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The present study aimed to assess mosquito species diversity, distribution, and ecological preferences in the Covè, Ouinhi, and Zangnanado communes, Southern Benin. Such information is critical to understand mosquito bio-ecology and to focus control efforts in high-risk areas for vector-borne diseases. Mosquito collections occurred quarterly in 60 clusters between June 2020 and April 2021, using human landing catches. In addition to the seasonal mosquito abundance, Shannon's diversity, Simpson, and Pielou's equitability indices were also evaluated to assess mosquito diversity. Ecological niche models were developed with MaxEnt using environmental variables to assess species distribution. Overall, mosquito density was higher in the wet season than in the dry season in all communes. A significantly higher Shannon's diversity index was also observed in the wet season than in the dry seasons in all communes (p < 0.05). Habitat suitability of An. gambiae s.s., An. coluzzii, Cx. quinquefasciatus and Ma. africana was highly influenced by slope, isothermality, site aspect, elevation, and precipitation seasonality in both wet and dry seasons. Overall, depending on the season, the ecological preferences of the four main mosquito species were variable across study communes. This emphasizes the impact of environmental conditions on mosquito species distribution. Moreover, mosquito populations were found to be more diverse in the wet season compared to the dry season., (© 2024. The Author(s).)

Background: The Dual-Active Ingredient long-lasting insecticidal nets (Dual-AI LLIN) have been developed to counteract the reduced efficacy of pyrethroid (PY)-only nets due to widespread pyrethroid insecticide resistance in malaria vector mosquitoes. They constitute half of the nets distributed in sub-Saharan Africa between 2022 and 2024. However, their effectiveness once they develop holes is unclear, particularly in pyrethroid-resistant settings. This study evaluates the textile integrity of three dual- AI LLINs compared to standard PY LLN, over 3 years of use in a community in Tanzania and the associated impact on malaria prevalence and incidence., Methods: A secondary analysis of data from a randomized controlled trial (RCT) in North-western Tanzania was conducted to evaluate the effectiveness of α-cypermethrin only; pyriproxyfen and α-cypermethrin (PPF-PY); chlorfenapyr and α-cypermethrin (chlorfenapyr-PY); and the synergist piperonyl butoxide and permethrin (PBO-PY) LLINs on malaria infection prevalence and case incidence. The association between the net textile condition and 1/malaria prevalence over 3 years of use between 2019 and 2022, and 2/malaria case incidence in a cohort of children over 2 years of follow-up was assessed between 2019 and 2021., Results: There was no significant association between damaged (OR 0.98, 95% CI 0.71-1.37, p-value  = 0.655) and too-torn (OR 1.07, 95% CI 0.77-1.47, p-value = 0.694) compared to intact nets on malaria prevalence for all net types. However, there were reduced rates of malaria case incidence in children sleeping under a net in good condition compared to too-torn nets (incidence rate ratio (IRR) 0.76 [95% CI 0.63-0.92], p = 0.005). Malaria incidence was also consistently lower in too-torn PBO-PY LLIN (IRR = 0.37 [95% CI 0.19-0.72], p = 0.003) and chlorfenapyr-PY LLIN (IRR = 0.45 [95% CI 0.33-0.97], p = 0.053) compared to an intact PY-only LLIN during the first year of follow up. In year 2, the incidence was only significantly lower in intact chlorfenapyr-PY LLIN (IRR = 0.49 [95% CI 0.29-0.81], p = 0.006) compared to intact PY LLIN., Conclusion: The study confirmed that sleeping under a chlorfenapyr-PY LLIN or PBO-PY LLIN offered superior protection to pyrethroid-only nets even when torn. Preventing the development of holes is essential as they impact the level of protection offered against malaria infection., Trial Registration: ClinicalTrials.gov, number (NCT03554616)., (© 2024. The Author(s).)

Background: Malaria continues to kill approximately 650 000 people each year. There is evidence that some second-generation insecticide-treated nets, which combine insecticide formulations with different modes of action, are protective against malaria while the nets are new; however, evidence for their impact over 3 years is scarce. In this study, we report the third-year results of a cluster-randomised controlled trial assessing the long-term effectiveness of dual-active ingredient long-lasting insecticidal nets (LLINs)., Methods: This is a secondary analysis of a cluster-randomised controlled trial, carried out between May 23, 2019, and April 30, 2023, in southern Benin. Restricted randomisation was used to assign 60 clusters (villages or groups of villages with a minimum of 100 households) to the three study groups (1:1:1) to evaluate the efficacy of pyriproxyfen-pyrethroid LLINs and chlorfenapyr-pyrethroid LLINs compared with pyrethroid-only LLINs (reference) against malaria transmission. The study staff and communities were masked to the group allocation. The primary outcome was malaria incidence measured over the third year after LLIN distribution, in a cohort of children aged 6 months to 9 years at the time of enrolment, in the intention-to-treat population. Here, we present the data of the third year post-LLIN distribution. The trial was registered with ClinicalTrials.gov, NCT03931473., Findings: Study net use declined over the 3 years and was consistently lowest in the pyriproxyfen-pyrethroid LLIN group (at 36 months: 889 [39·4%] of 2257 participants vs 1278 [52·2%] of 2450 participants for the chlorfenapyr-pyrethroid LLIN group and 1400 [57·6%] of 2430 participants for the pyrethroid-only LLIN group). The cohort of children for the third year of follow-up (600 per group) were enrolled between April 9 and 30, 2022. Mean malaria incidence during the third year after distribution was 1·19 cases per child-year (95% CI 1·09-1·29) in the pyrethroid-only LLIN reference group, 1·21 cases per child-year (1·12-1·31) in the pyriproxyfen-pyrethroid LLIN group (hazard ratio [HR] 1·02, 95% CI 0·71-1·44; p=0·92), and 0·96 cases per child-year (0·88-1·05) in the chlorfenapyr-pyrethroid LLIN group (HR 0·80, 0·56-1·17; p=0·25). No adverse events related to study nets were reported by participants., Interpretation: During the third year, as was also observed during the first 2 years, the pyriproxyfen-pyrethroid LLIN group did not have superior protection against malaria cases compared with the standard LLIN group. In the third year, people living in the chlorfenapyr-pyrethroid LLIN group no longer benefited from greater protection against malaria cases and infections than those living in the pyrethroid-only LLIN group. This was probably influenced by lower study net use than previous years and the declining concentration of partner insecticides in the nets., Funding: UNITAID, The Global Fund., Translation: For the French translation of the abstract see Supplementary Materials section., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: The massive scale-up of long-lasting insecticidal nets (LLIN) has led to a major reduction in malaria burden in many sub-Saharan African (SSA) countries. The World Health Organization (WHO) has recently issued a strong recommendation for the use of chlorfenapyr-pyrethroid LLINs compared to standard pyrethroid-only LLINs in areas of high insecticide resistance intensity. However, there is still a lack of conclusive evidence on the efficacy of piperonyl butoxide-pyrethroid (PBO-py) LLINs, especially in West Africa, where vector composition and resistance mechanisms may be different from vectors in East Africa., Methods: This is a three-arm, superiority, triple-blinded, cluster randomised trial, with village as the unit of randomisation. This study conducted in Côte d'Ivoire will evaluate the efficacy on epidemiological and entomological outcomes of (1) the control arm: MAGNet® LN, which contains the pyrethroid, alpha-cypermethrin, (2) VEERALIN® LN, a net combining the synergist PBO and alpha-cypermethrin, and (3) Interceptor® G2 LN, which incorporates chlorfenapyr and alpha-cypermethrin, two adulticides with different mechanisms of action. A total of 33 villages with an average of 200 households per village will be identified, mapped, and randomised in a ratio of 1:1:1. Nets will be distributed at a central point following national guidelines with 1 net for every 2 people. The primary outcome of the trial will be incidence of malaria cases (confirmed by rapid diagnostic test (RDT)) in a cohort of 50 children aged 6 months to 10 years in each cluster, followed for 12 months (active case detection). Secondary outcomes are cross-sectional community prevalence of malaria infection (confirmed by RDT) in the study population at 6 and 12 months post-intervention (50 randomly selected persons per cluster), vector density, entomological inoculation rate (EIR), and phenotypic and genotypic insecticide resistance at baseline and 12 months post-intervention in 3 sentinel villages in each treatment arm., Discussion: In addition to generating further evidence for next-generation LLINs, this study will also provide the first evidence for pyrethroid-PBO nets in a West African setting. This could further inform WHO recommendations on the pragmatic use of pyrethroid-PBO nets., Trial Registration: ClinicalTrials.gov NCT05796193. Registered on April 3, 2023., (© 2024. The Author(s).)

