Insecticide Resistance In Malaria Vectors Research Articles

Evidence of a multiple insecticide resistance in the malaria vector Anopheles funestus in South West Nigeria.

BackgroundKnowing the extent and spread of insecticide resistance in malaria vectors is vital to successfully manage insecticide resistance in Africa. This information in the main malaria vector, Anopheles funestus sensu stricto, is completely lacking in the most populous country in Africa, Nigeria. This study reports the insecticide susceptibility status and the molecular basis of resistance of An. funestus as well as its involvement in malaria transmission in Akaka-Remo, a farm settlement village in southwest Nigeria.Results Plasmodium infection analysis using TaqMan protocol coupled with a nested PCR revealed an infection rate of 8% in An. funestus s.s. from Akaka-Remo. WHO susceptibility tests showed this species has developed multiple resistance to insecticides in the study area. Anopheles funestus s.s. population in Akaka-Remo is highly resistant to organochlorines: dieldrin (8%) and DDT (10%). Resistance was also observed against pyrethroids: permethrin (68%) and deltamethrin (87%), and the carbamate bendiocarb (84%). Mortality rate with DDT slightly increased (from 10 to 30%, n = 45) after PBO pre-exposure indicating that cytochrome P450s play little role in DDT resistance while high mortalities were recorded after PBO pre-exposure with permethrin (from 68 to 100%, n = 70) and dieldrin (from 8 to 100%, n = 48) suggesting the implication of P450s in the observed permethrin and dieldrin resistance. High frequencies of resistant allele, 119F in F0 (77%) and F1 (80% in resistant and 72% in susceptible) populations with an odd ratio of 1.56 (P = 0.1859) show that L119F-GSTe2 mutation is almost fixed in the population. Genotyping of the A296S-RDL mutation in both F0 and F1 samples shows an association with dieldrin resistance with an odd ratio of 81 (P < 0.0001) (allelic frequency (R) = 76% for F0; for F1, 90 and 10% were observed in resistant and susceptible populations, respectively) as this mutation is not yet fixed in the population.ConclusionThe study reports multiple insecticide resistance in An. funestus from Akaka Remo. It is, therefore, necessary to pay more attention to this major malaria vector for effective malaria control in Nigeria.

Chlorfenapyr (A Pyrrole Insecticide) Applied Alone or as a Mixture with Alpha-Cypermethrin for Indoor Residual Spraying against Pyrethroid Resistant Anopheles gambiae sl: An Experimental Hut Study in Cove, Benin

BackgroundIndoor spraying of walls and ceilings with residual insecticide remains a primary method of malaria control. Insecticide resistance in malaria vectors is a growing problem. Novel insecticides for indoor residual spraying (IRS) which can improve the control of pyrethroid resistant malaria vectors are urgently needed. Insecticide mixtures have the potential to improve efficacy or even to manage resistance in some situations but this possibility remains underexplored experimentally. Chlorfenapyr is a novel pyrrole insecticide which has shown potential to improve the control of mosquitoes which are resistant to current WHO-approved insecticides.MethodThe efficacy of IRS with chlorfenapyr applied alone or as a mixture with alpha-cypermeththrin (a pyrethroid) was evaluated in experimental huts in Cove, Southern Benin against wild free flying pyrethroid resistant Anopheles gambiae sl. Comparison was made with IRS with alpha-cypermethrin alone. Fortnightly 30-minute in situ cone bioassays were performed to assess the residual efficacy of the insecticides on the treated hut walls.ResultsSurvival rates of wild An gambiae from the Cove hut site in WHO resistance bioassays performed during the trial were >90% with permethrin and deltamethrin treated papers. Mortality of free-flying mosquitoes entering the experimental huts was 4% in the control hut. Mortality with alpha-cypermethrin IRS did not differ from the control (5%, P>0.656). The highest mortality was achieved with chlorfenapyr alone (63%). The alpha-cypermethrin + chlorfenapyr mixture killed fewer mosquitoes than chlorfenapyr alone (43% vs. 63%, P<0.001). While the cone bioassays showed a more rapid decline in residual mortality with chlorfenapyr IRS to <30% after only 2 weeks, fortnightly mortality rates of wild free-flying An gambiae entering the chlorfenapyr IRS huts were consistently high (50–70%) and prolonged, lasting over 4 months.ConclusionIRS with chlorfenapyr shows potential to significantly improve the control of malaria transmission in pyrethroid resistant areas compared to pyrethroid IRS or the mixture. Thirty minute in situ cone bioassays are not predictive of the performance of chlorfenapyr IRS under field conditions.

The impact of long-lasting microbial larvicides in reducing malaria transmission and clinical malaria incidence: study protocol for a cluster randomized controlled trial.

