Key gene modules and hub genes associated with pyrethroid and organophosphate resistance in Anopheles mosquitoes: a systems biology approach.

Indoor residual spraying (IRS) and insecticide-treated nets (ITNs) are the main methods used to control mosquito populations for malaria prevention. The efficacy of these strategies is threatened by the spread of insecticide resistance (IR), limiting the success of malaria control. Studies of the genetic evolution leading to insecticide resistance could enable the identification of molecular markers that can be used for IR surveillance and an improved understanding of the molecular mechanisms associated with IR. This study used a weighted gene co-expression network analysis (WGCNA) algorithm, a systems biology approach, to identify genes with similar co-expression patterns (modules) and hub genes that are potential molecular markers for insecticide resistance surveillance in Kenya and Benin. A total of 20 and 26 gene co-expression modules were identified via average linkage hierarchical clustering from Anopheles arabiensis and An. gambiae, respectively, and hub genes (highly connected genes) were identified within each module. Three specific genes stood out: serine protease, E3 ubiquitin-protein ligase, and cuticular proteins, which were top hub genes in both species and could serve as potential markers and targets for monitoring IR in these malaria vectors. In addition to the identified markers, we explored molecular mechanisms using enrichment maps that revealed a complex process involving multiple steps, from odorant binding and neuronal signaling to cellular responses, immune modulation, cellular metabolism, and gene regulation. Incorporation of these dynamics into the development of new insecticides and the tracking of insecticide resistance could improve the sustainable and cost-effective deployment of interventions.

Parallel Evolution in Mosquito Vectors-A Duplicated Esterase Locus is Associated With Resistance to Pirimiphos-methyl in Anopheles gambiae.

The primary control methods for the African malaria mosquito, Anopheles gambiae, are based on insecticidal interventions. Emerging resistance to these compounds is therefore of major concern to malaria control programs. The organophosphate (OP), pirimiphos-methyl, is a relatively new chemical in the vector control armory but is now widely used in indoor-residual spray campaigns. While generally effective, phenotypic resistance has developed in some areas in malaria vectors. Here, we used a population genomic approach to identify novel mechanisms of resistance to pirimiphos-methyl in A. gambiae s.l mosquitoes. In multiple populations, we found large and repeated signals of selection at a locus containing a cluster of detoxification enzymes, some of whose orthologs are known to confer resistance to OPs in Culex pipiens. Close examination revealed a pair of alpha-esterases, Coeae1f and Coeae2f, and a complex and diverse pattern of haplotypes under selection in A. gambiae, A. coluzzii and A. arabiensis. As in C. pipiens, copy number variants have arisen at this locus. We used diplotype clustering to examine whether these signals arise from parallel evolution or adaptive introgression. Using whole-genome sequenced phenotyped samples, we found that in West Africa, a copy number variant in A. gambiae is associated with resistance to pirimiphos-methyl. Overall, we demonstrate a striking example of contemporary parallel evolution which has important implications for malaria control programs.

DNA Nanostructure-Templated Antibody Complexes Provide Insights into the Geometric Requirements of Human Complement Cascade Activation.

The classical complement pathway is activated by antigen-bound IgG antibodies. Monomeric IgG must oligomerize to activate complement via the hexameric C1q complex, and hexamerizing mutants of IgG appear as promising therapeutic candidates. However, structural data have shown that it is not necessary to bind all six C1q arms to initiate complement, revealing a symmetry mismatch between C1 and the hexameric IgG complex that has not been adequately explained. Here, we use DNA nanotechnology to produce specific nanostructures to template antigens and thereby spatially control IgG valency. These DNA-nanotemplated IgG complexes can activate complement on cell-mimetic lipid membranes, which enabled us to determine the effect of IgG valency on complement activation without the requirement to mutate antibodies. We investigated this using biophysical assays together with 3D cryo-electron tomography. Our data revealed the importance of interantigen distance on antibody-mediated complement activation, and that the cleavage of complement component C4 by the C1 complex is proportional to the number of ideally spaced antigens. Increased IgG valency also translated to better terminal pathway activation and membrane attack complex formation. Together, these data provide insights into how nanopatterning antigen-antibody complexes influence the activation of the C1 complex and suggest routes to modulate complement activation by antibody engineering. Furthermore, to our knowledge, this is the first time DNA nanotechnology has been used to study the activation of the complement system.

