) ensures the optimal camera angle and position
Article
Background The WHO cone test is one of three tests currently used to evaluate the efficacy of insecticide-treated bed nets (ITNs). It generates two test outputs, knockdown and 24-h mortality, both indicative of immediate toxicity but that reveal little about the nature of mosquito and ITN interaction or how results translate to real-world settings. Methods A human arm held 5 mm behind the net surface acted as a host attractant during cone tests and a smartphone was used to capture mosquito behaviour in the cone. Post-exposure blood feeding and survival for nine days were recorded; ingested blood meal size was determined by measuring excreted haematin. Four strains of Anopheles gambiae (insecticide susceptible: Kisumu and N’gousso; insecticide resistant: Banfora and VK7) were tested with and without the host attractant using untreated, Permanet 2.0 and Olyset nets. Video recordings were scan sampled every five seconds to record mosquito positions on either the net, in flight or in contact with the cone. Generalized estimating equations were used to analyse all data except survival within nine days which was analysed using Weighted Cox Regression. Results Net contact was the most frequently recorded behaviour in all Anopheles spp. strains on all nets. Adding the human host as attractant triggered excitatory behaviours: in all strains, the magnitude of net contact was significantly decreased compared to tests without a host. ITN exposure altered the observed behaviour of the two susceptible strains, which exhibited a decreased response to the host during ITN tests. The resistant strains did not alter their behaviour during ITN tests. Significantly less net contact was observed during Olyset Net tests compared to Permanet 2.0. The host presence affected survival after exposure: Banfora and VK7 mosquitoes exposed to Permanet 2.0 with a host lived longer compared to tests performed without a host. However, mosquitoes that blood-fed and survived long enough to digest the blood meal did not exhibit significantly reduced longevity regardless of the presence of the host attractant. Conclusions Simple modifications to the WHO cone test and extension of post-test monitoring beyond the current 24 h enable detailed behavioural characterizations of individual ITNs to be compiled. The effects observed from testing with a host and including blood feeding suggest that more representative estimates of true of ITN efficacy are gained with these modifications than when using the current testing protocol.
 
P. falciparum 3D7 parasite growth rate after 48 h of incubation with various concentration of ATA. The IC50 was detected at 634 μM
P. falciparum 3D7 parasite growth rate after 48 h of incubation with various concentration of artemisinin. The IC50 of artemisinin was detected at 2.34 nM
P. falciparum 3D7 parasite growth rate after 48 h of incubation with various concentration of resveratrol. The IC50 was detected at 0.231 µM
The effect of 30 mg/kg BW ATA on survival rate of experimental cerebral malaria model mouse infected with P. berghei ANKA treatment with 30 mg/kb BW. ATA showed longer survival rate whereas the other concentrations did not significantly differ than the control untreated group
Histogram of the Plasmodium yoelii growth rate in BALB/c mice in the presence or absence of PMCA inhibitor, resveratrol. Sulfadoxine is used as positive control
Article
Background Rapid emergence of Plasmodium resistance to anti-malarial drug mainstays has driven a continual effort to discover novel drugs that target different biochemical pathway (s) during infection. Plasma membrane Calcium + 2 ATPase (PMCA4), a novel plasma membrane protein that regulates Calcium levels in various cells, namely red blood cell (RBC), endothelial cell and platelets, represents a new biochemical pathway that may interfere with susceptibility to malaria and/or severe malaria. Methods This study identified several pharmacological inhibitors of PMCA4, namely ATA and Resveratrol, and tested for their anti-malarial activities in vitro and in vivo using the Plasmodium falciparum 3D7 strain, the Plasmodium berghei ANKA strain, and Plasmodium yoelii 17XL strain as model. Results In vitro propagation of P. falciparum 3D7 strain in the presence of a wide concentration range of the inhibitors revealed that the parasite growth was inhibited in a dose-dependent manner, with IC50s at 634 and 0.231 µM, respectively. Results The results confirmed that both compounds exhibit moderate to potent anti-malarial activities with the strongest parasite growth inhibition shown by resveratrol at 0.231 µM. In vivo models using P. berghei ANKA for experimental cerebral malaria and P. yoelii 17XL for the effect on parasite growth, showed that the highest dose of ATA, 30 mg/kg BW, increased survival of the mice. Likewise, resveratrol inhibited the parasite growth following 4 days intraperitoneal injection at the dose of 100 mg/kg BW. Conclusion The findings indicate that the PMCA4 of the human host may be a potential target for novel anti-malarials, either as single drug or in combination with the currently available effective anti-malarials.
 