BackgroundThe massive scale-up of insecticide-treated nets (ITNs) and indoor residual spraying (IRS) has led to a substantial increase in malaria vector insecticide resistance as well as in increased outdoor transmission, both of which hamper the effectiveness and efficiency of ITN and IRS. Long-lasting microbial larvicide can be a cost-effective new supplemental intervention tool for malaria control.Methods/designWe will implement the long-lasting microbial larvicide intervention in 28 clusters in two counties in western Kenya. We will test FourStar controlled release larvicide (6 % by weight Bacillus thuringiensis israelensis and 1 % Bacillus sphaerius) by applying FourStar controlled release granule formulation, 90-day briquettes, and 180-day briquettes in different habitat types. The primary endpoint is clinical malaria incidence rate and the secondary endpoint is malaria vector abundance and transmission intensity. The intervention will be conducted as a two-step approach. First, we will conduct a four-cluster trial (two clusters per county, with one of the two clusters randomly assigned to the intervention arm) to optimize the larvicide application scheme. Second, we will conduct an open-label, cluster-randomized trial to evaluate the effectiveness and cost-effectiveness of the larvicide. Fourteen clusters in each county will be assigned to intervention (treatment) or no intervention (control) by a block randomization on the basis of clinical malaria incidence, vector density, and human population size per site. We will treat each treatment cluster with larvicide for three rounds at 4-month intervals, followed by no treatment for the following 8 months. Next, we will switch the control and treatment sites. The former control sites will receive three rounds of larvicide treatment at appropriate time intervals, and former treatment sites will receive no larvicide. We will monitor indoor and outdoor vector abundance using CO2-baited CDC light traps equipped with collection bottle rotators. Clinical malaria data will be aggregated from government-run malaria treatment centers.DiscussionSince current first-line vector intervention methods do not target outdoor transmission and will select for higher insecticide resistance, new methods beyond bed nets and IRS should be considered. Long-lasting microbial larviciding represents a promising new tool that can target both indoor and outdoor transmission and alleviate the problem of pyrethroid resistance. It also has the potential to diminish costs by reducing larvicide reapplications. If successful, it could revolutionize malaria vector control in Africa, just as long-lasting bed nets have done.Trial registrationU.S. National Institute of Health, study ID NCT02392832. Registered on 3 February 2015.

Influence of the agrochemicals used for rice and vegetable cultivation on insecticide resistance in malaria vectors in southern Côte d'Ivoire.

BackgroundVector control can contribute to the development of resistance to insecticides in malaria vectors. As the swamps and wetlands used for some agricultural activities constitute productive breeding sites for many mosquito species, agricultural pest control may increase the selection pressure for insecticide resistance in mosquitoes. Understanding the use of agrochemicals by farmers is important to plan and initiate effective integrated pest and vector management interventions.MethodsA knowledge–attitude–practice study, using questionnaires, was undertaken with 102 rice farmers in Tiassalé and 106 vegetable farmers in Dabou (South Côte d’Ivoire) in order to generate information on pesticide usage. In addition, insecticide susceptibility bioassays were conducted using adult mosquitoes obtained from larvae collected within farms, and the persistence of agricultural pesticides in the farming environment, including sediment and mosquito breeding site water, was investigated by HPLC.ResultsHerbicides and insecticides appeared to be the most frequently used pesticides for both crops. Amino phosphonates (mostly glyphosate) represented the most used herbicides (45 % for rice up to 89 % for vegetables). Pyrethroids appeared to be the most used insecticides (accounting for 90 % of all the insecticide use reported). Approximately 75 % of respondents had not been to school and do not understand product labels. Only about 45 % of farmers respect the recommended pesticide dosage and about 10–15 % of pesticides used for rice and vegetable, respectively, are not recommended for these crops. As per WHO criteria, the mosquito local populations from the two localities were resistant to three of the four insecticides tested, as mortalities were less than 35 % for deltamethrin, DDT and bendiocarb. Higher susceptibility was observed for malathion, although the population was considered resistant in Dabou (80 % mortality) and susceptible in Tiassalé (98 % mortality). With the exception of glyphosate, residues from each of six chemicals tested for were detected in each of the sites visited in the two localities.ConclusionThe study describes the use of insecticides and herbicides on crops and highlights the importance of considering agriculture practices when attempting to manage resistance in malaria vectors. Inter-sectoral collaboration between agriculture and public health is required to develop efficient integrated pest and vector management interventions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-016-1481-5) contains supplementary material, which is available to authorized users.

Insecticide susceptibility of natural populations of Anopheles coluzzii and Anopheles gambiae (sensu stricto) from Okyereko irrigation site, Ghana, West Africa.

