Margaret Pinder

Despite compelling evidence that modern housing protects against malaria, houses in endemic areas are still commonly porous to mosquitoes. The protective efficacy of four prototype screened doors and two windows designs against mosquito house entry, their impact on indoor climate, as well as their use, durability and acceptability was assessed in a Gambian village. A baseline survey collected data on all the houses and discrete household units, each consisting of a front and back room, were selected and randomly allocated to the study arms. Each prototype self-closing screened door and window was installed in six and 12 units, respectively, with six unaltered units serving as controls. All prototype doors reduced the number of house-entering mosquitoes by 59–77% in comparison with the control houses. The indoor climate of houses with screened doors was similar to control houses. Seventy-nine percentage of door openings at night occurred from dusk to midnight, when malaria vectors begin entering houses. Ten weeks after installation the doors and windows were in good condition, although 38% of doors did not fully self-close and latch (snap shut). The new doors and windows were popular with residents. The prototype door with perforated concertinaed screening was the best performing door because it reduced mosquito entry, remained fully functional, and was preferred by the villagers. Screened doors and windows may be useful tools for reducing vector exposure and keeping areas malaria-free after elimination, when investment in routine vector control becomes difficult to maintain.

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. 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. 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: ISRCTN, ISRCTN01738840 , registered: 27/08/2010 (Retrospectively registered).

The effectiveness of pyrethroid-treated bednets for malaria control in sub-Saharan Africa is under threat because of high levels of resistance to pyrethroid insecticides in the vectors. Here we assess the durability of polyethylene nets with a novel combination of permethrin, a pyrethroid, with pyriproxyfen, an insect juvenile mimic (PPF-LLIN), in comparison with a typical permethrin-treated long-lasting insecticidal net (LLIN). This is a cluster randomised controlled trial of net durability in Burkina Faso, with clustering at the level of the compound and includes entomological outcome measurements. Half the compounds in each village will be randomly allocated PPF-LLIN and half the LLIN. All sleeping places in a compound will be provided with one type of net. We will distribute the nets at the start of the first transmission season and follow net use at the start and end of each transmission season for 3 years. In one village, bio-efficacy and chemical content will be recorded immediately after net distribution and then at 6, 12, 18, 24, 30 and 36 months. In the other village net survivorship and fabric integrity will be recorded immediately after distribution, and then at 6, 12, 18, 24, 30 and 36 months. Routine measurements of indoor temperature and relative humidity will be made in both villages during the study. Residents will be followed for possible side effects of the PPF-LLIN by surveillance of known asthmatic subjects during the first month post-distribution and pregnancy outcomes will be monitored from antenatal clinic records. The protocol is novel on two accounts. Firstly, it is the first to describe the procedure for measuring net durability following recent World Health Organisation (WHO) guidelines. Meeting the minimum requirements set in the guidelines is essential before a new type of net can be recommended by WHO’s Pesticide Evaluation Scheme (WHOPES). Secondly, it describes methods to monitor the persistence of an active ingredient that reduces vector fertility and fecundity. If the PPF-LLIN is both effective and persistent it will provide an alternative vector control strategy where pyrethroid-resistant vectors are present. ISRCTN30634670 assigned 13 August 2014.