Malaria and schistosomiasis are two important parasitic diseases that are a particular threat to young children and pregnant women in sub-Saharan Africa. Malaria and schistosomiasis prevention and control strategies primarily focus on the distribution of long-lasting insecticidal nets and the delivery of praziquantel tablets to at-risk populations in high burden settings through mass drug administration, respectively. The objective of this scoping review was to identify previous efforts to integrate malaria and schistosomiasis prevention and control programs in the literature and to summarize the strategies and approaches used in these programs following the PRISMA-ScR guidelines. We reviewed published and grey literature using a combination of keywords and search terms following themes surrounding "malaria", "Plasmodium falciparum", "Anopheles", "schistosomiasis", "Schistosoma haematobium", "Schistosoma mansoni", and "snails". Neither a date limit nor relevant terms for prevention and control were used. Out of 6374, eight articles were included in the scoping review-three articles investigated the integration of mass drug administration for schistosomiasis with the administration of antimalarials, four articles investigated the effect of administering antimalarials on malaria, schistosomiasis, and their co-infection, and one article assessed the impact of an educational intervention on malaria and schistosomiasis knowledge and preventative behaviors. Our findings suggest that there is an opportunity to link disease control programs to increase access and coverage of interventions to improve outcomes for malaria, schistosomiasis, and their co-infection. Further research is needed on the potential benefits, feasibility, and cost-effectiveness of integrating malaria and schistosomiasis prevention and control programs., Competing Interests: The authors declare that they have no competing interests., (Copyright: © 2024 Duguay et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Long-lasting insecticidal nets (LLINs) may have different impacts on distinct mosquito vector species. We assessed the efficacy of pyrethroid-pyriproxyfen and pyrethroid-chlorfenapyr LLINs on the density of Anopheles gambiae s.s. and An. coluzzii compared to pyrethroid-only nets in a three-arm cluster randomised control trial in Benin., Methods: Indoor and outdoor collections of adult mosquitoes took place in 60 clusters using human landing catches at baseline and every 3 months for 2 years. After morphological identification, around 15% of randomly selected samples of An. gambiae s.l. were dissected to determine parity, species (using PCR)., Results: Overall, a total of 46,613 mosquito specimens were collected at baseline and 259,250 in the eight quarterly collections post-net distribution. Post-net distribution, approximately 70% of the specimens of An. gambiae s.l. speciated were An. coluzzii, while the rest were mostly composed of An. gambiae s.s. with a small proportion (< 1%) of hybrids (An. gambiae/coluzzii). There was no evidence of a significant reduction in vector density indoors in either primary vector species [An. coluzzii: DR (density ratio) = 0.62 (95% CI 0.21-1.77), p = 0.3683 for the pyrethroid-pyriproxyfen LLIN and DR = 0.56 (95% CI 0.19-1.62), p = 0.2866 for the pyrethroid-chlorfenapyr LLIN, An. gambiae s.s.: DR = 0.52 (95% CI 0.18-1.46), p = 0.2192 for the pyrethroid-pyriproxyfen LLIN and DR = 0.53 (95% CI 0.19-1.46), p = 0.2222 for the pyrethroid-chlorfenapyr]. The same trend was observed outdoors. Parity rates of An. gambiae s.l. were also similar across study arms., Conclusions: Compared with pyrethroid-only LLINs, pyrethroid-chlorfenapyr LLINs and pyrethroid-pyriproxyfen LLINs performed similarly against the two primary mosquito species An. gambiae s.s. and An. coluzzii in Benin., (© 2024. The Author(s).)