Article
Background Independent emergence and spread of artemisinin-resistant Plasmodium falciparum malaria have recently been confirmed in Africa, with molecular markers associated with artemisinin resistance increasingly detected. Surveillance to promptly detect and effectively respond to anti-malarial resistance is generally suboptimal in Africa, especially in low transmission settings where therapeutic efficacy studies are often not feasible due to recruitment challenges. However, these communities may be at higher risk of anti-malarial resistance. Methods From March 2018 to February 2020, a sequential mixed-methods study was conducted to evaluate the feasibility of the near-real-time linkage of individual patient anti-malarial resistance profiles with their case notifications and treatment response reports, and map these to fine scales in Nkomazi sub-district, Mpumalanga, a pre-elimination area in South Africa. Results Plasmodium falciparum molecular marker resistance profiles were linked to 55.1% (2636/4787) of notified malaria cases, 85% (2240/2636) of which were mapped to healthcare facility, ward and locality levels. Over time, linkage of individual malaria case demographic and molecular data increased to 75.1%. No artemisinin resistant validated/associated Kelch-13 mutations were detected in the 2385 PCR positive samples. Almost all 2812 samples assessed for lumefantrine susceptibility carried the wildtype mdr86ASN and crt76LYS alleles, potentially associated with decreased lumefantrine susceptibility. Conclusion Routine near-real-time mapping of molecular markers associated with anti-malarial drug resistance on a fine spatial scale provides a rapid and efficient early warning system for emerging resistance. The lessons learnt here could inform scale-up to provincial, national and regional malaria elimination programmes, and may be relevant for other antimicrobial resistance surveillance.
 
Gel electrophoresis of P. vivax 18S rRNA and pvmdr-1 nested PCR. a18S rRNA nested PCR. Blood samples were collected from malaria patients who agreed to the study. All blood samples collected in three hospitals (Yangju, Koyang and Ildong), during 2016 and 2017, were screened using 18S rRNA nested PCR. Based on the screened result, 73 samples were identified as positive samples. 1100 bp or 120 bp was detected by 2nd PCR of 18S rRNA. b Nested PCR targeting pvmdr-1 amplicons ranged from 104 bp to 4,254 bp were observed in 73 P. vivax positive samples using pvmdr-1 nested PCR. M: 1 Kb Plus DNA Ladder
The malaria-endemic region and pvmdr-1 haplotypes of ROK in 2016 and 2017. Each red triangle indicates malaria-endemic region of haplotypes [Goseong (Type 1 and 4), Cheorwon (Type 1, 2, 3, 4, and 5), Yeonchen (Type 0, 1, 2, 3 and 4), Paju (Type 1, 2, 3 and 4), Gimpo (Type 2), and Hwacheon (Type 1 and 4)]. Six haplotypes were classified from 73 specimens collected in the Armed Forces Hospitals (Yangju, Koyang, and Ildong). Numeric letters mean 6 haplotypes. Reprinted from USGS National Map Viewer [36] under a CC BY license, with permission from U.S. Geological Survey, original copyright 2002
Phylogenetic analysis of 6 classified haplotypes via pvmdr-1 SNP analysis of 73 P. vivax clinical samples. The sequences of pvmdr-1 were aligned with that of Sal I strain (GenBank Accession# AY571984) and PlasmoDB by using BioEdit Sequence Alignment Editor. The aligned sequences were performed phylogenetic analysis by using 1000 bootstrap replications and the neighbour-joining method
Article
Background Malaria chemoprophylaxis using chloroquine (CQ) and primaquine (PQ) has been administered to resident soldiers in the 3rd Army of Republic of Korea (ROK) to prevent malaria infection since the year 1997. Due to mass chemoprophylaxis against malaria, concern exists about the occurrence of chloroquine resistance (CQR). This study aimed to investigate the single nucleotide polymorphisms (SNPs) of the Plasmodium vivax multi-drug resistance protein-1 (pvmdr-1) gene to monitor the risk of CQR. Methods SNPs of the pvmdr-1 gene were analysed in 73 soldiers of the 3rd Army of ROK diagnosed with infection by P. vivax. Results Quintuple mutations (G698S, L845F, M908L, T958M, and F1076L) were detected in 73 soldiers. A newly identified non-synonymous mutation in the Y541C position had been introduced into P. vivax malaria-endemic areas in ROK, at a frequency of 1.3% (1/73). In addition, synonymous mutations were detected at positions K44 (38.4%, 28/73), L493 (26%, 19/73), T529 (61.6%, 45/73), and E1233 (52.1%, 38/73). Based on these SNPs, pvmdr-1 sequences of ROK were classified into 6 haplotypes. The phylogenetic analysis closed to the type of North Korean showed that P. vivax malaria of ROK could be a reason of influx from North Korea. Conclusions This study showed that synonymous and non-synonymous mutations of pvmdr-1 were observed in the malaria chemoprophylaxis-executed regions of ROK from 2016 to 2017. Based on the rapid transition of pvmdr-1 SNPs, continuous surveillance for SNPs of pvmdr-1 related to CQR in the malaria-endemic regions of ROK is essential.
 
