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The scale-up of malaria interventions over the past two decades has resulted in substantial reductions in malaria burden, especially in sub-Saharan Africa, which accounts for 95% of the global malaria burden.1,2 Between 2000 and 2019, an estimated 1·5 billion malaria cases and 7·6 million malaria deaths were averted, with many of the early gains attributed to widescale deployment of insecticide-treated nets and availability of effective drugs.1 However, progress in global malaria control and elimination efforts have stagnated in recent years, the reasons for which are multifactorial and include the development and spread of insecticide resistance in malaria vectors. The massive scale-up of insecticide-treated nets and, later, long-lasting insecticidal nets (LLINs), which have an insecticide incorporated into the net fibre and can last longer under typical use, was accompanied by the emergence of vector resistance to pyrethroids.3 This resistance is now present in all primary vectors throughout much of sub-Saharan Africa.3–5 Studies have shown that a new generation of LLINs that include the synergist piperonyl butoxide have improved efficacy compared with pyrethroid-only nets.6,7 However, although piperonyl butoxide can restore susceptibility in pyrethroid-resistant mosquito populations wherein the primary resistance mechanism is metabolic detoxification by oxidase enzymes, it has minimal impact on other mechanisms of pyrethroid resistance. The Royal Guard (Disease Control Technologies, Greer, SC, USA), an LLIN treated with α-cypermethrin and pyriproxyfen, provided little evidence of an added benefit relative to the standard LLINs.

Scale-up of insecticide-based interventions has averted more than 500 million malaria cases since 2000. Increasing insecticide resistance could herald a rebound in disease and mortality. We aimed to investigate whether insecticide resistance was associated with loss of effectiveness of long-lasting insecticidal nets and increased malaria disease burden. This WHO-coordinated, prospective, observational cohort study was done at 279 clusters (villages or groups of villages in which phenotypic resistance was measurable) in Benin, Cameroon, India, Kenya, and Sudan. Pyrethroid long-lasting insecticidal nets were the principal form of malaria vector control in all study areas; in Sudan this approach was supplemented by indoor residual spraying. Cohorts of children from randomly selected households in each cluster were recruited and followed up by community health workers to measure incidence of clinical malaria and prevalence of infection. Mosquitoes were assessed for susceptibility to pyrethroids using the standard WHO bioassay test. Country-specific results were combined using meta-analysis. Between June 2, 2012, and Nov 4, 2016, 40 000 children were enrolled and assessed for clinical incidence during 1·4 million follow-up visits. 80 000 mosquitoes were assessed for insecticide resistance. Long-lasting insecticidal net users had lower infection prevalence (adjusted odds ratio [OR] 0·63, 95% CI 0·51–0·78) and disease incidence (adjusted rate ratio [RR] 0·62, 0·41–0·94) than did non-users across a range of resistance levels. We found no evidence of an association between insecticide resistance and infection prevalence (adjusted OR 0·86, 0·70–1·06) or incidence (adjusted RR 0·89, 0·72–1·10). Users of nets, although significantly better protected than non-users, were nevertheless subject to high malaria infection risk (ranging from an average incidence in net users of 0·023, [95% CI 0·016–0·033] per person-year in India, to 0·80 [0·65–0·97] per person year in Kenya; and an average infection prevalence in net users of 0·8% [0·5–1·3] in India to an average infection prevalence of 50·8% [43·4–58·2] in Benin). Irrespective of resistance, populations in malaria endemic areas should continue to use long-lasting insecticidal nets to reduce their risk of infection. As nets provide only partial protection, the development of additional vector control tools should be prioritised to reduce the unacceptably high malaria burden. Bill & Melinda Gates Foundation, UK Medical Research Council, and UK Department for International Development.

