Malaria Journal

Malaria remains a major global health burden, with traditional control methods facing challenges such as insecticide resistance and high operational costs. Genetic biocontrol offers a promising alternative for mosquito population suppression, but its field efficacy would require assessment. This study evaluates the role that population genomic statistics can play in detecting decreases in population size in the context of a cluster randomized control trial (cRCT), investigating the response of nucleotide diversity ({pi}), Tajimas D, segregating sites, and linkage disequilibrium (LD) under both constant and seasonal demographic scenarios. We simulated 90% and 99% population declines with various degrees of between-cluster heterogeneity, and assessed the detection power of each statistic over time and number of clusters per arm. Results show that Tajimas D is highly sensitive and robust across crash severity, seasonality and heterogeneity scenarios. Segregating sites has similar power to Tajimas D when baseline data are available. We further estimated that cRCTs require approximately 3 to 5 villages per treatment arm to achieve adequate statistical power. These findings provide recommendations for genetic monitoring of vector control interventions in wild populations.

Background Malaria remains a major public health challenge globally and in Tanzania, driven by persistent Plasmodium parasite transmission, environmental variability, and socio-economic inequalities. Despite targeted control strategies, transmission remains heterogeneous and under-captured by routine surveillance. This study utilised community cross-sectional surveys (CSS) data and spatial modelling to determine household-level risk estimates and identify micro-hotspots to guide more efficient, evidence-based malaria interventions in Mainland Tanzania. Methods The CSS data used in this study were collected in 13 villages across five regions with moderate to high malaria transmission in Mainland Tanzania between July and August 2023. Individuals aged 6 months and above, residing in the study villages for 3 months and above, were enrolled after providing informed consent and tested for malaria using rapid diagnostic tests (RDTs). Socio-demographic, clinical, anthropometric, parasitological and geo-coordinates data were collected using structured electronic tools. Household-level Plasmodium parasite prevalence was modelled using Bayesian geostatistical methods implemented through Integrated Nested Laplace Approximation within a Stochastic Partial Differential Equation framework, incorporating relevant environmental covariates. Model performance was evaluated using the Watanabe-Akaike Information Criterion (WAIC). Results Bayesian models with village specific covariates consistently outperformed null models, as indicated by lower WAIC values. In Kyerwa district (Kagera region), grass cover increased the risk of Plasmodium parasite prevalence (Posterior mean (PM)=0.076; 95percent credible interval [CrI]: 0.040 -- 0.112), while altitude had a protective effect (PM = -0.002; 9percent CrI: -0.003 to -0.001), with strong sub-village clustering of malaria infection (variance=0.485; 95percent CrI [0.333 -- 0.730]). In Buhigwe district (Kigoma region), shrub cover increased the risk of Plasmodium parasite prevalence (PM=0.119; 95percent CrI: 0.029 -- 0.210) while in Ludewa (Njombe), both shrub (PM=0.512; 95% CrI: 0.066 -- 0.989) and grass (PM=0.490; 95percent CrI: 0.117 -- 0.879) increased the risk of infection, with pronounced sub-village clustering (variance=0.84; 95percent CrI: [0.38 -- 2.40] ). In Nyasa district (Ruvuma), shrub cover had a modest positive effect (PM=0.070; 95percent CrI: 0.005 -- 0.135), in Muheza district (Tanga region), its effect was influential (PM=0.160; 95percent CrI: 0.056 -- 0.266). Risk maps revealed fine scale heterogeneity in the household level risk of Plasmodium parasite prevalence. Conclusion There was pronounced micro-scale heterogeneity in Plasmodium transmission across the study districts, driven by localised environmental factors and strong spatial dependence. Altitude had a protective effect, while vegetation cover increased the risk of infection. Geostatistical models effectively identified household-level hotspots, highlighting the limitations of aggregated surveillance, emphasising the need for locally precision-guided malaria control strategies to improve intervention efficiency and enhance the ongoing elimination strategies.

The intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) remains the main strategy to prevent malaria in pregnancy. However, continued drug pressure may also contribute to the emergence of resistant parasites and impact the gametocyte carriage and subsequent infectiousness. Pregnant women are thought to be a potential reservoir for malaria transmission due to the increased carriage of gametocytes following long-lasting infections. We used molecular methods to examine 100 Plasmodium falciparum (P. falciparum) isolates collected from Mozambican women at delivery in 2014-15, to determine SP resistance polymorphisms in P. falciparum dihydrofolate reductase (pfdhfr) and dihydropteroate synthetase (pfdhps) genes as well as the presence of gametocytes by RT-qPCR. Overall, 54% and 7% of parasites harbored quintuple and sextuple pfdhfr/pfdhps mutant haplotypes, respectively. Gametocytes were detected in 34% of isolates. Gametocyte carriage was significantly associated with quintuple mutant infections (AOR = 7.5, p = 0.001), which accounted for 80% of infections with detectable gametocytes. Results indicate the relevance of ongoing surveillance of SP resistance in Mozambique to guide future evaluation of alternative IPTp approaches as resistance levels evolve and to anticipate potential implications for parasite transmission and maternal-fetal health.

