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Abstract Normal respiratory rates (RR) for children under five in the tropics are well-documented, but data for older children are limited. This study tracked RR changes with age and examined associations with nutritional status and environmental factors. We monitored rural Gambian children aged 6 months to 14 years, recording RR during home visits twice weekly over two rainy seasons. Using a generalized additive model, we constructed RR reference curves, and a linear mixed-effect model identified factors influencing RR. A total of 830 children provided 67,512 RR measurements. Their median age was 6.07 years (interquartile range 4.21–8.55) and 400 (48.2%) were female. Age, stunting, ambient temperature, and time of RR measurement were independent predictors of respiratory rate. Strikingly, children showing signs of illness had greater variability in repeat RR measurements. We constructed a RR reference chart for children aged one to 13 years and proposed a cutoff of > 26 breaths/min for raised RR among children aged > 5 years bridging an important gap in this age group. Although the time of data collection, nutritional status, and ambient temperature were predictors of RR, their effect size is not clinically significant enough to warrant a change in the current WHO guidelines owing to the prevailing uncertainty in the measurement of RR. The finding that RRs between repeat measurements were more variable among children with signs of illness suggests that a single RR measurement may be inadequate to reliably assess the status of sick children—a population in which accurate diagnosis is essential to enable targeted interventions with lifesaving treatment.

Abstract Background The housing stock of rural sub-Saharan Africa is changing rapidly. With millions of new homes required over the coming decades, there is an opportunity to protect residents by screening homes from malaria mosquitoes. This study, undertaken in the Upper River Region of The Gambia, explores local perceptions of what a good house should provide for its inhabitants and responses to living in a house that has been modified as part of a randomized control trial designed to assess whether improved housing provided additional protection against clinical malaria in children (the RooPfs trial). Methods This descriptive, exploratory study was undertaken over 22 months using mixed-methods (informal conversations, observations, focus group discussions, photovoice, and a questionnaire survey) in a parallel convergent design. Analysis was conducted across the data sets using a framework approach. Following coding, the textual data were charted by a priori and emerging themes. These themes were compared with the quantitative survey results. The nature and range of views about housing and the RooPfs study modifications and the relationships among them were identified and described. Results The data were derived from a total of 35 sets of observations and informal conversations in 10 villages, 12 discussions with the photovoice photographers, 26 focus group discussions (across 13 villages) and 391 completed questionnaires. The study participants described a ‘good house’ as one with a corrugate-metal roof, cement walls (preferably cement block, but mud block covered with cement plaster was also an acceptable and cheaper substitute) and well-fitting doors. These features align with local perceptions of a modern house that provides social status and protection from physical harms. The RooPfs modifications were largely appreciated, although poor workmanship caused concerns that houses had become insecure. However, the long-term trusting relationship with the implementing institution and the actions taken to rectify problems provided reassurance and enhanced acceptability. Conclusion In developing housing to address population needs in Africa, attention should be paid to local perceptions of what is required to make a house secure for its inhabitants, as well as providing a healthy environment.

Abstract Background In rural sub-Saharan Africa, thatch roofs are being replaced by metal roofs. Metal roofing, however, increases indoor temperatures above human comfort levels, and thus makes it more likely that residents will not use an insecticide-treated bed net (ITN) at night. Whether the colour of a metal roof affects indoor temperature and human comfort was assessed. Methods Two identical, experimental houses were constructed with metal roofs in rural Gambia. Roof types were: (1) original bare-metal, (2) painted with red oxide primer or (3) white gloss, to reflect solar radiation. Pairwise comparisons were run in six, five-night blocks during the malaria season 2018. Indoor climate was measured in each house and multivariate analysis used to compare indoor temperatures during the day and night. Results From 21.00 to 23.59 h, when most residents decide whether to use an ITN or not, the indoor temperature of a house with a bare metal roof was 31.5 °C (95% CI 31.2–31.8 °C), a red roof, 30.3 °C (95% CI 30.0–30.6) and a white roof, 29.8 °C (95% CI 29.4–30.1). During the same period, red-roofed houses were 1.23 °C cooler (95% CI 1.22–1.23) and white roofs 1.74 °C cooler (95% CI 1.70–1.79) than bare-metal roofed houses (p

