345 research outputs found
Bionomics and insecticides resistance profiling of malaria vectors at a selected site for experimental hut trials in central Cameroon.
Get PDFBACKGROUND
Malaria vectors are increasingly developing resistance to insecticides across Africa. The impact of such resistance on the continued effectiveness of insecticide-based interventions remains unclear due to poor characterization of vector populations. This study reports the characterization of malaria vectors at Mibellon, a selected site in Cameroon for experimental hut study, including species composition, Plasmodium infection rate, resistance profiles and mechanisms.
METHODS
Indoor resting blood-fed Anopheles mosquitoes were collected from houses at Mibellon in 2017 and forced to lay eggs to generate F adult mosquitoes. Insecticides susceptibility bioassays were performed on the F adult mosquitoes following the WHO protocol to assess resistance profile to insecticides. The molecular basis of resistance and Plasmodium infection rate were investigated using TaqMan genotyping.
RESULTS
Anopheles funestus sensu stricto (s.s.) was predominant in Mibellon (80%) followed by Anopheles gambiae s.s. (20%). High levels of resistance to pyrethroids and organochlorides were observed for both species. Moderate resistance was observed against bendiocarb (carbamate) in both species, but relatively higher in An. gambiae s.s. In contrast, full susceptibility was recorded for the organophosphate malathion. The PBO synergist assays with permethrin and deltamethrin revealed a significant recovery of the susceptibility in Anopheles funestus s.s. population (48.8 to 98.1% mortality and 38.3 to 96.5% mortality, respectively). The DDT/pyrethroid 119F-GSTe2 resistant allele (28.1%) and the dieldrin 296S-RDL resistant (9.7%) were detected in An. funestus s.s. The high pyrethroid/DDT resistance in An. gambiae correlated with the high frequency of 1014F knockdown resistance allele (63.9%). The 1014S-kdr allele was detected at low frequency (1.97%). The Plasmodium infection rate was 20% in An. gambiae, whereas An. funestus exhibited an oocyst rate of 15 and 5% for the sporozoite rate.
CONCLUSION
These results highlight the increasing spread of insecticide resistance and the challenges that control programmes face to maintain the continued effectiveness of insecticide-based interventions
Genetic diversity and population structure of goliath frogs (Conraua goliath) from Cameroon.
Get PDFThe goliath frog (Conraua goliath) is an Endangered species exclusively found in Cameroon and Equatorial Guinea. Climate change, deforestation and overhunting are principal causes driving this species to extinction. Therefore, a better understanding of the genetic diversity and population structure of this species is necessary to improve conservation efforts. Here we used two mitochondrial genes (Cytochrome Oxidase subunit 1 (COI) and 16S) extracted from 54 C. goliath individuals from six localities in Cameroon to examine their genetic diversity. The result shows a low DNA substitution between the sequences. There were four 16S and two COI haplotypes in total. Overall, genetic diversity was very low for all the genes with nucleotide diversity of 0.00106 and 0.00007 for 16S and COI respectively. The Tajima D and Fu Fs statistics were negative. The TCS haplotype network revealed a predominant and ancestral haplotype (H1) for these genes which is distributed in the 6 populations. Pairwise genetic differentiation (F) generated between these populations using 16S revealed very high differentiation between populations from Nkam and Mungo Administrative Divisions in Cameroon. In contrast, we observed low differentiation among the geographically clustered Mungo and Nkam populations. Overall, human activities and perhaps climate change can appear to have depleted genetic diversity in the scattered populations that remain of this amphibian. To sustain the Goliath frog, we suggest to the Cameroonian government to implement more effective strategies to conserve and manage remnant populations of this iconic species through more effort against poaching which contribute to reduce the genetic diversity
Exploring the impact of glutathione S-transferase (GST)-based metabolic resistance to insecticide on vector competence of Anopheles funestus for Plasmodium falciparum
Get PDFBackground: Malaria control heavily relies on insecticide-based interventions against mosquito vectors. However, the increasing spread of insecticide resistance is a major threat. The extent to which such resistance, notably metabolic resistance, interferes with the development of the Plasmodium parasite and its impact on overall malaria transmission remains poorly characterized. Here, we investigated whether glutathione S-transferase-based resistance could influence Plasmodium falciparum development in Anopheles funestus. Methods: Anopheles funestus females were infected with P. falciparum gametocytes and midguts were dissected at day 7 post infection for detection/quantification of oocysts. Infection parameters were compared between individual with different L119F-GSTe2 genotypes, and the polymorphism of the GSTe2 gene was analyzed in infected and uninfected mosquito groups. Results: Overall, 403 mosquitoes were dissected and genotyped. The frequency of the L119F-GSTe2 resistance allele was significantly higher in non-infected (55.88%) compared to infected (40.99%) mosquitoes (Fisher's exact test, P<0.0001). Prevalence of infection was significantly higher in heterozygous and homozygous susceptible genotypes (P<0.001). However, homozygous resistant and heterozygous mosquitoes exhibited significantly higher infection intensity (P<0.01). No association was observed between the GSTe2 polymorphism and the infection status of mosquitoes. Conclusion: Altogether, these results suggest that GSTe2-based metabolic resistance may affect the vectorial competence of resistant An. funestus mosquitoes to P. falciparum infection, by increasing its permissiveness to Plasmodium infection
