The paradox of plant preference: The malaria vectors Anopheles gambiae and Anopheles coluzzii select suboptimal food sources for their survival and reproduction.

Anopheles gambiae and Anopheles coluzzii mosquitoes, two major malaria vectors in sub-Saharan Africa, exhibit selectivity among plant species as potential food sources. However, it remains unclear if their preference aligns with optimal nutrient intake and survival. Following an extensive screening of the effects of 31 plant species on An. coluzzii in Burkina Faso, we selected three species for their contrasting effects on mosquito survival, namely Ixora coccinea, Caesalpinia pulcherrima, and Combretum indicum. We assessed the sugar content of these plants and their impact on mosquito fructose positivity, survival, and insemination rate, using Anopheles coluzzii and Anopheles gambiae, with glucose 5% and water as controls. Plants displayed varying sugar content and differentially affected the survival, sugar intake, and insemination rate of mosquitoes. All three plants were more attractive to mosquitoes than controls, with An. gambiae being more responsive than An. coluzzii. Notably, C. indicum was the most attractive but had the lowest sugar content and offered the lowest survival, insemination rate, and fructose positivity. Our findings unveil a performance-preference mismatch in An. coluzzii and An. gambiae regarding plant food sources. Several possible reasons for this negative correlation between performance and preference are discussed.

Entomotherapy as an alternative treatment for diseases due to Gram-negative bacteria in Burkina Faso.

Insects are known for their harmful effects. However, they also benefit humans, animals, plants, and ecosystems. Its beneficial uses include entomophagy and entomotherapy. This study aimed to evaluate the antibacterial activity of insect extracts against Gram-negative bacteria. Antibacterial activities of thirteen crude extracts of medicinal insects were tested against twelve Gram-negative bacteria by diffusion on agar. Imipenem was used as an antibiotic for positive control. The thirteen extracts acted differently against certain Gram-negative bacteria. The largest inhibition diameter was for extracts of Cirina butyrospermi and Mylabris variabilis against Pseudomonas aeruginosa ATCC27853 and Salmonella enteritidis ATCC13076, respectively. The diameters of inhibition obtained using imipenem against these same bacterial strains were 13.0 ± 0.0 mm and 22 ± 1.0 mm, respectively. The lowest inhibition diameter (7.5 ± 0.0 mm) was obtained using Anopheles gambiae extract against Salmonella Typhimurium ATCC14028. Imipenem was active on all strains tested. The highest values of the index multi-resistance to insect's extracts were reported for Pseudomonas aeruginosa ATCC9027 and Serratia odorifera 652411. Overall, the results of this study confirmed the antibacterial activities of insects used by traditional health practitioners to treat different pathologies. Entomotherapy could be an alternative treatment for certain infectious pathologies caused by gram-negative bacteria.

Natural plant diet impacts phenotypic expression of pyrethroid resistance in Anopheles mosquitoes.

Success in reducing malaria transmission through vector control is threatened by insecticide resistance in mosquitoes. Although the proximal molecular mechanisms and genetic determinants involved are well documented, little is known about the influence of the environment on mosquito resistance to insecticides. The aim of this study was to assess the effect of plant sugar feeding on the response of Anopheles gambiae sensu lato to insecticides. Adults were fed with one of four treatments, namely a 5% glucose control solution, nectariferous flowers of Barleria lupulina, of Cascabela thevetia and a combination of both B. lupulina + C. thevetia. WHO tube tests were performed with 0.05% and 0.5% deltamethrin, and knockdown rate (KD) and the 24 h mosquito mortality were measured. Plant diet significantly influenced mosquito KD rate at both concentrations of deltamethrin. Following exposure to 0.05% deltamethrin, the B. lupulina diet induced a 2.5 fold-increase in mosquito mortality compared to 5% glucose. Species molecular identification confirmed the predominance of An. gambiae (60% of the samples) over An. coluzzii and An. arabiensis in our study area. The kdr mutation L1014F displayed an allelic frequency of 0.75 and was positively associated with increased phenotypic resistance to deltamethrin. Plant diet, particularly B. lupulina, increased the susceptibility of mosquitoes to insecticides. The finding that B. lupulina-fed control individuals (i.e. not exposed to deltamethrin) also displayed increased 24 h mortality suggests that plant-mediated effects may be driven by a direct effect of plant diet on mosquito survival rather than indirect effects through interference with insecticide-resistance mechanisms. Thus, some plant species may weaken mosquitoes, making them less vigorous and more vulnerable to the insecticide. There is a need for further investigation, using a wider range of plant species and insecticides, in combination with other relevant environmental factors, to better understand the expression and evolution of insecticide resistance.