Abstract Background African houses are frequently too hot and uncomfortable to use a bed net at night. Indoor thermal comfort is often evaluated by measuring temperature and humidity, ignoring ventilation. This study explored ways to measure ventilation in single-roomed rural Gambian houses during the malaria transmission season and evaluated building designs that could increase airflow at night and help keep the occupants comfortable. Methods Two identical mud-walled houses were constructed with a metal roof, three doors and closed eaves. Experiment 1 compared five methods for measuring ventilation in a building: (1) using a blower door, (2) increasing carbon dioxide (CO2) levels indoors using an artificial source of CO2 and then measuring the rate of gas decay, (3) using a similar approach with a natural source of CO2, (4) measuring the rise of CO2 when people enter a building and (5) using hot-wire anemometers. Experiment 2 used CO2 data loggers to compare ventilation in a reference metal-roofed house with closed eaves and badly-fitting doors with a similar house with (1) thatched roof and open eaves, (2) eaves tubes, (3) screened doors and (4) screened doors and windows. Results In experiment 1, CO2 data loggers placed indoors in two identical houses showed similar changes in airflow (p > 0.05) for all three methods recording either decreasing or increasing CO2. Blower doors were unable to measure airflow in houses with open eaves or screened windows and the anemometers broke down under field conditions. In experiment 2, open eaves in thatched houses, screened doors alone, and screened doors and windows increased indoor ventilation compared to the reference metal-roofed house with closed eaves and badly fitting doors (p

Abstract Background Recording behaviours that have the potential to impact health can be doubly challenging if the behaviour takes place in private spaces that cannot be observed directly, and where respondents answer what they think the recorder may want to hear. Sleeping under a long-lasting insecticidal net (LLIN) is an important intervention for malaria prevention, yet it is difficult to gauge the extent to which coverage (how many nets are in the community) differs from usage (how many people actually sleep under a net). List randomization, a novel method which partially obscures respondents’ answers to sensitive questions, was employed to estimate LLIN usage in The Gambia. Methods 802 heads-of-household from 15 villages were recruited into a randomized controlled trial assessing the effect of a housing intervention on malaria. These houses were randomly assigned to a housing intervention versus control, with stratification by village so as to ensure balance between arms. From these, 125 households (63 intervention, 52 control) were randomly selected for participation in the list randomization experiment, along with 68 households from the same villages but which were not part of the housing improvement study, resulting in a total of 196 households for the list randomization experiment. Approximately half (n = 97) of the 196 study participants were randomly assigned to the control group and received a four-question list about non-sensitive behaviours; the intervention group (n = 99) received the same list, with the addition of one question on a sensitive behaviour: whether or not they had used a bed net the previous night. Participants were read the list of questions and then said how many of the statements were true. Bed net usage was estimated by calculating the difference in means between the number of affirmative responses between the two groups. Results The mean number of affirmative responses in the control group was 2.60 of four statements (95% confidence interval, 95% CI 2.50–2.70), compared with 3.68 (95% CI 3.59–3.78) in the intervention group. Such difference (1.08; 95% CI 94.9–100%) suggests near universal bed net usage. Conclusions Bed net usage by household heads in these rural villages was found to be high. Though not entirely unexpected given other studies’ estimates of high bed net usage in the area, the list randomization method should be further validated in an area with lower coverage.

Abstract Background Long-lasting insecticidal nets (LLINs) treated with pyrethroids are the foundation of malaria control in sub-Saharan Africa. Rising pyrethroid resistance in vectors, however, has driven the development of alternative net formulations. Here the durability of polyethylene nets with a novel combination of a pyrethroid, permethrin, and the insect juvenile hormone mimic, pyriproxyfen (PPF), compared to a standard permethrin LLIN, was assessed in rural Burkina Faso. Methods A compound-randomized controlled trial was completed in two villages. In one village 326 of the PPF-permethrin nets (Olyset Duo) and 327 standard LLINs (Olyset) were distributed to assess bioefficacy. In a second village, 170 PPF-permethrin nets and 376 LLINs were distributed to assess survivorship. Nets were followed at 6-monthly intervals for 3 years. Bioefficacy was assessed by exposing permethrin-susceptible and resistant Anopheles gambiae sensu lato mosquito strains to standard World Health Organization (WHO) cone and tunnel tests with impacts on fertility measured in the resistant strain. Insecticide content was measured using high-performance liquid chromatography. LLIN survivorship was recorded with a questionnaire and assessed by comparing the physical integrity using the proportionate hole index (pHI). Results The PPF-permethrin net met WHO bioefficacy criteria (≥ 80% mortality or ≥ 95% knockdown) for the first 18 months, compared to 6 months for the standard LLIN. Mean mosquito mortality for PPF-permethrin nets, across all time points, was 8.6% (CI 2.6–14.6%) higher than the standard LLIN. Fertility rates were reduced after PPF-permethrin net exposure at 1-month post distribution, but not later. Permethrin content of both types of nets remained within the target range of 20 g/kg ± 25% for 242/248 nets tested. The pyriproxyfen content of PPF-permethrin nets declined by 54%, from 10.4 g/kg (CI 10.2–10.6) to 4.7 g/kg (CI 3.5–6.0, p