The mix of primers and probes used in this study, adapted from Schindler et al. [22]
Article
Background Malaria is an infectious disease considered as one of the biggest causes of mortality in endemic areas. This life-threatening disease needs to be quickly diagnosed and treated. The standard diagnostic tools recommended by the World Health Organization are thick blood smears microscopy and immuno-chromatographic rapid diagnostic tests. However, these methods lack sensitivity especially in cases of low parasitaemia and non-falciparum infections. Therefore, the need for more accurate and reliable diagnostic tools, such as real-time polymerase chain reaction based methods which have proven greater sensitivity particularly in the screening of malaria, is prominent. This study was conducted at the French National Malaria Reference Centre to assess sensitivity and specificity of two commercial malaria qPCR kits and two in-house developed qPCRs compared to LAMP. Methods 183 blood samples received for expertise at the FNMRC were included in this study and were subjected to four different qPCR methods: the Biosynex Ampliquick ® Malaria test, the BioEvolution Plasmodium Typage test, the in-house HRM and the in-house TaqMan qPCRs. The specificity and sensitivity of each method and their confidence intervals were determined with the LAMP-based assay Alethia® Malaria as the reference for malaria diagnosis. The accuracy of species diagnosis of the Ampliquick ® Malaria test and the two in-house qPCRs was also evaluated using the BioEvolution Plasmodium Typage test as the reference method for species identification. Results The main results showed that when compared to LAMP, a test with excellent diagnostic performances, the two in-house developed qPCRs were the most sensitive (sensitivity at 100% for the in-house TaqMan qPCR and 98.1% for the in-house HRM qPCR), followed by the two commercial kits: the Biosynex Ampliquick ® Malaria test (sensitivity at 97.2%) and the BioEvolution Plasmodium Typage (sensitivity at 95.4%). Additionally, with the in-house qPCRs we were able to confirm a Plasmodium falciparum infection in microscopically negative samples that were not detected by commercial qPCR kits. This demonstrates that the var genes of P. falciparum used in these in-house qPCRs are more reliable targets than the 18S sRNA commonly used in most of the developed qPCR methods for malaria diagnosis. Conclusion Overall, these results accentuate the role molecular methods could play in the screening of malaria. This may represent a helpful tool for other laboratories looking to implement molecular diagnosis methods in their routine analysis, which could be essential for the detection and treatment of malaria carriers and even for the eradication of this disease.
 
Cost-consequences framework for the Private Sector Malaria Prevention project. This provides the analytical framework for the consequences included in the catalysed expenditure analysis. Intermediate outcome and impact measures were not included
Ratio of annual non-donor to donor expenditure for 5 years after project end. Projected ratios presented for five scenarios ranging from best case to worst case according to assumptions for non-donor contributions (see Table 5 and Additional File 1 for details of scenario assumptions)
Article
Background An estimated 1.5 billion malaria cases and 7.6 million malaria deaths have been averted globally since 2000; long-lasting insecticidal nets (LLINs) have contributed an estimated 68% of this reduction. Insufficient funding at the international and domestic levels poses a significant threat to future progress and there is growing emphasis on the need for enhanced domestic resource mobilization. The Private Sector Malaria Prevention (PSMP) project was a 3-year intervention to catalyse private sector investment in malaria prevention in Ghana. Methods To assess value for money of the intervention, non-donor expenditure in the 5 years post-project catalysed by the initial donor investment was predicted. Non-donor expenditure catalysed by this investment included: workplace partner costs of malaria prevention activities; household costs in purchasing LLINs from retail outlets; domestic resource mobilization (public sector financing and private investors). Annual ratios of projected non-donor expenditure to annualized donor costs were calculated for the 5 years post-project. Alternative scenarios were constructed to explore uncertainty around future consequences of the intervention. Results The total donor financial cost of the 3-year PSMP project was USD 4,418,996. The average annual economic donor cost per LLIN distributed through retail sector and workplace partners was USD 21.17 and USD 7.55, respectively. Taking a 5-year post-project time horizon, the annualized donor investment costs were USD 735,805. In the best-case scenario, each USD of annualized donor investment led to USD 4.82 in annual projected non-donor expenditure by the fifth-year post-project. With increasingly conservative assumptions around the project consequences, this ratio decreased to 3.58, 2.16, 1.07 and 0.93 in the “very good”, “good”, “poor” and “worst” case scenarios, respectively. This suggests that in all but the worst-case scenario, donor investment would be exceeded by the non-donor expenditure it catalysed. Conclusions The unit cost per net delivered was high, reflecting considerable initial investment costs and relatively low volumes of LLINs sold during the short duration of the project. However, taking a longer time horizon and broader perspective on the consequences of this complex catalytic intervention suggests that considerable domestic resources for malaria control could be mobilized, exceeding the value of the initial donor investment.
 