Essential oils of plants and their combinations as an alternative adulticides against Anopheles gambiae (Diptera: Culicidae) populations.

The persistence of malaria and the increasing of resistance of Anopheles gambiae species to chemicals remain major public health concerns in sub-Saharan Africa. Faced to these concerns, the search for alternative vector control strategies as use of essential oils (EOs) need to be implemented. Here, the five EOs from Cymbopogon citratus, Cymbopogon nardus, Eucalyptus camaldulensis, Lippia multiflora, Ocimum americanum obtained by hydro distillation were tested according to World Health Organization procedures on An. gambiae "Kisumu" and field strains collected in "Vallée du Kou". Also, the binary combinations of C. nardus (Cn) and O. americanum (Oa) were examined. As results, among the EOs tested, L. multiflora was the most efficient on both An. gambiae strains regarding KDT50 (50% of mosquitoes knock down time) and KDT95 and rate of morality values. Our current study showed that C8 (Cn 80%: Oa 20%) and C9 (Cn 90%: Oa 10%), were the most toxic to An. gambiae strain "Vallée de Kou" (VK) with the mortality rates reaching 80.7 and 100% at 1% concentration, respectively. These two binary combinations shown a synergistic effect on the susceptible population. However, only C9 gave a synergistic effect on VK population. The bioactivity of the two EOs, C. nardus and O. americanum, was improved by the combinations at certain proportions. The resistance ratios of all EOs and of the combinations were low (< 5). The combinations of C. nardus and O. americanum EOs at 90: 10 ratio and to a lesser extent L. multiflora EO, could be used as alternative bio-insecticides against malaria vectors resistant to pyrethroids in vector control programmes.

Alternatives to Pyrethroid Resistance: Combinations of Cymbopogon nardus and Ocimum americanum Essential Oils Improve the Bioefficiency Control Against the Adults' Populations of Aedes aegypti (Diptera: Culicidae).

Dengue vector control strategies are mostly based on chemicals use against Aedes aegypti populations. The current study aimed at investigating the insecticidal effects of essential oils (EOs) obtained from five plant species, Cymbopogon citrates (D. C.) Stapf. (Poaceae), Cymbopogon nardus (Linn.) Rendle (Poaceae), Eucalyptus camaldulensis Linn. (Myrtaceae), Lippia multiflora Moldenke (Verbenaceae), and Ocimum americanum Linn. Lamiaceae, and combinations of Cymbopogon nardus and Ocimum americanum on Ae. aegypti populations from Bobo-Dioulasso. For this purpose, adults of the susceptible and field strains of Ae. aegypti were tested in WHO tubes with EO alone and binary combinations of O. americanum (OA) and C. nardus (CN; scored from C1 to C9). The extraction of the essential oils was done by hydrodistillation, and their components were determined by GC/MS. Among the 5 EOs tested, L. multiflora essential oil was the most efficient, with KDT50 values below 60 min on all Ae. aegypti strains tested, and also with a rate of mortality up to 100 and 85% for Bora Bora and Bobo-Dioulasso strains, respectively. This efficacy may be due to its major compounds which are with major compounds as ß-caryophyllene, p-cymene, thymol acetate, and 1.8 cineol. Interestingly, on all strains, C8 combination showed a synergistic effect, while C2 showed an additive effect. These combinations exhibit a rate of mortality varying from 80 to 100%. Their toxicity would be due to the major compounds and the putative combined effects of some major and minor compounds. More importanly, L. multiflora EO and combinations of C. nardus and O. americanum EO, may be used as alternatives against pyrethroid resistant of Ae. aegypti.