Summary: Background: In malaria-endemic areas, residents of modern houses have less malaria than those living in traditional houses. We aimed to assess whether children in The Gambia received an incremental benefit from improved housing, where current best practice of insecticide-treated nets, indoor residual spraying, seasonal malaria chemoprevention in children younger than 5 years, and prompt treatment against clinical malaria was in place. Methods: In this randomised controlled study, 800 households with traditional thatched-roofed houses were randomly selected from 91 villages in the Upper River Region of The Gambia. Within each village, equal numbers of houses were randomly allocated to the control and intervention groups using a sampling frame. Houses in the intervention group were modified with metal roofs and screened doors and windows, whereas houses in the control group received no modifications. In each group, clinical malaria in children aged 6 months to 13 years was monitored by active case detection over 2 years (2016–17). We did monthly collections from indoor light traps to estimate vector densities. Primary endpoints were the incidence of clinical malaria in study children with more than 50% of observations each year and household vector density. The trial is registered at ISRCTN02622179. Findings: In June, 2016, 785 houses had one child each recruited into the study (398 in unmodified houses and 402 in modified houses). 26 children in unmodified houses and 28 children in modified houses did not have at least 50% of visits in a year and so were excluded from analysis. 38 children in unmodified houses were recruited after study commencement, as were 21 children in modified houses, meaning 410 children in unmodified houses and 395 in modified houses were included in the parasitological analyses. At the end of the study, 659 (94%) of 702 children were reported to have slept under an insecticide-treated net; 662 (88%) of 755 children lived in houses that received indoor residual spraying; and 151 (90%) of 168 children younger than 5 years had seasonal malaria chemoprevention. Incidence of clinical malaria was 0·12 episodes per child-year in children in the unmodified houses and 0·20 episodes per child-year in the modified houses (unadjusted incidence rate ratio [RR] 1·68 [95% CI 1·11–2·55], p=0·014). Household vector density was 3·30 Anopheles gambiae per house per night in the unmodified houses compared with 3·60 in modified houses (unadjusted RR 1·28 [0·87–1·89], p=0·21). Interpretation: Improved housing did not provide protection against clinical malaria in this area of low seasonal transmission with high coverage of insecticide-treated nets, indoor residual spraying, and seasonal malaria chemoprevention. Funding: Global Health Trials funded by Medical Research Council, UK Department for International Development, and Wellcome Trust.

Abstract Background Malnutrition is common in children in sub-Saharan Africa and is thought to increase the risk of infectious diseases, including malaria. The relationship between malnutrition and malaria was examined in a cohort of 6–59 month-old children in rural Gambia, in an area of seasonal malaria transmission. The study used data from a clinical trial in which a cohort of children was established and followed for clinical malaria during the 2011 transmission season. A cross-sectional survey to determine the prevalence of malaria and anaemia, and measure the height and weight of these children was carried out at the beginning and end of the transmission season. Standard anthropometric indices (stunting, wasting and underweight) were calculated using z-scores. Results At the beginning of the transmission season, 31.7% of children were stunted, 10.8% wasted and 24.8% underweight. Stunting was more common in Fula children than other ethnicities and in children from traditionally constructed houses compared to more modern houses. Stunted children and underweight children were significantly more likely to have mild or moderate anaemia. During the transmission season, 13.7% of children had at least one episode of clinical malaria. There was no association between stunting and malaria incidence (odds ratio = 0.79, 95% CI: 0.60–1.05). Malaria was not associated with differences in weight or height gain. Conclusions Chronic malnutrition remains a problem in rural Gambia, particularly among the poor and Fula ethnic group, but it was not associated with an increased risk of malaria. Trial registration Trial registration: ISRCTN, ISRCTN01738840, registered: 27/08/2010 (Retrospectively registered).

