Background: The implementation of anti-larval strategies in the fight against malaria vectors requires fundamental knowledge of their oviposition sites. The aim of this study was to assess the spatial and temporal distribution of Anopheles breeding sites as well as the influence of abiotic and biotic factors on the proliferation of larvae in urban and non-urban areas of Benin. Methods: Sampling of Anopheles larvae was carried out during the rainy and dry seasons in urbanized and non-urbanized areas of the cities of Cotonou, Bohicon, Parakou, and Natitingou in Benin. The Anopheles larval breeding sites were georeferenced and characterized by their nature, type, physicochemical (pH, temperature, dissolved oxygen, conductivity, turbidity, salinity) and biological attributes (larval density and coliform density). Results: A total of 198 positive breeding sites for Anopheles larvae were identified, comprising 163 (82.3%) in the rainy season and 35 (17.7%) in the dry season. Out of these larval habitats, 61.9% were located in urbanized areas, and were predominantly puddles. Principal component analysis revealed a high positive correlation of larval density with temperature and dissolved oxygen, and with salinity in the coastal zone. In addition, cross-sectional analysis of the microbiological results with larval density showed a significant negative correlation between larval productivity and faecal coliform load. Conclusions: This study indicated the presence of multiple larval habitats of Anopheles in the urban areas which were created through human activities, and associations between larval density and intrinsic factors of the habitats such as temperature, dissolved oxygen and faecal coliform load. This type of information may be useful for the implementation of appropriate control strategies in urban areas, including regulation of the human activities that lead to the creation of breeding sites, proper environmental management and targeted larvicidal use. Graphical

Mosquito-borne diseases account for human morbidity and mortality worldwide, caused by the parasites (e.g., malaria) or viruses (e.g., dengue, Zika) transmitted through bites of infected female mosquitoes. Globally, billions of people are at risk of infection, imposing significant economic and public health burdens. As such, efficient methods to monitor mosquito populations and prevent the spread of these diseases are at a premium. One proposed technique is to apply acoustic monitoring to the challenge of identifying wingbeats of individual mosquitoes. Although researchers have successfully used wingbeats to survey mosquito populations, implementation of these techniques in areas most affected by mosquito-borne diseases remains challenging. Here, methods utilizing easily accessible equipment and encouraging community-scientist participation are more likely to provide sufficient monitoring. We present a practical, community-science-based method of monitoring mosquito populations using smartphones. We applied deep-learning algorithms (TensorFlow Inception v3) to spectrogram images generated from smartphone recordings associated with six mosquito species to develop a multiclass mosquito identification system, and flag potential invasive vectors not present in our sound reference library. Though TensorFlow did not flag potential invasive species with high accuracy, it was able to identify species present in the reference library at an 85% correct identification rate, an identification rate markedly higher than similar studies employing expensive recording devices. Given that we used smartphone recordings with limited sample sizes, these results are promising. With further optimization, we propose this novel technique as a way to accurately and efficiently monitor mosquito populations in areas where doing so is most critical.

LLINs containing an insecticide plus the synergist, piperonyl butoxide (PBO) have been designed for increased efficacy against pyrethroid-resistant malaria vectors. In this study, two LLINs with PBO, PermaNet^® 3.0 and Olyset^® Plus, and a pyrethroid-only LLIN, Yorkool^®, were evaluated in experimental huts against a free-flying, wild population of Anopheles gambiae s.l. in Kolokopé, a cotton cultivated area of Togo. WHO susceptibility tube tests and subsequent molecular assays determine the An. gambiae s.l. populations to be resistant to pyrethroids and DDT with both target site kdr and metabolic resistance mechanisms involved in the resistance observed. Anopheles gambiae s.s. and An. coluzzi were present in sympatry though the kdr (L1014F) mutation was observed at a higher frequency in An. gambiae s.s. The experimental hut results showed that both PermaNet^® 3.0 and Olyset^® Plus nets induced similar levels of deterrence, exophily, and reduced blood feeding rate against wild An. gambiae s.l. in contrast to the pyrethroid only LLIN, Yorkool^®. The proportion of wild An. gambiae s.l. killed by unwashed PermaNet^® 3.0 was significantly higher than unwashed Olyset^® Plus (corrected mortality 80.5% compared to 66.6%). Similar blood feeding inhibition rates were observed for unwashed PermaNet^® 3.0 and Olyset^® Plus; however, PermaNet^® 3.0 washed 20 times demonstrated significantly higher blood feeding inhibition rate than Olyset^® Plus washed 20 times (91.1% compared with 85.6% respectively). Yorkool^® performed the worst for all the parameters evaluated. In an area of pyrethroid resistance of An. gambiae s.l involving kdr target site and metabolic resistance mechanisms, LLINs with PBO can provide additional protection in terms of reduction in blood feeding and increase in mosquito mortality compared to a pyrethroid-only net, and should be considered in malaria vector control strategies.