Transcriptomic analysis of Anopheles gambiae from Benin reveals overexpression of salivary and cuticular proteins associated with cross-resistance to pyrethroids and organophosphates.

BACKGROUND: Insecticide resistance (IR) is one of the major threats to malaria vector control programs in endemic countries. However, the mechanisms underlying IR are poorly understood. Thus, investigating gene expression patterns related to IR can offer important insights into the molecular basis of IR in mosquitoes. In this study, RNA-Seq was used to characterize gene expression in Anopheles gambiae surviving exposure to pyrethroids (deltamethrin, alphacypermethrin) and an organophosphate (pirimiphos-methyl). RESULTS: Larvae of An. gambiae s.s. collected from Bassila and Djougou in Benin were reared to adulthood and phenotyped for IR using a modified CDC intensity bottle bioassay. The results showed that mosquitoes from Djougou were more resistant to pyrethroids (5X deltamethrin: 51.7% mortality; 2X alphacypermethrin: 47.4%) than Bassila (1X deltamethrin: 70.7%; 1X alphacypermethrin: 77.7%), while the latter were more resistant to pirimiphos-methyl (1.5X: 48.3% in Bassila and 1X: 21.5% in Djougou). RNA-seq was then conducted on resistant mosquitoes, non-exposed mosquitoes from the same locations and the laboratory-susceptible An. gambiae s.s. Kisumu strain. The results showed overexpression of detoxification genes, including cytochrome P450s (CYP12F2, CYP12F3, CYP4H15, CYP4H17, CYP6Z3, CYP9K1, CYP4G16, and CYP4D17), carboxylesterase genes (COEJHE5E, COE22933) and glutathione S-transferases (GSTE2 and GSTMS3) in all three resistant mosquito groups analyzed. Genes encoding cuticular proteins (CPR130, CPR10, CPR15, CPR16, CPR127, CPAP3-C, CPAP3-B, and CPR76) were also overexpressed in all the resistant groups, indicating their potential role in cross resistance in An. gambiae. Salivary gland protein genes related to 'salivary cysteine-rich peptide' and 'salivary secreted mucin 3' were also over-expressed and shared across all resistant groups. CONCLUSION: Our results suggest that in addition to metabolic enzymes, cuticular and salivary gland proteins could play an important role in cross-resistance to multiple classes of insecticides in Benin. These genes warrant further investigation to validate their functional role in An. gambiae resistance to insecticides.

Beninese Plant Extracts with Antiplasmodial Activity Select New Allele Variants Msp1 and Msp2 in Plasmodium falciparum.

Background: Natural medicinal products are commonly used as a remedy against malaria infections in African populations and have become a major source of information for the screening of new and more effective antiplasmodial molecules. Therefore, in vitro studies are needed to validate the efficacy of these medicinal products and to explore the potential effects of such drugs on the genetic diversity of Plasmodium falciparum. The current study has investigated the impact of some Beninese plant extracts with antiplasmodial activity on the genetic diversity of P. falciparum. Method: Five (5) ethanolic plant extracts (Dissotis rotundifolia, Ehretia cymosa Thonn, Hibiscus surattensis L., Cola millenii K. Shum, and Costus afer Ker Gawl) and a compound extracted from Ehretia cymosa Thonn (encoded CpE2) were tested against asexual stage parasites of a culture-adapted strain of P. falciparum. Subsequently, the P. falciparum Msp1 and Msp2 markers were genotyped, and the number of allelic variants and the multiplicity of infection (MOI) were compared between drug-exposed and unexposed parasites. Results: All plant extracts have shown inhibitory activity against asexual P. falciparum and selected new allelic variants of the Msp1 and Msp2 genes compared to unexposed parasites. The newly selected allelic variants were K1_100bp and RO33_300bp of the Msp1 gene and FC27_150bp, FC27_300bp, FC27_400bp, and FC27_600bp of the Msp2 gene. However, there was no significant difference in MOI between drug-exposed and unexposed parasites. Conclusion: Our study highlights a source for the selection of new Msp1 and Msp2 alleles after exposure to antimalarial drugs. These findings pave the way for further studies investigating the true roles of these newly selected alleles in P. falciparum.

Parallel evolution in mosquito vectors - a duplicated esterase locus is associated with resistance to pirimiphos-methyl in An. gambiae.

