Experimental Huts Research Articles

Implementing OECD GLP principles for the evaluation of novel vector control tools: a case study with two novel LLINs, SafeNet® and SafeNet NF®

BackgroundTo sustain high universal Long-Lasting Insecticidal Nets (LLINs) coverage, affordable nets that provide equivalent or better protection than standard LLINs, are required. Test facilities evaluating new LLINs require compliance to Good Laboratory Practice (GLP) standards to ensure the quality and integrity of test data. Following GLP principles allows for the reconstruction of activities during the conduct of a study and minimizes duplication of efficacy testing. This case study evaluated the efficacy of two LLINs: SafeNet NF® and SafeNet® LLIN.MethodsThe study was conducted according to GLP principles and followed World Health Organization guidelines for evaluating LLINs. The LLINs were assessed in experimental huts against wild, pyrethroid-resistant Anopheles arabiensis mosquitoes. Nets were either unwashed or washed 20 times and artificially holed to simulate a used torn net. Blood-feeding inhibition and mortality were compared with a positive control (Interceptor® LLIN) and an untreated net.ResultsMosquito entry in the huts was reduced compared to negative control for the unwashed SafeNet NF, washed Safenet LLIN and the positive control arms. Similar exiting rates were found for all the treatment arms. Significant blood-feeding inhibition was only found for the positive control, both when washed and unwashed. All insecticide treatments induced significantly higher mortality compared to an untreated net. Compared to the positive control, the washed and unwashed SafeNet NF® resulted in similar mortality. For the SafeNet® LLINs the unwashed net had an equivalent performance, but the mortality for the washed net was significantly lower than the positive control.Internal audits of the study confirmed that all critical phases complied with Standard Operating Procedures (SOPs) and the study plan. The external audit confirmed that the study complied with GLP standards.ConclusionsSafeNet NF® and SafeNet® LLIN offered equivalent protection to the positive control (Interceptor® LLIN). However, further research is needed to investigate the durability, acceptability, and residual efficacy of these nets in the community. This study demonstrated that GLP-compliant evaluation of LLINs can be successfully conducted by African research institutions.

Enhancing the Quality of Spray Application in IRS: Evaluation of the Micron Track Sprayer

Simple SummaryA key tool in the fight against mosquitoes, which transmit malaria, is the application of insecticidal indoor residual spray (IRS) to the internal walls of buildings where mosquitoes alight and rest to digest their blood-meals. When evaluating the effectiveness of IRS formulations for killing mosquitoes when applied to a wall, it is important that the insecticide is applied evenly at the target dose. Traditionally, IRS is applied using a hand-held pump, but this study showed that an automated track sprayer delivered the desired dose to wall surfaces more accurately and more evenly. This was first shown using a fluorescent tracer to measure spray deposit on the wall of a laboratory, and then by spraying different IRS formulations onto the walls of an experimental hut.Indoor residual spraying (IRS) has changed little since its introduction in the 1940s. Manual spraying is still prone to variation in insecticide dose. To improve the application of IRS in experimental hut trials, an automated track sprayer was developed, which regulates the speed of application and the distance of the nozzle from the wall, two key sources of variation. The automated track sprayer was compared to manual spraying, firstly using fluorescein solution in controlled indoor settings, and secondly in experimental huts in Tanzania using several IRS products. Manual spraying produced greater variation with both fluorescein and insecticide applications. Both manual and automated spray methods under-dosed the actual dose sprayed compared to the target dose. Overall, the track sprayer treats surfaces more consistently, offering a potential improvement over manual spraying for experimental hut evaluation of new IRS formulations.

Resistance intensity status of Anopheles gambiae s.l. species at KOLOKOPE, eastern plateau Togo: A potential site to assess new vector control tools

According to WHO recommendations, the deployment of the next generation of Long-Lasting Insecticidal Nets (LLINs) for malaria vector control requires appropriate investigations on the insecticide resistance profile of the vector. Most of the next generation of LLINs are impregnated with a combination of pyrethroid insecticides and piperonyl butoxide (PBO), a synergist with an additional impact on the increase in the mortality rate of Anopheles gambiae s.l. (Diptera: Culicidae). Kolokopé is a cotton-growing area in the central region of Togo characterized by an intensive use of agricultural pesticides and insecticides where there is a phase II experimental hut station. For the characterization of the site, WHO susceptibility tests using diagnostic doses of ten insecticides, PBO synergist assays and intensity assays of three pyrethroids (5x and 10x) were conducted on adult female mosquitoes obtained from larvae collected around the site. Anopheles gambiae s.l. from Kolokopé showed high resistance to pyrethroids and DDT, but to a lesser extent to carbamates and organophosphates. Likewise, high intensity of resistance to pyrethroid was observed with less than 40% mortality at 10x deltamethrin, 52 and 29% mortality at 10x permethrin and 10x alphacypermethrin, respectively. Also, PBO treatment resulted in increased mortality which was higher than the mortality rate at 10x doses of pyrethroids. The high pyrethroid intensity resistance recorded at Kolokopé could be mainly due to the selection pressure on An. gambiae s.l. caused by the excessive use of insecticide in agriculture. These results can be used to assess the next generation of LLINs either in experimental hut or at a community trial.