Recent reductions in malaria in sub-Saharan Africa have been associated with increased coverage with long-lasting insecticidal nets (LLINs). Pyrethroids are currently the only insecticide class used for treating nets, and the rapid increase in resistance to pyrethroids in vector mosquitoes may jeopardise future vector control. Nets containing a novel combination of permethrin, a pyrethroid, and pyriproxyfen, an insect juvenile hormone mimic, (PPF-LLIN) may enhance malaria control, as well as reducing the spread of pyrethroid-resistant mosquitoes. This trial will determine whether PPF-LLINs provide incremental protection against malaria over current best practice of LLINs and prompt treatment in an area with pyrethroid-resistant vectors. A 2 armed cluster-randomised controlled trial will be conducted in Burkina Faso to assess whether PPF-LLIN (containing 2% permethrin and 1% pyriproxyfen w/w) provide better protection against clinical malaria in children than 2% permethrin-treated LLINs. Study subjects will be recruited and provided with LLINs at the start of the study. The LLINs will be exchanged for PPF-LLIN by cluster in a step-wedge fashion so 3 months before the end of the 2 year trial all participants will have a PPF-LLIN. The primary endpoint will be clinical malaria incidence measured by passive case detection in a cohort of children, aged 6 months to 5 years. Anaemia and parasite prevalence will also be measured in children during cross-sectional surveys. Exposure to malaria parasites will be assessed using light traps followed by identification of common vector species and their sporozoite infection rates. Safety evaluation will include recording of adverse events and pregnancy outcomes. The main endpoint analysis will include adjusting for distance between village clusters with different types of nets, as the impact of PPF-LLIN is likely to increase as larger areas are dominated by PPF-LLIN, reducing the spill over of mosquitoes from villages with LLINs. The step-wedge design is to measure the impact of an incrementally delivered environmental intervention where we expect the degree of control to be improved as more people use PPF-LLIN over a larger area. Trial findings will help inform policy makers on the effectiveness of PPF-LLIN nets for malaria control in areas of pyrethroid resistance. ISRCTN21853394 – AvecNet, registered on 3 April 2013.

Malaria 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. Insecticide 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. Bendiocarb 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.

Qualitative studies suggest that bed nets affect the thermal comfort of users. To understand and reduce this discomfort the effect of bed nets on temperature, humidity, and airflow was measured in rural homes in Asia and Africa, as well as in an experimental wind tunnel. Two investigators with architectural training selected 60 houses in The Gambia, Tanzania, Philippines, and Thailand. Data-loggers were used to measure indoor temperatures in hourly intervals over a 12 months period. In a subgroup of 20 houses airflow, temperature and humidity were measured at five-minute intervals for one night from 21.00 to 6.00 hrs inside and outside of bed nets using sensors and omni-directional thermo-anemometers. An investigator set up a bed net with a mesh size of 220 holes per inch² in each study household and slept under the bed net to simulate a realistic environment. The attenuation of airflow caused by bed nets of different mesh sizes was also measured in an experimental wind tunnel. The highest indoor temperatures (49.0 C) were measured in The Gambia. During the hottest months of the year the mean temperature at night (9 pm) was between 33.1 C (The Gambia) and 26.2 C (Thailand). The bed net attenuated the airflow from a minimum of 27% (Philippines) to a maximum of 71% (The Gambia). Overall the bed nets reduced airflow compared to un-attenuated airflow from 9 to 4 cm sec^-1 or 52% (p < 0.001). In all sites, no statistically significant difference in temperature or humidity was detected between the inside and outside of the bed net. Wind tunnel experiments with 11 different mesh-sized bed nets showed an overall reduction in airflow of 64% (range 55 - 71%) compared to un-attenuated airflow. As expected, airflow decreased with increasing net mesh size. Nets with a mesh of 136 holes inch^-2 reduced airflow by 55% (mean; range 51 - 73%). A denser net (200 holes inch^-2) attenuated airflow by 59% (mean; range 56 - 74%). Despite concerted efforts to increase the uptake of this intervention in many areas uptake remains poor. Bed nets reduce airflow, but have no influence on temperature and humidity. The discomfort associated with bed nets is likely to be most intolerable during the hottest and most humid period of the year, which frequently coincides with the peak of malaria vector densities and the force of pathogen transmission. These observations suggest thermal discomfort is a factor limiting bed net use and open a range of architectural possibilities to overcome this limitation.

Background: Recently, there has been mounting interest in scaling-up vector control against malaria in Africa. It needs to be determined if indoor residual spraying (IRS with DDT) will provide significant marginal protection against malaria over current best practice of long-lasting insecticidal nets (LLINs) and prompt treatment in a controlled trial, given that DDT is currently the most persistent insecticide for IRS. Methods: A 2 armed cluster-randomised controlled trial will be conducted to assess whether DDT IRS and LLINs combined provide better protection against clinical malaria in children than LLINs alone in rural Gambia. Each cluster will be a village, or a group of small adjacent villages; all clusters will receive LLINs and half will receive IRS in addition. Study children, aged 6 months to 13 years, will be enrolled from all clusters and followed for clinical malaria using passive case detection to estimate malaria incidence for 2 malaria transmission seasons in 2010 and 2011. This will be the primary endpoint. Exposure to malaria parasites will be assessed using light and exit traps followed by detection of Anopheles gambiae species and sporozoite infection. Study children will be surveyed at the end of each transmission season to estimate the prevalence of Plasmodium falciparum infection and the prevalence of anaemia. Discussion: Practical issues concerning intervention implementation, as well as the potential benefits and risks of the study, are discussed. Trial Registration: ISRCTN01738840 - Spraying And Nets Towards malaria Elimination (SANTE)