Plasmodium falciparum merozoites invade erythrocytes using various ligand-receptor interactions. Important ligands encoded by the eba and Rh gene families have varying expression levels in different parasite isolates, affecting their vaccine candidacy. Analyses of clinical isolates from endemic areas in Africa have indicated that most variation in these expression profiles exists within each local area, and only minor differences are seen between areas, although comparisons with non-African populations have not previously been performed. To enable this, relative transcript levels of three eba genes and five Rh genes have been analysed in new population samples, Malaysian isolates sampled from Sabah State in Borneo prior to endemic malaria elimination, and Gambian isolates, cultured under the same conditions to harvest schizonts for reverse transcription quantitative PCR assays. Significant differences between these populations were seen for three of the ligand genes, levels of eba175 being higher in Malaysia, while levels of eba181 and Rh2b were lower in Malaysia. The gene transcript profiles did not differ between single genotype and or multiple-genotype isolates. The distinctness of the Malaysian population expression profile was also supported by comparing previous data on clinical isolates from Ghana. In tests for correlation with previously determined parasite multiplication rates, eba181 transcript levels correlated positively among Malaysian isolates but not among Gambian isolates. These findings suggest that expression of three P. falciparum merozoite ligands involved in invasion may be regionally differentiated, and further analysis of Asian parasite populations would be important if vaccines based on these candidates are to be considered for future use.

BackgroundIn chronic asymptomatic Plasmodium vivax infections, the spleen accounts for more than 98% of total-body parasite biomass. Whether this splenic tropism also exists in acute infection and how the spleen influences pathogenesis have not been systematically explored. Materials and MethodsIn Papua, Indonesia, we compared plasma levels of P. vivax lactate dehydrogenase [PvLDH]) and circulating parasitemia in 24 spleen-intact and 25 previously splenectomized patients with acute uncomplicated vivax malaria. Clinical and hematology data were collected and plasma markers of intravascular hemolysis (cell-free hemoglobin [CFHb]), endothelial activation (angiopoietin-2), inflammation (interleukin [IL]-1 beta, IL-6, IL-18, IL-10, tumor necrosis factor-alpha) and neutrophil activation (elastase) were measured by ELISA. Giemsa-based histology in one spleen from an untreated patient splenectomized for trauma during an episode of acute vivax malaria enabled direct assessment of splenic and circulating parasitemia and biomass microscopically. ResultsCirculating parasitemia was 4-times higher in splenectomized compared to spleen-intact patients (median 21,100 vs 4,820 parasites/{micro}L, p=0.0002) but total-body P. vivax biomass (PvLDH) was 3-times lower in patients without a spleen (median 721 vs 2,140 ng/mL, p=0.026). Parasite staging and greater organ-specific symptoms suggest redistribution of parasites in the absence of a spleen. Linear regression modeling, adjusting for circulating parasitemia, patient age, sex and duration of fever, demonstrated an 8.1-fold higher PvLDH concentration in spleen-intact patients (95% confidence interval [CI]: 3.4-19.5-fold, p<0.0001), indicating a splenic biomass accounting for 89% (95%CI: 77.3-95.1%) of total-body parasites. Histopathology revealed a spleen-to-blood biomass ratio of 10.7, in-line with the PvLDH-based estimate. In spleen-intact patients, splenic P. vivax biomass correlated strongly with markers of disease intensity, endothelial activation and systemic inflammation, whereas circulating parasitemia correlated weakly or not at all. Compared to spleen-intact patients, CFHb, endothelial activation and systemic inflammation were higher in splenectomized patients while inflammasome-dependent responses were lower. ConclusionsP. vivax is predominantly an infection of the spleen, even in acute clinical vivax malaria. We conservatively estimate that 89% of total-body parasite biomass in acute infection is splenic. While the size of this hidden population correlates with disease intensity, the spleen likely regulates inflammatory pathways and heme-associated pathology.

