Resistance to insecticides inAnophelesmosquitoes 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 femaleAn. gambiaeandAn. coluzziifrom 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 aroundCyp6aa1, this resistance was based on a combination of several independent CNVs inAn. coluzzii, and on a non-CNV bearing haplotype inAn. 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 ofAce1, more widespread signals included cytochrome P450s, glutathione S-transferases, and a subunit of the nAChR target site of neonicotinoid insecticides. The regions aroundCyp9k1and theTepfamily 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.

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.

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Background and aim: Hibiscus surattensis L. is a medicinal plant widely used traditionally in Benin to treat malaria. This study was designed to investigate antiplasmodial activity, hemolytic power, acute toxicity, antioxidant activity and phytochemical content of H. surattensis. Methodology: Bioassay-guided isolation approach was adopted and extracts and fractions collected were continuously tested in vitro against Chloroquine-sensitive and field isolate strains of Plasmodium falciparum by immune-dosage of Plasmodium lactate dehydrogenase. Hemolytic effect and acute toxicity of extracts were evaluated respectively on human erythrocytes and according to OECD guideline N°423. 2, 2-diphenyl-1- picrylhydrazyl) radical scavenging, Ferric reducing antioxidant power, superoxyde radical scavenging and hydrogen peroxide radical scavenging methods were used to investigate antioxidant activity. Results and conclusion:  The best antiplasmodial activity was obtained with ethanolic extract of H. surattensis L. against field isolate (IC50 = 5.03±0.31 µg/mL) and Chloroquine -sensitive (IC50 = 7.55±0.59 µg/mL) whereas aqueous extract exhibited moderate activity. Bioassay-guided fractionation of ethanolic extract shows progressive decrease of the antiplasmodial activity. Both extracts exhibited strong antioxidant activity, hemolytic power less than 1%. No mortality of rats was recorded with ethanolic extract at 2000 mg/kg body weight. Flavonoids, anthraquinones, coumarins, and triterpenes are present in both extracts with tannins in the ethanolic extract. In summary, the extracts of H. surattensis have interesting antiplasmodial and antioxidant properties probably resulting from a synergetic action of their secondary metabolites, without toxicity effect on rats and human erythrocytes. These findings strengthen the traditional use of H. surattensis as antimalarial plants. Keywords: Hibiscus surattensis; antiplasmodial; antioxidant; toxicity; phytoconstituents.

Aims: This study was designed to assess antiplasmodial and antioxidant activities in relation to phytochemical contents and toxicological profile of crude extracts of Cola millenii leaves. Place and Duration of Study: The study was carried out from April 2015 to November 2018 at University of Abomey-Calavi, Laboratory of Biochemistry and Bioactives Natural Products and Laboratory of Infectious Vector-borne Diseases. Methodology: Phytochemical screening of aqueous and ethanolic crude extracts was performed using standard methods. Estimation of total phenolics contains (TPC) and total flavonoids contains (TFC) was done using colorimetric methods. Antioxidant activity was assessed in vitro by 2,2-diphenyl-1-picrylhydrazyl (DPPH) Radical-Scavenging, Reducing power, superoxide radical scavenging and Hydrogen peroxide scavenging assay. Antiplasmodial activity was investigated in vitro using both field isolate and laboratory 3D7 strains of P. falciparum using Plasmodium lactate dehydrogenase (pLDH) assay. Hemoglobin release was measured spectrophotometrically to assess hemolytic power. Acute oral toxicity of extracts was evaluated on rats according to OECD 423 guideline. Results: Flavonoids, anthraquinones, coumarins, and triterpenes were recorded in both extracts while tannins were recorded only in ethanolic extract. High TPC and TFC were recorded with ethanolic extract with significate difference (P <0.01) when compared to aqueous extract. Ethanolic extract exhibited the highest parasite growth inhibition against both field isolate (41.62±1.78%) and 3D7 (45.89±1.66%) strains at 100 µg/mL. Both extracts exhibited strong antioxidant activity according to different methods used. Toxicological profile revealed no hemolytic effect on human red blood cells as well as acute toxicity signs in rats. Conclusion: This study demonstrated strong antioxidant and moderate antiplasmodial activities of C. millenii extracts without toxicity effect on rats and human erythrocytes. It would play an important role in malaria and oxidative damage control.

