Large-Scale Deployment of Pyriproxyfen-Treated Lehmann Funnel Entry Traps to Control Malaria Mosquito Populations

Background: The Lehmann Funnel Entry Trap has proven to be effective in catching and killing up to 70% of mosquitoes even in a high mosquito density setting. A medium-sized prototype was selected and deployed at large scale in Vallée du Kou 3 (VK3) in the Southwest of Burkina Faso to assess its entomological and sociological impact.Method: Overall, 1,313 traps impregnated with Pyriproxyfen (PPF), were deployed. Of them, 12 traps were randomly selected across the intervention village compared to houses without traps in the control village, Vallée du Kou 5 (VK5). Traps were placed at the windows while doors were blocked with curtains. Mosquitoes were collected in traps and matching houses in VK3 and in houses only in VK5, for nine days per month from July to October 2015. Collected mosquitoes were morphologically identified, counted, and preserved in 80% ethanol vials for subsequent analyses, including resistance genes and female mosquito age structure. The impact of the trap on mosquito density at community level was assessed by performing a pyrethrum spray catch (PSC) with bioassays to assess the effect of PPF. Results: Overall mosquito density was reduced by ~90% in all houses equipped with traps in VK3. At the community level, while mosquito density before intervention was 33% higher in VK3 than in VK5, it was 47% higher in VK5 after the intervention. Old female mosquito numbers increased in VK5 by 12% in October but not in VK3, indicating that the traps were cumulatively killing old females. The additional effect of PPF was to limit egg-laying, with a smaller number of eggs counted, and with a low hatching rate. Mosquitoes were highly resistant to pyrethroids with ~0.9 frequency of the kdr mutation. The trap was well accepted by the communities as 85.4% and 93.8% of interviewees in VK3 found the traps reducing mosquito bites with peaceful sleep respectively. Conclusion: The Lehmann Funnel Entry Trap has real potential to control malaria mosquito populations and can be widely used to sustain the global effort of malaria elimination.

Citizen science for monitoring the spatial and temporal dynamics of malaria vectors in relation to environmental risk factors in Ruhuha, Rwanda

Background As part of malaria prevention and control efforts, the distribution and density of malaria mosquitoes requires continuous monitoring. Resources for long-term surveillance of malaria vectors, however, are often limited. The aim of the research was to evaluate the value of citizen science in providing insight into potential malaria vector hotspots and other malaria relevant information, and to determine predictors of malaria vector abundance in a region where routine mosquito monitoring has not been established to support vector surveillance. Methods A 1-year citizen science programme for malaria mosquito surveillance was implemented in five villages of the Ruhuha sector in Bugesera district, Rwanda. In total, 112 volunteer citizens were enrolled and reported monthly data on mosquitoes collected in their peridomestic environment using handmade carbon-dioxide baited traps. Additionally, they reported mosquito nuisance experienced as well as the number of confirmed malaria cases in their household. Results In total, 3793 female mosquitoes were collected, of which 10.8% were anophelines. For the entire period, 16% of the volunteers reported having at least one confirmed malaria case per month, but this varied by village and month. During the study year 66% of the households reported at least one malaria case. From a sector perspective, a higher mosquito and malaria vector abundance was observed in the two villages in the south of the study area. The findings revealed significant positive correlations among nuisance reported and confirmed malaria cases, and also between total number of Culicidae and confirmed malaria cases, but not between the numbers of the malaria vector Anopheles gambiae and malaria cases. At the sector level, of thirteen geographical risk factors considered for inclusion in multiple regression, distance to the river network and elevation played a role in explaining mosquito and malaria mosquito abundance. Conclusions The study demonstrates that a citizen science approach can contribute to mosquito monitoring, and can help to identify areas that, in view of limited resources for control, are at higher risk of malaria.

Wolbachia cifB induces cytoplasmic incompatibility in the malaria mosquito vector

Wolbachia, a maternally inherited intracellular bacterial species, can manipulate host insect reproduction by cytoplasmic incompatibility (CI), which results in embryo lethality in crosses between infected males and uninfected females. CI is encoded by two prophage genes, cifA and cifB. Wolbachia, coupled with the sterile insect technique, has been used in field trials to control populations of the dengue vector Aedes albopictus, but CI-inducing strains are not known to infect the malaria vector Anopheles gambiae. Here we show that cifA and cifB can induce conditional sterility in the malaria vector An. gambiae. We used transgenic expression of these Wolbachia-derived genes in the An. gambiae germline to show that cifB is sufficient to cause embryonic lethality and that cifB-induced sterility is rescued by cifA expression in females. When we co-expressed cifA and cifB in male mosquitoes, the CI phenotype was attenuated. In female mosquitoes, cifB impaired fertility, which was overcome by co-expression of cifA. Our findings pave the way towards using CI to control malaria mosquito vectors.