BackgroundThe World Health Organization (WHO) has raised concerns over increasing Pfhrp2/3 deletions, undermining the sensitivity of Pfhrp2-based rapid diagnostic tests (RDTs). Close monitoring of the population and a change in diagnostic methods are recommended if the prevalence of parasites with Pfhrp2/3 deletions exceeds 5%. In high transmission settings, accurate estimates are hampered by the frequent occurrence of mixed-clone infections (multiplicity of infection; MOI). Objective and MethodsIf parasites with and without deletions are present in an infection, standard molecular assays cannot detect the presence of the former. To accurately estimate frequencies of haplotypes with Pfhrp2/3 deletions in the presence of mixed infections, a novel statistical model that combines genetic/molecular information from Pfhrp2/3 with that from neutral markers is introduced. Maximum-likelihood estimates (MLEs) are obtained for haplotype frequencies characterized by markers at Phrp2/3 loci and loci for neutral markers. The expectation-maximization algorithm is used to derive the MLEs. The adequacy of the method (precision and accuracy) is assessed by numerical simulations. ResultsThe method was applied to an active surveillance study conducted in a tribal community in Jagdalpur, India, which enrolled febrile community members (n = 432) between October and November 2021. Four markers each at Pfhrp2 and Pfhrp3 are combined with one marker each at Pfmsp1 (which encodes P. falciparum merozoite surface protein 1) and Pfmsp2. Data from a total of 117 patients who had both P. falciparum infections and genetic information for the molecular markers underwent further analysis with the novel statistical method. ConclusionResults indicate that this novel method has promising statistical properties (asymptotic and in finite samples) and can be readily applied to real-world situations. A stable implementation of the method in R is provided. This novel approach enables accurate estimation of Pfhrp2/3 deletion frequencies in complex P. falciparum infections, addressing a key limitation of current molecular surveillance methods. Author summaryPlasmodium falciparum (Pf) causes the most severe form of human malaria, accounting for over 90% of cases. Rapid diagnostic tests (RDTs) have become a cornerstone of malaria control. These RDTs detect Pf-specific antigens in a blood drop. HRP2/3 emerged as the best antigen for such tests because it is Pf-specific and expressed in abundance. However, some parasites lack the genes that code for HRP2/3 proteins. If parasites in an infection have such gene deletions, RDT results can be false negative. The WHO considers the containment of such deletions a public health priority and recommends monitoring their prevalence. The detection of HRP deletions is challenging if parasites with and without deletions co-occur in infections because standard molecular assays cannot detect deletions in this situation. To overcome this challenge, we introduce a novel statistical method to estimate the frequency distribution of parasite variants with deletions. The method combines information from neutral molecular markers and from HRP-related markers to correct for unobservable information. Here we provide a derivation of the statistical model, a stable implementation, and test its statistical properties with synthetic and real data, thereby showing that our method is well-suited for the underlying problem.

Entomological surveillance plays a crucial role in areas where malaria remains endemic, yet gathering data on mosquito populations is often expensive and complicated, particularly in remote locations with challenging logistics and inconsistent sampling schedules. Access to extensive time series data on mosquito species at specific sites would greatly enhance insights into seasonal trends and the biting habits of vectors of malaria parasites. Gaps in mosquito count records pose a significant challenge for researchers and public health officials seeking to establish early warning systems and effective vector control programs. In this study, we apply quantitative machine learning techniques to address missing data in estimates of mosquito abundance collected from 2009 to 2016 in Bolivar State, Venezuela. We evaluated Linear Regression, Stochastic Linear Regression, K Nearest-Neighbor, and Gradient Boosting methods for imputing missing counts of Anopheles mosquitoes, employing a leave-one-out cross-validation strategy. Additionally, we developed a predictive malaria transmission model incorporating mosquito abundance and climate variables (El Nino 3.4 Index, rainfall, and mean air temperature) as covariates. Our generalized time series model forecasts malaria incidence of Plasmodium vivax and Plasmodium falciparum based on climate dynamics and imputed mosquito data. Model performance was assessed using root mean square error, mean absolute error, and mean absolute percentage error. The final results demonstrated that machine learning imputation significantly improved the accuracy and reliability of P. vivax malaria incidence predictions but failed to predict P. falciparum incidence. The study demonstrates that method choice significantly influences the reconstruction of seasonal abundance patterns and the performance of malaria incidence models. Nevertheless, the proposed models strengthen the foundation for targeted interventions and surveillance in endemic regions. Despite limitations in data continuity and coverage, the findings highlight the value of combining multiyear entomological data sets with robust imputation and sensitivity analyses to improve predictive modeling in resource-constrained, malaria-endemic settings.

Rwanda is a malaria endemic country and a focal point for emerging Plasmodium falciparum artemisinin partial resistance (ART-R). While Demographic and Health Surveys (DHS) provide both national and province-level representative data, malaria testing in Rwandan DHS (RDHS) studies has been limited to a subset of adult women and children under 5 years using RDT and/or microscopy. Recent work using ultra-sensitive quantitative real time PCR on residual dried blood spots (DBS) from the 2014-15 RDHS revealed a significantly higher P. falciparum prevalence than detected by standard DHS diagnostics. Building on this study, we analyzed 7,127 adult DBS samples collected for HIV testing in the 2019-20 RDHS to generate updated prevalence measures. We found a national P. falciparum infection prevalence of 7.7% (95%CI [6.8%, 8.7%]), with predominantly low-density infections (median parasitemia: 7.3 parasites/uL). We assessed covariates of P. falciparum malaria infection, identifying male sex, lower household wealth, lower educational achievement, and residence at lower elevation as significant predictors. Notably, national P. falciparum prevalence decreased 53% relative to the parallel 2014-15 RDHS study, despite reports of increasing ART-R-associated mutations in Rwanda. These findings demonstrate the utility of ultra-sensitive molecular surveillance, and suggest that national malaria control efforts have substantially reduced malaria burden in Rwanda even amid rising antimalarial parasite prevalence. Subsequent studies on this data set will provide measures of minor Plasmodium species prevalence, as well as temporospatial analysis of antimalarial resistance markers in P. falciparum positive samples.