Unfortunately, the original article [1] contained an error mistakenly carried forward by the Production department handling this article whereby some figures and their captions were interchanged. The correct figures (Figs. 1, 2, 3, 4, 5) and captions are presented in this erratum. The original article has also been updated to reflect this correction.

Abstract Background Insecticide resistance threatens malaria control in sub-Saharan Africa. Knockdown resistance to pyrethroids and organochlorines in Anopheles gambiae sensu lato (s.l.) is commonly caused by mutations in the gene encoding a voltage-gated sodium channel which is the target site for the insecticide. The study aimed to examine risk factors for knockdown resistance in An. gambiae s.l. and its relationship with malaria infection in children in rural Gambia. Point mutations at the Vgsc-1014 locus, were measured in An. gambiae s.l. during a 2-year trial. Cross-sectional surveys were conducted at the end of the transmission season to measure malaria infection in children aged 6 months–14 years. Results Whilst few Anopheles arabiensis and Anopheles coluzzii had Vgsc-1014 mutations, the proportion of An. gambiae sensu stricto (s.s.) mosquitoes homozygous for the Vgsc-1014F mutation increased from 64.8 to 90.9% during the study. The Vgsc-1014S or 1014F mutation was 80% higher in 2011 compared to 2010, and 27% higher in the villages with indoor residual spraying compared to those without. An increase in the proportion of An. gambiae s.l. mosquitoes with homozygous Vgsc-1014F mutations and an increase in the proportion of An. gambiae s.s. in a cluster were each associated with increased childhood malaria infection. Homozygous Vgsc-1014F mutations were, however, most common in An. gambiae s.s. and almost reached saturation during the study meaning that the two variables were colinear. Conclusions As a result of colinearity between homozygous Vgsc-1014F mutations and An. gambiae s.s., it was not possible to determine whether insecticide resistance or species composition increased the risk of childhood malaria infection.

An amendment to this paper has been published and can be accessed via the original article.

Summary: Introduction: Unprecedented improvements in housing are occurring across much of rural sub-Saharan Africa, but the consequences of these changes on malaria transmission remain poorly explored. We examined how different typologies of rural housing affect mosquito house entry and indoor climate. Methods: Five typologies of mud-block houses were constructed in rural Gambia: four were traditional designs with poorly fitted doors and one was a novel design with gable windows to improve ventilation. In each house, one male volunteer slept under a bednet and mosquitoes were collected indoors with a light trap. Typologies were rotated between houses weekly. Indoor conditions were monitored with data loggers and the perceived comfort of sleepers recorded with questionnaires. We used pyschrometric modelling to quantify the comfort of the indoor climate using the logger data. Primary measurements were mean number of Anopheles gambiae and mean temperature for each house typology. Findings: In thatched-roofed houses, closing the eaves reduced A gambiae house entry by 94% (95% CI 89–97) but increased the temperature compared with thatched-roofed houses with open eaves. In houses with closed eaves, those with metal roofs had more A gambiae, were hotter (1·5°C hotter [95% CI 1·3–1·7]) between 2100h and 2300 h, and had 25% higher concentrations of carbon dioxide (211·1 ppm higher [117·8–304·6]) than those with thatched roofs. In metal-roofed houses with closed eaves, mosquito house entry was reduced by 96% (91–98) by well fitted screened doors. Improved ventilation of metal-roofed houses made them as cool as thatched houses with open eaves. Metal-roofed houses with closed eaves were considered more uncomfortable than thatched ones with closed eaves. In metal-roofed houses, ventilated houses were more comfortable than unventilated houses before midnight, when people retired to bed. Interpretation: Closing the eaves reduced vector entry in thatched houses but increased entry in metal-roofed houses. Metal-roofed houses with closed eaves were, however, protected against malaria vectors by well fitted screened doors and were made comfortable by increasing ventilation. House designs that exclude mosquitoes and are comfortable to live in should be a priority in sub-Saharan Africa. Funding: Sir Halley Stewart Trust, Global Clinical Trials, and Global Challenges Research Fund.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is believed to confer protection against Plasmodium falciparum malaria, but the precise nature of the protective effect has proved difficult to define as G6PD deficiency has multiple allelic variants with different effects in males and females, and it has heterogeneous effects on the clinical outcome of P. falciparum infection. Here we report an analysis of multiple allelic forms of G6PD deficiency in a large multi-centre case-control study of severe malaria, using the WHO classification of G6PD mutations to estimate each individual’s level of enzyme activity from their genotype. Aggregated across all genotypes, we find that increasing levels of G6PD deficiency are associated with decreasing risk of cerebral malaria, but with increased risk of severe malarial anaemia. Models of balancing selection based on these findings indicate that an evolutionary trade-off between different clinical outcomes of P. falciparum infection could have been a major cause of the high levels of G6PD polymorphism seen in human populations.