Experimental Hut Trials Reveal That CYP6P9a/b P450 Alleles Are Reducing the Efficacy of Pyrethroid-Only Olyset Net against the Malaria Vector Anopheles funestus but PBO-Based Olyset Plus Net Remains Effective.

Malaria remains a major public health concern in Africa. Metabolic resistance in major malaria vectors such as An. funestus is jeopardizing the effectiveness of long-lasting insecticidal nets (LLINs) to control malaria. Here, we used experimental hut trials (EHTs) to investigate the impact of cytochrome P450-based resistance on the efficacy of PBO-based net (Olyset Plus) compared to a permethrin-only net (Olyset), revealing a greater loss of efficacy for the latter. EHT performed with progenies of F5 crossing between the An. funestus pyrethroid-resistant strain FUMOZ and the pyrethroid-susceptible strain FANG revealed that PBO-based nets (Olyset Plus) induced a significantly higher mortality rate (99.1%) than pyrethroid-only nets (Olyset) (56.7%) (p < 0.0001). The blood-feeding rate was higher in Olyset compared to Olyset Plus (11.6% vs. 5.6%; p = 0.013). Genotyping the CYP6P9a/b and the intergenic 6.5 kb structural variant (SV) resistance alleles showed that, for both nets, homozygote-resistant mosquitoes have a greater ability to blood-feed than the susceptible mosquitoes. Homozygote-resistant genotypes significantly survived more with Olyset after cone assays (e.g., CYP6P9a OR = 34.6; p < 0.0001) than homozygote-susceptible mosquitoes. A similar but lower correlation was seen with Olyset Plus (OR = 6.4; p < 0.001). Genotyping EHT samples confirmed that CYP6P9a/b and 6.5 kb_SV homozygote-resistant mosquitoes survive and blood-feed significantly better than homozygote-susceptible mosquitoes when exposed to Olyset. Our findings highlight the negative impact of P450-based resistance on pyrethroid-only nets, further supporting that PBO nets, such as Olyset Plus, are a better solution in areas of P450-mediated resistance to pyrethroids.

Simplified Procedures for Managing Insecticidal Waste: A Case Report of KCMUCo-PAMVERC Vector Control Product Testing Facility in North-Eastern Tanzania

Insecticide testing facilities that evaluate a variety of vector control products may generate a large number of hazardous wastes from routine operations. These wastes originate from degraded technical grade materials, sprayed substrates with Indoor Residual Spraying (IRS), and redundant stock or working insecticidal solutions. The washing of Long-Lasting Insecticidal Nets (LLINs) during preparation for laboratory and experimental hut trials also contribute to wastewater with insecticide content. Human and environmental exposure to insecticidal waste can occur during transport, categorization, storage, and disposal, resulting in environmental pollution and potential health effects. Various national and international guidelines have been devised for safe disposal and should be strictly followed to avoid adverse effects on humans or environment. To facilitate proper insecticidal waste management, this paper outlines simple but safe practices derived from international and national guidelines that can be adopted by other similar facilities. National and international policies related to chemical management were reviewed and translated into Standard Operating Procedures (SOPs), training pathways, and manuals. National Environmental Management Council (NEMC) assessed the test facility and recommended disposal procedures. NEMC recommendations were followed to construct the soak pits for liquid waste disposal while chemical transporters and incinerators were contracted for solid waste disposal. An environmental expert was contracted for environmental audit and annual monitoring. Studies and activities at the facility were evaluated for their waste generation index. Safety manual and SOPs on risk assessment, waste management and disposal, handling hazardous materials, health and safety procedures, and chemical usage were written. At KCMUCo-PAMVERC, an annual average of 0.02 and 24.9 cubic meters of liquid waste, which mainly includes a mixture of water, insecticides, and solvents, were generated from laboratory experiments (phase I) and semi-field LLIN experiments (phase II), respectively, while the IRS semi-field experiments generated an annual average of 88 L of liquid waste and 6.3 tons of solid waste. An annual average of 0.18 cubic meters of liquid waste results from other sources, including expired laboratory reagents. Well-translated national and international policies/regulations may be adopted by insecticide test facilities for proper and effective waste disposal.

Pyrethroid-piperonyl butoxide (PBO) nets reduce the efficacy of indoor residual spraying with pirimiphos-methyl against pyrethroid-resistant malaria vectors

Pirimiphos-methyl is a pro-insecticide requiring activation by mosquito cytochrome P450 enzymes to induce toxicity while PBO blocks activation of these enzymes in pyrethroid-resistant vector mosquitoes. PBO may thus antagonise the toxicity of pirimiphos-methyl IRS when combined with pyrethroid-PBO ITNs. The impact of combining Olyset Plus and PermaNet 3.0 with Actellic 300CS IRS was evaluated against pyrethroid-resistant Anopheles gambiae s.l. in two parallel experimental hut trials in southern Benin. The vector population was resistant to pyrethroids and PBO pre-exposure partially restored deltamethrin toxicity but not permethrin. Mosquito mortality in experimental huts was significantly improved in the combinations of bendiocarb IRS with pyrethroid-PBO ITNs (33–38%) compared to bendiocarb IRS alone (14–16%, p < 0.001), demonstrating an additive effect. Conversely, mortality was significantly reduced in the combinations of pirimiphos-methyl IRS with pyrethroid-PBO ITNs (55–59%) compared to pirimiphos-methyl IRS alone (77–78%, p < 0.001), demonstrating evidence of an antagonistic effect when both interventions are applied in the same household. Mosquito mortality in the combination was significantly higher compared to the pyrethroid-PBO ITNs alone (55–59% vs. 22–26% p < 0.001) showing potential of pirimiphos-methyl IRS to enhance vector control when deployed to complement pyrethroid-PBO ITNs in an area where PBO fails to fully restore susceptibility to pyrethroids.