Severe malaria disproportionately affects children during their earliest Plasmodium falciparum infections, when immunopathology rather than parasite burden often drives clinical deterioration. Because direct investigation of host-parasite interactions during severe disease is ethically impossible, we developed a two-dimensional ex vivo co-culture system that recapitulates key physiological features of malaria pathogenesis. Using PBMCs from malaria-naive and malaria-exposed adults co-cultured with a freshly adapted P. falciparum isolate, we modelled the combined effects of febrile temperature, pipecolic acid (PA), and lysophosphatidylcholine (LPC) depletion on IL-6 secretion. We also integrated clinical data from children with severe malaria in Anambra State, Nigeria. Across conditions, IL-6 output was not driven by temperature alone but by a metabolically gated interaction: febrile temperature amplified IL-6 only when PA was present, and LPC was not limiting. LPC depletion suppressed IL-6 to near-baseline levels regardless of temperature or PA, indicating that lipid availability constrains inflammatory signalling. Clinical data showed that adverse outcomes clustered with markers of multi-organ dysfunction. Together, these findings support a model in which IL-6 is a context-dependent mediator - participating in inflammatory pathways but not acting as a singular causal driver - and in which metabolic stress, febrile cues, and host tolerance mechanisms jointly shape cytokine production. Ongoing bioinformatics analysis will define the transcriptional responses of both parasite and host cells under these malaria-relevant conditions.

Plasmodium vivax transmission from humans to mosquitoes depends on the density of gametocytes that in turn depends on asexual parasite replication and gametocyte commitment. These processes are often analyzed separately, despite being biologically linked and measured with substantial uncertainty. We used a joint Bayesian latent-variable model to simultaneously analyze parasite density, gametocyte density, and mosquito infectivity while accounting for measurement error and propagating uncertainty across linked processes. The model was applied to individual-level data from three P. vivax transmission studies conducted in Ethiopia (n = 455). A tenfold increase in gametocyte density was associated with more than a twofold increase in the odds of mosquito infection (odds ratio [OR] = 2.32, 95% credible interval [CrI]: 2.12-2.54). Asexual parasite density was also positively associated with infectivity after accounting for gametocyte density (OR = 1.74, 95% CrI: 1.60-1.90), and inclusion of parasite density improved predictive performance. Pathway decomposition within the joint model indicated that approximately 41% of the parasite-infectivity association operated through gametocyte density. Increasing age was associated with lower asexual parasite density but higher gametocyte density, resulting in minimal overall association with infectivity. Predicted infection probability increased sigmoidally with gametocyte density, remaining low at lower densities before increasing sharply and approaching a plateau at higher densities. Gametocyte density produced the largest predicted changes in the proportion of infected mosquitoes, while asexual parasite density added predictive information not fully captured by measured gametocyte density alone. This approach links molecular parasite measurements with mosquito infection risk while accounting for measurement uncertainty and provides an interpretable framework for studying the P. vivax infectious reservoir. Author SummaryMalaria transmission occurs when mosquitoes ingest sexual-stage parasites, called gametocytes, during a blood meal. In Plasmodium vivax infections, human-to-mosquito transmission depends on linked biological stages, including asexual parasite replication, gametocyte production, and mosquito infection. These processes are closely connected and often measured with uncertainty, making them difficult to study using standard approaches that analyze them separately. In this study, we applied a joint Bayesian model that analyzes parasite density, gametocyte density, and mosquito infectivity together while accounting for uncertainty in laboratory measurements. Using data from three studies in Ethiopia, we quantified how parasite density, gametocyte density, and host characteristics relate to mosquito infection. The analysis showed that measured gametocyte density alone did not fully explain variation in infectivity, and that asexual parasite density provided additional predictive information. We also found that age was associated differently with asexual parasite and gametocyte densities, resulting in little overall association with infectivity. This approach helps link molecular parasite measurements with transmission outcomes and improves understanding of the P. vivax infectious reservoir in endemic settings.