No evidence for long-range male sex pheromones in two malaria mosquitoes.

Cues involved in mate seeking and recognition prevent hybridization and can be involved in speciation processes. In malaria mosquitoes, females of the two sibling species Anopheles gambiae s.s. and An. coluzzii mate in monospecific male swarms and hybrids are rare. Long-range sex pheromones driving this behaviour have been debated in literature but so far, no study has proven their existence or their absence. Here, we attempted to bring to light their existence. To put all the odds in our favour, we used different chemical ecology methods such as behavioural and electrophysiological assays as well chemical analyses, and we worked with mosquitoes at their optimal physiological mating state that is with swarming males during their natural swarming windows. Despite all our efforts, our results support the absence of long-range sex pheromones involved in swarm detection and recognition by females. We briefly discuss the implications of this finding in ecology, evolution and for control strategies.

Nutritional and Functional Properties of Defatted Flour, Protein Concentrates, and Isolates of Brachytrupes membranaceus (Orthoptera: Gryllidae) (Drury: 1773) and Macrotermes subhyalinus (Isoptera: Blattodea) (Rambur: 1842) from Burkina Faso.

Brachytrupes membranaceus and Macrotermes subhyalinus are edible insects in Burkina Faso. Our research aimed to evaluate the nutritional composition and functional properties of the defatted flours, protein concentrates, and isolates of Brachytrupes membranaceus and Macrotermes subhyalinus. Proximate and mineral composition were determined according to AOAC methods. The amino acid and fatty acid composition were determined by high-performance liquid chromatography (HPLC) and gas chromatography, respectively. The protein concentrates and isolates were obtained by solubilization, precipitation, and lyophilization. Macrotermes subhyalinus showed the highest protein (45.75 g/100 g), iron (11.76 mg/100 g), and zinc (13.18 mg/100 g) contents. The highest isoleucine and lysine contents, the best fat absorption (10.87 g/g), and foaming capacities (49.60%) were obtained with the isolate of Brachytrupes membranaceus. Consumption of Macrotermes subhyalinus could be used to fight or correct iron and zinc deficiencies. Macrotermes subhyalinus was a source of macronutrients and micronutrients, while the protein concentrates and isolates of Brachytrupes membranaceus were endowed with functional properties (fat absorption and foaming capacities).

Diversity and Role of Entomological Fauna Associated With Annona senegalensis (Magnoliales: Annonaceae) in Burkina Faso, West Africa.

Annona senegalensis Pers. is a shrub of tropical countries that, during the fruiting period, harbor many insects. All parts of the plant are used and exploited in traditional medicine, food, and firewood. Our study aimed at evaluating the diversity of insects associated with the different phenological stages of A. senegalensis fruits in two phytogeographic zones of Burkina Faso. Sampling was carried out on flowers, green fruits, ripe fruits, and decayed fruits of A. senegalensis. For the first time, a total of 48 insects species belonging to 6 orders and 23 families were identified. These orders were Orthoptera, Hemiptera, Hymenoptera, Coleoptera, Lepidoptera, and Diptera. Our data indicated that the diversity of insect species varies according to the stages of development of the fruit (P = 0.017) and according to the site (P = 2.2e-16). Among these insects, Curculionidae (Endaeus spp.) predominate on flowers, are known to be pollinators, and Formicidae (Messor galla Mayr, [Hymenoptera: Formicidae], Trichomyrmex abyssinicus Forel, [Hymenoptera: Formicidae], and Crematogaster sp.) and Tettigometridae (Hilda undata Walker, [Hemiptera: Tettigometridae]) are suspected to have mutualistic relationships on green fruits. Potential pests belonging to the Scarabaeidae (Pachnoda spp., Polybaphes spp., and Xeloma Maura Boheman, [Coleoptera: Scarabaeidae]), Drosophilidae (Zaprionus indianus Gupta, [Diptera: Drosophilidae]), and Nitidulidae (Carpophilus nepos Murray, [Coleoptera: Nitidulidae]) are associated with ripe and decayed fruits. The data in this study highlight the diversity in terms of pollinators that ensure the fruit production and Formicidae known to protect A. senegalensis against potential pests. These data provide valuable information in terms of valuation of this plant.