RNAi by Soaking Aedes aegypti Pupae in dsRNA

RNA-interference (RNAi) is a standard technique for functional genomics in adult mosquitoes. However, RNAi in immature, aquatic mosquito stages has been challenging. Several studies have shown successful larval RNAi, usually in combination with a carrier molecule. Except for one study in malaria mosquito, Anopheles gambiae, none of the previous studies has explored RNAi in mosquito pupae. Even in the study that used RNAi in pupae, double stranded RNA (dsRNA) was introduced by microinjection. Here, we describe a successful method by soaking pupae in water containing dsRNA without any carrier or osmotic challenge. The knockdown persisted into adulthood. We expect that this simple procedure will be useful in the functional analysis of genes that highly express in pupae or newly emerged adults.

Functional Characterization of Ecdysis Triggering Hormone Receptors (AgETHR-A and AgETHR-B) in the African Malaria Mosquito, Anopheles gambiae

Insect ecdysis behavior, shedding off the old cuticle, is under the control of specific neuropeptides with the top command by the ecdysis triggering hormone (ETH). We characterized the ETH receptor (ETHR) of the malaria mosquito, Anopheles gambiae, by manual annotation of the NCBI gene (AGAP002881) and functional analysis, using a heterologous expression system in a mammalian cell line. The two splicing variants of ETHRs, ecdysis triggering hormone receptors (AgETHR-A and AgETHR-B), a conserved feature among insects, included of four (552 aa) and five exons (635 aa), respectively. The main feature of manual annotation of the receptor was a correction of N-terminal and exon-intron boundaries of an annotated gene (AGAP002881). Interestingly, the functional expression of the receptor in Chinese hamster ovary cells required modification of the transcription initiation site for mammalian Kozak consensus. In the calcium mobilization assay using the heterologous expression of each receptor, AgETHR-B showed a higher sensitivity to AgETH-1 (28 times) and AgETH-2 (320 times) than AgETHR-A. The AgETHRs showed specificity only to the ETH group of peptides but not to other groups carrying the C-termini motifs as PRXamide, such as pyrokinin1/DH and pyrokinin2/PBAN. Ecdysis triggering hormone receptors (AgETHR-B) responded to different ETH variants of other insect species more promiscuously than AgETHR-A.

Molecular evolution and the decline of purifying selection with age

Life history theory predicts that the intensity of selection declines with age, and this trend should impact how genes expressed at different ages evolve. Here we find consistent relationships between a gene’s age of expression and patterns of molecular evolution in two mammals (the human Homo sapiens and the mouse Mus musculus) and two insects (the malaria mosquito Anopheles gambiae and the fruit fly Drosophila melanogaster). When expressed later in life, genes fix nonsynonymous mutations more frequently, are more polymorphic for nonsynonymous mutations, and have shorter evolutionary lifespans, relative to those expressed early. The latter pattern is explained by a simple evolutionary model. Further, early-expressed genes tend to be enriched in similar gene ontology terms across species, while late-expressed genes show no such consistency. In humans, late-expressed genes are more likely to be linked to cancer and to segregate for dominant disease-causing mutations. Last, the effective strength of selection (Nes) decreases and the fraction of beneficial mutations increases with a gene’s age of expression. These results are consistent with the diminishing efficacy of purifying selection with age, as proposed by Medawar’s classic hypothesis for the evolution of senescence, and provide links between life history theory and molecular evolution.

Impact of increased ventilation on indoor temperature and malaria mosquito density: an experimental study in The Gambia

In sub-Saharan Africa, cooler houses would increase the coverage of insecticide-treated bednets, the primary malaria control tool. We examined whether improved ventilation, using windows screened with netting, cools houses at night and reduces malaria mosquito house entry in The Gambia. Identical houses were constructed, with badly fitting doors the only mosquito entry points. Two men slept in each house and mosquitoes captured using light traps. First, temperature and mosquito density were compared in four houses with 0, 1, 2 and 3 screened windows. Second, carbon dioxide (CO 2 ), a major mosquito attractant, was measured in houses with (i) no windows, (ii) screened windows and (iii) screened windows and screened doors. Computational fluid dynamic modelling captured the spatial movement of CO 2 . Increasing ventilation made houses cooler, more comfortable and reduced malaria mosquito house entry; with three windows reducing mosquito densities by 95% (95%CI = 90–98%). Screened windows and doors reduced the indoor temperature by 0.6°C (95%CI = 0.5–0.7°C), indoor CO 2 concentrations by 31% between 21.00 and 00.00 h and malaria mosquito entry by 76% (95%CI = 69–82%). Modelling shows screening reduces CO 2 plumes from houses. Under our experimental conditions, cross-ventilation not only reduced indoor temperature, but reduced the density of house-entering malaria mosquitoes, by weakening CO 2 plumes emanating from houses.