The primary control methods for the African malaria mosquito, Anopheles gambiae, are based on insecticidal interventions. Emerging resistance to these compounds is therefore of major concern to malaria control programmes. The organophosphate, pirimiphos-methyl, is a relatively new chemical in the vector control armoury but is now widely used in indoor residual spray campaigns. Whilst generally effective, phenotypic resistance has developed in some areas in malaria vectors. Here, we used a population genomic approach to identify novel mechanisms of resistance to pirimiphos-methyl in Anopheles gambiae s.l mosquitoes. In multiple populations, we found large and repeated signals of selection at a locus containing a cluster of detoxification enzymes, some of whose orthologs are known to confer resistance to organophosphates in Culex pipiens. Close examination revealed a pair of alpha-esterases, Coeae1f and Coeae2f, and a complex and diverse pattern of haplotypes under selection in An. gambiae, An. coluzzii and An. arabiensis. As in Cx. pipiens, copy number variation seems to play a role in the evolution of insecticide resistance at this locus. We used diplotype clustering to examine whether these signals arise from parallel evolution or adaptive introgression. Using whole-genome sequenced phenotyped samples, we found that in West Africa, a copy number variant in Anopheles gambiae is associated with resistance to pirimiphos-methyl. Overall, we demonstrate a striking example of contemporary parallel evolution which has important implications for malaria control programmes.

Vectorial competence, insecticide resistance in Anopheles funestus and operational implications for malaria vector control strategies in Benin Republic.

The primary reason for the failure of malaria vector control across endemic regions is the widespread insecticide resistance observed in Anopheles vectors. The most dominant African vectors of malaria parasites are Anopheles gambiae and Anopheles funestus mosquitoes. These species often exhibit divergent behaviours and adaptive changes underscoring the importance of deploying active and effective measures in their control. Unlike An. gambiae, An. funestus mosquitoes are poorly studied in Benin Republic. However, recent reports indicated that An. funestus can adapt and colonize various ecological niches owing to its resistance against insecticides and adaptation to changing breeding habitats. Unfortunately, scientific investigations on the contribution of An. funestus to malaria transmission, their susceptibility to insecticide and resistance mechanism developed are currently insufficient for the design of better control strategies. In an attempt to gather valuable information on An. funestus, the present review examines the progress made on this malaria vector species in Benin Republic and highlights future research perspectives on insecticide resistance profiles and related mechanisms, as well as new potential control strategies against An. funestus. Literature analysis revealed that An. funestus is distributed all over the country, although present in low density compared to other dominant malaria vectors. Interestingly, An. funestus is being found in abundance during the dry seasons, suggesting an adaptation to desiccation. Among the An. funestus group, only An. funestus sensu stricto (s.s.) and Anopheles leesoni were found in the country with An. funestus s.s. being the most abundant species. Furthermore, An. funestus s.s. is the only one species in the group contributing to malaria transmission and have adapted biting times that allow them to bite at dawn. In addition, across the country, An. funestus were found resistant to pyrethroid insecticides used for bed nets impregnation and also resistant to bendiocarb which is currently being introduced in indoor residual spraying formulation in malaria endemic regions. All these findings highlight the challenges faced in controlling this malaria vector. Therefore, advancing the knowledge of vectorial competence of An. funestus, understanding the dynamics of insecticide resistance in this malaria vector, and exploring alternative vector control measures, are critical for sustainable malaria control efforts in Benin Republic.

Emplacement of the Argyle diamond deposit into an ancient rift zone triggered by supercontinent breakup.

Argyle is the world's largest source of natural diamonds, yet one of only a few economic deposits hosted in a Paleoproterozoic orogen. The geodynamic triggers responsible for its alkaline ultramafic volcanic host are unknown. Here we show, using U-Pb and (U-Th)/He geochronology of detrital apatite and detrital zircon, and U-Pb dating of hydrothermal titanite, that emplacement of the Argyle lamproite is bracketed between 1311 ± 9 Ma and 1257 ± 15 Ma (2σ), older than previously known. To form the Argyle lamproite diatreme complex, emplacement was likely driven by lithospheric extension related to the breakup of the supercontinent Nuna. Extension facilitated production of low-degree partial melts and their migration through transcrustal corridors in the Paleoproterozoic Halls Creek Orogen, a rheologically-weak rift zone adjacent to the Kimberley Craton. Diamondiferous diatreme emplacement during (super)continental breakup may be prevalent but hitherto under-recognized in rift zones at the edges of ancient continental blocks.