Combining data from genome-wide association studies (GWAS) conducted at different locations, using genotype imputation and fixed-effects meta-analysis, has been a powerful approach for dissecting complex disease genetics in populations of European ancestry. Here we investigate the feasibility of applying the same approach in Africa, where genetic diversity, both within and between populations, is far more extensive. We analyse genome-wide data from approximately 5,000 individuals with severe malaria and 7,000 population controls from three different locations in Africa. Our results show that the standard approach is well powered to detect known malaria susceptibility loci when sample sizes are large, and that modern methods for association analysis can control the potential confounding effects of population structure. We show that pattern of association around the haemoglobin S allele differs substantially across populations due to differences in haplotype structure. Motivated by these observations we consider new approaches to association analysis that might prove valuable for multicentre GWAS in Africa: we relax the assumptions of SNP-based fixed effect analysis; we apply Bayesian approaches to allow for heterogeneity in the effect of an allele on risk across studies; and we introduce a region-based test to allow for heterogeneity in the location of causal alleles.

BackgroundMalaria vector control is threatened by resistance to pyrethroids, the only class of insecticides used for treating bed nets. The second major vector control method is indoor residual spraying with pyrethroids or the organochloride DDT. However, resistance to pyrethroids frequently confers resistance to DDT. Therefore, alternative insecticides are urgently needed.Methodology/principal findingsInsecticide resistance and the efficacy of indoor residual spraying with different insecticides was determined in a Gambian village. Resistance of local vectors to pyrethroids and DDT was high (31% and 46% mortality, respectively) while resistance to bendiocarb and pirimiphos methyl was low (88% and 100% mortality, respectively). The vectors were predominantly Anopheles gambiae s.s. with 94% of them having the putative resistant genotype kdr 1014F. Four groups of eight residential compounds were each sprayed with either (1) bendiocarb, a carbamate, (2) DDT, an organochlorine, (3) microencapsulated pirimiphos methyl, an organophosphate, or (4) left unsprayed. All insecticides tested showed high residual activity up to five months after application. Mosquito house entry, estimated by light traps, was similar in all houses with metal roofs, but was significantly less in IRS houses with thatched roofs (p=0.02). Residents participating in focus group discussions indicated that IRS was considered a necessary nuisance and also may decrease the use of long-lasting insecticidal nets.Conclusion/significanceBendiocarb and microencapsulated pirimiphos methyl are viable alternatives for indoor residual spraying where resistance to pyrethroids and DDT is high and may assist in the management of pyrethroid resistance.

BACKGROUND:In areas of declining malaria transmission such as in The Gambia, the identification of malaria infected individuals becomes increasingly harder. School surveys may be used to identify foci of malaria transmission in the community. METHODS:The survey was carried out in May-June 2011, before the beginning of the malaria transmission season. Thirty two schools in the Upper River Region of The Gambia were selected with probability proportional to size; in each school approximately 100 children were randomly chosen for inclusion in the study. Each child had a finger prick blood sample collected for the determination of antimalarial antibodies by ELISA, malaria infection by microscopy and PCR, and for haemoglobin measurement. In addition, a simple questionnaire on socio-demographic variables and the use of insecticide-treated bed nets was completed. The cut-off for positivity for antimalarial antibodies was obtained using finite mixture models. The clustered nature of the data was taken into account in the analyses. RESULTS:A total of 3,277 children were included in the survey. The mean age was 10 years (SD = 2.7) [range 4-21], with males and females evenly distributed. The prevalence of malaria infection as determined by PCR was 13.6% (426/3124) [95% CI = 12.2-16.3] with marked variation between schools (range 3-25%, p

