Wolbachia are maternally transmitted intracellular bacteria that invade insect populations by manipulating their reproduction and immunity and thus limiting the spread of numerous human pathogens. Experimental Wolbachia infections can reduce Plasmodium numbers in Anopheles mosquitoes in the laboratory, however, natural Wolbachia infections in field anophelines have never been reported. Here we show evidence of Wolbachia infections in Anopheles gambiae in Burkina Faso, West Africa. Sequencing of the 16S rRNA gene identified Wolbachia sequences in both female and male germlines across two seasons, and determined that these sequences are vertically transmitted from mother to offspring. Whole-genome sequencing of positive samples suggests that the genetic material identified in An. gambiae belongs to a novel Wolbachia strain, related to but distinct from strains infecting other arthropods. The evidence of Wolbachia infections in natural Anopheles populations promotes further investigations on the possible use of natural Wolbachia–Anopheles associations to limit malaria transmission.

The maternally inherited alpha-proteobacterium Wolbachia has been proposed as a tool to block transmission of devastating mosquito-borne infectious diseases like dengue and malaria. Here we study the reproductive manipulations induced by a recently identified Wolbachia strain that stably infects natural mosquito populations of a major malaria vector, Anopheles coluzzii, in Burkina Faso. We determine that these infections significantly accelerate egg laying but do not induce cytoplasmic incompatibility or sex-ratio distortion, two parasitic reproductive phenotypes that facilitate the spread of other Wolbachia strains within insect hosts. Analysis of 221 blood-fed A. coluzzii females collected from houses shows a negative correlation between the presence of Plasmodium parasites and Wolbachia infection. A mathematical model incorporating these results predicts that infection with these endosymbionts may reduce malaria prevalence in human populations. These data suggest that Wolbachia may be an important player in malaria transmission dynamics in Sub-Saharan Africa.

This new resistance will have serious effects on malaria control.

Growing problems of pyrethroid resistance in Anopheles funestus have intensified efforts to identify alternative insecticides. Many agrochemicals target the GABA receptors, but cross-resistance from dieldrin resistance may preclude their introduction.Dieldrin resistance was detected in An. funestus populations from West (Burkina Faso) and central (Cameroon) Africa, but populations from East (Uganda) and Southern Africa (Mozambique and Malawi) were fully susceptible to this insecticide. Partial sequencing of the dieldrin target site, the γ-aminobutyric acid (GABA) receptor, identified two amino acid substitutions, A296S and V327I. The A296S mutation has been associated with dieldrin resistance in other species. The V327I mutations was detected in the resistant sample from Burkina Faso and Cameroon and consistently associated with the A296S substitution. The full-length of the An. funestus GABA-receptor gene, amplified by RT-PCR, generated a sequence of 1674 bp encoding 557 amino acid of the protein in An. funestus with 98% similarity to that of Anopheles gambiae. Two diagnostic assays were developed to genotype the A296S mutation (pyrosequencing and PCR-RFLP), and use of these assays revealed high frequency of the resistant allele in Burkina Faso (60%) and Cameroon (82%), moderate level in Benin (16%) while low frequency or absence of the mutation was observed respectively in Uganda (7.5%) or 0% in Malawi and Mozambique.The distribution of the RdlR mutation in An. funestus populations in Africa suggests extensive barriers to gene flow between populations from different regions.

BackgroundThe molecular forms of Anopheles gambiae are undergoing speciation. They are characterized by a strong assortative mating and they display partial habitat segregation. The M form is mostly found in flooded/irrigated areas whereas the S form dominates in the surrounding areas, but the ecological factors that shape this habitat segregation are not known. Resource competition has been demonstrated between species undergoing divergent selection, but resource competition is not the only factor that can lead to divergence.ResultsIn a field experiment using transplantation of first instar larvae, we evaluated the role of larval predators in mediating habitat segregation between the forms. We found a significant difference in the ability of the molecular forms to exploit the different larval sites conditioned on the presence of predators. In absence of predation, the molecular forms outcompeted each other in their respective natural habitats however, the developmental success of the M form was significantly higher than that of the S form in both habitats under predator pressure.ConclusionOur results provide the first empirical evidence for specific adaptive differences between the molecular forms and stress the role of larval predation as one of the mechanisms contributing to their divergence.