The insecticide resistance in Anopheles gambiae mosquitoes has remained the major threat for vector control programs but the fitness effects conferred by these mechanisms are poorly understood. To fill this knowledge gap, the present study aimed at testing the hypothesis that antibiotic oxytetracycline could have an interaction with insecticide resistance genotypes and consequently inhibit the fecundity in An. gambiae. Four strains of An. gambiae: Kisumu (susceptible), KisKdr (kdr (L1014F) resistant), AcerKis (ace-1 (G119S) resistant) and AcerKdrKis (both kdr (L1014F) and ace-1 (G119S) resistant) were used in this study. The different strains were allowed to bloodfeed on a rabbit previously treated with antibiotic oxytetracycline at a concentration of 39·10–5 M. Three days later, ovarian follicles were dissected from individual mosquito ovaries into physiological saline solution (0.9% NaCl) under a stereomicroscope and the eggs were counted. Fecundity was substantially lower in oxytetracycline-exposed KisKdr females when compared to that of the untreated individuals and oxytetracycline-exposed Kisumu females. The exposed AcerKis females displayed an increased fecundity compared to their nontreated counterparts whereas they had reduced fecundity compared to that of oxytetracycline-exposed Kisumu females. There was no substantial difference between the fecundity in the treated and untreated AcerKdrKis females. The oxytetracycline-exposed AcerKdrKis mosquitoes had an increased fecundity compared to that of the exposed Kisumu females. Our data indicate an indirect effect of oxytetracycline in reducing fecundity of An. gambiae mosquitoes carrying kdrR (L1014F) genotype. These findings could be useful for designing new integrated approaches for malaria vector control in endemic countries.

Aim: Dissotis rotundifolia were selected after an ethnopharmacological survey conducted on plants used traditionally for malaria treatment in South Benin, with the aim of discovering new natural active extracts against malaria parasites. Place and Duration of Study: Laboratory of Biochemistry and Bioactive Natural Substances, University of Abomey-Calavi (Benin)/ Laboratory of Infectious Vector Borne Diseases, Regional Institute of Public Health (Benin)/ Laboratoire d’Histologie, de Cytogénétique et d’Embryologie, Faculté des Sciences de la Santé (Benin). The study was conduct from October 2018 to June 2019 in Benin. Methodology: The antiplasmodial activity of the plant extracts was evaluated using the parasite lactate dehydrogenase (pLDH) immunodetection assay. The extract with the best antiplasmodial activity were used on Wistar rats for acute toxicity. Results: Ethanolic extract of Dissotis rotundifolia showed promising activity (Isolate: IC50 = 22.58 ± 1.12 µg/mL; 3D7: IC50 = 6.81 ± 0.85 µg/mL) on Plasmodium falciparum compared to the aqueous extract (Isolate: IC50 > 100 µg/mL; 3D7: IC50> 100 µg/mL). The aqueous fraction of D. rotundifolia exhibit highly potent activity against P. falciparum strain (Isolate: IC50 > 100 µg/mL μg/mL; 3D7: IC50 = 4.05 ± 0.72 μg/mL). Haemolytic effect of actives extracts and fractions is less than 5%. Ethanolic extract of D. rotundifolia revealed no obvious acute toxicity in rat up to the highest dose administered (2000 mg/kg). Conclusion: This study justifies traditional uses of D. rotundifolia against malaria. A bioguided fractionation of these extracts would identify molecules responsible for their antiplasmodial activity. Moreover, these results could lead to the design of improved traditional medicines in the basis of this plant.

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.