This study evaluated the effects of different Arachis hypogaea dietary preparations on parasite load, haematological indices, and physiological responses in Plasmodium berghei-infected mice. Forty-five albino mice were randomly assigned to five groups: normal control, infected untreated control, roasted groundnut, boiled groundnut, and a combination of roasted and boiled groundnut diets. Data were analyzed using one-way ANOVA at p > 0.05. Infection resulted in a high parasite load in the untreated group, with no significant difference compared to the boiled and combined diet groups. However, the roasted groundnut group showed a reduction in parasite load and relatively higher chemosuppressive activity, although differences were not statistically significant. White blood cell counts increased significantly following infection, and dietary treatments did not restore normal levels. Similarly, red blood cell counts and packed cell volume were significantly reduced in infected mice. The roasted groundnut diet moderately improved PCV compared to other treatments but did not restore it to normal levels. Weight loss was most pronounced in untreated mice, while roasted groundnut intake showed slight mitigation. No significant effects on temperature regulation were observed. Overall, A. hypogaea diets did not significantly improve parasitemia or haematological parameters, indicating limited therapeutic value in malaria management. ImportanceThis study is of significant importance due to its contribution to the ongoing search for accessible, affordable, and nutritionally based supportive interventions in malaria management. Malaria remains a major public health burden, particularly in sub-Saharan Africa, where increasing resistance of Plasmodium species to conventional antimalarial drugs continues to undermine control efforts. By investigating the effects of Arachis hypogaea (groundnut), a widely consumed and locally available food resource, this research explores a practical dietary approach that could complement existing malaria treatment strategies. In summary, this research is important because it bridges nutrition, parasitology, and public health, offering practical insights that could inform both scientific advancement and real-world malaria management strategies.

Background Malaria rapid diagnostic tests (RDTs) are widely used to detect and treat malaria infections, yet a diagnostic gap remains. With turnaround times of ~15 minutes, RDTs may be too slow to enable broad-scale implementation in certain contexts. Novel non-invasive diagnostics (NIDs) have potential to provide faster (<5 minutes), sensitive (90% for symptomatic, 65% for asymptomatic carriage), and cost-effective alternatives, which may increase testing throughput, enhance case detection, guide appropriate antimicrobial use, and reduce waste by using fewer consumables. Their potential impact has yet to be investigated. Methods We modeled a country-agnostic population of 10 million individuals to assess the impact of population-level scale-up of four malaria testing strategies for active case-finding: 1) current practice (50% syndromic diagnosis and 50% RDTs), 2) full RDT scale-up, 3) full NID implementation, and 4) NID screening plus confirmatory RDT, using a decision-tree model of the malaria diagnostic and care cascade. We varied prevalence (0.02-0.25) and proportion of cases with symptoms (0.05-0.60) to evaluate strategy performance across epidemiological contexts. We investigated case detection rates, antimicrobial use, incremental cost-effectiveness ratios (ICERs) per disability adjusted life year (DALY) averted, net positive treatment outcomes, and threshold performance levels at which an NID would outperform RDTs. Results Full NID implementation (strategy 3) yielded the highest case detection rates (up to 85%), followed by strategies 2, 4, and 1 (45%, 38%, 36% respectively). NID-based methods (strategies 3 and 4) saved costs and RDT scale-up was cost-effective at averting DALYs compared to current practice (ICERs: $60-1,270). Despite high case detection, universal NID testing spiked unnecessary antimicrobial use. Overall, our results suggest that an NID with 55% asymptomatic sensitivity and 84% specificity, followed by RDT confirmation (strategy 4), could simultaneously improve case detection, reduce antimicrobial overuse, and limit costs. Conclusions This modeling analysis suggests that NIDs can sustainably optimize malaria case detection in symptomatic and asymptomatic cases and reduce costs, potentially making them a valuable addition to the diagnostic toolbox. When paired with confirmatory RDTs, they could help reduce inappropriate antimicrobial use, supporting drug efficacy amid rising resistance. Further research should assess their real-world utility, feasibility, and scalability for malaria surveillance and elimination efforts.