BackgroundDue to the spread of pyrethroid-resistance in malaria vectors in Africa, new strategies and tools are urgently needed to better control malaria transmission. The aim of this study was to evaluate the performances of a new mosaic long-lasting insecticidal net (LLIN), i.e. PermaNet® 3.0, against wild pyrethroid-resistant Anopheles gambiae s.l. in West and Central Africa.MethodsA multi centre experimental hut trial was conducted in Malanville (Benin), Vallée du Kou (Burkina Faso) and Pitoa (Cameroon) to investigate the exophily, blood feeding inhibition and mortality induced by PermaNet® 3.0 (i.e. a mosaic net containing piperonyl butoxide and deltamethrin on the roof) comparatively to the WHO recommended PermaNet® 2.0 (unwashed and washed 20-times) and a conventionally deltamethrin-treated net (CTN).ResultsThe personal protection and insecticidal activity of PermaNet 3.0 and PermaNet® 2.0 were excellent (>80%) in the "pyrethroid-tolerant" area of Malanville. In the pyrethroid-resistance areas of Pitoa (metabolic resistance) and Vallée du Kou (presence of the L1014F kdr mutation), PermaNet® 3.0 showed equal or better performances than PermaNet® 2.0. It should be noted however that the deltamethrin content on PermaNet® 3.0 was up to twice higher than that of PermaNet® 2.0. Significant reduction of efficacy of both LLIN was noted after 20 washes although PermaNet® 3.0 still fulfilled the WHO requirement for LLIN.ConclusionThe use of combination nets for malaria control offers promising prospects. However, further investigations are needed to demonstrate the benefits of using PermaNet® 3.0 for the control of pyrethroid resistant mosquito populations in Africa.

In West Africa, lineage splitting between the M and S molecular forms of the major Afro-tropical malaria mosquito, Anopheles gambiae is thought to be driven by ecological divergence, occurring mainly at the larval stage. Here, we present evidences for habitat segregation between the two molecular forms in and around irrigated rice-fields located within the humid savannahs background of western Burkina Faso. Longitudinal sampling of adult mosquitoes emerging from a range of breeding sites distributed along a transect extending from the heart of the rice-fields area into the surrounding savannahs was conducted from June to November 2009. Analysis revealed that the two molecular forms and their sibling species An. arabiensis are not randomly distributed in the area. A major ecological gradient was extracted, in relation to the rice-fields perimeter. The M form was associated with larger breeding sites, which were mainly represented by rice field paddies whereas the S form and An. arabiensis were found to depend upon temporary, rain-filled breeding sites. These results support hypotheses about larval habitat segregation and confirm that both forms have different larval habitat requirement. Segregation appears clearly linked to anthropogenic permanent habitats and the community structure they support.

The widespread insecticide resistance raises concerns for vector control implementation and sustainability particularly for the control of the main vector of human malaria, Anopheles gambiae sensu stricto. However, the extent to which insecticide resistance mechanisms interfere with the development of the malignant malaria parasite in its vector and their impact on overall malaria transmission remains unknown. We explore the impact of insecticide resistance on the outcome of Plasmodium falciparum infection in its natural vector using three An. gambiae strains sharing a common genetic background, one susceptible to insecticides and two resistant, one homozygous for the ace-1R mutation and one for the kdr mutation. Experimental infections of the three strains were conducted in parallel with field isolates of P. falciparum from Burkina Faso (West Africa) by direct membrane feeding assays. Both insecticide resistant mutations influence the outcome of malaria infection by increasing the prevalence of infection. In contrast, the kdr resistant allele is associated with reduced parasite burden in infected individuals at the oocyst stage, when compared to the susceptible strain, while the ace-1 R resistant allele showing no such association. Thus insecticide resistance, which is particularly problematic for malaria control efforts, impacts vector competence towards P. falciparum and probably parasite transmission through increased sporozoite prevalence in kdr resistant mosquitoes. These results are of great concern for the epidemiology of malaria considering the widespread pyrethroid resistance currently observed in Sub-Saharan Africa and the efforts deployed to control the disease.