Indigenous knowledge system associated with the uses of insects for therapeutic or medicinal purposes in two main provinces of Burkina Faso, West Africa.

BACKGROUND: Some insects are harmful to humans, plants and animals, but some of them can also be a source of proteins, fats, vitamins and minerals and be of therapeutic value. The therapeutic potential requires that medicinal insects and their derived products need to be scrutinized. This study highlights the indigenous knowledge related to their use of medicinal insects in peri-urban and urban areas of Burkina Faso. METHODS: The survey was carried out among 60 traditional healers spread across two phytogeographical zones of Burkina Faso. The questionnaire focused on medicinal insects used by experienced traditional healers. Chi-square tests and principal component analysis were performed to test for significant differences regarding knowledge of how insects in phytogeographically different areas were used therapeutically in connection with different disease categories. RESULTS: A total of 19 species of medicinal insects belonging to 6 orders were cited in connection with treatments of at least 78 pathologies and symptoms. Most frequently mentioned was gastroenteritis. Our study showed that 48.78% of the insects and their products were associated with 46 plant species for the treatment of pathologies. In addition, honey, beeswax and nests were the most widely insect products used. CONCLUSION: The current study allows us to identify medicinal insects as well as their products used in the treatment of pathologies and symptoms, suggesting the presence of a considerable diversity of therapeutically important insect species. These insects are used alone and/or with their products but often in association with medicinal plants. The results constitute a useful database for future studies of medicinal insects in central and western parts of Burkina Faso.

Lack of robust evidence for a Wolbachia infection in Anopheles gambiae from Burkina Faso.

The endosymbiont Wolbachia can have major effects on the reproductive fitness, and vectorial capacity of host insects and may provide new avenues to control mosquito-borne pathogens. Anopheles gambiae s.l is the major vector of malaria in Africa but the use of Wolbachia in this species has been limited by challenges in establishing stable transinfected lines and uncertainty around native infections. High frequencies of infection of Wolbachia have been previously reported in An. gambiae collected from the Valle du Kou region of Burkina Faso in 2011 and 2014. Here, we re-evaluated the occurrence of Wolbachia in natural samples, collected from Valle du Kou over a 12-year time span, and in addition, expanded sampling to other sites in Burkina Faso. Our results showed that, in contrast to earlier reports, Wolbachia is present at an extremely low prevalence in natural population of An. gambiae. From 5341 samples analysed, only 29 were positive for Wolbachia by nested PCR representing 0.54% of prevalence. No positive samples were found with regular PCR. Phylogenetic analysis of 16S rRNA gene amplicons clustered across supergroup B, with some having similarity to sequences previously found in Anopheles from Burkina Faso. However, we cannot discount the possibility that the amplicon positive samples we detected were due to environmental contamination or were false positives. Regardless, the lack of a prominent native infection in An. gambiae s.l. is encouraging for applications utilizing Wolbachia transinfected mosquitoes for malaria control.

(GTG)5-PCR fingerprinting of multi-drug resistant Escherichia coli bacteria isolates from hospital in Ouagadougou, Burkina Faso.