Article
Background In Burkina Faso, malaria remains the first cause of medical consultation and hospitalization in health centres. First-line case management of malaria in the country’s health facilities is based on the use of artemisinin-based combination therapy (ACT). To optimize the use of these anti-malarial drugs in the perspective of mitigating the emergence of artemisinin resistance, which is a serious threat to malaria control and elimination, a pilot programme using multiple first-line therapies (MFTs) [three artemisinin-based combinations—pyronaridine–artesunate, dihydroartemisinin–piperaquine and artemether-lumefantrine] has been designed for implementation. As the success of this MFT pilot programme depends on the perceptions of key stakeholders in the health system and community members, the study aimed to assess their perceptions on the implementation of this strategy. Methods Semi-structured interviews, including 27 individual in-depth interviews and 41 focus groups discussions, were conducted with key stakeholders including malaria control policymakers and implementers, health system managers, health workers and community members. Volunteers from targets stakeholder groups were randomly selected. All interviews were recorded, transcribed and translated. Content analysis was performed using the qualitative software programme QDA Miner. Results The interviews revealed a positive perception of stakeholders on the implementation of the planned MFT programme. They saw the strategy as an opportunity to strengthen the supply of anti-malarial drugs and improve the management of fever and malaria. However, due to lack of experience with the products, health workers and care givers expressed some reservations about the effectiveness and side-effect profiles of the two anti-malarial drugs included as first-line therapy in the MFT programme (pyronaridine–artesunate, dihydroartemisinin–piperaquine). Questions were raised about the appropriateness of segmenting the population into three groups and assigning a specific drug to each group. Conclusion The adherence of both populations and key stakeholders to the MFT implementation strategy will likely depend on the efficacy of the proposed drugs, the absence of, or low frequency of, side-effects, the cost of drugs and availability of the different combinations.
 
Flow diagram for phenotypic and genotypic testing for confirming pfhrp2 and pfhrp3 gene status. Following collection of blood sample, tests can be performed immediately or later by laboratory assays. Antigen and gene data can be gathered for appropriate interpretation decision making. aPoint-of-contact (POC) assays will require whole blood, whereas laboratory assays can accommodate whole blood, fractionated blood, or dried blood on filter paper as appropriate starting sample types
Article
Rapid diagnostic tests (RDTs) detecting Plasmodium falciparum histidine-rich protein 2 (HRP2) have been an important tool for malaria diagnosis, especially in resource-limited settings lacking quality microscopy. Plasmodium falciparum parasites with deletion of the pfhrp2 gene encoding this antigen have now been identified in dozens of countries across Asia, Africa, and South America, with new reports revealing a high prevalence of deletions in some selected regions. To determine whether HRP2-based RDTs are appropriate for continued use in a locality, focused surveys and/or surveillance activities of the endemic P. falciparum population are needed. Various survey and laboratory methods have been used to determine parasite HRP2 phenotype and pfhrp2 genotype, and the data collected by these different methods need to be interpreted in the appropriate context of survey and assay utilized. Expression of the HRP2 antigen can be evaluated using point-of-care RDTs or laboratory-based immunoassays, but confirmation of a deletion (or mutation) of pfhrp2 requires more intensive laboratory molecular assays, and new tools and strategies for rigorous but practical data collection are particularly needed for large surveys. Because malaria diagnostic strategies are typically developed at the national level, nationally representative surveys and/or surveillance that encompass broad geographical areas and large populations may be required. Here is discussed contemporary assays for the phenotypic and genotypic evaluation of P. falciparum HRP2 status, consider their strengths and weaknesses, and highlight key concepts relevant to timely and resource-conscious workflows required for efficient diagnostic policy decision making.
 
Location of Meghalaya and the study sites. The pins in the detailed map present the PHCs. Each colour represents the PHCs in the different districts. The PHCs selected for the study are circled
Article
Background India has made considerable progress in malaria reduction over the past two decades, with government-sponsored indoor residual spraying (IRS) and insecticide-treated bed net (ITN) or long-lasting insecticidal nets (LLIN) distribution being the main vector-related prevention efforts. Few investigations have used non-participant observational methods to assess malaria control measures while they were being implemented, nor documented people’s perceptions and acceptance of IRS or LLINs in India, and none have done so in the northeast region. This study evaluated household (HH)-level operation of IRS and distribution of LLINs by India’s National Vector Borne Disease Control Programme (NVBDCP) in 50 villages of Meghalaya state, and documented their acceptance and use. Methods Study field teams accompanied the government health system teams during August-October, 2019 and 2020 to observe deployment of LLINs, and record HH-level data on LLIN numbers and use. In addition, NVBDCP spray teams were followed during 2019–2021 to observe IRS preparation and administration. HH members were interviewed to better understand reasons for acceptance or refusal of spraying. Results A total of 8386 LLINs were distributed to 2727 HHs in 24 villages from five Primary Health Centres, representing 99.5% of planned coverage. Interviews with 80 HH residents indicated that they appreciated the LLIN dissemination programme, and generally made regular and appropriate use of LLINs, except during overnight travel or when working in agricultural fields. However, HH-level IRS application, which was observed at 632 HHs, did not always follow standard insecticide preparation and safety protocols. Of 1,079 occupied HHs visited by the spray team, 632 (58.6%) refused to allow any spraying. Only 198 (18.4%) HHs agreed to be sprayed, comprising 152 (14.1%) that were only partly sprayed, and 46 (4.3%) that were fully sprayed. Reasons for refusal included: inadequate time to rearrange HH items, young children were present, annoying smell, staining of walls, and threat to bee-keeping or Eri silk moth cultivation. Conclusions These findings are among the first in India that independently evaluate people's perceptions and acceptance of ongoing government-sponsored IRS and LLIN programmes for malaria prevention. They represent important insights for achieving India's goal of malaria elimination by 2030.
 