Genome-wide association studies reveal novel loci associated with pyrethroid and organophosphate resistance in Anopheles gambiae and Anopheles coluzzii.

Resistance to insecticides in Anopheles mosquitoes threatens the effectiveness of malaria control, but the genetics of resistance are only partially understood. We performed a large scale multi-country genome-wide association study of resistance to two widely used insecticides: deltamethrin and pirimiphos-methyl, using sequencing data from An. gambiae and An. coluzzii from ten locations in West Africa. Resistance was highly multi-genic, multi-allelic and variable between populations. While the strongest and most consistent association with deltamethrin resistance came from Cyp6aa1, this was based on several independent copy number variants (CNVs) in An. coluzzii, and on a non-CNV haplotype in An. gambiae. For pirimiphos-methyl, signals included Ace1, cytochrome P450s, glutathione S-transferases and the nAChR target site of neonicotinoid insecticides. The regions around Cyp9k1 and the Tep family of immune genes showed evidence of cross-resistance to both insecticides. These locally-varying, multi-allelic patterns highlight the challenges involved in genomic monitoring of resistance, and may form the basis for improved surveillance methods.

Relative expression of key genes involved in nucleic acids methylation in Anopheles gambiae sensu stricto.

In vertebrates, enzymes responsible for DNA methylation, one of the epigenetic mechanisms, are encoded by genes falling into the cytosine methyltransferases genes family (Dnmt1, Dnmt3a,b and Dnmt3L). However, in Diptera, only the methyltransferase Dnmt2 was found, suggesting that DNA methylation might act differently for species in this order. Moreover, genes involved in epigenetic dynamics, such as Ten-eleven Translocation dioxygenases (TET) and Methyl-CpG-binding domain (MBDs), present in vertebrates, might play a role in insects. This work aimed at investigating nucleic acids methylation in the malaria vector Anopheles gambiae (Diptera: Culicidae) by analysing the expression of Dnmt2, TET2 and MBDs genes using quantitative real-time polymerase chain reaction (qRT-PCR) at pre-immature stages and in reproductive tissues of adult mosquitoes. In addition, the effect of two DNA methylation inhibitors on larval survival was evaluated. The qPCR results showed an overall low expression of Dnmt2 at all developmental stages and in adult reproductive tissues. In contrast, MBD and TET2 showed an overall higher expression. In adult mosquito reproductive tissues, the expression level of the three genes in males' testes was significantly higher than that in females' ovaries. The chemical treatments did not affect larval survival. The findings suggest that mechanisms other than DNA methylation underlie epigenetic regulation in An. gambiae.

Asymptomatic Plasmodium infection among primary schoolchildren and Anopheles-mediated malaria transmission: A cross-sectional study in Ouidah; south-western Benin.

Understanding the contribution of asymptomatic Plasmodium carriers in malaria transmission might be helpful to design and implement new control measures. The present study explored the prevalence of asymptomatic and symptomatic Plasmodium infections (asexual and sexual stages) and the contribution of asymptomatic P. falciparum carriers to Anopheles-mediated malaria transmission in Ouidah (Benin). Thick and thin blood smears were examined from finger-prick blood specimens using light microscopy, and the density of both asexual and sexual stages of Plasmodium species was calculated. Infectivity of gametocyte-infected blood samples to Anopheles gambiae was assessed through direct membrane feeding assays. The prevalence of asymptomatic Plasmodium infections was 28.73% (289/1006). All the asymptomatic gametocyte-carriers (19/19), with gametocytaemia ranging from 10 - 1200 gametocytes/µL of blood, were infectious to An. gambiae mosquitoes. The mean oocyst prevalences varied significantly (χ 2  = 16.42, df = 7, p = 0.02) among laboratory mosquito strains (6.9 - 39.4%) and near-field mosquitoes (4.9 - 27.2%). Likewise, significant variation (χ 2  = 56.85, df = 7, p = 6.39 × 10-10) was observed in oocyst intensity. Our findings indicate that asymptomatic Plasmodium carriers could significantly contribute to malaria transmission. Overall, this study highlights the importance of diagnosing and treating asymptomatic and symptomatic infection carriers during malaria control programmes.