BACKGROUND: Escherichia coli (E. coli) is the most common bacterial species implicated in various types of infections including septicemia, gastroenteritis, urinary tract infections, meningitis and others pathologies. These involve several bacterial clones with multidrug resistance making them difficult to treat. The aims of this study was to perform molecular typing of E. coli strains using universal primer (GTG)5. In this study, 53 E. coli strains were collected from inpatients and outpatients. The test of antimicrobial sensibility was realized using CA-SFM /EUCAST method and strains were identified by conventional microbiological tests. The carbapenemase-producing strains were demonstrated by phenotypic method. Bacterial DNA was extracted by boiling method. (GTG)5-PCR was used for strain subtyping. The DendroUPGMA software was used for grouping of strains from the genetic fingerprints obtained by (GTG)5-PCR. RESULTS: Antibiotic susceptibility test revealed that all strains were multi-drug resistant (MDR). Its strains showed resistance to at least three different families of antibiotics. Of this MDR strains, only one was a metallo-ß-lactamase producer. The dendrogram obtained using genetic fingerprinting allowed the E. coli strains to be grouped into 22 clusters (G1 to G22). CONCLUSION: The (GTG) 5-PCR assay enabled rapid molecular typing of E. coli strains. The strains of E. coli typed in this study would belong to different clones.

Local adaptation and colonization are potential factors affecting sexual competitiveness and mating choice in Anopheles coluzzii populations.

The mating behaviour of the malaria vector Anopheles gambiae complex is an important aspect of its reproduction biology. The success of mosquito release programmes based on genetic control of malaria crucially depends on competitive mating between both laboratory-reared and wild individuals, and populations from different localities. It is known that intrinsic and extrinsic factors can influence the mating success. This study addressed some of the knowledge gaps about factors influcencing mosquito mating success. In semi-field conditions, the study compared the mating success of three laboratory-reared and wild allopatric An. coluzzii populations originating from ecologically different locations in Burkina Faso. Overall, it was found that colonization reduced the mating competitiveness of both males and females compared to that of wild type individuals. More importly, females were more likely to mate with males of their own population of origin, be it wild or colonised, suggesting that local adaptation affected mate choice. The observations of mating behaviour of colonized and local wild populations revealed that subtle differences in behaviour lead to significant levels of population-specific mating. This is the first study to highlight the importance of local adaptation in the mating success, thereby highlighting the importance of using local strains for mass-rearing and release of An. coluzzii in vector control programmes.

Contrasting effects of the alkaloid ricinine on the capacity of Anopheles gambiae and Anopheles coluzzii to transmit Plasmodium falciparum.

BACKGROUND: Besides feeding on blood, females of the malaria vector Anopheles gambiae sensu lato readily feed on natural sources of plant sugars. The impact of toxic secondary phytochemicals contained in plant-derived sugars on mosquito physiology and the development of Plasmodium parasites remains elusive. The focus of this study was to explore the influence of the alkaloid ricinine, found in the nectar of the castor bean Ricinus communis, on the ability of mosquitoes to transmit Plasmodium falciparum. METHODS: Females of Anopheles gambiae and its sibling species Anopheles coluzzii were exposed to ricinine through sugar feeding assays to assess the effect of this phytochemical on mosquito survival, level of P. falciparum infection and growth rate of the parasite. RESULTS: Ricinine induced a significant reduction in the longevity of both Anopheles species. Ricinine caused acceleration in the parasite growth rate with an earlier invasion of the salivary glands in both species. At a concentration of 0.04 g l-1 in An. coluzzii, ricinine had no effect on mosquito infection, while 0.08 g l-1 ricinine-5% glucose solution induced a 14% increase in An. gambiae infection rate. CONCLUSIONS: Overall, our findings reveal that consumption of certain nectar phytochemicals can have unexpected and contrasting effects on key phenotypic traits that govern the intensity of malaria transmission. Further studies will be required before concluding on the putative role of ricinine as a novel control agent, including the development of ricinine-based toxic and transmission-blocking sugar baits. Testing other secondary phytochemicals in plant nectar will provide a broader understanding of the impact which plants can have on the transmission of vector-borne diseases.

Chemical composition, energy and nutritional values, digestibility and functional properties of defatted flour, protein concentrates and isolates from Carbula marginella (Hemiptera: Pentatomidae) and Cirina butyrospermi (Lepidoptera: Saturniidae).