Severe malaria (SM) is a major cause of death in sub-Saharan Africa. Identification of both specific and sensitive clinical features to predict death is needed to improve clinical management.A 13-year observational study was conducted from 1997 through 2009 of 2,901 children with SM enrolled at the Royal Victoria Teaching Hospital in The Gambia to identify sensitive and specific predictors of poor outcome in Gambian children with severe malaria between the ages 4 months to 14 years. We have measured the sensitivity and specificity of clinical features that predict death or development of neurological sequelae.Impaired consciousness (odds ratio {OR} 4.4 [95% confidence interval {CI}, 2.7-7.3]), respiratory distress (OR 2.4 [95%CI, 1.7-3.2]), hypoglycemia (OR 1.7 [95%CI, 1.2-2.3]), jaundice (OR 1.9 [95%CI, 1.2-2.9]) and renal failure (OR 11.1 [95%CI, 3.3-36.5]) were independently associated with death in children with SM. The clinical features that showed the highest sensitivity and specificity to predict death were respiratory distress (area under the curve 0.63 [95%CI, 0.60-0.65]) and impaired consciousness (AUC 0.61[95%CI, 0.59-0.63]), which were comparable to the ability of hyperlactatemia (blood lactate>5 mM) to predict death (AUC 0.64 [95%CI, 0.55-0.72]). A Blantyre coma score (BCS) of 2 or less had a sensitivity of 74% and specificity of 67% to predict death (AUC 0.70 [95% C.I. 0.68-0.72]), and sensitivity and specificity of 74% and 69%, respectively to predict development of neurological sequelae (AUC 0.72 [95% CI, 0.67-0.76]).The specificity of this BCS threshold to identify children at risk of dying improved in children less than 3 years of age (AUC 0.74, [95% C.I 0.71-0.76]).The BCS is a quantitative predictor of death. A BCS of 2 or less is the most sensitive and specific clinical feature to predict death or development of neurological sequelae in children with SM.

BACKGROUND:Chrysomya spp are common blowflies in Africa, Asia and parts of South America and some species can reproduce in prodigious numbers in pit latrines. Because of their strong association with human feces and their synanthropic nature, we examined whether these flies are likely to be vectors of diarrheal pathogens. METHODOLOGY/PRINCIPAL FINDINGS:Flies were sampled using exit traps placed over the drop holes of latrines in Gambian villages. Odor-baited fly traps were used to determine the relative attractiveness of different breeding and feeding media. The presence of bacteria on flies was confirmed by culture and bacterial DNA identified using PCR. A median of 7.00 flies/latrine/day (IQR = 0.0-25.25) was collected, of which 95% were Chrysomya spp, and of these nearly all were Chrysomya putoria (99%). More flies were collected from traps with feces from young children (median = 3.0, IQR = 1.75-10.75) and dogs (median = 1.50, IQR = 0.0-13.25) than from herbivores (median = 0.0, IQR = 0.0-0.0; goat, horse, cow and calf; p

African pit latrines produce prodigious numbers of the latrine fly, Chrysomya putoria, a putative vector of diarrhoeal pathogens. We set out to develop a simple, low-cost odour-baited trap for collecting C. putoria in the field. A series of field experiments was carried out in The Gambia to assess the catching-efficiency of different trap designs. The basic trap was a transparent 3L polypropylene box baited with 50 g of fish, with a white opaque lid with circular entrance holes. We tested variations of the number, diameter, position and shape of the entrance holes, the height of the trap above ground, degree of transparency of the box, its shape, volume, colour, and the attractiveness of gridded surfaces on or under the trap. Traps were rotated between positions on different sampling occasions using a Latin Square design. The optimal trapping features were incorporated into a final trap that was tested against commercially available traps. Features of the trap that increased the number of flies caught included: larger entrance holes (compared with smaller ones, p