During malaria infection the Toll-like receptor 9 (TLR9) is activated through induction with plasmodium DNA or another malaria motif not yet identified. Although TLR9 activation by malaria parasites is well reported, the implication to the susceptibility to severe malaria is not clear. The aim of this study was to assess the contribution of genetic variation at TLR9 to severe malaria. This study explores the contribution of TLR9 genetic variants to severe malaria using two approaches. First, an association study of four common single nucleotide polymorphisms was performed on both family- and population-based studies from Malawian and Gambian populations (n>6000 individual). Subsequently, it was assessed whether TLR9 expression is affected by cis-acting variants and if these variants could be mapped. For this work, an allele specific expression (ASE) assay on a panel of HapMap cell lines was carried out. No convincing association was found with polymorphisms in TLR9 for malaria severity, in either Gambian or Malawian populations, using both case-control and family based study designs. Using an allele specific expression assay it was observed that TLR9 expression is affected by cis-acting variants, these results were replicated in a second experiment using biological replicates. By using the largest cohorts analysed to date, as well as a standardized phenotype definition and study design, no association of TLR9 genetic variants with severe malaria was found. This analysis considered all common variants in the region, but it is remains possible that there are rare variants with association signals. This report also shows that TLR9 expression is potentially modulated through cis-regulatory variants, which may lead to differential inflammatory responses to infection between individuals.

Background: Malaria in The Gambia is highly seasonal, with transmission occurring as Anopheles gambiae s.l. populations expand during and immediately after a single annual rainy season that lasts from June to October. There has been very limited investigation of the ecology of vectors during the dry season, when numbers are very limited and distributions may be restricted. Methods: Weekly adult mosquito collections (pyrethrum spray, light trap, and search collections from rooms, as well as light trap collections from animal shelters, abandoned wells and grain stores), and artificial sentinel breeding site surveys were performed in four villages near the upper tidal and partially saline part of the Gambia River in the last four months of an annual dry season (March to June). Mosquito species were identified by morphological and DNA analysis, and ELISA assays were performed to test for Plasmodium falciparum sporozoites and human blood meal components. Results: Adults of An. gambiae s.l. were collected throughout the period, numbers increasing towards the end of the dry season when humidity was increasing. Adult collections were dominated by An. melas (86%), with An. gambiae s.s. (10%) and An. arabiensis (3%) also present throughout. Most females collected in room search and spray collections contained blood meals, but most from light traps were unfed. None of the females tested (n = 1709) contained sporozoites. Larvae (mostly An. gambiae s.s.) were recovered from artificial sentinel breeding sites in the two villages that had freshwater pools. These two villages had the highest proportions of An. gambiae s.s. adults, and experienced the most substantial increase in proportions of An. gambiae s.s. after the onset of rains. Conclusion: During the dry season population minimum, An. melas was the predominant vector species, but differences among villages in availability of fresh-water breeding sites correlate with egg laying activity and relative numbers of An. gambiae s.s. adults, and with the increase in this species immediately after the beginning of the rains. Local variation in dry season vector persistence is thus likely to influence spatial heterogeneity of transmission intensity in the early part of the rainy season.

The sickle (βs) mutation in the beta-globin gene (HBB) occurs on five "classical" βs haplotype backgrounds in ethnic groups of African ancestry. Strong selection in favour of the βs allele – a consequence of protection from severe malarial infection afforded by heterozygotes – has been associated with a high degree of extended haplotype similarity. The relationship between classical βs haplotypes and long-range haplotype similarity may have both anthropological and clinical implications, but to date has not been explored. Here we evaluate the haplotype similarity of classical βs haplotypes over 400 kb in population samples from Jamaica, The Gambia, and among the Yoruba of Nigeria (Hapmap YRI).