Genome-wide association studies reveal novel loci associated with pyrethroid and organophosphate resistance in Anopheles gambiae s.l.

Resistance to insecticides in Anopheles mosquitoes threatens the effectiveness of the most widespread tools currently used to control malaria. The genetic underpinnings of resistance are still only partially understood, with much of the variance in resistance phenotype left unexplained. We performed a multi-country large scale genome-wide association study of resistance to two insecticides widely used in malaria control: deltamethrin and pirimiphos-methyl. Using a bioassay methodology designed to maximise the phenotypic difference between resistant and susceptible samples, we sequenced 969 phenotyped female An. gambiae and An. coluzzii from ten locations across four countries in West Africa (Benin, Côte d'Ivoire, Ghana and Togo), identifying single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) segregating in the populations. The patterns of resistance association were highly multiallelic and variable between populations, with different genomic regions contributing to resistance, as well as different mutations within a given region. While the strongest and most consistent association with deltamethrin resistance came from the region around Cyp6aa1 , this resistance was based on a combination of several independent CNVs in An. coluzzii , and on a non-CNV bearing haplotype in An. gambiae . Further signals involved a range of cytochrome P450, mitochondrial, and immunity genes. Similarly, for pirimiphos-methyl, while the strongest signal came from the region of Ace1 , more widespread signals included cytochrome P450s, glutathione S-transferases, and a subunit of the nAChR target site of neonicotinoid insecticides. The regions around Cyp9k1 and the Tep family of immune genes were associated with resistance to both insecticide classes, suggesting possible cross-resistance mechanisms. These locally-varying, multigenic and multiallelic patterns highlight the challenges involved in genomic monitoring and surveillance of resistance, and form the basis for improvement of methods used to detect and predict resistance. Based on simulations of resistance variants, we recommend that yet larger scale studies, exceeding 500 phenotyped samples per population, are required to better identify associated genomic regions.

Malaria-Transmitting Vectors Microbiota: Overview and Interactions With Anopheles Mosquito Biology.

Malaria remains a vector-borne infectious disease that is still a major public health concern worldwide, especially in tropical regions. Malaria is caused by a protozoan parasite of the genus Plasmodium and transmitted through the bite of infected female Anopheles mosquitoes. The control interventions targeting mosquito vectors have achieved significant success during the last two decades and rely mainly on the use of chemical insecticides through the insecticide-treated nets (ITNs) and indoor residual spraying (IRS). Unfortunately, resistance to conventional insecticides currently being used in public health is spreading in the natural mosquito populations, hampering the long-term success of the current vector control strategies. Thus, to achieve the goal of malaria elimination, it appears necessary to improve vector control approaches through the development of novel environment-friendly tools. Mosquito microbiota has by now given rise to the expansion of innovative control tools, such as the use of endosymbionts to target insect vectors, known as "symbiotic control." In this review, we will present the viral, fungal and bacterial diversity of Anopheles mosquitoes, including the bacteriophages. This review discusses the likely interactions between the vector microbiota and its fitness and resistance to insecticides.

P. falciparum msp1 and msp2 genetic diversity in P. falciparum single and mixed infection with P. malariae among the asymptomatic population in Southern Benin.

Plasmodium falciparum and Plasmodium malariae infections are prevalent in malaria-endemic countries. However, very little is known about their interactions especially the effect of P. malariae on P. falciparum genetic diversity. This study aimed to assess P. falciparum genetic diversity in P. falciparum and mixed infection P. falciparum/P. malariae isolates among the asymptomatic populations in Southern Benin. Two hundred and fifty blood samples (125 of P. falciparum and 125 P. falciparum/P. malariae isolates) were analysed by a nested PCR amplification of msp1 and msp2 genes. The R033 allelic family was the most represented for the msp1 gene in mono and mixed infection isolates (99.2% vs 86.4%), while the K1 family had the lowest frequency (38.3% vs 20.4%). However, with the msp2 gene, the two allelic families displayed similar frequencies in P. falciparum isolates while the 3D7 allelic family was more represented in P. falciparum/P. malariae isolates (88.7%). Polyclonal infections were also lower (62.9%) in P. falciparum/P. malariae isolates (p < 0.05). Overall, 96 individual alleles were identified (47 for msp1 and 49 for msp2) in P. falciparum isolates while a total of 50 individual alleles were identified (23 for msp1 and 27 for msp2) in P. falciparum/P. malariae isolates. The Multiplicity of Infection (MOI) was lower in P. falciparum/P. malariae isolates (p < 0.05). This study revealed a lower genetic diversity of P. falciparum in P. falciparum/P. malariae isolates using msp1 and msp2 genes among the asymptomatic population in Southern Benin.