Article
Background Coronavirus disease 2019 (COVID-19) often causes atypical clinical manifestations similar to other infectious diseases. In malaria-endemic areas, the pandemic situation will very likely result in co-infection of COVID-19 and malaria, although reports to date are still few. Meanwhile, this disease will be challenging to diagnose in areas with low malaria prevalence because the symptoms closely resemble COVID-19. Case presentation A 23-year-old male patient presented to the hospital with fever, anosmia, headache, and nausea 1 week before. He was diagnosed with COVID-19 and treated for approximately 10 days, then discharged to continue self-quarantine at home. 2 weeks later, he returned to the hospital with a fever raised intermittently every 2 days and marked by a chilling-fever-sweating cycle. A laboratory test for malaria and a nasopharyngeal swab for SARS CoV-2 PCR were conducted, confirming both diagnoses. The laboratory examination showed markedly elevated D-dimer. He was treated with dihydroartemisinin-piperaquine (DHP) 4 tablets per day for 3 days and primaquine 2 tablets per day for 14 days according to Indonesian National Anti-malarial Treatment Guidelines. After 6 days of treatment, the patient had no complaints, and the results of laboratory tests had improved. This report describes the key points in considering the differential diagnosis and prompt treatment of malaria infection during the pandemic of COVID-19 in an endemic country to prevent the worse clinical outcomes. COVID-19 and malaria may also cause a hypercoagulable state, so a co-infection of those diseases may impact the prognosis of the disease. Conclusion This case report shows that considering the possibility of a co-infection in a COVID-19 patient who presents with fever can prevent delayed treatment that can worsen the disease outcome. Paying more attention to a history of travel to malaria-endemic areas, a history of previous malaria infection, and exploring anamnesis regarding the fever patterns in patients are important points in making a differential diagnosis of malaria infection during the COVID-19 pandemic.
 
Article
Background: Important knowledge gaps exist in the understanding of the management of the risks of imported malaria in Canada among Francophone immigrants from sub-Saharan Africa (FISSA). The aim of this cross-sectional study was to investigate the malaria related-knowledge, attitude and practices (KAP) of FISSA in Edmonton, where these immigrants are in an official minority language situation and the impact of language barriers on these factors. Methods: A structured survey was used to examine the KAP of 382 FISSA in the Edmonton area from 2018 to 2019. Fisher's Exact Test was applied to determine if there were associations between knowledge of malaria and different risk factors. Results: Almost all FISSA (97%) had an accurate knowledge of fever as the key symptom of malaria. Interestingly, 60% of participants identified bed nets as a preventive method and only 19% of participants had accurate knowledge of malaria transmission. An accurate knowledge of symptoms was significantly associated with a high perceived risk of contracting malaria [odds ratio (OR) 4.33, 95% confidence interval (CI) 1.07-20.62]. Furthermore, even though 70% of FISSA had a high perceived risk of contracting malaria in endemic regions, only 52% of travellers had a pre-travel medical encounter. Importantly, language was not the predominant reason for not seeking pre-travel medical advice, although 84% of respondents chose French as their official language of preference when seeking medical advice. Having a French-speaking physician was correlated with satisfactory prevention knowledge (OR 1.96, 95% CI 1.16-3.35). With respect to health-seeking behaviour, 88% of respondents with a child < 5 years of age would seek medical care for fever in the child after travel to sub-Saharan Africa (SSA). Conclusion: This study highlights that factors other than knowledge, risk assessment, and language might determine the lack of compliance with pre-travel medical encounters. It underscores the need for effective strategies to improve this adherence in minority settings.
 
Map of the surveyed districts highlighting the location of included health facilities
Source for intermittent preventative treatment for malaria in pregnancy (IPTp) as reported by women in the intervention arm, endline only (N = 343). CHW community health worker
Source for intermittent preventative treatment for malaria in pregnancy (IPTp), January 2019 to July 2020, routine service data. ANC antenatal care, CHW community health worker
Frequency of community health worker (CHW) visits with pregnant women by visit number, January 2019 to July 2020
Article
Background Malaria in pregnancy doubles the risk of low birthweight; up to 11% of all neonatal deaths in sub-Saharan Africa are associated with malaria in pregnancy. To prevent these and other adverse health consequences, the World Health Organization recommends administering intermittent preventive treatment in pregnancy (IPTp) with sulfadoxine–pyrimethamine for all pregnant women at each antenatal care (ANC) visit, starting as early as possible in the second trimester. The target is for countries to administer a minimum of three doses (IPTp3+) to at least 85% of pregnant women. Methods A cluster randomized, controlled trial was conducted to assess the effect of delivery of IPTp by community health workers on the coverage of IPTp3 + and ANC visits in Malawi. Community delivery of IPTp was implemented within two districts in Malawi over a 21-month period, from November 2018 to July 2020. In control sites, IPTp was delivered at health facilities. Representative samples of women who delivered in the prior 12 months were surveyed at baseline (n = 370, December 2017) and endline (n = 687, August 2020). A difference in differences analysis was conducted to assess the change in coverage of IPTp and ANC over time, accounting for clustering at the health facility level. Results Overall IPTp coverage increased over the study period. At baseline, women received a mean of 2.3 IPTp doses (range 0–5 doses) across both arms, and at endline, women received a mean of 2.8 doses (range 0–9 doses). Despite overall increases, the change in IPTp3 + coverage was not significantly different between intervention and control groups (6.9%, 95% CI: -5.9%, 19.6%). ANC4 + coverage increased significantly in the intervention group compared with the control group, with a difference-in-differences of 25.3% points (95% CI: 1.3%, 49.3%). Conclusions In order to reduce the burden of malaria in pregnancy, new strategies are needed to improve uptake of effective interventions such as IPTp. While community health workers’ delivery of IPTp did not increase uptake in this study, they may be effective in other settings or circumstances. Further research can help identify the health systems characteristics that are conducive to community delivery of IPTp and the operational requirements for effective implementation. Trial registration : ClinicalTrials.gov Identifier: NCT03376217. Registered December 6, 2017, https://clinicaltrials.gov/ct2/show/NCT03376217 .
 