Laboratory and experimental hut trial evaluation of VECTRON™ T500 for indoor residual spraying (IRS) against insecticide resistant malaria vectors in Burkina Faso

Background: Malaria cases in some areas could be attributed to vector resistant to the insecticide. World Health Organization recommended insecticides for vector control are limited in number. It is essential to find rotational partners for existing Indoor Residual Spraying (IRS) products. VECTRON™ T500 is a novel insecticide with broflanilide as active ingredient. It has a mode of action on mosquitoes completely different to usually used. The aim of this study was to determine the optimum effective dose and efficacy of VECTRONTM against susceptible and resistant strains of Anopheles in Burkina Faso. Methods: VECTRON™ was sprayed at 50, 100 and 200 mg/m² doses onto mud and concrete blocks using Potter Spray Tower. The residual activity of broflanilide was assessed through cone bioassays 1 week and then monthly up to 14 months post spraying. Its efficacy was evaluated at 100 and 150 mg/m² against wild free-flying mosquitoes in experimental huts on both substrates. Actellic 300CS was applied at 1000 mg/m² as reference product. Cone assays were conducted monthly, using susceptible and resistant mosquito strains. Results: In the laboratory, VECTRON™ showed residual efficacy (≥80% mortality) on An. gambiae Kisumu up to 12 and 14 months, respectively, on concrete and mud blocks. Similar results were found with 100 and 200 mg/m² using An. coluzzii pyrethroid resistant strain. In experimental huts, a total of 19,552 An. gambiae s.l. were collected. Deterrence, blood-feeding inhibition and exophily with VECTRON™ treated huts were very low. At 100 and 150 mg/m², mortality of wild An. gambiae s.l. ranged between 55% and 73%. Monthly cone bioassay mortality remained &gt;80% up to 9 months. Conclusions: VECTRON™ shows great potential as IRS formulation for malaria vector control. It can be added to the arsenal of IRS products for use in rotations to control malaria and manage mosquito insecticide resistance.

Influence of testing modality on bioefficacy for the evaluation of Interceptor\xae G2 mosquito nets to combat malaria mosquitoes in Tanzania

BackgroundInsecticide-treated net (ITN) durability is evaluated using longitudinal bioefficacy and fabric integrity sampling post-distribution. Interceptor® G2 was developed for resistance management and contains two adulticides: alpha-cypermethrin and chlorfenapyr; it is a pro-insecticide that is metabolized into its active form by mosquito-detoxifying enzymes and may be enhanced when the mosquito is physiologically active. To elucidate the impact of bioassay modality, mosquito exposures of the alphacypermethrin ITN Interceptor® and dual adulticide Interceptor® G2 were investigated.MethodsThis study evaluated the performance of Interceptor® G2 compared to Interceptor® against local strains of mosquitoes in Tanzania. Unwashed and 20× times washed nets were tested. Efficacy of ITNs was measured by four bioassay types: (1) World Health Organisation (WHO) cone test (cone), (2) WHO tunnel test (tunnel), (3) Ifakara ambient chamber test (I-ACT) and (4) the WHO gold standard experimental hut test (hut). Hut tests were conducted against free-flying wild pyrethroid metabolically resistant Anopheles arabiensis and Culex quinquefasciatus. Cone, tunnel and I-ACT bioassays used laboratory-reared metabolically resistant An. arabiensis and Cx. quinquefasciatus and pyrethroid susceptible Anopheles gambiae sensu stricto and Aedes aegypti.ResultsAgainst resistant strains, superiority of Interceptor® G2 over Interceptor® was observed in all “free-flying bioassays”. In cone tests (which restrict mosquito flight), superiority of Interceptor® over Interceptor® G2 was recorded. Mortality of unwashed Interceptor® G2 among An. arabiensis was lowest in hut tests at 42.9% (95% CI: 37.3–48.5), although this increased to 66.7% (95% CI: 47.1–86.3) by blocking hut exit traps so mosquitoes presumably increased frequencies of contact with ITNs. Higher odds of mortality were consistently observed in Interceptor® G2 compared to Interceptor® in “free-flying” bioassays using An. arabiensis: tunnel (OR = 1.42 [95% CI:1.19–1.70], p < 0.001), I-ACT (OR = 1.61 [95% CI: 1.05–2.49], p = 0.031) and hut (OR = 2.53 [95% CI: 1.96–3.26], p < 0.001). Interceptor® and Interceptor® G2 showed high blood-feeding inhibition against all strains.ConclusionBoth free-flying laboratory bioassays (WHO Tunnel and I-ACT) consistently measured similarly, and both predicted the results of the experimental hut test. For bioefficacy monitoring and upstream product evaluation of ITNs in situ, the I-ACT may provide an alternative bioassay modality with improved statistical power. Interceptor G2® outperformed Interceptor ® against pyrethroid-resistant strains, demonstrating the usefulness of chlorfenapyr in mitigation of malaria.Graphical

An evidence synthesis approach for combining different data sources illustrated using entomological efficacy of insecticides for indoor residual spraying.