Overexpression of FcεRI on Bone Marrow Mast Cells, but Not MRGPRX2, in Clonal Mast Cell Disorders With Wasp Venom Anaphylaxis.

Background: Uncertainties remain about the molecular mechanisms governing clonal mast cell disorders (CMCD) and anaphylaxis. Objective: This study aims at comparing the burden, phenotype and behavior of mast cells (MCs) and basophils in patients with CMCD with wasp venom anaphylaxis (CMCD/WVA+), CMCD patients without anaphylaxis (CMCD/ANA-), patients with an elevated baseline serum tryptase (EBST), patients with wasp venom anaphylaxis without CMCD (WVA+) and patients with a non-mast cell haematological pathology (NMHP). Methods: This study included 20 patients with CMCD/WVA+, 24 with CMCD/ANA-, 19 with WVA+, 6 with EBST and 5 with NMHP. We immunophenotyped MCs and basophils and compared baseline serum tryptase (bST) and both total and venom specific IgE in the different groups. For basophil studies, 13 healthy controls were also included. Results: Higher levels of bST were found in CMCD patients with wasp venom anaphylaxis, CMCD patients without anaphylaxis and EBST patients. Total IgE levels were highest in patients with wasp venom anaphylaxis with and without CMCD. Bone marrow MCs of patients with CMCD showed lower CD117 expression and higher expression of CD45, CD203c, CD63, CD300a and FcεRI. Within the CMCD population, patients with wasp venom anaphylaxis showed a higher expression of FcεRI as compared to patients without anaphylaxis. Expression of MRGPRX2 on MCs did not differ between the study populations. Basophils are phenotypically and functionally comparable between the different patient populations. Conclusion: Patients with CMCD show an elevated burden of aberrant activated MCs with a significant overexpression of FcεRI in patients with a wasp venom anaphylaxis.

Surveillance of Plasmodium malariae infection among inhabitants of rural areas in Ouidah-Kpomasse-Tori Bossito health district, Benin.

Among the Plasmodium species that infect humans, P. falciparum has been largely studied in malaria endemic areas. However, P. malariae infection is less documented among the human population. This study aimed to monitor the prevalence and distribution of P. malariae in Southern Benin. A cross-sectional survey was conducted in rural localities in the Ouidah-Kpomasse-Tori Bossito (OKT) health district in Southern Benin from June to October 2019. Socio-demographic data were collected using a questionnaire, while malaria infection data were obtained on the one hand by microscopy diagnosis and, on the other, by nested polymerase chain reaction (PCR). Based on microscopy, the prevalence of P. malariae mono-infection and coinfection of P. falciparum, P. malariae was respectively 2.3% and 1.2% in the OKT health district. This prevalence was higher (P < 0.01) than that reported by Damien et al. (2010) 10 years ago in the same study area with 0.7% and 0.3% of P. malariae and P. falciparum/P. malariae, respectively. Based on PCR analysis, P. malariae prevalence was 14.1%, including 5.2% of mono-infection and 8.9% of mixed infection with P. falciparum. Sub-microscopic Plasmodium infections were high (30.6%) and more pronounced in older participants (>20 years). The present study revealed that P. malariae increased in the OKT health district with a high prevalence of submicroscopic infection. Since our results provide valuable evidence of increasing P. malariae infection, the National Malaria Control Programs (NMCPs) must consider P. malariae when designing future measures for effective control and malaria treatment.

Single nucleotide polymorphism (SNP) in the doublesex ( dsx) gene splice sites and relevance for its alternative splicing in the malaria vector Anopheles gambiae.