Background: New classes of long-lasting insecticidal nets (LLINs) containing two active ingredients have been recently recommended by WHO in areas where malaria vectors are resistant to pyrethroids. This policy was based on evidence generated by the first 2 years of our recently published trial in Tanzania. In this Article, we report the final third-year trial findings, which are necessary for assessing the long-term effectiveness of new classes of LLIN in the community and the replacement intervals required., Methods: A third year of follow-up of a four-arm, single-blind, cluster-randomised controlled trial of dual active ingredient LLINs was conducted between July 14, 2021, and Feb 10, 2022, in Misungwi, Tanzania. Restricted randomisation was used to assign 84 clusters to the four LLIN groups (1:1:1:1) to receive either standard pyrethroid (PY) LLINs (reference), chlorfenapyr-PY LLINs, pyriproxyfen-PY LLINs, or piperonyl butoxide (PBO)-PY LLINs. All households received one LLIN for every two people. Data collection was done in consenting households in the cluster core area with at least one child between 6 months and 15 years of age who permanently resided in the selected household. Exclusion criteria were householders absent during the visit, living in the cluster buffer area, no adult caregiver capable of giving informed consent, or eligible children who were severely ill. Field staff and study participants were masked to allocation, and those analysing data were not. The primary 24-month endpoint was reported previously; here, we present the secondary outcome, malaria infection prevalence in children at 36 months post LLIN distribution, reported in the intention-to-treat analysis. The trial was registered with ClinicalTrials.gov (NCT03554616) and is now complete., Findings: Overall usage of study nets was 1023 (22·3%) of 4587 people at 36 months post distribution. In the standard PY LLIN group, malaria infection was prevalent in 407 (37·4%) of 1088 participants, compared with 261 (22·8%) of 1145 in the chlorfenapyr-PY LLIN group (odds ratio 0·57, 95% CI 0·38-0·86; p=0·0069), 338 (32·2%) of 1048 in the PBO-PY LLIN group (0·95, 0·64-1·42; p=0·80), and 302 (28·8%) of 1050 in the pyriproxyfen-PY LLIN group (0·82, 0·55-1·23; p=0·34). None of the participants or caregivers reported side-effects., Interpretation: Despite low coverage, the protective efficacy against malaria offered by chlorfenapyr-PY LLINs was superior to that provided by standard PY LLINs over a 3-year LLIN lifespan. Appropriate LLIN replacement strategies to maintain adequate usage of nets will be necessary to maximise the full potential of these nets., Funding: Department for International Development, UK Medical Research Council, Wellcome Trust, Department of Health and Social Care, and Bill & Melinda Gates Foundation via the Innovative Vector Control Consortium., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: After a decade of successful control, malaria is on the rise again. The prevalence of malaria in Tanzania has increased from 7% in 2017 to 8% in 2022 and reached 18% in Kagera region in the North West of Tanzania. Malaria vectors in Muleba district Kagera have high level of pyrethroid resistance. The aim of this paper is to explore factors associated with malaria infection prevalence in children aged 6 months to 14 years in Muleba, where Long Lasting Insecticidal Net (LLIN) combining a pyrethroid insecticide and synergist piperonyl butoxide (PBO) that counteract resistance in the mosquitoes, was first distributed under trial conditions in 2015., Methods: The trial was a community randomized control in which there were two malaria prevalence cross-sectional household surveys each year (June and December) from 2015 to 2017 in Muleba. In this study we conducted a secondary data analysis of the December surveys only. Multilevel Poisson regression analysis was used to assess factors associated with malaria infection., Results: A total of 10,941 children and 4,611 households were included in this study. Overall malaria prevalence was 35.8%, 53.3% and 54.4% in the year 2015, 2016 and 2017 respectively. Living in an area with standard LLIN as opposed to the novel PBO synergist LLIN, being a male child, above 5 years of age, living in a house with open eaves, living in house without IRS, having head of household with no formal education, lower socioeconomic status and survey year were associated with increased risk of malaria infection., Conclusions: Using PBO LLIN reduced the risk of malaria infection. However, additional measures could further reduce malaria infection in areas of insecticide resistance such as housing improvement., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Kibondo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: After decades of success in reducing malaria through the scale-up of pyrethroid long-lasting insecticidal nets (LLINs), the decline in the malaria burden has stalled, coinciding with the rapid spread of pyrethroid resistance. In a previously reported study, nets treated with a pyrethroid and a synergist, piperonyl butoxide (PBO), demonstrated superior efficacy compared to standard pyrethroid LLINs (std-LLINs) against malaria. Evidence was used to support the public health recommendation of PBO-Pyrethroid-LLIN by the World Health Organization in 2018. This study looks at the third year of rollout of these nets in Muleba district, Tanzania to inform whether policy guidelines need to be updated., Methods: A four-group cluster randomized trial (CRT) using a two-by-two factorial design was carried out between January 2014 and December 2017. A total of 48 clusters, were randomized in a 1:1:1:1 ratio to the following treatment groups, each intervention being provided once in 2015: 1/std-LLIN; 2/PBO-pyrethroid LLIN; 3/std-LLIN + Indoor Residual Spraying (IRS) and 4/PBO-Pyrethroid-LLIN + IRS. During the third year follow-up, malaria infection prevalence in 80 children per cluster, aged 6 months to 14 years, was measured at 28- and 33-months post-intervention and analysed as intention-to-treat (ITT) and per protocol (PP). Mosquito collections were performed monthly in all clusters, using CDC light traps in 7 randomly selected houses per cluster., Results: At 28 and 33 months, study net usage among household participants was only 47% and 31%, respectively. In ITT analysis, after 28 months malaria infection prevalence among 7471 children was 80.9% in the two std-LLIN groups compared to 69.3% in the two PBO-Pyrethroid-LLIN (Odds Ratio: 0.45, 95% Confidence Interval: 0.21-0.95, p-value: 0.0364). After 33 months the effect was weaker in the ITT analysis (prevalence 59.6% versus 49.9%, OR: 0.60, 95%CI:0.32-1.13, p-value: 0.1131) but still evident in the PP analysis (57.2% versus 44.2%, OR: 0.34, 95%CI: 0.16-0.71, p-value: 0.0051). Mean number of Anopheles per night collected per house was similar between PBO-Pyrethroid-LLIN groups (5.48) and std-LLIN groups (5.24) during the third year., Conclusions: Despite low usage of PBO- Pyrethroid LLIN, a small impact of those nets on malaria infection prevalence was still observed in the 3rd year with the most protection offered to children still using them. To maximize impact, it is essential that net re-distribution cycles are aligned with this LLIN lifespan to maintain maximum coverage., Trial Registration: The trial was registered with ClinicalTrials.gov (registration number NCT02288637)., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: Insecticide resistance among malaria-vector species is a pervasive problem that might jeopardise global disease-control efforts. Novel vector-control tools with different modes of action, including long-lasting insecticidal nets (LLINs) incorporating new active ingredients, are urgently needed to delay the evolution and spread of insecticide resistance. We aimed to measure phenotypic and genotypic insecticide-resistance profiles among wild Anopheles collected over 3 years to assess the longitudinal effects of dual-active-ingredient LLINs on insecticide resistance., Methods: For this analysis, data nested in a 3-year, four parallel-arm, superiority cluster-randomised controlled trial (cRCT) in Tanzania, collected from 84 clusters (39 307 households) formed of 72 villages in the Misungwi district, were used to measure insecticide-resistance profiles among female Anopheles mosquitoes via insecticide-resistance bioassays and quantitative RT-PCR of metabolic-resistance genes. Wild, blood-fed, indoor-resting mosquitoes were collected annually during the rainy seasons from house walls in clusters from all four trial groups. Mosquitoes were morphologically identified as An gambiae sensu lato (SL) or An funestus SL before separate bioassay testing. The primary outcomes were lethal-dose values for α-cypermethrin, permethrin, and piperonyl butoxide pre-exposure plus permethrin-resistance intensity bioassays, mortality 72 h after insecticidal exposure for chlorfenapyr bioassays, fertility reduction 72 h after insecticidal exposure for pyriproxyfen bioassays, and fold change in metabolic-enzyme expression relative to an insecticide-susceptible laboratory strain. All primary outcomes were measured in An funestus SL 1 year, 2 years, and 3 years after LLIN distribution. Primary outcomes were also assessed in An gambiae SL if enough mosquitoes were collected. The cRCT is registered with ClinicalTrials.gov (NCT03554616)., Findings: Between May 24, 2019, and Oct 25, 2021, 47 224 female Anopheles were collected for resistance monitoring. In the pyrethroid (PY)-LLIN group, there were significant increases in α-cypermethrin-resistance intensity (year 1 LD50=9·52 vs year 2 76·20, p<0·0001) and permethrin-resistance intensity (year 1 13·27 vs year 2 35·83, p=0·0019) in An funestus SL. In the pyriproxyfen PY-LLIN group, there was similar increase in α-cypermethrin-resistance intensity (year 1 0·71 vs year 2 81·56, p<0·0001) and permethrin-resistance intensity (year 1 5·68 vs year 2 50·14, p<0·0001). In the piperonyl butoxide PY-LLIN group, α-cypermethrin-resistance intensity (year 1 33·26 vs year 3 70·22, p=0·0071) and permethrin-resistance intensity (year 1 47·09 vs year 3 2635·29, p<0·0001) also increased over time. In the chlorfenapyr PY-LLIN group, there were no effects on α-cypermethrin-resistance intensity (year 1 0·42 vs year 3 0·99, p=0·54) or permethrin-resistance intensity (data were not estimable due to nearly 100% mortality). There were also minimal reductions in chlorfenapyr susceptibility. However, in the chlorfenapyr PY-LLIN group, a significant decline in piperonyl-butoxide synergy was seen by year 3 (year 1 0·02 vs year 3 0·26, p=0·020). Highly over-expressed detoxification enzymes showed dynamic patterns of selection throughout the trial., Interpretation: Our phenotypic data supports trial epidemiological findings; chlorfenapyr PY-LLINs provided superior protection from malaria across multiple transmission seasons, with few effects on insecticide-resistance selection. Rapid pyrethroid-resistance intensification in the piperonyl butoxide PY-LLIN group and pre-existing tolerance of pyriproxyfen in vector populations might explain the poorer performance of these two interventions regarding malaria outcomes. Further work is required to elucidate the potential mechanisms driving cross-resistance between pyrethroids and novel active ingredients to better inform the design of pre-emptive resistance-management strategies., Funding: UK Department for International Development; UK Medical Research Council; Wellcome Trust; UK Department of Health and Social Care; UK Foreign, Commonwealth and Development Office; and The Bill and Melinda Gates Foundation via the Innovative Vector Control Consortium., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: Gains in malaria control are threatened by widespread pyrethroid resistance in malaria vectors across sub-Saharan Africa. New long-lasting insecticidal nets (LLINs) containing two active ingredients (dual active-ingredient LLINs) have been developed to interrupt transmission in areas of pyrethroid resistance. We aimed to evaluate the effectiveness of three dual active-ingredient LLINs compared with standard pyrethroid LLINs against pyrethroid-resistant malaria vectors in rural Tanzania., Methods: In this study, we did a secondary analysis of entomological data from a four-group, 3 year, single-blind, cluster-randomised controlled trial carried out between Feb 18, 2019, and Dec 6, 2021. We conducted quarterly indoor mosquito collections using the Centers for Disease Control and Prevention light trap, in eight houses in each of the 84 study clusters in the Misungwi district, northwestern Tanzania. Anopheles vectors were then tested for malaria parasites and identified at species level, to distinguish between sibling species of the Anopheles gambiae and Anopheles funestus groups, using molecular laboratory techniques. The primary outcomes were density of different malaria vector species measured as the number of female Anopheles collected per household per night, the entomological inoculation rate (EIR), an indicator of malaria transmission, and sporozoite rate. Entomological outcomes were assessed on the basis of intention to treat, and the effect of the three dual active-ingredient LLINs was compared with the standard pyrethroid LLINs at household level., Findings: Dual active-ingredient LLINs had the greatest effect on Anopheles funestus sl, the most efficient vector in the study area, with comparatively weak effect on An arabiensis. An funestus density was 3∙1 per house per night in the pyrethroid LLIN group, 1∙2 in the chlorfenapyr pyrethroid LLIN group (adjusted density ratio [aDR]=0∙26, 95% CI 0∙17-0∙14, p<0∙0001), 1∙4 in the piperonyl-butoxide pyrethroid LLIN group (aDR=0∙49, 0∙32-0∙76, p=0∙0012), and 3∙0 in the pyriproxyfen pyrethroid LLIN group (aDR=0∙72, 0∙47-1∙11, p=0∙15). Malaria transmission intensity was also significantly lower in the chlorfenapyr pyrethroid group, with 0∙01 versus 0∙06 infective bites per household per night in the pyrethroid LLIN group (aDR=0∙21, 0∙14-0∙33, p<0∙0001). Ecological niche models indicated that vector-species distribution was stable following LLIN intervention despite the reductions observed in An funestus sl density., Interpretation: Chlorfenapyr pyrethroid LLINs were the most effective intervention against the main malaria vector An funestus sl over 3 years of community use, whereas the effect of piperonyl-butoxide pyrethroid LLIN was sustained for 2 years. The other vector, An arabiensis, was not controlled by any of the dual active-ingredient LLINs. Additional vector control tools and strategies targeted to locally prevalent vector species evading dual active-ingredient LLINs should be deployed to further reduce malaria transmission and achieve elimination., Funding: The Department for International Development, UK Medical Research Council, Wellcome Trust, the Department of Health and Social Care, and The Bill & Melinda Gates Foundation via the Innovative Vector Control Consortium., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: The effectiveness of malaria vector control interventions is often evaluated using cluster randomized trials (CRT) with outcomes assessed using repeated cross-sectional surveys. A key requirement for appropriate design and analysis of longitudinal CRTs is accounting for the intra-cluster correlation coefficient (ICC). In addition to exchangeable correlation (constant ICC over time), correlation structures proposed for longitudinal CRT are block exchangeable (allows a different within- and between-period ICC) and exponential decay (allows between-period ICC to decay exponentially). More flexible correlation structures are available in statistical software packages and, although not formally proposed for longitudinal CRTs, may offer some advantages. Our objectives were to empirically explore the impact of these correlation structures on treatment effect inferences, identify gaps in the methodological literature, and make practical recommendations., Methods: We obtained data from a parallel-arm CRT conducted in Tanzania to compare four different types of insecticide-treated bed-nets. Malaria prevalence was assessed in cross-sectional surveys of 45 households in each of 84 villages at baseline, 12-, 18- and 24-months post-randomization. We re-analyzed the data using mixed-effects logistic regression according to a prespecified analysis plan but under five different correlation structures as well as a robust variance estimator under exchangeable correlation and compared the estimated correlations and treatment effects. A proof-of-concept simulation was conducted to explore general conclusions., Results: The estimated correlation structures varied substantially across different models. The unstructured model was the best-fitting model based on information criteria. Although point estimates and confidence intervals for the treatment effect were similar, allowing for more flexible correlation structures led to different conclusions based on statistical significance. Use of robust variance estimators generally led to wider confidence intervals. Simulation results showed that under-specification can lead to coverage probabilities much lower than nominal levels, but over-specification is more likely to maintain nominal coverage., Conclusion: More flexible correlation structures should not be ruled out in longitudinal CRTs. This may be particularly important in malaria trials where outcomes may fluctuate over time. In the absence of robust methods for selecting the best-fitting correlation structure, researchers should examine sensitivity of results to different assumptions about the ICC and consider robust variance estimators., (© 2023. The Author(s).)