Prospective malaria public health interventions are initially tested for entomological impact using standardised experimental hut trials. In some cases, data are collated as aggregated counts of potential outcomes from mosquito feeding attempts given the presence of an insecticidal intervention. Comprehensive data i.e. full breakdowns of probable outcomes of mosquito feeding attempts, are more rarely available. Bayesian evidence synthesis is a framework that explicitly combines data sources to enable the joint estimation of parameters and their uncertainties. The aggregated and comprehensive data can be combined using an evidence synthesis approach to enhance our inference about the potential impact of vector control products across different settings over time. Aggregated and comprehensive data from a meta-analysis of the impact of Pirimiphos-methyl, an indoor residual spray (IRS) product active ingredient, used on wall surfaces to kill mosquitoes and reduce malaria transmission, were analysed using a series of statistical models to understand the benefits and limitations of each. Many more data are available in aggregated format (N = 23 datasets, 4 studies) relative to comprehensive format (N = 2 datasets, 1 study). The evidence synthesis model had the smallest uncertainty at predicting the probability of mosquitoes dying or surviving and blood-feeding. Generating odds ratios from the correlated Bernoulli random sample indicates that when mortality and blood-feeding are positively correlated, as exhibited in our data, the number of successfully fed mosquitoes will be under-estimated. Analysis of either dataset alone is problematic because aggregated data require an assumption of independence and there are few and variable data in the comprehensive format. We developed an approach to combine sources from trials to maximise the inference that can be made from such data and that is applicable to other systems. Bayesian evidence synthesis enables inference from multiple datasets simultaneously to give a more informative result and highlight conflicts between sources. Advantages and limitations of these models are discussed.

Durability of three types of dual active ingredient long-lasting insecticidal net compared to a pyrethroid-only LLIN in Tanzania: methodology for a prospective cohort study nested in a cluster randomized controlled trial

BackgroundProgress achieved by long-lasting insecticidal nets (LLINs) against malaria is threatened by widespread selection of pyrethroid resistance among vector populations. LLINs with non-pyrethroid insecticides are urgently needed. This study aims to assess the insecticide and textile durability of three classes of dual-active ingredient (A.I.) LLINs using techniques derived from established WHO LLIN testing methods to set new standards of evaluation.MethodsA WHO Phase 3 active ingredients and textile durability study will be carried out within a cluster randomized controlled trial in 40 clusters in Misungwi district, Tanzania. The following treatments will be evaluated: (1) Interceptor®G2 combining chlorfenapyr and the pyrethroid alpha-cypermethrin, (2) Royal Guard® treated with pyriproxyfen and alpha-cypermethrin, (3) Olyset™ Plus which incorporates a synergist piperonyl butoxide and the pyrethroid permethrin, and (4) a reference standard alpha-cypermethrin only LLIN (Interceptor®). 750 nets will be followed in 5 clusters per intervention arm at 6, 12, 24 and 36 months post distribution for survivorship and hole index assessment. A second cohort of 1950 nets per net type will be identified in 10 clusters, of which 30 LLINs will be withdrawn for bio-efficacy and chemical analysis every 6 months up to 36 months and another 30 collected for experimental hut trials every year. Bio-efficacy will be assessed using cone bioassays and tunnel tests against susceptible and resistant laboratory strains of Anopheles gambiae sensu stricto. Efficacy of field-collected nets will be compared in six experimental huts. The main outcomes will be Anopheles mortality up to 72 h post exposure, blood feeding and egg maturation using ovary dissection to assess impact on fecundity.ConclusionsStudy findings will help develop bio-efficacy and physical durability criteria for partner A.I., in relation to the cRCT epidemiological and entomological outcomes, and refine preferred product characteristics of each class of LLIN. If suitable, the bioassay and hut outcomes will be fitted to transmission models to estimate correlation with cRCT outcomes.Trial registration number: NCT03554616.