Background High coverage of insecticide-treated bed nets in Asembo and low coverage in Seme, two adjacent communities in western Nyanza Province, Kenya; followed by expanded coverage of bed nets in Seme, as the Kenya national malaria programme rolled out; provided a natural experiment for quantification of changes in relative abundance of two primary malaria vectors in this holoendemic region. Both belong to the Anopheles gambiae sensu lato (s.l.) species complex, namely A. gambiae sensu stricto (s.s.) and Anopheles arabiensis . Historically, the former species was proportionately dominant in indoor resting collections of females. Methods Data of the relative abundance of adult A. gambiae s.s. and A. arabiensis sampled from inside houses were obtained from the literature from 1970 to 2002 for sites west of Kisumu, Kenya, to the region of Asembo ca. 50 km from the city. A sampling transect was established from Asembo (where bed net use was high due to presence of a managed bed net distribution programme) eastward to Seme, where no bed net programme was in place. Adults of A. gambiae s.l. were sampled from inside houses along the transect from 2003 to 2009, as were larvae from nearby aquatic habitats, providing data over a nearly 40 year period of the relative abundance of the two species. Relative proportions of A. gambiae s.s. and A. arabiensis were determined for each stage by identifying species by the polymerase chain reaction method. Household bed net ownership was measured with surveys during mosquito collections. Data of blood host choice, parity rate, and infection rate for Plasmodium falciparum in A. gambiae s.s. and A. arabiensis were obtained for a sample from Asembo and Seme from 2005. Results Anopheles gambiae s.s. adult females from indoor collections predominated from 1970 to 1998 (ca. 85%). Beginning in 1999, A. gambiae s.s decreased proportionately relative to A. arabiensis , then precipitously declined to rarity coincident with increased bed net ownership as national bed net distribution programmes commenced in 2004 and 2006. By 2009, A. gambiae s.s. comprised proportionately ca. 1% of indoor collections and A. arabiensis 99%. In Seme compared to Asembo in 2003, proportionately more larvae were A. gambiae s.s. , larval density was higher, and more larval habitats were occupied. As bed net use rose in Seme, the proportion of A. gambiae larvae declined as well. These trends continued to 2009. Parity and malaria infection rates were lower in both species in Asembo (high bed net use) compared to Seme (low bed net use), but host choice did not vary within species in both communities (predominantly cattle for A. arabiensis , humans for A. gambiae s.s. ). Conclusions A marked decline of the A. gambiae s.s. population occurred as household ownership of bed nets rose in a region of western Kenya over a 10 year period. The increased bed net coverage likely caused a mass effect on the composition of the A. gambiae s.l. species complex, resulting in the observed proportionate increase in A. arabiensis compared to its closely related sibling species, A. gambiae s.s. These observations are important in evaluating the process of regional malaria elimination, which requires sustained vector control as a primary intervention.

The Anopheles stephensi mosquito is an invasive malaria vector recently reported in Djibouti, Ethiopia, Sudan, Somalia, Nigeria, and Ghana. The World Health Organization has called on countries in Africa to increase surveillance efforts to detect and report this vector and institute appropriate and effective control mechanisms. In Kenya, the Division of National Malaria Program conducted entomological surveillance in counties at risk for An. stephensi mosquito invasion. In addition, the Kenya Medical Research Institute conducted molecular surveillance of all sampled Anopheles mosquitoes from other studies to identify An. stephensi mosquitoes. We report the detection and confirmation of An. stephensi mosquitoes in Marsabit and Turkana Counties by using endpoint PCR and morphological and sequence identification. We demonstrate the urgent need for intensified entomological surveillance in all areas at risk for An. stephensi mosquito invasion, to clarify its occurrence and distribution and develop tailored approaches to prevent further spread.

Aedes aegypti is the primary global vector to humans of yellow fever and dengue flaviviruses. Over the past 50 years, many population genetic studies have documented large genetic differences among global populations of this species. These studies initially used morphological polymorphisms, followed later by allozymes, and most recently various molecular genetic markers including microsatellites and mitochondrial markers. In particular, since 2000, fourteen publications and four unpublished datasets have used sequence data from the NADH dehydrogenase subunit 4 mitochondrial gene to compare Ae. aegypti collections and collectively 95 unique mtDNA haplotypes have been found. Phylogenetic analyses in these many studies consistently resolved two clades but no comprehensive study of mtDNA haplotypes have been made in Africa, the continent in which the species originated. ND4 haplotypes were sequenced in 426 Ae. aegypti s.l. from Senegal, West Africa and Kenya, East Africa. In Senegal 15 and in Kenya 7 new haplotypes were discovered. When added to the 95 published haplotypes and including 6 African Aedes species as outgroups, phylogenetic analyses showed that all but one Senegal haplotype occurred in a basal clade while most East African haplotypes occurred in a second clade arising from the basal clade. Globally distributed haplotypes occurred in both clades demonstrating that populations outside Africa consist of mixtures of mosquitoes from both clades. Populations of Ae. aegypti outside Africa consist of mosquitoes arising from one of two ancestral clades. One clade is basal and primarily associated with West Africa while the second arises from the first and contains primarily mosquitoes from East Africa.

Background Long-lasting insecticide nets (LLINs) are a core malaria intervention. LLINs should retain efficacy against mosquito vectors for a minimum of three years. Efficacy and durability of Olyset ® Plus, a permethrin and piperonyl butoxide (PBO) treated LLIN, was evaluated versus permethrin treated Olyset ® Net. In the absence of WHO guidelines of how to evaluate PBO nets, and considering the manufacturer’s product claim, Olyset ® Plus was evaluated as a pyrethroid LLIN. Methods This was a household randomized controlled trial in a malaria endemic rice cultivation zone of Kirinyaga County, Kenya between 2014 and 2017. Cone bioassays and tunnel tests were done against Anopheles gambiae Kisumu. The chemical content, fabric integrity and LLIN survivorship were monitored. Comparisons between nets were tested for significance using the Chi-square test. Exact binomial distribution with 95% confidence intervals (95% CI) was used for percentages. The WHO efficacy criteria used were ≥ 95% knockdown and/or ≥ 80% mortality rate in cone bioassays and ≥ 80% mortality and/or ≥ 90% blood-feeding inhibition in tunnel tests. Results At 36 months, Olyset ® Plus lost 52% permethrin and 87% PBO content; Olyset ® Net lost 24% permethrin. Over 80% of Olyset ® Plus and Olyset ® Net passed the WHO efficacy criteria for LLINs up to 18 and 12 months, respectively. At month 36, 91.2% Olyset ® Plus and 86.4% Olyset ® Net survived, while 72% and 63% developed at least one hole. The proportionate Hole Index (pHI) values representing nets in good, serviceable and torn condition were 49.6%, 27.1% and 23.2%, respectively for Olyset ® Plus, and 44.9%, 32.8% and 22.2%, respectively for Olyset ® Net but were not significantly different. Conclusions Olyset ® Plus retained efficacy above or close to the WHO efficacy criteria for about 2 years than Olyset ® Net (1–1.5 years). Both nets did not meet the 3-year WHO efficacy criteria, and showed little attrition, comparable physical durability and survivorship, with 50% of Olyset ® Plus having good and serviceable condition after 3 years. Better community education on appropriate use and upkeep of LLINs is essential to ensure effectiveness of LLIN based malaria interventions. Graphical Abstract