Edible insects constitute a potential source of alternative proteins as a food supplement. The present study aimed to investigate the chemical composition, energy and nutritional values, the digestibility and functional properties of Carbula marginella (Thunberg) and Cirina butyrospermi (Vuillet) defatted flour, protein concentrates, and isolates. Carbula marginella has shown the highest content of protein (41.44%), lipid (51.92%), calcium (33.92 mg/100 g) and sodium (185.84 mg/100 g) while the highest contents of carbohydrate (34.54%), ash (4.77%), iron (31.27 mg/100 g), magnesium (150.09 mg/100 g), and potassium (1277 mg/100 g) have been observed for C. butyrospermi. Linoleic (30.23%), palmitic (27.54%), oleic (26.41%) and stearic (8.90%) acids were the most dominant fatty acids found in C. marginella. Cirina butyrospermi was characterized by high levels of oleic (27.01%), stearic (21.02%), linolenic (20.42%), palmitic (13.06%), and linoleic (8.01%) acids. Protein and essential amino acid contents of the protein isolates in both insect species were 1.7-2 times higher than that of their defatted flours. The protein isolate of C. marginella exhibited the highest protein digestibility (87.63%), while the highest fat absorption capacity (8.84 g/g) and foaming capacity (48.40%) have been obtained from the protein isolate of C. butyrospermi. These findings indicate that the protein concentrates and isolates of C. marginella and C. butyrospermi have great potential for industrial applications.

Comparison of swarming, mating performance and longevity of males Anopheles coluzzii between individuals fed with different natural fruit juices in laboratory and semi-field conditions.

BACKGROUND: It is assumed that malaria vectors feed on locally available nectar sources to obtain energy. Sugar feeding is energetically critical for the Anopheles male swarming and mating activities. However, little is known about the impact of local nectar feeding on male physiological development and its consequences on male mosquito life traits in the malaria control context. This study aimed to evaluate the influence of local fruit juices on the life traits of males Anopheles coluzzii. METHODS: Swarming characteristics (number of males in swarm, number of mating pairs, and swarm duration) in semi-field conditions; mating rate and longevity in a laboratory setting were compared between males An. coluzzii fed exclusively with mango, papaya or banana juices. The trophic preference was investigated in semi-field conditions. RESULTS: The results of this study showed that in the laboratory, mosquitoes fed with papaya juices lived on average longer (10 days) than those fed with banana or mango juices (5 days) and had higher a mating rate (53%) than those fed with banana juice (40%). In the semi-field, the swarm size of mosquitoes fed with banana juice (85 males) was larger than that of mosquitoes fed with mango juice (60 males). The number of mating pairs formed from banana-fed male swarms (17 mating pairs) was higher than that formed from mango-fed male swarm (8 mating pairs). There was no difference in swarming duration between male treatments. Male mosquitoes had a preference for papaya and banana juices. CONCLUSIONS: The results indicate that the origin of plant-derived feeding is an important factor in the survival and reproduction of mosquitoes. This calls for further investigations of chemical contents of nectars and their impact on the physiological development of mosquitoes.

Monitoring Insecticide Susceptibility in Aedes Aegypti Populations from the Two Biggest Cities, Ouagadougou and Bobo-Dioulasso, in Burkina Faso: Implication of Metabolic Resistance.