Odour baited traps have potential use in population surveillance of insect vectors of disease, and in some cases for vector population reduction. Established attractants for human host-seeking mosquitoes include a combination of CO(2) with L-lactic acid and ammonia, on top of which additional candidate compounds are being tested. In this field study in rural Gambia, using Latin square experiments with thorough randomization and replication, we tested nine different leading candidate combinations of chemical odorants for attractiveness to wild mosquitoes including anthropophilic malaria vectors, using modified Mosquito Magnet-X (MM-X) counterflow traps outside experimental huts containing male human sleepers. Highest catches of female mosquitoes, particularly of An. gambiae s.l. and Mansonia species, were obtained by incorporation of tetradecanoic acid. As additional carboxylic acids did not increase the trap catches further, this 'reference blend' (tetradecanoic acid with L-lactic acid, ammonia and CO(2)) was used in subsequent experiments. MM-X traps with this blend caught similar numbers of An. gambiae s.l. and slightly more Mansonia and Culex mosquitoes than a standard CDC light trap, and these numbers were not significantly affected by the presence or absence of human sleepers in the huts. Experiments with CO(2) produced from overnight yeast cultures showed that this organic source was effective in enabling trap attractiveness for all mosquito species, although at a slightly lower efficiency than obtained with use of CO(2) gas cylinders. Although further studies are needed to discover additional chemicals that increase attractiveness, as well as to optimise trap design and CO(2) source for broader practical use, the odour-baited traps described here are safe and effective for sampling host-seeking mosquitoes outdoors and can be incorporated into studies of malaria vector ecology.

With the functional demonstration of a role in erythrocyte invasion by Plasmodium falciparum parasites, implications in the aetiology of common conditions that prevail in individuals of African origin, and a wealth of pharmacological knowledge, the stimulatory G protein (Gs) signal transduction pathway presents an exciting target for anti-malarial drug intervention. Having previously demonstrated a role for the G-alpha-s gene, GNAS, in severe malaria disease, we sought to identify other important components of the Gs pathway. Using meta-analysis across case-control and family trio (affected child and parental controls) studies of severe malaria from The Gambia and Malawi, we sought evidence of association in six Gs pathway candidate genes: adenosine receptor 2A (ADORA2A) and 2B (ADORA2B), beta-adrenergic receptor kinase 1 (ADRBK1), adenylyl cyclase 9 (ADCY9), G protein beta subunit 3 (GNB3), and regulator of G protein signalling 2 (RGS2). Our study amassed a total of 2278 cases and 2364 controls. Allele-based models of association were investigated in all genes, and genotype and haplotype-based models were investigated where significant allelic associations were identified. Although no significant associations were observed in the other genes, several were identified in ADORA2A. The most significant association was observed at the rs9624472 locus, where the G allele (approximately 20% frequency) appeared to confer enhanced risk to severe malaria [OR = 1.22 (1.09-1.37); P = 0.001]. Further investigation of the ADORA2A gene region is required to validate the associations identified here, and to identify and functionally characterize the responsible causal variant(s). Our results provide further evidence supporting a role of the Gs signal transduction pathway in the regulation of severe malaria, and request further exploration of this pathway in future studies.

Interferon Regulatory Factor 1 (IRF-1) is a member of the IRF family of transcription factors, which have key and diverse roles in the gene-regulatory networks of the immune system. IRF-1 has been described as a critical mediator of IFN-gamma signalling and as the major player in driving TH1 type responses. It is therefore likely to be crucial in both innate and adaptive responses against intracellular pathogens such as Plasmodium falciparum. Polymorphisms at the human IRF1 locus have been previously found to be associated with the ability to control P. falciparum infection in populations naturally exposed to malaria. In order to test whether genetic variation at the IRF1 locus also affects the risk of developing severe malaria, we performed a family-based test of association for 18 Single Nucleotide Polymorphisms (SNPs) across the gene in three African populations, using genotype data from 961 trios consisting of one affected child and his/her two parents (555 from The Gambia, 204 from Kenya and 202 from Malawi). No significant association with severe malaria or severe malaria subphenotypes (cerebral malaria and severe malaria anaemia) was observed for any of the SNPs/haplotypes tested in any of the study populations. Our results offer no evidence that the molecular pathways regulated by the transcription factor IRF-1 are involved in the immune-based pathogenesis of severe malaria.

Congenital cytomegalovirus (CMV) infection is the most prevalent congenital infection worldwide. Epidemiology and clinical outcomes are known to vary with socio-economic background, but few data are available from developing countries, where the overall burden of infectious diseases is frequently high.As part of an ongoing birth cohort study in The Gambia among term infants, urine samples were collected at birth and tested by PCR for the presence of CMV DNA. Risk factors for transmission and clinical outcome were assessed, including placental malaria infection. Babies were followed up at home monthly for morbidity and anthropometry, and at one year of age a clinical evaluation was performed. The prevalence of congenital CMV infection was 5.4% (40/741). A higher prevalence of hepatomegaly was the only significant clinical difference at birth. Congenitally infected children were more often first born babies (adjusted odds ratio (OR) 5.3, 95% confidence interval (CI) 2.0-13.7), more frequently born in crowded compounds (adjusted OR 2.9, 95%CI 1.0-8.3) and active placental malaria was more prevalent (adjusted OR 2.9, 95%CI 1.0-8.4). These associations were corrected for maternal age, bed net use and season of birth. During the first year of follow up, mothers of congenitally infected children reported more health complaints for their child.In this study, the prevalence of congenital CMV among healthy neonates was much higher than previously reported in industrialised countries, and was associated with active placental malaria infection. There were no obvious clinical implications during the first year of life. The effect of early life CMV on the developing infant in the Gambia could be mitigated by environmental factors, such as the high burden of other infections.