Background: New classes of long-lasting insecticidal nets (LLINs) combining mixtures of insecticides with different modes of action could put malaria control back on track after rebounds in transmission across sub-Saharan Africa. We evaluated the relative efficacy of pyriproxyfen-pyrethroid LLINs and chlorfenapyr-pyrethroid LLINs compared with standard LLINs against malaria transmission in an area of high pyrethroid resistance in Benin., Methods: We conducted a cluster-randomised, superiority trial in Zou Department, Benin. Clusters were villages or groups of villages with a minimum of 100 houses. We used restricted randomisation to randomly assign 60 clusters to one of three LLIN groups (1:1:1): to receive nets containing either pyriproxyfen and alpha-cypermethrin (pyrethroid), chlorfenapyr and alpha-cypermethrin, or alpha-cypermethrin only (reference). Households received one LLIN for every two people. The field team, laboratory staff, analyses team, and community members were masked to the group allocation. The primary outcome was malaria case incidence measured over 2 years after net distribution in a cohort of children aged 6 months-10 years, in the intention-to-treat population. This study is ongoing and is registered with ClinicalTrials.gov, NCT03931473., Findings: Between May 23 and June 24, 2019, 53 854 households and 216 289 inhabitants were accounted for in the initial census and included in the study. Between March 19 and 22, 2020, 115 323 LLINs were distributed to 54 030 households in an updated census. A cross-sectional survey showed that study LLIN usage was highest at 9 months after distribution (5532 [76·8%] of 7206 participants), but decreased by 24 months (4032 [60·6%] of 6654). Mean malaria incidence over 2 years after LLIN distribution was 1·03 cases per child-year (95% CI 0·96-1·09) in the pyrethroid-only LLIN reference group, 0·84 cases per child-year (0·78-0·90) in the pyriproxyfen-pyrethroid LLIN group (hazard ratio [HR] 0·86, 95% CI 0·65-1·14; p=0·28), and 0·56 cases per child-year (0·51-0·61) in the chlorfenapyr-pyrethroid LLIN group (HR 0·54, 95% CI 0·42-0·70; p<0·0001)., Interpretation: Over 2 years, chlorfenapyr-pyrethroid LLINs provided greater protection from malaria than pyrethroid-only LLINs in an area with pyrethroid-resistant mosquitoes. Pyriproxyfen-pyrethroid LLINs conferred protection similar to pyrethroid-only LLINs. These findings provide crucial second-trial evidence to enable WHO to make policy recommendations on these new LLIN classes. This study confirms the importance of chlorfenapyr as an LLIN treatment to control malaria in areas with pyrethroid-resistant vectors. However, an arsenal of new active ingredients is required for successful long-term resistance management, and additional innovations, including pyriproxyfen, need to be further investigated for effective vector control strategies., Funding: UNITAID, The Global Fund., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

To control pyrethroid-resistant malaria vectors, Indoor Residual Spraying (IRS) and Long-Lasting Insecticidal Nets (LLINs) that include additional ingredients to pyrethroid are being developed. Same progress needs to be made to the pyrethroid-treated blankets, which are more compatible with shelter structures found in emergency settings such as displaced populations. In the current study, efficacy of blankets treated with permethrin and piperonyl butoxide (PBO) was evaluated against pyrethroid-resistant Anopheles gambiae sensu stricto. Efficacy was compared with that of Olyset LLIN, Olyset Plus LLIN and untreated blanket in terms of mortality and blood-feeding inhibition against pyrethroid-resistant Anopheles gambiae mosquitoes. The current study indicates that, in emergency shelters such as migrant and refugee camps where LLINs cannot be used, PBO-permethrin blankets may provide protection against resistant mosquitoes if widely used. No side effects related to the use of the treated blankets were reported from the participants. These results need validation in a large-scale field trial to assess the epidemiological impact of the intervention, durability and acceptability of this new vector control strategy for malaria vector control., (© 2022. The Author(s).)