Large-scale (Phase III) evaluation of broflanilide 50WP (VECTRON\u2122 T500) for indoor residual spraying for malaria vector control in Northeast Tanzania: study protocol for a two-arm, non-inferiority, cluster-randomised community trial

BackgroundIndoor residual spraying (IRS) is a major method of malaria vector control across sub-Saharan Africa. Effective control is being undermined by the rapid spread of insecticide resistance. There is major investment in development of new insecticides for IRS that possess novel modes of action, long residual activity, low mammalian toxicity and minimal cross-resistance. VECTRON™ T500, a new IRS product containing the active ingredient broflanilide as a 50% wettable powder (WP), has been shown to be efficacious against pyrethroid susceptible and resistant vector species on mud and concrete substrates in experimental hut (Phase II) trials.MethodsA two-arm non-inferiority cluster randomized controlled trial (Phase III) will be undertaken in Muheza District, Tanga Region, Tanzania. VECTRON™ T500 will be compared to the IRS product Fludora® Fusion (clothianidin 50% WP + deltamethrin 6.25% WP). The predominant malaria vectors in the study area are pyrethroid-resistant Anopheles gambiae s.s., An. arabiensis and An. funestus s.s. Sixteen village clusters will be pair-matched on baseline vector densities and allocated to reference and intervention arms. Consenting households in the intervention arm will be sprayed with VECTRON™ T500 and those in the reference arm will be sprayed with Fludora® Fusion. Each month, CDC light traps will collect mosquitoes to estimate changes in vector density, indoor biting, sporozoite and entomological inoculation rates (EIR). Susceptibility to IRS active ingredients will be assessed using World Health Organisation (WHO) bottle bioassays. Target site and metabolic resistance mechanisms will be characterised among Anopheles field populations from both trial arms. Residual efficacy of both IRS products will be monitored for 12 months post intervention. Questionnaire and focus group discussions will explore factors that influence adherence, adverse effects and benefits of IRS.DiscussionThis protocol describes a large-scale non-inferiority evaluation of a novel IRS product to reduce the density and EIR of pyrethroid-resistant Anopheles vectors. If VECTRON™ T500 proves non-inferior to Fludora® Fusion, it will be considered as an additional vector control product for malaria prevention and insecticide resistance management.Trial registration: ClinicalTrials.gov, NCT05150808, registered on 26 November 2021. Retrospectively registered.

Assessing the Effects of Cooking Fuels on Anopheles Mosquito Behavior: An Experimental Study in Rural Rwanda.

ABSTRACT.Globally, cleaner cooking fuels are increasingly promoted to reduce household air pollution. However, there is concern that reductions in smoke from biomass fuels could lead to more favorable conditions for mosquitoes and potentially increase vectorborne disease risk. We investigated household entry, host-seeking, household exit, and mortality among Anopheles mosquitoes across three cooking fuel types: wood, charcoal, and liquid petroleum gas (LPG) in six experimental huts in Rwanda. Fifty laboratory-reared Anopheles gambiae mosquitoes were released each night in entry compartments outside each hut, and fuels were burned for 1 hour in the hut verandas. Collectors conducted human landing catch during cooking and for 2 hours afterward, and CDC light traps were used for the rest of the night to measure host-seeking. Differences in each outcome were assessed using generalized linear mixed models with random effects for hut, collector, and day. Cooking with LPG compared with wood and charcoal was associated with substantial increases in household entry and host-seeking. Household exit was not significantly different across fuels, and mortality was lower in LPG-burning huts compared with wood. Although these results are not directly generalizable to field conditions, they indicate a potential for clean fuel adoption to increase exposure to Anopheles mosquitoes compared with traditional biomass fuels. Additional entomological and epidemiological studies are needed to investigate changes in disease vector exposure associated with clean fuel adoption, and evaluate whether enhanced vector control interventions should be promoted in tandem with cleaner cooking fuels.

Scientific achievements and reflections after 20 years of vector biology and control research at the Pu Teuy mosquito field research station, Thailand

Additional vector control tools are needed to supplement current strategies to achieve malaria elimination and control of Aedes-borne diseases in many settings in Thailand and the Greater Mekong Sub-region. Within the next decade, the vector control community, Kasetsart University (KU), and the Ministry of Higher Education, Science, Research and Innovation must take full advantage of these tools that combine different active ingredients with different modes of action. Pu Teuy Mosquito Field Research Station (MFRS), Department of Entomology, Faculty of Agriculture, Kasetsart University (KU), Thailand was established in 2001 and has grown into a leading facility for performing high-quality vector biology and control studies and evaluation of public health insecticides that are operationally relevant. Several onsite mosquito research platforms have been established including experimental huts, a 40-m long semi-field screening enclosure, mosquito insectary, field-laboratory, and living quarters for students and researchers. Field research and assessments ranged from ‘basic’ investigations on mosquito biology, taxonomy and genetics to more ‘applied’ studies on responses of mosquitoes to insecticides including repellency, behavioural avoidance and toxicity. In the course of two decades, 51 peer-reviewed articles have been published, and 7 masters and 16 doctoral degrees in Entomology have been awarded to national and international students. Continued support of key national stakeholders will sustain MFRS as a Greater Mekong Subregion centre of excellence and a resource for both insecticide trials and entomological research.