Anopheles albimanus (Nyssorhynchus) is featured as the main malaria vector on Hispaniola. However, five other Anopheles species have been reported circulating in the area; four of them belonging to the subgenus Anopheles (An. crucians, An. grabhamii, An. pseudopunctipennis, and An. vestitipennis) and another one to the Nyssorhynchus subgenus (An. argyritarsis). Previous studies on mosquitoes in the genus Anopheles have identified and characterized peptides from immunogenic salivary proteins, with several of these peptides being unique to the Nyssorhynchus and Anopheles subgenera. This underscores their potential use as biomarkers for differentiating exposure to Anopheles mosquitoes in both the Old World and New World. Since both Nyssorhynchus and Anopheles subgenera have been reported in Haiti, a series of ELISAs were conducted to quantify IgG antibody titers against three published antigenic anopheline salivary peptides (gSG6-P1, Peroxi-P3, and Apy-2) in 348 participants registered in Haitis multi-partner/multidisciplinary Malaria Zero Program. This study aimed to evaluate the intensity of human-vector contact with Anopheles from both subgenera in Grand Anse, Haiti. In addition, the study measured antibodies against a panel of Plasmodium falciparum antigens to determine any association between anti-parasite and anti-peptide antibodies. Significantly elevated IgG responses to Peroxi-P3 in comparison to Apy2 and gSG6-P1 in the total study population (p < 0.001) were observed. Additionally, immune responses to Peroxi-P3 and gSG6-P1 differed significantly between [&le;]18-year-olds and >18-year-olds (p = 0.004 and p = 0.002), whereas no sex-based differences were observed for any peptide. Correlation analyses observed a greater number of significant positive associations in immune response between gSG6-P1 and Plasmodium antigens than any other salivary peptide, an occurrence which was more pronounced in [&le;]18-year-olds than >18-year-olds. A marked reduction in IgG responses to Apy2 and Peroxi-P3, but not gSG6-P1, among participants who kept a single household animal species compared with those who owned two or more species or those who did not have household animals was also demonstrated. Spatial analysis revealed heterogenous geographic overlap of high antibody responses among Peroxi-P3, Apy2, and gSG6-P1, alongside geographically overlapping clusters of low antibody responses to Peroxi-P3 and Apy2. These results provide additional data on the utility of anopheline salivary peptides to characterize human-vector-parasite exposure dynamics in low-transmission areas, such as Haiti.

Gene drives are genetic elements that bias their own inheritance to spread desired traits in target populations, enabling population modification or suppression. Although homing-based drives can propagate efficiently, their potential for uncontrolled spread may present a challenge for field deployment. Thus, confined drive systems are needed. Here, we developed a confined modification drive, called Toxin-Antidote Recessive Embryo (TARE) drive, in the globally important malaria vector Anopheles stephensi. This drive works by cleaving and disrupting wild-type alleles in the germline or early embryo from maternally deposited Cas9. Disrupted alleles are recessive lethal, thus increasing the drive in a frequency-dependent manner. Inheritance bias was moderate in crosses between drive heterozygote mosquitoes, possibly due to low gRNA activity and thus moderate germline cleavage rates. Single-release cage trials confirmed the TARE drives ability to spread, although the drive ultimately declined due to fitness costs and resistance alleles associated with repetitive elements. Nonetheless our modelling analysis indicate that this TARE system could achieve population spread if the resistance issue is addressed. These findings demonstrate a functional prototype TARE drive in Anopheles stephensi and highlight key parameters governing its performance. Minor design optimizations could substantially improve efficiency and integrity, enabling rapid but confined population modification.

Background Malaria remains a critical global health challenge, with over 68% of Ethiopias population living in at-risk areas. While Long-Lasting Insecticidal Nets (LLINs) are a cornerstone of prevention, their effectiveness depends on consistent use. This study aimed to assess LLIN ownership and utilization patterns and identify socio-behavioral and physical determinants of their effectiveness in endemic communities. Methods A community-based, cross-sectional survey was conducted from October 2024 to January 2025 across 11 administrative regions in Ethiopia. Using a two-stage stratified cluster sampling technique, data were collected from 9,222 households (34,427 individuals) through face-to-face interviews and direct physical observations. Data analysis was performed using the SPSS Complex Samples module and hierarchical multivariable logistic regression. Results The survey found a household LLIN ownership rate of 71.5%, while the proportion of sufficient LLINs for every two people was 58.3%. Among those who owned nets, the overall utilization rate was 59.9%, with significantly higher rates in rural areas (72.7%) than in urban areas. Vulnerable groups achieved higher usage levels, specifically pregnant women (78.5%) and children under five (67.2%). Multivariable analysis indicated that age and pregnancy status were the strongest predictors of LLIN use, with ORs of 0.258 (p < 0.001) and 0.662 (p < 0.001), respectively. Major barriers identified included a 60.5% lack of confidence in hanging nets (p < 0.001) and a widespread misconception (64.1%) that malaria risk is restricted to the rainy season. Conclusion Although Ethiopia has made strides in LLIN ownership and prioritized protection for vulnerable demographics, overall utilization remains below the 80% threshold required for community-wide protection. To bridge the gap between ownership and consistent use, national strategies should transition toward skill-based interventions and targeted communication to address practical barriers and seasonal misconceptions.