Background: Long-lasting insecticidal nets (LLINs) have successfully reduced malaria in sub-Saharan Africa, but their effectiveness is now partly compromised by widespread resistance to insecticides among vectors. We evaluated new classes of LLINs with two active ingredients with differing modes of action against resistant malaria vectors., Methods: We did a four-arm, cluster-randomised trial in Misungwi, Tanzania. Clusters were villages, or groups of hamlets, with at least 119 households containing children aged 6 months to 14 years living in the cluster's core area. Constrained randomisation was used to allocate clusters (1:1:1:1) to receive one of four types of LLIN treated with the following: α-cypermethrin only (pyrethroid-only [reference] group); pyriproxyfen and α-cypermethrin (pyriproxyfen group); chlorfenapyr and α-cypermethrin (chlorfenapyr group); or the synergist piperonyl butoxide and permethrin (piperonyl butoxide group). At least one LLIN was distributed for every two people. Community members and the field team were masked to group allocation. Malaria prevalence data were collected through cross-sectional surveys of randomly selected households from each cluster, in which children aged 6 months to 14 years were assessed for Plasmodium falciparum malaria infection by rapid diagnostic tests. The primary outcome was malaria infection prevalence at 24 months after LLIN distribution, comparing each of the dual-active-ingredient LLINs to the standard pyrethroid-only LLINs in the intention-to-treat population. The primary economic outcome was cost-effectiveness of dual-active-ingredient LLINs, based on incremental cost per disability-adjusted life-year (DALY) averted compared with pyrethroid-only LLINs, modelled over a 2-year period; we included costs of net procurement and malaria diagnosis and treatment, and estimated DALYs in all age groups. This study is registered with ClinicalTrials.gov (NCT03554616), and is ongoing but no longer recruiting., Findings: 84 clusters comprising 39 307 households were included in the study between May 11 and July 2, 2018. 147 230 LLINs were distributed among households between Jan 26 and Jan 28, 2019. Use of study LLINs was reported in 3155 (72·1%) of 4378 participants surveyed at 3 months post-distribution and decreased to 8694 (40·9%) of 21 246 at 24 months, with varying rates of decline between groups. Malaria infection prevalence at 24 months was 549 (45·8%) of 1199 children in the pyrethroid-only reference group, 472 (37·5%) of 1258 in the pyriproxyfen group (adjusted odds ratio 0·79 [95% CI 0·54-1·17], p=0·2354), 512 (40·7%) of 1259 in the piperonyl butoxide group (0·99 [0·67-1·45], p=0·9607), and 326 [25·6%] of 1272 in the chlorfenapyr group (0·45 [0·30-0·67], p=0·0001). Skin irritation or paraesthesia was the most commonly reported side-effect in all groups. Chlorfenapyr LLINs were the most cost-effective LLINs, costing only US$19 (95% uncertainty interval 1-105) more to public providers or $28 (11-120) more to donors per DALY averted over a 2-year period compared with pyrethroid-only LLINs, and saving costs from societal and household perspectives., Interpretation: After 2 years, chlorfenapyr LLINs provided significantly better protection than pyrethroid-only LLINs against malaria in an area with pyrethroid-resistant mosquitoes, and the additional cost of these nets would be considerably below plausible cost-effectiveness thresholds ($292-393 per DALY averted). Before scale-up of chlorfenapyr LLINs, resistance management strategies are needed to preserve their effectiveness. Poor textile and active ingredient durability in the piperonyl butoxide and pyriproxyfen LLINs might have contributed to their relative lack of effectiveness compared with standard LLINs., Funding: Joint Global Health Trials scheme (UK Foreign, Commonwealth and Development Office; UK Medical Research Council; Wellcome; UK Department of Health and Social Care), US Agency for International Development, President's Malaria Initiative., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Anopheles funestus is playing an increasing role in malaria transmission in parts of sub-Saharan Africa, where An. gambiae s.s. has been effectively controlled by long-lasting insecticidal nets. We investigated vector population bionomics, insecticide resistance and malaria transmission dynamics in 86 study clusters in North-West Tanzania. An. funestus s.l. represented 94.5% (4740/5016) of all vectors and was responsible for the majority of malaria transmission (96.5%), with a sporozoite rate of 3.4% and average monthly entomological inoculation rate (EIR) of 4.57 per house. Micro-geographical heterogeneity in species composition, abundance and transmission was observed across the study district in relation to key ecological differences between northern and southern clusters, with significantly higher densities, proportions and EIR of An. funestus s.l. collected from the South. An. gambiae s.l. (5.5%) density, principally An. arabiensis (81.1%) and An. gambiae s.s. (18.9%), was much lower and closely correlated with seasonal rainfall. Both An. funestus s.l. and An. gambiae s.l. were similarly resistant to alpha-cypermethrin and permethrin. Overexpression of CYP9K1, CYP6P3, CYP6P4 and CYP6M2 and high L1014S-kdr mutation frequency were detected in An. gambiae s.s. populations. Study findings highlight the urgent need for novel vector control tools to tackle persistent malaria transmission in the Lake Region of Tanzania.

Introduction: The massive scale-up of long-lasting insecticidal nets (LLINs) has led to major reductions in malaria burden in many sub-Saharan African countries. This progress is threatened by widespread insecticide resistance among malaria vectors. This cluster-randomised controlled trial (c-RCT) compares three of the most promising dual active ingredients LLINs (dual-AI LLINs), which incorporate mixtures of insecticides or insecticide synergists to standard LLINs in an area of pyrethroid insecticide resistance., Methods: A four-arm, single-blinded, c-RCT will evaluate the effectiveness of three types of dual-AI LLINs (1) Royal Guard, combining two insecticides, pyriproxyfen and the pyrethroid alpha-cypermethrin; (2) Interceptor G2, combining chlorfenapyr and alpha-cypermethrin; (3) Olyset Plus, an LLIN combining a synergist, piperonyl butoxide and the pyrethroid permethrin, compared with; (4) Interceptor LN, a standard LLIN containing the pyrethroid alpha-cypermethrin as the sole AI. The primary outcomes are malaria infection prevalence in children aged 6 months-14 years and entomological inoculation rate (EIR), as a standard measure of malaria transmission at 24 months postintervention and cost-effectiveness., Ethics and Dissemination: Ethical approval was received from the institutional review boards of the Tanzanian National Institute for Medical Research, Kilimanjaro Christian Medical University College, London School of Hygiene and Tropical Medicine, and University of Ottawa. Study findings will be actively disseminated via reports and presentations to stakeholders, local community leaders, and relevant national and international policy makers as well as through conferences, and peer-reviewed publications., Trial Registration Number: NCT03554616., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY. Published by BMJ.)

Background: Long-lasting insecticidal nets (LLINs) are currently the primary method of malaria control in sub-Saharan Africa and have contributed to a significant reduction in malaria burden over the past 15 years. However, this progress is threatened by the wide-scale selection of insecticide-resistant malaria vectors. It is, therefore, important to accelerate the generation of evidence for new classes of LLINs., Methods: This protocol presents a three-arm superiority, single-blinded, cluster randomized controlled trial to evaluate the impact of 2 novel dual-active ingredient LLINs on epidemiological and entomological outcomes in Benin, a malaria-endemic area with highly pyrethroid-resistant vector populations. The study arms consist of (i) Royal Guard® LLIN, a net combining a pyrethroid (alpha-cypermethrin) plus an insect growth regulator (pyriproxyfen), which in the adult female is known to disrupt reproduction and egg fertility; (ii) Interceptor G2® LLIN, a net incorporating two adulticides (alpha-cypermethrin and chlorfenapyr) with different modes of action; and (iii) the control arm, Interceptor® LLIN, a pyrethroid (alpha-cypermethrin) only LLIN. In all arms, one net for every 2 people will be distributed to each household. Sixty clusters were identified and randomised 1:1:1 to each study arm. The primary outcome is malaria case incidence measured over 24 months through active case detection in a cohort of 25 children aged 6 months to 10 years, randomly selected from each cluster. Secondary outcomes include 1) malaria infection prevalence (all ages) and prevalence of moderate to severe anaemia in children under 5 years old, measured at 6 and 18 months post-intervention; 2) entomological indices measured every 3 months using human landing catches over 24 months. Insecticide resistance intensity will also be monitored over the study period., Discussion: This study is the second cluster randomised controlled trial to evaluate the efficacy of these next-generation LLINs to control malaria transmitted by insecticide-resistant mosquitoes. The results of this study will form part of the WHO evidence-based review to support potential public health recommendations of these nets and shape malaria control strategies of sub-Saharan Africa for the next decade., Trial Registration: ClinicalTrials.gov, NCT03931473 , registered on 30 April 2019.