Identification and analysis of Single Nucleotide Polymorphisms (SNPs) in the mosquito Anopheles funestus, malaria vector
Get PDFBACKGROUND: Single nucleotide polymorphisms (SNPs) are the most common source of genetic variation in eukaryotic species and have become an important marker for genetic studies. The mosquito Anopheles funestus is one of the major malaria vectors in Africa and yet, prior to this study, no SNPs have been described for this species. Here we report a genome-wide set of SNP markers for use in genetic studies on this important human disease vector. RESULTS: DNA fragments from 50 genes were amplified and sequenced from 21 specimens of An. funestus. A third of specimens were field collected in Malawi, a third from a colony of Mozambican origin and a third form a colony of Angolan origin. A total of 494 SNPs including 303 within the coding regions of genes and 5 indels were identified. The physical positions of these SNPs in the genome are known. There were on average 7 SNPs per kilobase similar to that observed in An. gambiae and Drosophila melanogaster. Transitions outnumbered transversions, at a ratio of 2:1. The increased frequency of transition substitutions in coding regions is likely due to the structure of the genetic code and selective constraints. Synonymous sites within coding regions showed a higher polymorphism rate than non-coding introns or 3' and 5'flanking DNA with most of the substitutions in coding regions being observed at the 3(rd )codon position. A positive correlation in the level of polymorphism was observed between coding and non-coding regions within a gene. By genotyping a subset of 30 SNPs, we confirmed the validity of the SNPs identified during this study. CONCLUSION: This set of SNP markers represents a useful tool for genetic studies in An. funestus, and will be useful in identifying candidate genes that affect diverse ranges of phenotypes that impact on vector control, such as resistance insecticide, mosquito behavior and vector competence
Genome-Wide Transcription and Functional Analyses Reveal Heterogeneous Molecular Mechanisms Driving Pyrethroids Resistance in the Major Malaria Vector Anopheles funestus Across Africa.
Get PDFPyrethroid resistance in malaria vector, An. funestus is increasingly reported across Africa, threatening the sustainability of pyrethroid-based control interventions, including long lasting insecticidal nets (LLINs). Managing this problem requires understanding of the molecular basis of the resistance from different regions of the continent, to establish whether it is being driven by a single or independent selective events. Here, using a genome-wide transcription profiling of pyrethroid resistant populations from southern (Malawi), East (Uganda), and West Africa (Benin), we investigated the molecular basis of resistance, revealing strong differences between the different African regions. The duplicated cytochrome P450 genes (CYP6P9a and CYP6P9b) which were highly overexpressed in southern Africa are not the most upregulated in other regions, where other genes are more overexpressed, including GSTe2 in West (Benin) and CYP9K1 in East (Uganda). The lack of directional selection on both CYP6P9a and CYP6P9b in Uganda in contrast to southern Africa further supports the limited role of these genes outside southern Africa. However, other genes such as the P450 CYP9J11 are commonly overexpressed in all countries across Africa. Here, CYP9J11 is functionally characterized and shown to confer resistance to pyrethroids and moderate cross-resistance to carbamates (bendiocarb). The consistent overexpression of GSTe2 in Benin is coupled with a role of allelic variation at this gene as GAL4-UAS transgenic expression in Drosophila flies showed that the resistant 119F allele is highly efficient in conferring both DDT and permethrin resistance than the L119. The heterogeneity in the molecular basis of resistance and cross-resistance to insecticides in An. funestus populations throughout sub-Saharan African should be taken into account in designing resistance management strategies
Exposure to the insecticide-treated bednet PermaNet 2.0 reduces the longevity of the wild African malaria vector Anopheles funestus but GSTe2-resistant mosquitoes lives longer
Get PDFBackground: Despite the increased report of insecticide resistance in malaria vectors, its impact on mosquito’s life-traits after exposure to insecticide-treated nets remains under investigated. Here, we assessed the effects of exposure to PermaNet 2.0 on several life traits of An. gambiae s.l. and An. funestus s.l. field mosquitoes in Cameroon.