Flow diagram of patient screening and enrollment
Frequency and time to normalization of each Transcranial Doppler Ultrasound phenotype. Only survivors are included in the time to normalization graph. TCD transcranial doppler ultrasound, MO microvascular obstruction, IPH isolated posterior circulation high flow
Representative images of Transcranial Doppler Ultrasound phenotypes in children with cerebral malaria. A Normal middle cerebral artery (MCA) TCD flow velocities and waveform for a 3-year-old child. B TCD with increased systolic flow velocity, increased diastolic flow velocity, Lindegaard ratio (LR) < 3. These findings represent a child categorized as having hyperaemia. C TCD with decreased systolic flow velocity, decreased diastolic flow velocity, decreased mean flow velocity. These findings represent a child categorized as having low flow. D TCD with normal systolic flow velocity, reduced diastolic flow velocity, increased pulsatility index. These findings represent a child categorized as having microvascular obstruction. E TCD with increased systolic flow velocity, increased diastolic flow velocity, LR > 3. These findings represent a child categorized as having cerebral vasospasm. F TCD with increased systolic flow velocity, increased diastolic flow velocity, increased mean flow velocity in the basilar artery. At the same time, all measurements in the MCAs were normal. These findings represent a child categorized as having isolated posterior circulation high flow
Physiological or pathological factors that contribute to Transcranial Doppler Ultrasound (TCD) flow velocity or waveform alterations
Article
Background: Cerebral malaria (CM) results in significant paediatric death and neurodisability in sub-Saharan Africa. Several different alterations to typical Transcranial Doppler Ultrasound (TCD) flow velocities and waveforms in CM have been described, but mechanistic contributors to these abnormalities are unknown. If identified, targeted, TCD-guided adjunctive therapy in CM may improve outcomes. Methods: This was a prospective, observational study of children 6 months to 12 years with CM in Blantyre, Malawi recruited between January 2018 and June 2021. Medical history, physical examination, laboratory analysis, electroencephalogram, and magnetic resonance imaging were undertaken on presentation. Admission TCD results determined phenotypic grouping following a priori definitions. Evaluation of the relationship between haemodynamic, metabolic, or intracranial perturbations that lead to these observed phenotypes in other diseases was undertaken. Neurological outcomes at hospital discharge were evaluated using the Paediatric Cerebral Performance Categorization (PCPC) score. Results: One hundred seventy-four patients were enrolled. Seven (4%) had a normal TCD examination, 57 (33%) met criteria for hyperaemia, 50 (29%) for low flow, 14 (8%) for microvascular obstruction, 11 (6%) for vasospasm, and 35 (20%) for isolated posterior circulation high flow. A lower cardiac index (CI) and higher systemic vascular resistive index (SVRI) were present in those with low flow than other groups (p < 0.003), though these values are normal for age (CI 4.4 [3.7,5] l/min/m2, SVRI 1552 [1197,1961] dscm-5m2). Other parameters were largely not significantly different between phenotypes. Overall, 118 children (68%) had a good neurological outcome. Twenty-three (13%) died, and 33 (19%) had neurological deficits. Outcomes were best for participants with hyperaemia and isolated posterior high flow (PCPC 1-2 in 77 and 89% respectively). Participants with low flow had the least likelihood of a good outcome (PCPC 1-2 in 42%) (p < 0.001). Cerebral autoregulation was significantly better in children with good outcome (transient hyperemic response ratio (THRR) 1.12 [1.04,1.2]) compared to a poor outcome (THRR 1.05 [0.98,1.02], p = 0.05). Conclusions: Common pathophysiological mechanisms leading to TCD phenotypes in non-malarial illness are not causative in children with CM. Alternative mechanistic contributors, including mechanical factors of the cerebrovasculature and biologically active regulators of vascular tone should be explored.
 