Background: Progress in malaria control is under threat by wide-scale insecticide resistance in malaria vectors. Two recent vector control products have been developed: a long-lasting insecticidal net that incorporates a synergist piperonyl butoxide (PBO) and a long-lasting indoor residual spraying formulation of the insecticide pirimiphos-methyl. We evaluated the effectiveness of PBO long-lasting insecticidal nets versus standard long-lasting insecticidal nets as single interventions and in combination with the indoor residual spraying of pirimiphos-methyl., Methods: We did a four-group cluster randomised controlled trial using a two-by-two factorial design of 48 clusters derived from 40 villages in Muleba (Kagera, Tanzania). We randomly assigned these clusters using restricted randomisation to four groups: standard long-lasting insecticidal nets, PBO long-lasting insecticidal nets, standard long-lasting insecticidal nets plus indoor residual spraying, or PBO long-lasting insecticidal nets plus indoor residual spraying. Both standard and PBO nets were distributed in 2015. Indoor residual spraying was applied only once in 2015. We masked the inhabitants of each cluster to the type of nets received, as well as field staff who took blood samples. Neither the investigators nor the participants were masked to indoor residual spraying. The primary outcome was the prevalence of malaria infection in children aged 6 months to 14 years assessed by cross-sectional surveys at 4, 9, 16, and 21 months after intervention. The endpoint for assessment of indoor residual spraying was 9 months and PBO long-lasting insecticidal nets was 21 months. This trial is registered with ClinicalTrials.gov, number NCT02288637., Findings: 7184 (68·0%) of 10 560 households were selected for post-intervention survey, and 15 469 (89·0%) of 17 377 eligible children from the four surveys were included in the intention-to-treat analysis. Of the 878 households visited in the two indoor residual spraying groups, 827 (94%) had been sprayed. Reported use of long-lasting insecticidal nets, across all groups, was 15 341 (77·3%) of 19 852 residents after 1 year, decreasing to 12 503 (59·2%) of 21 105 in the second year. Malaria infection prevalence after 9 months was lower in the two groups that received PBO long-lasting insecticidal nets than in the two groups that received standard long-lasting insecticidal nets (531 [29%] of 1852 children vs 767 [42%] of 1809; odds ratio [OR] 0·37, 95% CI 0·21-0·65; p=0·0011). At the same timepoint, malaria prevalence in the two groups that received indoor residual spraying was lower than in groups that did not receive indoor residual spraying (508 [28%] of 1846 children vs 790 [44%] of 1815; OR 0·33, 95% CI 0·19-0·55; p<0·0001) and there was evidence of an interaction between PBO long-lasting insecticidal nets and indoor residual spraying (OR 2·43, 95% CI 1·19-4·97; p=0·0158), indicating redundancy when combined. The PBO long-lasting insecticidal net effect was sustained after 21 months with a lower malaria prevalence than the standard long-lasting insecticidal net (865 [45%] of 1930 children vs 1255 [62%] of 2034; OR 0·40, 0·20-0·81; p=0·0122)., Interpretation: The PBO long-lasting insecticidal net and non-pyrethroid indoor residual spraying interventions showed improved control of malaria transmission compared with standard long-lasting insecticidal nets where pyrethroid resistance is prevalent and either intervention could be deployed to good effect. As a result, WHO has since recommended to increase coverage of PBO long-lasting insecticidal nets. Combining indoor residual spraying with pirimiphos-methyl and PBO long-lasting insecticidal nets provided no additional benefit compared with PBO long-lasting insecticidal nets alone or standard long-lasting insecticidal nets plus indoor residual spraying., Funding: UK Department for International Development, Medical Research Council, and Wellcome Trust., (Copyright © 2018 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: Insecticide-treated nets (ITNs) and indoor residual spraying (IRS) of houses provide effective malaria transmission control. There is conflicting evidence about whether it is more beneficial to provide both interventions in combination. A cluster randomised controlled trial was conducted to investigate whether the combination provides added protection compared to ITNs alone., Methods and Findings: In northwest Tanzania, 50 clusters (village areas) were randomly allocated to ITNs only or ITNs and IRS. Dwellings in the ITN+IRS arm were sprayed with two rounds of bendiocarb in 2012. Plasmodium falciparum prevalence rate (PfPR) in children 0.5-14 y old (primary outcome) and anaemia in children <5 y old (secondary outcome) were compared between study arms using three cross-sectional household surveys in 2012. Entomological inoculation rate (secondary outcome) was compared between study arms. IRS coverage was approximately 90%. ITN use ranged from 36% to 50%. In intention-to-treat analysis, mean PfPR was 13% in the ITN+IRS arm and 26% in the ITN only arm, odds ratio = 0.43 (95% CI 0.19-0.97, n = 13,146). The strongest effect was observed in the peak transmission season, 6 mo after the first IRS. Subgroup analysis showed that ITN users were additionally protected if their houses were sprayed. Mean monthly entomological inoculation rate was non-significantly lower in the ITN+IRS arm than in the ITN only arm, rate ratio = 0.17 (95% CI 0.03-1.08)., Conclusions: This is the first randomised trial to our knowledge that reports significant added protection from combining IRS and ITNs compared to ITNs alone. The effect is likely to be attributable to IRS providing added protection to ITN users as well as compensating for inadequate ITN use. Policy makers should consider deploying IRS in combination with ITNs to control transmission if local ITN strategies on their own are insufficiently effective. Given the uncertain generalisability of these findings, it would be prudent for malaria control programmes to evaluate the cost-effectiveness of deploying the combination., Trial Registration: www.ClinicalTrials.gov NCT01697852 Please see later in the article for the Editors' Summary.

Background: Insecticide-treated nets (ITN) are one of the most effective measures for preventing malaria. Mass distribution campaigns are being used to rapidly increase net coverage in at-risk populations. This study had two purposes: to evaluate the impact of a universal coverage campaign (UCC) of long-lasting insecticidal nets (LLINs) on LLIN ownership and usage, and to identify factors that may be associated with inadequate coverage., Methods: In 2011 two cross-sectional household surveys were conducted in 50 clusters in Muleba district, north-west Tanzania. Prior to the UCC 3,246 households were surveyed and 2,499 afterwards. Data on bed net ownership and usage, demographics of household members and household characteristics including factors related to socio-economic status were gathered, using an adapted version of the standard Malaria Indicator Survey. Specific questions relating to the UCC process were asked., Results: The proportion of households with at least one ITN increased from 62.6% (95% Confidence Interval (CI) = 60.9-64.2) before the UCC to 90.8% (95% CI = 89.0-92.3) afterwards. ITN usage in all residents rose from 40.8% to 55.7%. After the UCC 58.4% (95% CI = 54.7-62.1) of households had sufficient ITNs to cover all their sleeping places. Households with children under five years (OR = 2.4, 95% CI = 1.9-2.9) and small households (OR = 1.9, 95% CI = 1.5-2.4) were most likely to reach universal coverage. Poverty was not associated with net coverage. Eighty percent of households surveyed received LLINs from the campaign., Conclusions: The UCC in Muleba district of Tanzania was equitable, greatly improving LLIN ownership and, more moderately, usage. However, the goal of universal coverage in terms of the adequate provision of nets was not achieved. Multiple, continuous delivery systems and education activities are required to maintain and improve bed net ownership and usage.