An amendment to this paper has been published and can be accessed via a link at the top of the paper.An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Background Industrial wastewater is a human health hazard upon exposure. Aquatic organisms in contaminated wastewater may accumulate the toxic elements with time. Human population living in informal settlements in Nairobi industrial area risk exposure to such toxic elements. Biomonitoring using aquatic organisms in open drainage channels can be key in metal exposure assessment. Levels of Mercury (Hg), Lead (Pb), Chromium (Cr), Cadmium (Cd), Thallium (Tl), and Nickel (Ni) were established in samples of wastewater, filamentous green algae ( Spirogyra ) and mosquitoes obtained from open drainage channels in Nairobi industrial area, Kenya. Results Pb, Cr, & Ni levels ranged from 3.08 to 15.31 µg/l while Tl, Hg, & Cd ranged from 0.05 to 0.12 µg/l in wastewater. The Pb, Cr, Ni, & Cd levels were above WHO, Kenya & US EPA limits for wastewater but Hg was not. Pb, Cr, Tl, & Ni levels in assorted field mosquitoes were 1.3–2.4 times higher than in assorted laboratory-reared mosquitoes. Hg & Cd concentrations in laboratory-reared mosquitoes (0.26 mg/kg & 1.8 mg/kg respectively) were higher than in field mosquitoes (0.048 mg/kg & 0.12 mg/kg respectively). The levels of Pb, Cr, & Ni were distinctively higher in field mosquito samples than in wastewater samples from the same site. Pb, Cr, Ni, Cd & Hg levels in green filamentous Spirogyra algae were 110.62, 29.75, 14.45, 0.44, & 0.057 mg/kg respectively. Correlation for Pb & Hg (r (2) = 0.957; P < 0.05); Cd & Cr (r (2) = 0.985; P < 0.05) in algae samples was noted. The metal concentrations in the samples analyzed were highest in filamentous green algae and least in wastewater. Conclusion Wastewater, mosquitoes, and filamentous green algae from open drainage channels and immediate vicinity, in Nairobi industrial area (Kenya) contained Hg, Pb, Cr, Cd, Tl, and Ni. Mosquitoes in urban areas and filamentous green algae in open drainage channels can play a role of metal biomonitoring in wastewater. The potential of urban mosquitoes transferring heavy metals to human population from the contaminated wastewater should be investigated.

Background Anopheles coluzzii is a primary vector of malaria found in West and Central Africa, but its presence has hitherto never been documented in Kenya. A thorough understanding of vector bionomics is important as it enables the implementation of targeted and effective vector control interventions. Malaria vector surveillance efforts in the country have tended to focus on historically known primary vectors. The current study sought to determine the taxonomic status of samples collected from five different malaria epidemiological zones in Kenya as well as describe the population genetic structure and insecticide resistance profiles in relation to other An. coluzzii populations. Methods Mosquitoes were sampled as larvae from Busia, Kwale, Turkana, Kirinyaga and Kiambu counties, representing the range of malaria endemicities in Kenya, in 2019 and 2021 and emergent adults analysed using Whole Genome Sequencing (WGS) data processed in accordance with the Anopheles gambiae 1000 Genomes Project phase 3. Where available, historical samples from the same sites were included for WGS. Comparisons were made with An. coluzzii cohorts from West and Central Africa. Results This study reports the detection of An. coluzzii for the first time in Kenya. The species was detected in Turkana County across all three time points from which samples were analyzed and its presence confirmed through taxonomic analysis. Additionally, there was a lack of strong population genetic differentiation between An. coluzzii from Kenya and those from the more northerly regions of West and Central Africa, suggesting they represent a connected extension to the known species range. Mutations associated with target-site resistance to DDT and pyrethroids and metabolic resistance to DDT were found at high frequencies up to 64%. The profile and frequencies of the variants observed were similar to An. coluzzii from West and Central Africa but the ace-1 mutation linked to organophosphate and carbamate resistance present in An. coluzzii from coastal West Africa was absent in Kenya. Conclusions These findings emphasize the need for the incorporation of genomics in comprehensive and routine vector surveillance to inform on the range of malaria vector species, and their insecticide resistance status to inform the choice of effective vector control approaches.