Background: Malaria burden continues to be significant in tropical regions, and conventional vector control methods are faced with challenges such as insecticide resistance. To overcome these challenges, additional vector control interventions are vital and include modern genetic approaches as well as classical methods like the sterile insect technique (SIT). In the major human malaria vector Anopheles gambiae, a candidate gene favourable for sterility induction is the doublesex ( dsx) gene, involved in mosquitos' somatic sexually dimorphic traits determination. However, the pathways that trigger the signal of dsx gene exon skipping alternative splicing mechanism in anopheline mosquitoes are not well characterized. This study aims to screen the An. gambiae dsx gene splice site sequences for single-nucleotide polymorphisms (SNPs) that could be critical to its alternative splicing. Methods: Variant annotation data from Ag1000G project phase 2 was analysed, in order to identify splice-relevant SNPs within acceptor and donor splice sites of the An. gambiae dsx gene ( Agdsx). Results: SNPs were found in both donor and acceptor sites of the Agdsx. No splice-relevant SNPs were identified in the female-specific intron 4 acceptor site and the corresponding region in males. Two SNPs (rs48712947, rs48712962) were found in the female-specific donor site of exon 5. They were not specific to either males or females as the rs48712947 was found in female mosquitoes from Cameroon, and in both males and females from Burkina Faso. In the other splice sites, the intron 3 acceptor site carried the greatest abundance of SNPs.   Conclusions: There were no gender association between the identified SNPs and the random distribution of these SNPs in mosquito populations. The SNPs in Agdsx splice sites are not critical for the alternative splicing. Other molecular mechanisms should be considered and investigated.

Putative pleiotropic effects of the knockdown resistance (L1014F) allele on the life-history traits of Anopheles gambiae.

BACKGROUND: Existing mechanisms of insecticide resistance are known to help the survival of mosquitoes following contact with chemical compounds, even though they could negatively affect the life-history traits of resistant malaria vectors. In West Africa, the knockdown resistance mechanism kdrR (L1014F) is the most common. However, little knowledge is available on its effects on mosquito life-history traits. The fitness effects associated with this knockdown resistance allele in Anopheles gambiae sensu stricto (s.s.) were investigated in an insecticide-free laboratory environment. METHODS: The life-history traits of Kisumu (susceptible) and KisKdr (kdr resistant) strains of An. gambiae s.s. were compared. Larval survivorship and pupation rate were assessed as well as fecundity and fertility of adult females. Female mosquitoes of both strains were directly blood fed through artificial membrane assays and then the blood-feeding success, blood volume and adult survivorship post-blood meal were assessed. RESULTS: The An. gambiae mosquitoes carrying the kdrR allele (KisKdr) laid a reduced number of eggs. The mean number of larvae in the susceptible strain Kisumu was three-fold overall higher than that seen in the KisKdr strain with a significant difference in hatching rates (81.89% in Kisumu vs 72.89% in KisKdr). The KisKdr larvae had a significant higher survivorship than that of Kisumu. The blood-feeding success was significantly higher in the resistant mosquitoes (84%) compared to the susceptible ones (34.75%). However, the mean blood volume was 1.36 µL/mg, 1.45 µL/mg and 1.68 µL/mg in Kisumu, homozygote and heterozygote KisKdr mosquitoes, respectively. After blood-feeding, the heterozygote KisKdr mosquitoes displayed highest survivorship when compared to that of Kisumu. CONCLUSIONS: The presence of the knockdown resistance allele appears to impact the life-history traits, such as fecundity, fertility, larval survivorship, and blood-feeding behaviour in An. gambiae. These data could help to guide the implementation of more reliable strategies for the control of malaria vectors.

Oceanic and super-deep continental diamonds share a transition zone origin and mantle plume transportation.

Rare oceanic diamonds are believed to have a mantle transition zone origin like super-deep continental diamonds. However, oceanic diamonds have a homogeneous and organic-like light carbon isotope signature (δ13C - 28 to - 20‰) instead of the extremely variable organic to lithospheric mantle signature of super-deep continental diamonds (δ13C - 25‰ to + 3.5‰). Here, we show that with rare exceptions, oceanic diamonds and the isotopically lighter cores of super-deep continental diamonds share a common organic δ13C composition reflecting carbon brought down to the transition zone by subduction, whereas the rims of such super-deep continental diamonds have the same δ13C as peridotitic diamonds from the lithospheric mantle. Like lithospheric continental diamonds, almost all the known occurrences of oceanic diamonds are linked to plume-induced large igneous provinces or ocean islands, suggesting a common connection to mantle plumes. We argue that mantle plumes bring the transition zone diamonds to shallower levels, where only those emplaced at the base of the continental lithosphere might grow rims with lithospheric mantle carbon isotope signatures.