Background: Long-lasting insecticidal nets (LLIN) are now standard for the prevention of malaria. However, only products with recommendation for public use from the World Health Organization should be used and this evaluation includes the assessment of net effectiveness after three years of field use. Results for one of the polyester-based products, Interceptor is presented., Methods: In five villages, 190 LLIN and 90 nets conventionally treated with the insecticide alpha-cypermethrin at 25 mg/m2 were distributed randomly and used by the families. Following a baseline household survey a net survey was carried out every six months to capture use, washing habits and physical condition of the nets. Randomly selected nets were collected after 6, 12, 24, 36 and 42 months and tested for remaining insecticide content and ability to knock-down and kill malaria transmitting mosquitoes., Results: During the three and a half years of observation only 16 nets were lost to follow-up resulting in an estimated attrition rate of 12% after three and 20/% after 3.5 years. Nets were used regularly and washed on average 1.5 times per year. After three and a half years 29% of the nets were still in good condition while 13% were seriously torn with no difference between the LLIN and control nets. The conventionally treated nets quickly lost insecticide and after 24 months only 7% of the original dose remained (1.6 mg/m2). Baseline median concentration of alpha-cypermethrin for LLIN was 194.5 mg/m2 or 97% of the target dose with between and within net variation of 11% and 4% respectively (relative standard deviation). On the LLIN 73.8 mg/m2 alpha-cypermethrin remained after three years of use and 56.2 mg/m2 after three and a half and 94% and 81% of the LLIN still had > 15 mg/m2 left respectively. Optimal effectiveness in bio-assays (≥ 95% 60 minute knock-down or ≥ 80% 24 hour mortality) was found in 83% of the sampled LLIN after three and 71% after three and a half years., Conclusions: Under conditions in Western Uganda the tested long-lasting insecticidal net Interceptor fulfilled the criteria for phase III of WHO evaluations and, based on preliminary criteria of the useful life, this product is estimated to last on average between three and four years.

Introduction: Malaria is re-emerging in most of the African highlands exposing the non immune population to deadly epidemics. A better understanding of the factors impacting transmission in the highlands is crucial to improve well targeted malaria control strategies., Methods and Findings: A conceptual model of potential malaria risk factors in the highlands was built based on the available literature. Furthermore, the relative importance of these factors on malaria can be estimated through "classification and regression trees", an unexploited statistical method in the malaria field. This CART method was used to analyse the malaria risk factors in the Burundi highlands. The results showed that Anopheles density was the best predictor for high malaria prevalence. Then lower rainfall, no vector control, higher minimum temperature and houses near breeding sites were associated by order of importance to higher Anopheles density., Conclusions: In Burundi highlands monitoring Anopheles densities when rainfall is low may be able to predict epidemics. The conceptual model combined with the CART analysis is a decision support tool that could provide an important contribution toward the prevention and control of malaria by identifying major risk factors.

In a highland province of Burundi, indoor residual spraying and long-lasting insecticidal net distribution were targeted in the valley, aiming also to protect the population living on the hilltops. The impact on malaria indicators was assessed, and the potential additional effect of nets evaluated. After the intervention--and compared with the control valleys--children 1-9 years old in the treated valleys had lower risks of malaria infection (odds ratio, OR: 0.55), high parasite density (OR: 0.48), and clinical malaria (OR: 0.57). The impact on malaria prevalence was even higher in infants (OR: 0.14). Using nets did not confer an additional protective effect to spraying. Targeted vector control had a major impact on malaria in the high-risk valleys but not in the less-exposed hilltops. Investment in targeted and regular control measures associated with effective case management should be able to control malaria in the highlands.

Background: Prevention of malaria epidemics is a priority for African countries. The 2000 malaria epidemic in Burundi prompted the government to implement measures for preventing future outbreaks. Case management with artemisinin-based combination therapy and malaria surveillance were nationally improved. A vector control programme was initiated in one of the most affected highland provinces. The focal distribution of malaria vectors in the highlands was the starting point for designing a targeted vector control strategy. The objective of this study was to present the results of this strategy on malaria transmission in an African highland region., Methods: In Karuzi, in 2002-2005, vector control activities combining indoor residual spraying and long-lasting insecticidal nets were implemented. The interventions were done before the expected malaria transmission period and targeted the valleys between hills, with the expectation that this would also protect the populations living at higher altitudes. The impact on the Anopheles population and on malaria transmission was determined by nine cross-sectional surveys carried out at regular intervals throughout the study period., Results: Anopheles gambiae s.l. and Anopheles funestus represented 95% of the collected anopheline species. In the valleys, where the vector control activities were implemented, Anopheles density was reduced by 82% (95% CI: 69-90). Similarly, transmission was decreased by 90% (95% CI: 63%-97%, p = 0.001). In the sprayed valleys, Anopheles density was further reduced by 79.5% (95% CI: 51.7-91.3, p < 0.001) in the houses with nets as compared to houses without them. No significant impact on vector density and malaria transmission was observed in the hill tops. However, the intervention focused on the high risk areas near the valley floor, where 93% of the vectors are found and 90% of the transmission occurs., Conclusion: Spatial targeted vector control effectively reduced Anopheles density and transmission in this highland district. Bed nets have an additional effect on Anopheles density though this did not translate in an additional impact on transmission. Though no impact was observed in the hilltops, the programme successfully covered the areas most at risk. Such a targeted strategy could prevent the emergence and spread of an epidemic from these high risk foci.

Background: African highlands often suffer of devastating malaria epidemics, sometimes in conjunction with complex emergencies, making their control even more difficult. In 2000, Burundian highlands experienced a large malaria outbreak at a time of civil unrest, constant insecurity and nutritional emergency. Because of suspected high resistance to the first and second line treatments, the provincial health authority and Médecins Sans Frontières (Belgium) decided to implement vector control activities in an attempt to curtail the epidemic. There are few reported interventions of this type to control malaria epidemics in complex emergency contexts. Here, decisions and actions taken to control this epidemic, their impact and the lessons learned from this experience are reported., Case Description: Twenty nine hills (administrative areas) were selected in collaboration with the provincial health authorities for the vector control interventions combining indoor residual spraying with deltamethrin and insecticide-treated nets. Impact was evaluated by entomological and parasitological surveys. Almost all houses (99%) were sprayed and nets use varied between 48% and 63%. Anopheles indoor resting density was significantly lower in treated as compared to untreated hills, the latter taken as controls. Despite this impact on the vector, malaria prevalence was not significantly lower in treated hills except for people sleeping under a net., Discussion: Indoor spraying was feasible and resulted in high coverage despite being a logistically complex intervention in the Burundian context (scattered houses and emergency situation). However, it had little impact on the prevalence of malaria infection, possibly because it was implemented after the epidemic's peak. Nevertheless, after this outbreak the Ministry of Health improved the surveillance system, changed its policy with introduction of effective drugs and implementation of vector control to prevent new malaria epidemics., Conclusion: In the absence of effective drugs and sufficient preparedness, present study failed to demonstrate any impact of vector control activities upon the course of a short-duration malaria epidemic. However, the experience gained lead to increased preparedness and demonstrated the feasibility of vector control measures in this specific context.