Methodology: Female Anopheles mosquitoes were collected indoor using electric aspirators in south-ern Cameroon (Obout) in 2016. After assessing the resistance status of F1 from the field collected-mosquitoes, progeny of the first generation (An. funestus s.l.) and seventh generation (An. gambiae s.l.) were used to assess the long-term effect of exposure to PermaNet 2.0 on several life-traits of these vectors (longevity, blood feeding ability, fecundity and fertility) in comparison to untreated net. In addition, the L119F-GSTe2 mutation associated with DDT/pyrethroids resistance in An. fu-nestus was genotyped to assess its association with increased life-span post-exposure.
Principal Findings: Both An. funestus and An. gambiae were resistant to pyrethroids and DDT with a greater level in the latter. Pyrethroid-only nets PermaNet 2.0 (17.5% mortality) and Olyset (0% mortality) exhibited a significantly reduced efficacy against An. funestus in contrast to a greater effi-cacy for PBO-based Nets Olyset Plus (65% mortality), PermaNet 3.0 top (100% mortality). In both species, mosquitoes that survived exposure to PermaNet 2.0 exhibited a significantly reduced longev-ity than those non-exposed (6.95 days vs 12.46 for An. funestus P<0.001; 8.87 vs 11.25 days for An. gambiae; P<0.001). However, no significant difference was observed for blood feeding and fecun-dity in both species. In addition, molecular analysis of the L119F-GSTe2 mutation revealed that this mutation is associated with an increase in the chance of surviving after exposure to this net in An. funestus.
Conclusions: These results show that although the PermaNet 2.0 presents a reduced efficacy against resistant populations, it remains efficient after exposure by reducing the life expectancy of the vec-tors which could contribute in the reduction of malaria incidence
Contrasting patterns of insecticide resistance and knockdown resistance (kdr) in the dengue vectors Aedes aegypti and Aedes albopictus from Malaysia
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Knowledge on the extent, distribution and mechanisms of insecticide resistance is essential for successful insecticide-based dengue control interventions. Here, we report an extensive resistance profiling of the dengue vectors Aedes aegypti and Aedes albopictus across Malaysia and establish the contribution of knockdown resistance mechanism revealing significant contrast between both species.
Methods
Aedes mosquitoes were collected from four states in Malaysia in 2010 using ovitraps and tested against six major insecticides using WHO bioassays. Knockdown resistance (kdr) was investigated in both species.
Results
A moderate resistance to temephos was detected from samples collected in 2010 in Penang, Kuala Lumpur, Johor Bharu and Kota Bharu (1.5 < RR < 3.3). A widespread and multiple resistances was observed in Ae. aegypti particularly against pyrethroids, DDT and bendiocarb. Mosquitoes from Kuala Lumpur consistently had the highest resistance levels and was the only population showing a moderate resistance to malathion (91% mortality). The resistance profile of Ae. albopictus contrasted to Ae. aegypti with full susceptibility to pyrethroids except in Kuala Lumpur where moderate resistance is observed. PBO synergist assays suggest metabolic resistance mechanisms play a major role in resistance in both species. Two kdr mutations, F1534C and V1016G, were detected in Ae. aegypti across Malaysia but neither of these mutations were found in Ae. albopictus. Additionally, signatures of selection were detected on the Voltage-gated sodium channel gene in Ae. aegypti but not in Ae. albopictus. The presence of the 1534C allele was significantly associated with pyrethroid resistance and an additive effect to pyrethroid resistance was observed in individuals containing both kdr alleles.
Conclusions
Findings from this study will help to design and implement successful insecticide-based interventions against Ae. aegypti and Ae. albopictus to improve dengue control across Malaysia
The P450 CYP6Z1 confers carbamate/pyrethroid cross-resistance in a major African malaria vector beside a novel carbamate-insensitive N485I acetylcholinesterase-1 mutation
Get PDFCarbamates are increasingly used for vector control notably in areas with pyrethroid resistance. However, a cross-resistance between these insecticides in major malaria vectors such as Anopheles funestus could severely limit available resistance management options. Unfortunately, the molecular basis of such cross-resistance remains uncharacterized in An. funestus, preventing effective resistance management.