The effect of light and ventilation on house entry by Anopheles arabiensis sampled using light traps in Tanzania: an experimental hut study

BackgroundIn sub-Saharan Africa, house design and ventilation affects the number of malaria mosquito vectors entering houses. This study hypothesized that indoor light from a CDC-light trap, visible from outside a hut, would increase entry of Anopheles arabiensis, an important malaria vector, and examined whether ventilation modifies this effect.MethodsFour inhabited experimental huts, each situated within a large chamber, were used to assess how light and ventilation affect the number of hut-entering mosquitoes in Tanzania. Each night, 300 female laboratory-reared An. arabiensis were released inside each chamber for 72 nights. Nightly mosquito collections were made using light traps placed indoors. Temperature and carbon dioxide concentrations were measured using data loggers. Treatments and sleepers were rotated between huts using a randomized block design.ResultsWhen indoor light was visible outside the huts, there was an 84% increase in the odds of collecting mosquitoes indoors (Odds ratio, OR = 1.84, 95% confidence intervals, 95% CI 1.74–1.95, p < 0.001) compared with when it was not. Although the odds of collecting mosquitoes in huts with closed eaves (OR = 0.54, 95% CI 0.41–0.72, p < 0.001) was less than those with open eaves, few mosquitoes entered either type of well-ventilated hut. The odds of collecting mosquitoes was 99% less in well-ventilated huts, compared with poorly-ventilated traditional huts (OR = 0.01, 95% CI 0.01–0.03, p < 0.001). In well-ventilated huts, indoor temperatures were 1.3 °C (95% CI 0.9–1.7, p < 0.001) cooler, with lower carbon dioxide (CO2) levels (mean difference = 97 ppm, 77.8–116.2, p < 0.001) than in poorly-ventilated huts.ConclusionAlthough light visible from outside a hut increased mosquito house entry, good natural ventilation reduces indoor carbon dioxide concentrations, a major mosquito attractant, thereby reducing mosquito-hut entry.

Optimising the deployment of vector control tools against malaria: a data-informed modelling study

Concern that insecticide resistant mosquitoes are threatening malaria control has driven the development of new types of insecticide treated nets (ITNs) and indoor residual spraying (IRS) of insecticide. Malaria control programmes have a choice of vector control interventions although it is unclear which controls should be used to combat the disease. The study aimed at producing a framework to easily compare the public health impact and cost-effectiveness of different malaria prevention measures currently in widespread use. We used published data from experimental hut trials conducted across Africa to characterise the entomological effect of pyrethroid-only ITNs versus ITNs combining a pyrethroid insecticide with the synergist piperonyl butoxide (PBO). We use these estimates to parameterise a dynamic mathematical model of Plasmodium falciparum malaria which is validated for two sites by comparing simulated results to empirical data from randomised control trials (RCTs) in Tanzania and Uganda. We extrapolated model simulations for a series of potential scenarios likely across the sub-Saharan African region and include results in an online tool (Malaria INtervention Tool [MINT]) that aims to identify optimum vector control intervention packages for scenarios with varying budget, price, entomological and epidemiological factors. Our model indicates that switching from pyrethroid-only to pyrethroid-PBO ITNs could averted up to twice as many cases, although the additional benefit is highly variable and depends on the setting conditions. We project that annual delivery of long-lasting, non-pyrethroid IRS would prevent substantially more cases over 3-years, while pyrethroid-PBO ITNs tend to be the most cost-effective intervention per case averted. The model was able to predict prevalence and efficacy against prevalence in both RCTs for the intervention types tested. MINT is applicable to regions of sub-Saharan Africa with endemic malaria and provides users with a method of designing intervention packages given their setting and budget. The most cost-effective vector control package will vary locally. Models able to recreate results of RCTs can be used to extrapolate outcomes elsewhere to support evidence-based decision making for investment in vector control. Medical Research Council, IVCC, Wellcome Trust. For the French translation of the abstract see Supplementary Materials section.

Comparative efficacy of two pyrethroid-piperonyl butoxide nets (Olyset Plus and PermaNet 3.0) against pyrethroid resistant malaria vectors: a non-inferiority assessment

BackgroundPyrethroid-PBO nets were conditionally recommended for control of malaria transmitted by mosquitoes with oxidase-based pyrethroid-resistance based on epidemiological evidence of additional protective effect with Olyset Plus compared to a pyrethroid-only net (Olyset Net). Entomological studies can be used to assess the comparative performance of other brands of pyrethroid-PBO ITNs to Olyset Plus.MethodsAn experimental hut trial was performed in Cové, Benin to compare PermaNet 3.0 (deltamethrin plus PBO on roof panel only) to Olyset Plus (permethrin plus PBO on all panels) against wild pyrethroid-resistant Anopheles gambiae sensu lato (s.l.) following World Health Organization (WHO) guidelines. Both nets were tested unwashed and after 20 standardized washes compared to Olyset Net. Laboratory bioassays were also performed to help explain findings in the experimental huts.ResultsWith unwashed nets, mosquito mortality was higher in huts with PermaNet 3.0 compared to Olyset Plus (41% vs. 28%, P < 0.001). After 20 washes, mortality declined significantly with PermaNet 3.0 (41% unwashed vs. 17% after washing P < 0.001), but not with Olyset Plus (28% unwashed vs. 24% after washing P = 0.433); Olyset Plus induced significantly higher mortality than PermaNet 3.0 and Olyset Net after 20 washes. PermaNet 3.0 showed a higher wash retention of PBO compared to Olyset Plus. A non-inferiority analysis performed with data from unwashed and washed nets together using a margin recommended by the WHO, showed that PermaNet 3.0 was non-inferior to Olyset Plus in terms of mosquito mortality (25% with Olyset Plus vs. 27% with PermaNet 3.0, OR = 1.528, 95%CI = 1.02–2.29) but not in reducing mosquito feeding (25% with Olyset Plus vs. 30% with PermaNet 3.0, OR = 1.192, 95%CI = 0.77–1.84). Both pyrethroid-PBO nets were superior to Olyset Net.ConclusionOlyset Plus outperformed PermaNet 3.0 in terms of its ability to cause greater margins of improved mosquito mortality compared to a standard pyrethroid net, after multiple standardized washes. However, using a margin of non-inferiority defined by the WHO, PermaNet 3.0 was non-inferior to Olyset Plus in inducing mosquito mortality. Considering the low levels of mortality observed and increasing pyrethroid-resistance in West Africa, it is unclear whether either of these nets would demonstrate the same epidemiological impact observed in community trials in East Africa.