Simple Summary: New types of bed nets are being developed which contain a pyrethroid plus a second chemical because of the development and increased frequency of mosquito mechanisms to avoid being killed by pyrethroids. When insecticide-treated bed nets are being trialed for efficacy or released onto the market it is essential to measure how effective the net is in killing mosquitoes, which includes testing how quickly insecticide is lost or degraded due to routine wear and tear. Pyrethroid-resistant mosquitoes are needed to test the effectiveness and insecticidal persistence of the second chemical, because they are not all killed by the pyrethroid, allowing the killing effect of the two chemicals to be evaluated independently. However, because resistance status varies between populations of mosquitoes that possess different resistance mechanisms, and because resistance intensity in a population can change over time, a method is needed to characterise the resistant mosquitoes. A focus group of experts discussed how this should best be done, considering pros and cons of different approaches, and co-wrote a protocol, which will be valuable for malaria control programmes and stakeholders wanting to test the effective lifespan of a new bed net in terms of the active ingredient bioefficacy. Durability monitoring of insecticide-treated nets (ITNs) containing a pyrethroid in combination with a second active ingredient (AI) must be adapted so that the insecticidal bioefficacy of each AI can be monitored independently. An effective way to do this is to measure rapid knock down of a pyrethroid-susceptible strain of mosquitoes to assess the bioefficacy of the pyrethroid component and to use a pyrethroid-resistant strain to measure the bioefficacy of the second ingredient. To allow robust comparison of results across tests within and between test facilities, and over time, protocols for bioefficacy testing must include either characterisation of the resistant strain, standardisation of the mosquitoes used for bioassays, or a combination of the two. Through a series of virtual meetings, key stakeholders and practitioners explored different approaches to achieving these goals. Via an iterative process we decided on the preferred approach and produced a protocol consisting of characterising mosquitoes used for bioefficacy testing before and after a round of bioassays, for example at each time point in a durability monitoring study. We present the final protocol and justify our approach to establishing a standard methodology for durability monitoring of ITNs containing pyrethroid and a second AI. [ABSTRACT FROM AUTHOR]

Long lasting insecticidal nets (LLINs) are a proven tool to reduce malaria transmission, but in Africa efficacy is being reduced by pyrethroid resistance in the major vectors. A previous study that was conducted in Muleba district, Tanzania indicated possible involvement of cytochrome P450 monooxygenases in a pyrethroid resistance in An. gambiae population where pre-exposure to piperonyl butoxide (PBO) followed by permethrin exposure in CDC bottle bioassays led to partial restoration of susceptibility. PBO is a synergist that can block pyrethroid-metabolizing enzymes in a mosquito. Insecticide resistance profiles and underlying mechanisms were investigated in Anopheles gambiae and An. funestus from Muleba during a cluster randomized trial. Diagnostic dose bioassays using permethrin, together with intensity assays, suggest pyrethroid resistance that is both strong and very common, but not extreme. Transcriptomic analysis found multiple P450 genes over expressed including CYP6M2, CYP6Z3, CYP6P3, CYP6P4, CYP6AA1 and CYP9K1 in An. gambiae and CYP6N1, CYP6M7, CYP6M1 and CYP6Z1 in An. funestus. Indeed, very similar suites of P450 enzymes commonly associated with resistant populations elsewhere in Africa were detected as over expressed suggesting a convergence of mechanisms across Sub-Saharan African malaria vectors. The findings give insight into factors that may correlate with pyrethroid PBO LLIN success, broadly supporting model predictions, but revision to guidelines previously issued by the World Health Organization is warranted. [ABSTRACT FROM AUTHOR]

Outdoor transmission of malaria and other vector borne diseases remains a problem. The WHO has recently recognized the need for suitable methods for assessing vector density outdoors and a number of tent-traps have been developed. Only one such trap, the Furvela tent-trap, does not require an 'entry' behavior on the part of the mosquito. It remains the cheapest and lightest tent-trap described. It takes less than two minutes to install and is the only trap that uses readily available components. Here we describe recent modifications to the trap, which make it even easier to set up in the field, provide a standard operating procedure (SOP) and describe some recent experiments examining the effect of the addition of light and door placement to working of the trap. The trap provides the closest approximation to CDC light-traps, widely used to collect indoor biting mosquitoes. This enables the effect of both indoor and outdoor interventions on mosquito density and behavior to be determined. [ABSTRACT FROM AUTHOR]

Background: Southwestern Uganda has high malaria heterogeneity despite moderate vector control and other interventions. Moreover, the early biting transmission and increased resistance to insecticides might compromise strategies relying on vector control. Consequently, monitoring of vector behaviour and insecticide efficacy is needed to assess the effectiveness of strategies aiming at malaria control. This eventually led to an entomological survey in two villages with high malaria prevalence in this region. Methods: During rainy, 2011 and dry season 2012, mosquitoes were collected in Engari and Kigorogoro, Kazo subcounty, using human landing collection, morning indoor resting collection, pyrethrum spray collection and larval collection. Circumsporozoite protein of Plasmodium falciparum sporozoites in female Anopheles mosquitoes was detected using ELISA assay. Bioassays to monitor Anopheles resistance to insecticides were performed. Results: Of the 1,021 female Anopheles species captured, 62% (632) were Anopheles funestus and 36% (371) were Anopheles gambiae s.l. The most common species were Anopheles gambiae s.l. in Engari (75%) and A. funestus in Kigorogoro (83%). Overall, P. falciparum prevalence was 2.9% by ELISA. The daily entomological inoculation rates were estimated at 0.17 and 0.58 infected bites/person/night during rainy and dry season respectively in Engari, and 0.81 infected bites/person/night in Kigorogoro during dry season. In both areas and seasons, an unusually early evening biting peak was observed between 6 - 8 p.m. In Engari, insecticide bioassays showed 85%, 34% and 12% resistance to DDT during the rainy season, dry season and to deltamethrin during the dry season, respectively. In Kigorogoro, 13% resistance to DDT and to deltamethrin was recorded. There was no resistance observed to bendiocarb and pirimiphos methyl. Conclusions: The heterogeneity of mosquito distribution, entomological indicators and resistance to insecticides in villages with high malaria prevalence highlight the need for a long-term vector control programme and monitoring of insecticide resistance in Uganda. The early evening biting habits of Anopheles combined with resistance to DDT and deltamethrin observed in this study suggest that use of impregnated bed nets alone is insufficient as a malaria control strategy, urging the need for additional interventions in this area of high transmission. [ABSTRACT FROM AUTHOR]

Malaria prevalence remains high in many African countries despite massive scaling-up of insecticide treated nets (ITN) and indoor residual spraying (IRS). This paper evaluates the protective effect of pyrethroid IRS and ITNs in relation to risk factors for malaria based on a study conducted in North-West Tanzania, where IRS has been conducted since 2007 and universal coverage of ITNs has been carried out recently. In 2011 community-based cross-sectional surveys were conducted in the two main malaria transmission periods that occur after the short and long rainy seasons. These included 5,152 and 4,325 children aged 0.5–14 years, respectively. Data on IRS and ITN coverage, household demographics and socio-economic status were collected using an adapted version of the Malaria Indicator Survey. Children were screened for malaria by rapid diagnostic test. In the second survey, haemoglobin density was measured and filter paper blood spots were collected to determine age-specific sero-prevalence in each community surveyed. Plasmodium falciparum infection prevalence in children 0.5–14 years old was 9.3% (95%CI:5.9–14.5) and 22.8% (95%CI:17.3–29.4) in the two surveys. Risk factors for infection after the short rains included households not being sprayed (OR = 0.39; 95%CI:0.20–0.75); low community net ownership (OR = 0.45; 95%CI:0.21–0.95); and low community SES (least poor vs. poorest tertile: OR = 0.13, 95%CI:0.05–0.34). Risk factors after the long rains included household poverty (per quintile increase: OR = 0.89; 95%CI:0.82–0.97) and community poverty (least poor vs. poorest tertile: OR = 0.26, 95%CI:0.15–0.44); household IRS or high community ITN ownership were not protective. Despite high IRS coverage and equitable LLIN distribution, poverty was an important risk factor for malaria suggesting it could be beneficial to target additional malaria control activities to poor households and communities. High malaria prevalence in some clusters and the limited protection given by pyrethroid IRS and LLINs suggest that it may be necessary to enhance established vector control activities and consider additional interventions. [ABSTRACT FROM AUTHOR]