Plasmodium falciparum genetic diversity is high in asymptomatic infections. A Distribution of COI in asymptomatic and first-febrile infections. The COI of majority of asymptomatic samples was distributed around 2 while for first-febrile infections it was distributed around 1. B Proportion of individuals with different number of P. falciparum genotypes per every 2 years in asymptomatic infections and C in first-febrile infections. In B and C, n represents the total number of successfully genotyped samples. While the various colours represent the different number of clones: dark grey (1), orange (2), light blue (3), green (4), yellow (5), blue (6), brown (7), pink (8) and red (10)
Article
Background High levels of genetic diversity are common characteristics of Plasmodium falciparum parasite populations in high malaria transmission regions. There has been a decline in malaria transmission intensity over 12 years of surveillance in the community in Kilifi, Kenya. This study sought to investigate whether there was a corresponding reduction in P. falciparum genetic diversity, using msp2 as a genetic marker. Methods Blood samples were obtained from children (< 15 years) enrolled into a cohort with active weekly surveillance between 2007 and 2018 in Kilifi, Kenya. Asymptomatic infections were defined during the annual cross-sectional blood survey and the first-febrile malaria episode was detected during the weekly follow-up. Parasite DNA was extracted and successfully genotyped using allele-specific nested polymerase chain reactions for msp2 and capillary electrophoresis fragment analysis. Results Based on cross-sectional surveys conducted in 2007–2018, there was a significant reduction in malaria prevalence (16.2–5.5%: P-value < 0.001), however msp2 genetic diversity remained high. A high heterozygosity index (He) (> 0.95) was observed in both asymptomatic infections and febrile malaria over time. About 281 (68.5%) asymptomatic infections were polyclonal (> 2 variants per infection) compared to 46 (56%) polyclonal first-febrile infections. There was significant difference in complexity of infection (COI) between asymptomatic 2.3 [95% confidence interval (CI) 2.2–2.5] and febrile infections 2.0 (95% CI 1.7–2.3) (P = 0.016). Majority of asymptomatic infections (44.2%) carried mixed alleles (i.e., both FC27 and IC/3D7), while FC27 alleles were more frequent (53.3%) among the first-febrile infections. Conclusions Plasmodium falciparum infections in Kilifi are still highly diverse and polyclonal, despite the reduction in malaria transmission in the community.
 
Factors associated with malaria among 15-19 years-old adolescents
Article
Background In Senegal, malaria morbidity has sharply decreased over these past years. However, malaria epidemiology remains heterogeneous with persistent transmission in the southeastern part of the country and many cases among older children and adolescents. Little is known about factors associated with clinical malaria among this group. A better understanding of malaria transmission among this newly emerging vulnerable group will guide future interventions targeting this population group. This study aimed to identify factors associated with clinical malaria among adolescents in Senegal. Methods A case–control study was conducted from November to December 2020 in four health posts located in the Saraya district. Cases were defined as adolescents (10–19 years) with an uncomplicated malaria episode with fever (temperature > 37.5°) or a history of fever and positive malaria rapid diagnostic test (RDT). Controls were from the same age group, living in the neighbourhood of the case, presenting a negative RDT. A standardized, pre-tested questionnaire was administered to each study participant followed by a home visit to assess the participant's living conditions. Factors associated with clinical malaria were assessed using stepwise logistic regression analysis. Results In total, 492 individuals were recruited (246 cases and 246 controls). In a multivariate analysis, factors associated with clinical malaria included non-use of long-lasting insecticidal net (LLIN) (aOR = 2.65; 95% CI 1.58–4.45), non-use of other preventive measures (aOR = 2.51; 95% CI 1.53–4.11) and indoor sleeping (aOR = 3.22; 95% CI 1.66–6.23). Protective factors included 15–19 years of age (aOR = 0.38; 95% CI 0.23–0.62), absence of stagnant water around the house (aOR = 0.27; 95% CI 0.16–0.44), having a female as head of household (aOR = 0.47; 95% CI 0.25–0.90), occupation such as apprentice (OR = 0.24; 95% CI 0.11–0.52). Conclusions The study revealed that environmental factors and non-use of malaria preventive measures are the main determinants of malaria transmission among adolescents living in areas with persistent malaria transmission in Senegal. Strategies aimed at improving disease awareness and access to healthcare interventions, such as LLINs, are needed to improve malaria control and prevention among these vulnerable groups.
 
Article
Background Antibody and cellular memory responses following vaccination are important measures of immunogenicity. These immune markers were quantified in the framework of a vaccine trial investigating the malaria vaccine candidate GMZ2. Methods Fifty Gabonese adults were vaccinated with two formulations (aluminum Alhydrogel and CAF01) of GMZ2 or a control vaccine (Verorab). Vaccine efficacy was assessed using controlled human malaria infection (CHMI) by direct venous inoculation of 3200 live Plasmodium falciparum sporozoites (PfSPZ Challenge). GMZ2-stimulated T and specific B-cell responses were estimated by flow cytometry before and after vaccination. Additionally, the antibody response against 212 P. falciparum antigens was estimated before CHMI by protein microarray. Results Frequencies of pro- and anti-inflammatory CD4 ⁺ T cells stimulated with the vaccine antigen GMZ2 as well as B cell profiles did not change after vaccination. IL-10-producing CD4 ⁺ T cells and CD20 ⁺ IgG ⁺ B cells were increased post-vaccination regardless of the intervention, thus could not be specifically attributed to any malaria vaccine regimen. In contrast, GMZ2-specific antibody response increased after the vaccination, but was not correlated to protection. Antibody responses to several P. falciparum blood and liver stage antigens (MSP1, MSP4, MSP8, PfEMP1, STARP) as well as the breadth of the malaria-specific antibody response were significantly higher in protected study participants. Conclusions In lifelong malaria exposed adults, the main marker of protection against CHMI is a broad antibody pattern recognizing multiple stages of the plasmodial life cycle. Despite vaccination with GMZ2 using a novel formulation, expansion of the GMZ2-stimulated T cells or the GMZ2-specific B cell response was limited, and the vaccine response could not be identified as a marker of protection against malaria. Trial registration PACTR; PACTR201503001038304; Registered 17 February 2015; https://pactr.samrc.ac.za/TrialDisplay.aspx?TrialID=1038
 