Background: Malaria remains a major global health threat, with 249 million cases and 609,000 deaths reported in 2022. The Ashanti Region of Ghana bears a disproportionate burden, with severe malaria accounting for 24% of hospital admissions in 2021, above the national average of 19%. Despite intensified control efforts, inpatient mortality patterns remain poorly understood. This study identifies key determinants of severe malaria mortality among hospitalized patients in the Ashanti Region. Methods: We analyzed inpatient surveillance data from the District Health Information Management System 2 (DHIMS2) for severe malaria admissions from 2018 to 2022. Descriptive statistics, bivariate analyses with robust survey estimation (accounting for design effects), and multivariable Firth penalized logistic regression were used to identify mortality predictors. Survey-adjusted logistic regression served as a sensitivity analysis to validate findings. Results: Among 54,544 severe malaria admissions, females comprised 51.1% and children under five 39.4%. The case fatality rate was 0.4% (200 deaths). Mortality was significantly associated with age, occupation, insurance status, facility ownership, admitting department, length of stay, and comorbidities. Males had 1.4 times higher mortality odds than females. Compared to children under five, patients aged 5 to 17 years had 44% lower odds of mortality (aOR = 0.56, 95% CI: 0.33, 0.94). Active NHIS membership had lower mortality odds by 67% (aOR=0.33, 95% CI: 0.25, 0.45) compared to inactive membership. Admissions to faith-based facilities showed lower mortality odds (aOR=0.38, 95% CI: 0.23, 0.65) than government facilities, while medical wards had higher odds (aOR=2.38, 95% CI: 1.48, 3.84) than paediatric wards. Stays of 3 to 5 days were associated with lower mortality odds (aOR=0.67, 95% CI: 0.47, 0.97) compared to stays <3 days. Those with comorbidities had twice the mortality odds versus those without. Sensitivity analysis confirmed consistent direction and significance. Conclusion: Age, comorbidities, insurance coverage, facility type, and admission practices strongly influence severe malaria mortality in Ashanti. Strengthening NHIS enrollment, extending inpatient monitoring beyond three days, and adopting best practices from paediatric and faith based facilities could improve survival. Integrating comorbidity screening and management into malaria protocols is essential to reducing preventable deaths.

BackgroundInsecticides remain the cornerstone of mosquito vector control for malaria, dengue, and other mosquito-borne diseases, yet global patterns of deployment and their socioeconomic and environmental drivers are poorly characterized. Understanding where and why insecticides are used is essential for better targeting control efforts and ensuring they are effective, equitable, and efficient. MethodsWe analyzed annual country-level insecticide-use data from 122 countries (1990-2019), reported as standard spray coverage for insecticide-treated nets (ITNs), residual spraying (RS), spatial spraying (SS), and larviciding (LA). Generalized linear mixed models and hurdle models quantified associations between deployment and disease incidence, human development index (HDI), human population density, temperature, and precipitation. Models were evaluated using repeated cross-validation and applied to generate downscaled predictions of insecticide use at subnational administrative region level 2 (ADM2) globally. FindingsInsecticide deployment increased with malaria and dengue incidence, but this response was substantially stronger in higher-HDI countries, indicating that deployment depends on socioeconomic capacity as well as disease burden that leads to weaker scaling in lower-resource settings. Intervention types exhibited distinct patterns; ITN use tracked malaria burden, whereas infrastructure-intensive approaches (e.g., RS and SS) were concentrated in higher-HDI settings and increased with Aedes-borne disease incidence. Downscaled ADM2-level maps uncovered substantial within-country heterogeneity that is obscured at the national scale, highlighting regions where predicted deployment remains low relative to disease risk across sub-Saharan Africa, South Asia, and parts of Latin America. InterpretationGlobal insecticide deployment reflects not only epidemiological need but also economic and logistical capacity, creating mismatches between risk and control. High-resolution mapping can support more equitable allocation of interventions, guide insecticide resistance stewardship, and improve strategic planning as climate and urbanization reshape mosquito-borne disease risk.

Abstract Background: To influence malaria-related behaviours, it is important to understand key behavioural drivers, encourage enablers and address barriers to individuals and communities adoption of interventions to prevent malaria. The capability(C), opportunity(O), and motivation(M) Behaviour(B) model (COM-B model) was used to inform development of perennial malaria chemoprevention (PMC) social, and behaviour change (SBC) message delivered through routine immunization (RI) platform. This paper presents how the COM-B model was used for designing the SBC messages for PMC using the findings from a qualitative study. Methodology The COM-B model provided the theoretical framework for designing the PMC SBC intervention by identifying, capability, opportunity motivation for PMC as well as the barriers, and possible enablers for PMC uptake. A qualitative study was conducted as key source of information. Twelve focus group discussions (FGDs) were conducted with the target audience comprising of mothers of children under two years, pregnant women, men, ward development committee members, community mobilizers and health workers. A total of 120 people participated in the study. An SBC workshop was conducted to develop key messages and content for a community dialogue flipbook and facilitators' guide. Results Knowledge of malaria signs that prompt mothers to seek health care for their children as well as awareness about malaria prevalence and severity, were identified as capabilities that could drive behaviour change, while forgetting the time to visit the health facility was noted as a hindrance. Opportunities and social influencers included spousal support, the positive influence of health workers, accessibility and affordability of the intervention, and the availability of transportation. Motivation was shaped by the perceived seriousness of malaria as a health problem that could lead to the death of children. Fathers were motivated when they observed reduced malaria burden and improved child health, although a lack of perceived urgency remained a demotivating factor for seeking care. Mothers' motivation was strengthened by encouragement from husbands, community mobilisers and health workers. Conclusion The COM-B model provided an effective framework for identifying and developing key messages that informed changes needed to improve capability, opportunities, motivation of individuals and communities towards increased uptake of PMC during PMC pilot study in Osun state Nigeria.

