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Background: Understanding the mechanisms used by Anopheles mosquitoes to survive insecticide exposure is key to manage existing insecticide resistance and develop more suitable insecticide-based malaria vector control interventions as well as other alternative integrated tools. To this regard, the molecular basis of permethrin, DDT and dieldrin resistance in Anopheles funestus (sensu stricto) at Akaka-Remo was investigated., Methods: Bioassays were conducted on 3-5-day-old adult An. funestus (s.s.) mosquitoes for permethrin, DDT and dieldrin susceptibility test. The molecular mechanisms of mosquito resistance to these insecticides were investigated using microarray and reverse transcriptase PCR techniques. The voltage-gated sodium channel region of mosquitoes was also screened for the presence of knockdown resistance mutations (kdr west and east) by sequencing method., Results: Anopheles funestus (s.s.) population was resistant to permethrin (mortality rate of 68%), DDT (mortality rate of 10%) and dieldrin (mortality rate of 8%) insecticides. Microarray and RT-PCR analyses revealed the overexpression of glutathione S-transferase genes, cytochrome P450s, esterase, trypsin and cuticle proteins in resistant mosquitoes compared to control. The GSTe2 was the most upregulated detoxification gene in permethrin-resistant (FC = 44.89), DDT-resistant (FC = 57.39) and dieldrin-resistant (FC = 41.10) mosquitoes compared to control population (FC = 22.34). The cytochrome P450 gene, CYP6P9b was also upregulated in both permethrin- and DDT-resistant mosquitoes. The digestive enzyme, trypsin (hydrolytic processes) and the cuticle proteins (inducing cuticle thickening leading to reduced insecticides penetration) also showed high involvement in insecticide resistance, through their overexpression in resistant mosquitoes compared to control. The kdr east and west were absent in all mosquitoes analysed, suggesting their non-involvement in the observed mosquito resistance., Conclusions: The upregulation of metabolic genes, especially the GSTe2 and trypsin, as well as the cuticle proteins is driving insecticide resistance of An. funestus (s.s.) population. However, additional molecular analyses, including functional metabolic assays of these genes as well as screening for a possible higher cuticular hydrocarbon and lipid contents, and increased procuticle thickness in resistant mosquitoes are needed to further describe their distinct roles in mosquito resistance.

Background: Malaria is endemic in sub-Saharan Africa with considerable burden for human health. Major insecticide resistance mechanisms such as kdr(R) and ace-1(R) alleles constitute a hindrance to malaria vector control programs. Anopheles gambiae bearing both kdr and ace-1 resistant alleles are increasingly recorded in wild populations. In order to maintain the efficacy of vector control strategies, the characterization of concomitant kdr and ace-1 resistance, and their pleiotropic effects on malaria vector phenotype on insecticide efficacy are important., Methods: Larval and adult bioassays were performed with different insecticide classes used in public health following WHO standard guidelines on four laboratory Anopheles gambiae strains, sharing the same genetic background but harboring distinct resistance status: KISUMU with no resistance allele; ACERKIS with ace-1(R) allele; KISKDR with kdr(R) allele and ACERKDRKIS with both resistance alleles' ace-1(R) and kdr(R) ., Results: Larval bioassays indicate that the homozygote resistant strain harboring both alleles (ACERKDRKIS) displayed slightly but significantly higher resistance level to various insecticides like carbamates (bendiocarb, p < 0.001; propoxur, p = 0.02) and organophosphates (chlorpyriphos-methyl, p = 0.002; fenitrothion, p < 0.001) when compared to ACERKIS strain. However, no differences were recorded between ACERKDRKIS and KISKDR resistance level against permethrin (Pyrethroid, p = 0.7) and DDT (Organochlorine, p = 0.24). For adult bioassays, the percentages of mosquitoes knocked down were significantly lower for ACERKDRKIS than for KISKDR with permethrin (p = 0.003) but not with deltamethrin. The percentage of mortality from adult bioassays was similar between ACERKDRKIS and ACERKIS with carbamates and organophosphates, or between ACERKDRKIS and KISKDR with pyrethroid and DDT. Concerning acetylcholinesterase enzyme, ACERKDRKIS strain showed similarAChE1 activity than that of ACERKIS., Conclusion: The presence of both kdr(R) and ace-1(R) alleles seems to increase the resistance levels to both carbamate and organophosphate insecticides and at operational level, may represent an important threat to malaria vector control programs in West Africa.

Background: The Culex quinquefasciatus mosquito, a major pest and vector of filariasis and arboviruses in the tropics, has developed multiple resistance mechanisms to the main insecticide classes currently available in public health. Among them, the insensitive acetylcholinesterase (ace-1(R) allele) is widespread worldwide and confers cross-resistance to organophosphates and carbamates. Fortunately, in an insecticide-free environment, this mutation is associated with a severe genetic cost that can affect various life history traits. Salivary proteins are directly involved in human-vector contact during biting and therefore play a key role in pathogen transmission., Methods and Results: An original proteomic approach combining 2D-electrophoresis and mass spectrometry was adopted to compare the salivary expression profiles of two strains of C. quinquefasciatus with the same genetic background but carrying either the ace-1(R) resistance allele or not (wild type). Four salivary proteins were differentially expressed (>2 fold, P<0.05) in susceptible (SLAB) and resistant (SR) mosquito strains. Protein identification indicated that the D7 long form, a major salivary protein involved in blood feeding success, presented lower expression in the resistant strain than the susceptible strain. In contrast, three other proteins, including metabolic enzymes (endoplasmin, triosephosphate isomerase) were significantly over-expressed in the salivary gland of ace-1(R) resistant mosquitoes. A catalogue of 67 salivary proteins of C. quinquefasciatus sialotranscriptome was also identified and described., Conclusion: The "resistance"-dependent expression of salivary proteins in mosquitoes may have considerable impact on biting behaviour and hence on the capacity to transmit parasites/viruses to humans. The behaviour of susceptible and insecticide-resistant mosquitoes in the presence of vertebrate hosts and its impact on pathogen transmission urgently requires further investigation., Data Deposition: All proteomic data will be deposited at PRIDE (http://www.ebi.ac.uk/pride/).