Availability of mRDTs in each outlet type A overall, B by region and C by area. *p = 0.01, **p < 0.01, ***p < 0.001
How HH respondents would diagnose malaria for a family member. Multiple responses were available. A Respondents stated their preferred method of diagnosis depending on whether the family member was an adult or child. Considering adults only, HH diagnostic behaviour is disaggregated by B region, C area and D HH monthly income. *p ≤ 0.05, **p ≤ 0.01, ***p < 0.001
Article
Background To avoid misuse of anti-malarials, correct diagnosis of fever prior to drug prescription is essential. Presumptive treatment in the private healthcare sector is a concern in Nigeria, where availability of affordable artemisinin-based combination therapy (ACT) is high following the implementation of subsidy schemes from 2010 to 2017. Similar subsidies have not, however, been implemented for malaria rapid diagnostic tests (RDTs). A market survey in 2018 predominantly designed to assess the ACT market in the private sector also collected data related to RDTs, results of which are presented herein. Methods A 2018 market survey consisted of (i) an outlet survey targeting private pharmacies and Proprietary and Patent Medicine Vendors (PPMVs) across different regions of Nigeria to assess supply-side market factors related to availability of RDTs (defined as having stock available for purchase at the time of the survey) and (ii) a household survey to determine demand-side factors related to knowledge of RDTs, healthcare-seeking practices and affordability. Results Availability of RDTs at the time of the survey was low in both outlet types and significantly lower in PPMVs (22.1%, 95% CI) among pharmacies versus (13.6%, 95% CI) among PPMVs (p < 0.01). Reasons for not restocking RDTs included low demand and no supply. The majority of households diagnose malaria based on experience, while one-third would visit a PPMV or pharmacy. Half of households had heard of RDTs (48.4%) and 38.6% thought they were affordable. Conclusions Low availability of RDTs among PPMVs and pharmacies may be attributed to lack of demand, supply-side issues and cost. Increasing household knowledge of RDTs may aid increasing demand, while subsidized RDTs may address supply and price issues. Addressing the deficit in RDT provision is important for targeting of ACT medicines.
 
Article
Abstract Background: To sustain high universal Long-Lasting Insecticidal Nets (LLINs) coverage, affordable nets that provide equivalent or better protection than standard LLINs, are required. Test facilities evaluating new LLINs require compliance to Good Laboratory Practice (GLP) standards to ensure the quality and integrity of test data. Following GLP principles allows for the reconstruction of activities during the conduct of a study and minimizes duplication of efficacy testing. This case study evaluated the efficacy of two LLINs: SafeNet NF® and SafeNet® LLIN. Methods: The study was conducted according to GLP principles and followed World Health Organization guidelines for evaluating LLINs. The LLINs were assessed in experimental huts against wild, pyrethroid-resistant Anopheles arabiensis mosquitoes. Nets were either unwashed or washed 20 times and artificially holed to simulate a used torn net. Blood-feeding inhibition and mortality were compared with a positive control (Interceptor® LLIN) and an untreated net. Results: Mosquito entry in the huts was reduced compared to negative control for the unwashed SafeNet NF, washed Safenet LLIN and the positive control arms. Similar exiting rates were found for all the treatment arms. Significant blood-feeding inhibition was only found for the positive control, both when washed and unwashed. All insecticide treatments induced significantly higher mortality compared to an untreated net. Compared to the positive control, the washed and unwashed SafeNet NF® resulted in similar mortality. For the SafeNet® LLINs the unwashed net had an equivalent performance, but the mortality for the washed net was significantly lower than the positive control. Internal audits of the study confirmed that all critical phases complied with Standard Operating Procedures (SOPs) and the study plan. The external audit confirmed that the study complied with GLP standards. Conclusions: SafeNet NF® and SafeNet® LLIN offered equivalent protection to the positive control (Interceptor® LLIN). However, further research is needed to investigate the durability, acceptability, and residual efficacy of these
 
Top-cited authors
Chris Drakeley
  • London School of Hygiene and Tropical Medicine
Gerry F Killeen
  • Liverpool School of Tropical Medicine
David L Smith
  • University of Washington Seattle
Robert W Snow
  • KEMRI-Wellcome Trust Research Programme
Bart GJ Knols