Background:To optimize the success of insecticide-based malaria control intervention, knowledge of the distribution ofAnopheles gambiaespecies and insecticide resistance mechanisms is necessary. This paper reported an updated data on pyrethroids/DDT resistance in theAn. gambiae s.lpopulation from Togo. Methods:From December 2013 to April 2015, females of indoor-restingAn. gambiae s.lwere captured in three locations belonging to three different ecological zones. Resistance to DDT, permethrin and deltamethrin was screened in F1 progeny of collected mosquitoes using WHO susceptibility tests. The identification of species ofAn. gambiaecomplex and the detection ofkdrandace.1R allele were carried out using DNA-based molecular techniques. Results: An. gambiaefrom Kovié and Nangbéto were highly resistant to DDT and permethrin with mortalities rate ranging from 0.83% to 1.58% for DDT and zero to 8.54% for permethrin. Mosquitoes collected in Nangbéto displayed 81.53% mortality with deltamethrin.An. coluzziiandAn. gambiaes.s were found in sympatry in Nangbéto and Mango.The allelic frequency of L1014F was high, ranging from 66 to 100% in bothAn. coluzziiandAn. gambiae s.s. For the first time we detected the L1014S allele in bothAn. coluzziiandAn. gambiae s.s.from Togo at the frequency ranging from 5% to 13% in all the sites. ThekdrN1575Y was present at various frequencies in both species ranging from 10% to 45%. BothAn. gambiae s.s.andAn. coluzziishared theace1R mutation in all investigated sites with allelic frequency ranging from 4% to 16%. Conclusion:These results showed that multiple mutations are involved in insecticides resistance inAn. gambiaepopulations from Togo including the kdr L1014F, L1014S, and N1575Y andace.1R G119S mutations.

Malaria, one of the world’s greatest public health challenges, is an endemic disease with stable transmission in Togo. Combating malaria requires an effective vector control. This study provides temporal data on insecticide resistance status in the major malaria vector Anopheles gambiae sensu lato (s.l.) from Togo. Two to 5 days old females of An. gambiae s.l., originating from three localities (Baguida, Kovié, Kolokopé) were subjected to insecticide-impregnated papers during 3 years (2012, 2013, 2016) as follows: organochlorides (4% DDT), pyrethroids (0.05% deltamethrin, 0.75% permethrin, 0.05% lambdacyhalothrin), carbamates (0.4% bendiocarb and 0.1% propoxur), and organophosphates (5% malathion, 0.4% chlorpyrifos methyl, 1% fenitrothion) following the WHO standard protocol. Dead and surviving mosquitoes were stored separately in Eppendorf tubes containing silica gel for DNA extraction, species identification, and kdr and ace-1 genotyping. Knockdown times (KDT50 and KDT95) were high in An. gambiae s.l. The lowest KDTs were recorded at Baguida in 2013 for deltamethrin (KDT50 = 24.7, CI [22.4–27.12] and KDT95 = 90.78, CI [76.35–113.49]). No KDTs were recorded for DDT and in some instances for permethrin. In general, An. gambiae s.l. was resistant to most of the four classes of insecticides during the survey periods regardless of locality and year, except to chlorpyrifos methyl. In some instances, mosquitoes were fully susceptible to fenitrothion (Kolokopé: 100% and Kovié: 98.05%, CI [95.82–100.26]) and malathion (100% at both Kolokopé and Kovié) in 2013, and malathion only (Kolokopé; 100%) in 2016. Anopheles coluzzii, An. gambiae and Anopheles arabiensis were the three sibling species identified at the three localities with some hybrids at Baguida (2013), and Kovié (2012 and 2016), respectively. Anopheles gambiae was relatively dominant (61.6%). The kdr 1014F allele frequency was > 0.9 in most of the cases, except at Kolokopé (f (1014F) = 0.63, CI [0.55–0.71]) in 2013. The kdr 1014S allele frequency was below 0.02. The highest ace-1 frequencies were identified in An. gambiae at Baguida (2012: 0.52, CI [0.34–0.69] and 2013: 0.66, CI [0.46–0.86]). The resistance status is worrying in Togo and should be considered in future malaria vector resistance management programmes by decision-makers.