Here, using a genome-wide transcription profiling, we revealed that metabolic resistance through up-regulation of cytochrome P450 genes is driving carbamate resistance. The P450s CYP6P9a, CYP6P9b and CYP6Z1 were the most up-regulated detoxification genes in the multiple resistant mosquitoes. However, in silico docking simulations predicted CYP6Z1 to metabolise both pyrethroids and carbamates, whereas CYP6P9a and CYP6P9b were predicted to metabolise only the pyrethroids. Using recombinant enzyme metabolism and inhibition assays we demonstrated that CYP6Z1 metabolizes bendiocarb and pyrethroids, whereas CYP6P9a and CYP6P9b metabolise only the pyrethroids. Other up-regulated gene families in resistant mosquitoes included several cuticular protein genes suggesting a possible reduced penetration resistance mechanism. Investigation of the target-site resistance in acetylcholinesterase 1 (ace-1) gene detected and established the association between the new N485I mutation and bendiocarb resistance (Odds ratio 7.3; P<0.0001). The detection of multiple haplotypes in single mosquitoes after cloning suggested the duplication of ace-1. A TaqMan genotyping of the N485I in nine countries revealed that the mutation is located only in Southern Africa with frequency of 10-15% suggesting its recent occurrence.
These findings will help in monitoring the spread and evolution of carbamate resistance and improve the design of effective resistance management strategies to control this malaria vector
Combined over-expression of two cytochrome P450 genes exacerbates the fitness cost of pyrethroid resistance in the major African malaria vector Anopheles funestus
Get PDFMetabolic resistance driven by multiple P450 genes is worsening insecticide resistance in malaria vectors. However, it remains unclear whether such multiple over-expression imposes an additive fitness cost in the vectors. Here, we showed that two highly over-expressed P450 genes (CYP6P9a and CYP6P9b) combine to impose additive fitness costs in pyrethroid-resistant Anopheles funestus. Genotyping of the CYP6P9b resistance allele in hybrid mosquitoes from a pyrethroid-resistant FUMOZ-R and the susceptible FANG strains revealed that this gene imposes a fitness cost in resistant mosquitoes similar to CYP6P9a. Homozygote susceptible CYP6P9b_S (SS) significantly lay more eggs than the resistant (OR = 2.2, P = 0.04) and with greater hatching rate (p < 0.04). Homozygote resistant larvae CYP6P9b_R (RR) developed significantly slower than homozygote susceptible from L1-L4 (χ2 = 7.2; P = 0.007) with a late pupation observed for RR compared to both heterozygotes and homozygotes susceptible (χ 2 = 11.17; P = 0.0008). No difference was observed between genotypes for adult longevity with no change in allele frequency and gene expression across the lifespan. Furthermore, we established that CYP6P9b combines with CYP6P9a to additively exacerbate the fitness cost of pyrethroid resistance with a greater reduction in fecundity/fertility and increased developmental time of double homozygote resistant mosquitoes. Moreover, an increased proportion of double homozygote susceptible individuals was noted over 10 generations in the insecticide-free environment (χ2 = 6.3; P = 0.01) suggesting a reversal to susceptibility in the absence of selection. Such greater fitness cost imposed by multiple P450 genes shows that resistance management strategy based on rotation could help slow the spread of resistance
Multiple insecticide resistance in the major malaria vector Anopheles funestus in southern Ghana: implications for malaria control
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Understanding the dynamics of insecticide resistance in African malaria vectors is crucial for successful implementation of resistance management strategies in the continent. This study reports a high and multiple insecticide resistance in Anopheles funestus from southern Ghana which could compromise the Malaria Operational Plan in this country, if not tackled. Adult Anopheles mosquitoes were collected in Obuasi and Adawukwa, in southern Ghana. Plasmodium infection rates, susceptibility to the main insecticides used in public health and the molecular basis of insecticide resistance were established.
Results
An. funestus (sensu stricto) (s.s.) was the predominant mosquito species found resting inside the houses in Obuasi, while at Adawukwa it was found together with An. coluzzii. Parasite rates were high in An. funestus (s.s.) populations from both localities, with Plasmodium infection rates greater than 12.5 %. Both, An. funestus (s.s.) and An. coluzzii, from the two sites exhibited high resistance to the insecticide from various classes including the pyrethroids, carbamates and DDT, but remained fully susceptible to the organophosphates. A preliminary characterization of the underlying molecular mechanisms of resistance in An. funestus (s.s.) populations from both sites revealed that CYP6P9a, CYP6P9b, CYP6M7 and GSTe2 genes are upregulated, markedly higher in Obuasi (between 3.35 and 1.83 times) than in Adawukwa population. The frequency of L119F-GSTe2 and A296S-RDL resistance markers were also higher in Obuasi (42.5 and 68.95 % higher), compared with An. funestus (s.s.) populations from Adawukwa. These findings suggest that the similar resistance pattern observed in both An. funestus (s.s.) populations are driven by different mechanisms.
Conclusions
Resistance to multiple insecticides in public health use is present in malaria vectors from Ghana with major resistance genes already operating in the field. This should be taken into consideration in the design of resistance management strategies to avoid operational failure
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