Wild populations of malaria vectors can mate both inside and outside human dwellings

BackgroundWild populations of Anopheles mosquitoes are generally thought to mate outdoors in swarms, although once colonized, they also mate readily inside laboratory cages. This study investigated whether the malaria vectors Anopheles funestus and Anopheles arabiensis can also naturally mate inside human dwellings.MethodMosquitoes were sampled from three volunteer-occupied experimental huts in a rural Tanzanian village at 6:00 p.m. each evening, after which the huts were completely sealed and sampling was repeated at 11:00 p.m and 6 a.m. the next morning to compare the proportions of inseminated females. Similarly timed collections were done inside local unsealed village houses. Lastly, wild-caught larvae and pupae were introduced inside or outside experimental huts constructed inside two semi-field screened chambers. The huts were then sealed and fitted with exit traps, allowing mosquito egress but not entry. Mating was assessed in subsequent days by sampling and dissecting emergent adults caught indoors, outdoors and in exit traps.ResultsProportions of inseminated females inside the experimental huts in the village increased from approximately 60% at 6 p.m. to approximately 90% the following morning despite no new mosquitoes entering the huts after 6 p.m. Insemination in the local homes increased from approximately 78% to approximately 93% over the same time points. In the semi-field observations of wild-caught captive mosquitoes, the proportions of inseminated An. funestus were 20.9% (95% confidence interval [CI]: ± 2.8) outdoors, 25.2% (95% CI: ± 3.4) indoors and 16.8% (± 8.3) in exit traps, while the proportions of inseminated An. arabiensis were 42.3% (95% CI: ± 5.5) outdoors, 47.4% (95% CI: ± 4.7) indoors and 37.1% (CI: ± 6.8) in exit traps.ConclusionWild populations of An. funestus and An. arabiensis in these study villages can mate both inside and outside human dwellings. Most of the mating clearly happens before the mosquitoes enter houses, but additional mating happens indoors. The ecological significance of such indoor mating remains to be determined. The observed insemination inside the experimental huts fitted with exit traps and in the unsealed village houses suggests that the indoor mating happens voluntarily even under unrestricted egress. These findings may inspire improved vector control, such as by targeting males indoors, and potentially inform alternative methods for colonizing strongly eurygamic Anopheles species (e.g. An. funestus) inside laboratories or semi-field chambers.Graphical

Partial indoor residual spraying with pirimiphos-methyl as an effective and cost-saving measure for the control of Anopheles gambiae s.l. in northern Ghana

The scale up of indoor residual spraying (IRS) and insecticide treated nets have contributed significantly to global reductions in malaria prevalence over the last two decades. However, widespread pyrethroid resistance has necessitated the use of new and more expensive insecticides for IRS. Partial IRS with pirimiphos-methyl in experimental huts and houses in a village-wide trial was evaluated against Anopheles gambiae s.l. in northern Ghana. Four different scenarios in which either only the top or bottom half of the walls of experimental huts were sprayed, with or without also spraying the ceiling were compared. Mortality of An. gambiae s.l. on partially sprayed walls was compared with the standard procedures in which all walls and ceiling surfaces are sprayed. A small-scale trial was then conducted to assess the effectiveness, feasibility, and cost of spraying only the upper walls and ceiling as compared to full IRS and no spraying in northern Ghana. Human landing catches were conducted to estimate entomological indices and determine the effectiveness of partial IRS. An established transmission dynamics model was parameterized by an analysis of the experimental hut data and used to predict the epidemiological impact and cost effectiveness of partial IRS for malaria control in northern Ghana. In the experimental huts, partial IRS of the top (IRR 0.89, p = 0.13) or bottom (IRR 0.90, p = 0.15) half of walls and the ceiling was not significantly less effective than full IRS in terms of mosquito mortality. In the village trial, the annual entomological inoculation rate was higher for the unsprayed control (217 infective bites/person/year (ib/p/yr)) compared with the fully and partially sprayed sites, with 28 and 38 ib/p/yr, respectively. The transmission model predicts that the efficacy of partial IRS against all-age prevalence of malaria after six months would be broadly equivalent to a full IRS campaign in which 40% reduction is expected relative to no spray campaign. At scale, partial IRS in northern Ghana would have resulted in a 33% cost savings ($496,426) that would enable spraying of 36,000 additional rooms. These findings suggest that partial IRS is an effective, feasible, and cost saving approach to IRS that could be adopted to sustain and expand implementation of this key malaria control intervention.