In West Africa, Aedes aegypti remains the major vector of dengue virus. Since 2013, dengue fever has been reemerging in Burkina Faso with annual outbreaks, thus becoming a major public health problem. Its control relies on vector control, which is unfortunately facing the problem of insecticide resistance. At the time of this study, although data on phenotypic resistance were available, information related to the metabolic resistance in Aedes populations from Burkina Faso remained very scarce. Here, we assessed the phenotypic and the metabolic resistance of Ae. aegypti populations sampled from the two main urban areas (Ouagadougou and Bobo-Dioulasso) of Burkina Faso. Insecticide susceptibility bioassays to chlorpyriphos-methyl 0.4%, bendiocarb 0.1% and deltamethrin 0.05% were performed on natural populations of Ae. aegypti using the WHO protocol. The activity of enzymes involved in the rapid detoxification of insecticides, especially non-specific esterases, oxidases (cytochrome P450) and glutathione-S-transferases, was measured on individual mosquitos. The mortality rates for deltamethrin 0.05% were low and ranged from 20.72% to 89.62% in the Bobo-Dioulasso and Ouagadougou sites, respectively. When bendiocarb 0.1% was tested, the mortality rates ranged from 7.73% to 71.23%. Interestingly, in the two urban areas, mosquitoes were found to be fully susceptible to chlorpyriphos-methyl 0.4%. Elevated activity of non-specific esterases and glutathione-S-transferases was reported, suggesting multiple resistance mechanisms involved in Ae. aegypti populations from Bobo-Dioulasso and Ouagadougou (including cytochrome P450). This update to the insecticide resistance status within Ae. aegypti populations in the two biggest cities is important to better plan dengue vectors control in the country and provides valuable information for improving vector control strategies in Burkina Faso, West Africa.

Sex aggregation and species segregation cues in swarming mosquitoes: role of ground visual markers.

BACKGROUND: Mating swarm segregation in closely related insect species may contribute to reproductive isolation. Visual markers are used for swarm formation; however, it is unknown whether they play a key role in swarm location, species segregation and sex aggregation. METHODS: Using two sympatric closely related species of the Anopheles gambiae complex, An. coluzzii and An. gambiae (s.s.), we investigated in both laboratory and semi-field conditions (i) whether males of the two species use visual markers (black cloths) to locate their swarm; and (ii) whether the presence/absence and size of the marker may differentially affect swarm characteristics. We also investigated whether conspecific virgin females use these markers to join male swarm sites. RESULTS: We showed that males of the two species used visual markers but in different ways: An. coluzzii swarm right above the marker whereas An. gambiae (s.s.) locate their swarm at a constant distance of 76.4 ± 0.6 cm from a 20 × 20 cm marker in the laboratory setup and at 206 ± 6 cm from a 60 × 60 cm marker in the semi-field setup. Although increased marker size recruited more mosquitoes and consequently increased the swarm size in the two species, An. coluzzii swarms flew higher and were stretched both vertically and horizontally, while An. gambiae (s.s.) swarms were only stretched horizontally. Virgin females displayed a swarm-like behavior with similar characteristics to their conspecific males. CONCLUSIONS: Our results provided experimental evidence that both An. coluzzii and An. gambiae (s.s.) males use ground visual markers to form and locate their swarm at species-specific locations. Moreover, the marker size differentially affected swarm characteristics in the two species. Our results also showed that virgin females displayed a swarm-like behavior. However, these "swarms" could be due to the absence of males in our experimental conditions. Nevertheless, the fact that females displayed these "swarms" with the same characteristics as their respective males provided evidence that visual markers are used by the two sexes to join mating spots. Altogether, this suggests that visual markers and the way species and sexes use them could be key cues in species segregation, swarm location and recognition.

Semi-field and indoor setups to study malaria mosquito swarming behavior.

BACKGROUND: The recent resurgence of interest in sterile insect techniques to control vector mosquitoes has renewed interest in novel methods for observing mating behavior. Malarial vectors of the Anopheles gambiae complex are known to mate in swarms at specific locations at dawn and dusk. Most knowledge of mosquito swarming behavior is derived from field observations and a few experimental studies designed to assess critical parameters that affect mosquito swarming. However, such studies are difficult to implement in the field because of uncontrollable environmental factors and mosquito conditions. Here, we present two experimental setups specifically designed to analyze mosquito swarming behavior and provide evidence that swarming behavior of mosquitoes can be generated and accurately assessed under both semi-field and laboratory conditions. METHODS: The Mosquito Ecology Research Facility setup is a semi-field enclosure made of 12 compartments (10.0 × 6.0 × 4.5 m L × W × H each) exposed to ambient meteorological and lighting conditions. The laboratory setup consists of a windowless room (5.1 × 4.7 × 3.0 m) in which both environmental and mosquito conditions can be controlled. In the two setups, 300 3-6-days-old An. coluzzii virgin males were released and some swarm characteristics were recorded such as the time at which the swarm started, the number of mosquitoes in the swarm and the height. Climatic conditions in the semi-field setup were also recorded. RESULTS: In both setups, An. coluzzii males displayed stereotyped and consistent swarming behavior day after day; males gradually gather into a swarm over a ground marker at sunset, flying in loops in relation to specific visual features on the ground. Although semi-field climatic conditions were slightly different from outdoors conditions, they did not impede swarming behavior and swarm characteristics were similar to those observed in the field. CONCLUSIONS: Swarm characteristics and their consistency across days provide evidences that these facilities can be used confidently to study swarming behavior. These facilities come to complement existing semi-field setups and pave the way for new experimental studies which will enhance our understanding of mating behavior but also mosquito ecology and evolution, a prerequisite for application of genetic approaches to malaria control.