BackgroundResistance of malaria parasites to chloroquine (CQ) and sulphadoxine-pyrimethamine (SP) is increasing in prevalence in Africa. Combination therapy can both improve treatment and provide important public health benefits if it curbs the spread of parasites harbouring resistance genes. Thus, drug combinations must be identified which minimise gametocyte emergence in treated cases, and so prevent selective transmission of parasites resistant to any of the partner drugs.Methods and findingsIn a randomised controlled trial, 497 children with uncomplicated falciparum malaria were treated with CQ and SP (three doses and one dose respectively; n = 91), or six doses of artemether in fixed combination with lumefantrine (co-artemether [Coartem, Riamet]) (n = 406). Carriage rates of Plasmodium falciparum gametocytes and trophozoites were measured 7, 14, and 28 d after treatment. The infectiousness of venous blood from 29 children carrying P. falciparum gametocytes 7 d after treatment was tested by membrane-feeding of Anopheles mosquitoes. Children treated with co-artemether were significantly less likely to carry gametocytes within the 4 weeks following treatment than those receiving CQ/SP (30 of 378 [7.94%] versus 42 of 86 [48.8%]; p < 0.0001). Carriers in the co-artemether group harboured gametocytes at significantly lower densities, for shorter periods (0.3 d versus 4.2 d; p < 0.0001) and were less infectious to mosquitoes at day 7 (p < 0.001) than carriers who had received CQ/SP.ConclusionsCo-artemether is highly effective at preventing post-treatment transmission of P. falciparum. Our results suggest that co-artemether has specific activity against immature sequestered gametocytes, and has the capacity to minimise transmission of drug-resistant parasites.

Many malaria vaccines are currently in development, although very few have been evaluated for efficacy in the field. Plasmodium falciparum multiple epitope (ME)- thrombospondin-related adhesion protein (TRAP) candidate vaccines are designed to potently induce effector T cells and so are a departure from earlier malaria vaccines evaluated in the field in terms of their mechanism of action. ME-TRAP vaccines encode a polyepitope string and the TRAP sporozoite antigen. Two vaccine vectors encoding ME-TRAP, plasmid DNA and modified vaccinia virus Ankara (MVA), when used sequentially in a prime-boost immunisation regime, induce high frequencies of effector T cells and partial protection, manifest as delay in time to parasitaemia, in a clinical challenge model.A total of 372 Gambian men aged 15-45 y were randomised to receive either DNA ME-TRAP followed by MVA ME-TRAP or rabies vaccine (control). Of these men, 296 received three doses of vaccine timed to coincide with the beginning of the transmission season (141 in the DNA/MVA group and 155 in the rabies group) and were followed up. Volunteers were given sulphadoxine/pyrimethamine 2 wk before the final vaccination. Blood smears were collected weekly for 11 wk and whenever a volunteer developed symptoms compatible with malaria during the transmission season. The primary endpoint was time to first infection with asexual P. falciparum. Analysis was per protocol. DNA ME-TRAP and MVA ME-TRAP were safe and well-tolerated. Effector T cell responses to a non-vaccine strain of TRAP were 50-fold higher postvaccination in the malaria vaccine group than in the rabies vaccine group. Vaccine efficacy, adjusted for confounding factors, was 10.3% (95% confidence interval, -22% to +34%; p = 0.49). Incidence of malaria infection decreased with increasing age and was associated with ethnicity.DNA/MVA heterologous prime-boost vaccination is safe and highly immunogenic for effector T cell induction in a malaria-endemic area. But despite having produced a substantial reduction in liver-stage parasites in challenge studies of non-immune volunteers, this first generation T cell-inducing vaccine was ineffective at reducing the natural infection rate in semi-immune African adults.