Transfluthrin eave-positioned targeted insecticide (EPTI) reduces human landing rate (HLR) of pyrethroid resistant and susceptible malaria vectors in a semi-field simulated peridomestic space

BackgroundVolatile pyrethroids (VPs) are proven to reduce human–vector contact for mosquito vectors. With increasing resistance to pyrethroids in mosquitoes, the efficacy of VPs, such as transfluthrin, may be compromised. Therefore, experiments were conducted to determine if the efficacy of transfluthrin eave-positioned targeted insecticide (EPTI) depends on the resistance status of malaria vectors.MethodsRibbons treated with 5.25 g transfluthrin or untreated controls were used around the eaves of an experimental hut as EPTI inside a semi-field system. Mosquito strains with different levels of pyrethroid resistance were released simultaneously, recaptured by means of human landing catches (HLCs) and monitored for 24-h mortality. Technical-grade (TG) transfluthrin was used, followed by emulsifiable concentrate (EC) transfluthrin and additional mosquito strains. Generalized linear mixed models with binomial distribution were used to determine the impact of transfluthrin and mosquito strain on mosquito landing rates and 24-h mortality.ResultsEPTI treated with 5.25 g of either TG or EC transfluthrin significantly reduced HLR of all susceptible and resistant Anopheles mosquitoes (Odds Ratio (OR) ranging from 0.14 (95% Confidence Interval (CI) [0.11–0.17], P < 0.001) to 0.57, (CI [0.42–0.78] P < 0.001). Both TG and EC EPTI had less impact on landing for the resistant Anopheles arabiensis (Mbita strain) compared to the susceptible Anopheles gambiae (Ifakara strain) (OR 1.50 [95% CI 1.18–1.91] P < 0.001) and (OR 1.67 [95% CI 1.29–2.17] P < 0.001), respectively. The EC EPTI also had less impact on the resistant An. arabiensis (Kingani strain) (OR 2.29 [95% CI 1.78–2.94] P < 0.001) compared to the control however the TG EPTI was equally effective against the resistant Kingani strain and susceptible Ifakara strain (OR 1.03 [95% CI 0.82–1.32] P = 0.75). Finally the EC EPTI was equally effective against the susceptible An. gambiae (Kisumu strain) and the resistant An. gambiae (Kisumu-kdr strain) (OR 0.98 [95% CI 0.74–1.30] P = 0.90).ConclusionsTransfluthrin-treated EPTI could be useful in areas with pyrethroid-resistant mosquitoes, but it remains unclear whether stronger resistance to pyrethroids will undermine the efficacy of transfluthrin. At this dosage, transfluthrin EPTI cannot be used to kill exposed mosquitoes.

Effectiveness of a long-lasting insecticide treatment kit (ICON\xae Maxx) for polyester nets over three years of household use: a WHO phase III trial in Tanzania

BackgroundICON® Maxx (Syngenta) is an insecticide treatment kit of pyrethroid and binding agent for long-lasting treatment of mosquito nets. Interim recommendation for use on nets was granted by the World Health Organization (WHO) after successful evaluation in experimental huts following multiple washes. A full WHO recommendation is contingent upon demonstration of continued bio-efficacy after 3 years of use.MethodsA household-randomized prospective study design was used to assess ICON Maxx-treated nets over 3 years in north-eastern Tanzania. Conventional treated nets (with lambda-cyhalothrin, but without binder) served as a positive control. At 6-monthly intervals, cross-sectional household surveys monitored net use and physical integrity, while cone and tunnel tests assessed insecticidal efficacy. Pyrethroid content was determined after 12 and 36 months. A parallel cohort of nets was monitored annually for evidence of net deterioration and attrition.ResultsAfter 12 months’ use, 97% of ICON Maxx-treated nets but only 67% of CTN passed the WHO efficacy threshold for insecticidal durability (> 80% mortality in cone or tunnel or 90% feeding inhibition in tunnel). After 24- and 36-months use, 67% and 26% of ICON Maxx treated nets met the cone criteria, respectively, and over 90% met the combined cone and tunnel criteria. Lambda-cyhalothrin content after 36 months was 17% (15.8 ± 4.3 mg/m2) of initial content. ICON Maxx nets were used year-round and washed approximately 4 times per year. In cross-sectional survey after 36 months the average number of holes was 20 and hole index was 740 cm2 per net. Cohort nets had fewer holes and smaller hole index than cross-sectional nets. However, only 15% (40/264) of cohort nets were not lost to follow-up or not worn out after 36 months.ConclusionsBecause more than 80% of nets met the WHO efficacy criteria after 36 months use, ICON Maxx was granted WHO full recommendation. Cross-sectional and cohort surveys were complementary and gave a fuller understanding of net durability. To improve net usage and retention, stronger incentives and health messaging should be introduced in WHO LLIN longitudinal trials. Untreated polyester nets may be made long-lastingly insecticidal in Africa through simple household treatment using ICON Maxx pyrethroid-binder kits.