Insects to feed insects - feeding Aedes mosquitoes with flies for laboratory rearing.

The black soldier fly, yellow mealworm and house fly are known for their wide distribution, ease of breeding, and environmental and nutritional attributes. Diets based on these fly proteins for the rearing of mosquito larvae are more accessible and affordable when compared to the reference IAEA diet which consists largely of costly livestock products such as bovine liver powder. Following a step-by-step assessment, we developed diet mixtures based on insect meal for the optimal mass production of Aedes albopictus and Ae. aegypti. Based on the assessed parameters including mosquito egg hatch, body size, flight ability, longevity and diet cost reduction, two mixtures are recommended: 1/2 tuna meal (TM) + 7/20 black soldier fly (BSF) + 3/20 brewer's yeast and 1/2 TM + 1/2 BSF. These findings, which could be adapted to other mosquito species, provide alternative protein sources for mass rearing insects for genetic control strategies.

Insecticide resistance mechanisms in Anopheles gambiae complex populations from Burkina Faso, West Africa.

Vector control constitutes a fundamental approach in reducing vector density and the efficient option to break malaria transmission in Africa. Malaria vectors developed resistance to almost all classes of insecticides recommended by WHO for vector control in most places of African countries and may compromise the vector control strategies. This study updated the resistance status of Anopheles gambiae complex populations to insecticides recommended for vector control in the western part of Burkina Faso. Insecticide susceptibility bioassays were performed on seven natural populations of An. gambiae complex from western Burkina Faso in the 2016 rainy season using the WHO protocol. Biochemical assays were carried out according to the WHO protocol on the same populations to estimate detoxifying enzymes activities including non-specific esterases (NSEs), oxidases (cytochrome P450) and Glutathione-S-Transferases (GSTs). Polymerase Chain Reactions (PCRs) were performed for the identification of the An. gambiae complex species as well as the detection of kdr-west and ace-1 mutations. Susceptibility bioassays showed that An. gambiae complex was multi-resistant to pyrethroids, DDT and carbamates in almost all areas. The mortality rates ranged from 10 to 38%, 2.67 to 59.57% and 64.38 to 98.02% for Deltamethrin, DDT and Bendiocarb respectively. A full susceptibility (100%) to an organophosphate, the Chlorpyrifos-methyl, was observed at the different sites. Three (3) species of the An. gambiae complex were identified: An. gambiae s.s, An. coluzzii and An. arabiensis. The frequencies of the kdr-w mutation were highly widespread (0.66 to 0.98) among the three species of the complex. The ace-1 mutation was detected at low frequencies (0 to 0.12) in An. gambiae s.s and An. coluzzii. A high level of GSTs and NSEs were observed within the different populations of the An. gambiae complex. Several mechanisms of insecticide resistance were found simultaneously in the same populations of An. gambiae complex conferring high multi-resistance to DDT, Carbamate and Pyrethroids. The full susceptibility of An. gambiae complex to organophosphates is a useful data for the national malaria control program in selecting the most appropriate products to both maintain the effectiveness of vector control strategies and best manage insecticide resistance as well as developing new alternative strategies for the control of major malaria vectors in Burkina Faso.