Background: Partial resistance to artemisinins (ART-R) has emerged in East Africa, associated with mutations in the Plasmodium falciparum kelch13 gene. It is currently unclear whether ART-R has implications for gametocyte production or for onward transmission to mosquitoes. Methods: In a cohort of uncomplicated malaria patients attending Kalongo Hospital in northern Uganda, we quantified carriage of PfKelch13 mutant parasites by conventional sequencing and droplet digital PCR (ddPCR) for the C469Y and A675V mutations. Prevalence and density of gametocytes and ring-stage parasites were assessed by microscopy and quantitative reverse-transcriptase PCR (qRT-PCR). Lumefantrine concentrations, indicative of prior malaria treatment, were determined by ultra-high performance liquid chromatography-tandem mass spectrometry. Transmission potential of wild-type and PfKelch13 mutant parasites was assessed by mosquito feeding assays and complemented with molecular characterization of parasites in wild-caught mosquitoes from household resting catches. Findings: We enrolled 235 patients with symptomatic P. falciparum infection; PfKelch13 C469Y or A675V mutations were detected in 35.8% (78/218) of infections by sequencing and 59.1% (136/230) by ddPCR. Gametocyte carriage was 24.0% (56/233) by microscopy and 56.6% (133/235) by qRT-PCR and not associated with the abundance of PfKelch13 mutant parasites by ddPCR (p=0.603). Among a total of 227 mosquito feeds with patient whole blood, 1.4% (120/8745) of mosquitoes became infected. Mosquito infection rates were positively associated with gametocyte density ({beta} = 0.39, 95% CI = 0.23-0.59, p < 0.001) without an observed interaction with the abundance of PfKelch13 mutant parasites (p = 0.452). PfKelch13 C469Y or A675V mutations were detected in 40.1% (21/52) of malaria-infected bloodmeals of field-caught mosquitoes and in 28.0% (7/25) of sporozoite-positive mosquitoes. Interpretation: We conclude that pfkelch13 mutations are very common in patients in northern Uganda with uncomplicated malaria, mostly in multiclonal infections. We observed no evidence that ART-R affected gametocyte production or transmission to mosquitoes. Funding: Dutch Research Council (NWO)

Abstract Background In many countries, particularly in sub-Saharan Africa, medical pluralism and utilization of multiple therapeutic approaches for managing diseases, including malaria, are common. Ghana's antimalarial medicine policy has recommended herbal medications for treating uncomplicated malaria. While this is in line with complementary and alternative medicine (CAM) use for the treatment of malaria, exploring CAM for malaria prevention could be an important consideration for public health initiatives towards malaria elimination efforts. This study assessed the prevalence, perceptions, and attitudes on CAM use for malaria prevention in the general population and associated factors. Methods A community-based cross-sectional analytic survey was conducted among 3064 adult residents sampled between September and November 2023. A multi-stage sampling method was used to select participants from 18 sub-districts drawn from 6 districts and 6 regions in Ghana. Data on CAM use for malaria prevention, attitudes about CAM, perceptions about malaria, and sociodemographic factors were collected. The primary outcome was reported CAM use for malaria prevention within the 12months preceding the survey, measured as a binary outcome. Multiple logistic regression analyses were performed to identify the predictors of CAM use for malaria prevention. Results A total of 3,064 household respondents were involved in the analysis, with 51.2% (n=1,570) females and a median age of 31 years (IQR: 24-42 years). The use of CAM for malaria prevention in the last 12 months was 31.6% (95% CI: 30.0-33.3%). The most common types of CAM used included botanical/herbal medicine (21.8%), vitamin supplements (12.3%), mineral supplements (10.7%), and spiritual healing/prayers (9.6%). Increased CAM use for malaria prevention was associated with education and perceptions, such as concerns and consequences about malaria. Factors associated with decreased odds of CAM use included formal employment and having a skeptical and indifferent attitude about CAM. Conclusion Over a third of the population used CAM for malaria prevention in the last year, highlighting its role in public health. Integrating herbal medicine into prevention strategies could enhance community acceptance and help with efforts toward malaria elimination. However, further research is needed to validate clinical efficacy, establish potential drug-herb interactions, and isolate lead compounds for optimized malaria prevention therapy.