BackgroundThe increasing spread of insecticide resistance in malaria vectors has been well documented across sub-Saharan Africa countries. The influence of irrigation on increasing vector resistance is poorly understood, and is critical to successful and ethical implementation of food security policies. This study investigated the insecticide resistance status of An. gambiae (s.l.) mosquitoes collected from the irrigated rice area of Okyereko, a village containing about 42 hectares of irrigated field within an irrigation project plan in the Central Region of Ghana. Large amounts of insecticides, herbicides and fertilizers are commonly used in the area to boost the annual production of the rice.MethodsMosquito larvae were collected and adults were assayed from the F1 progeny. The resistance status, allele and genotype were characterized using WHO susceptibility testing and PCR methods respectively.ResultsThe An. gambiae (s.l.) populations from Okyereko are highly resistant to DDT and pyrethroid insecticides, with possible involvement of metabolic mechanisms including the elevation of P450 and GST enzyme as well as P-gp activity. The population was mostly composed of An. coluzzii specimens (more than 96 %) with kdr and ace-1 frequencies of 0.9 and 0.2 %, respectively.ConclusionThis study brings additional information on insecticide resistance and the characterization of An. gambiae (s.l.) mosquitoes from Okyereko, which can be helpful in decision making for vector control programmes in the region.

The contribution of agricultural insecticide use to increasing insecticide resistance in African malaria vectors.

The fight against malaria is increasingly threatened by failures in vector control due to growing insecticide resistance. This review examines the recent primary research that addresses the putative relationship between agricultural insecticide use and trends in insecticide resistance. To do so, descriptive evidence offered by the new research was categorized, and additional factors that impact the relationship between agricultural insecticide use and observed insecticide resistance in malaria vectors were identified. In 23 of the 25 relevant recent publications from across Africa, higher resistance in mosquito populations was associated with agricultural insecticide use. This association appears to be affected by crop type, farm pest management strategy and urban development.

Status of insecticide resistance in high-risk malaria provinces in Afghanistan.

BackgroundInsecticide resistance seriously threatens the efficacy of vector control interventions in malaria endemic countries. In Afghanistan, the status of insecticide resistance is largely unknown while distribution of long-lasting insecticidal nets has intensified in recent years. The main objective of this study was thus to measure the level of resistance to four classes of insecticides in provinces with medium to high risk of malaria transmission.MethodsAdult female mosquitoes were reared from larvae successively collected in the provinces of Nangarhar, Kunar, Badakhshan, Ghazni and Laghman from August to October 2014. WHO insecticide susceptibility tests were performed with DDT (4 %), malathion (5 %), bendiocarb (0.1 %), permethrin (0.75 %) and deltamethrin (0.05 %). In addition, the presence of kdr mutations was investigated in deltamethrin resistant and susceptible Anopheles stephensi mosquitoes collected in the eastern provinces of Nangarhar and Kunar.ResultsAnalyses of mortality rates revealed emerging resistance against all four classes of insecticides in the provinces located east and south of the Hindu Kush mountain range. Resistance is observed in both An. stephensi and Anopheles culicifacies, the two dominant malaria vectors in these provinces. Anopheles superpictus in the northern province of Badakhshan shows a different pattern of susceptibility with suspected resistance observed only for deltamethrin and bendiocarb. Genotype analysis of knock down resistance (kdr) mutations at the voltage-gated channel gene from An. stephensi mosquitoes shows the presence of the known resistant alleles L1014S and L1014F. However, a significant fraction of deltamethrin-resistant mosquitoes were homozygous for the 1014L wild type allele indicating that other mechanisms must be considered to account for the observed pyrethroid resistance.ConclusionsThis study confirms the importance of monitoring insecticide resistance for the development of an integrated vector management in Afghanistan. The validation of the kdr genotyping PCR assay applied to An. stephensi collected in Afghanistan paves the way for further studies into the mechanisms of insecticide resistance of malaria vectors in this region.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-016-1149-1) contains supplementary material, which is available to authorized users.

Patterns of insecticide resistance and knock down resistance (kdr) in malaria vectors An. arabiensis, An. coluzzii and An. gambiae from sympatric areas in Senegal

BackgroundMalaria vector control in Africa relies on insecticides targeting adult mosquito vectors via insecticide treated nets or indoor residual spraying. Despite the proven efficacy of these strategies, the emergence and rapid rise in insecticide resistance in malaria vectors raises many concerns about their sustainability. Therefore, the monitoring of insecticide resistance is essential for resistance management strategies implementation. We investigated the kdr mutation frequencies in 20 sympatric sites of An. arabiensis Patton, An. coluzzii Coetzee & Wilkerson and An. gambiae Giles and its importance in malaria vector control by evaluating the susceptibility to insecticides in four representative sites in Senegal.MethodsSibling species identification and kdr mutation detection were determined using polymerase chain reaction on mosquitoes collected using pyrethrum sprays collection in 20 sites belonging to two transects with differential insecticide selection pressure. The World Health Organization (WHO) tube test was used to determine phenotypic resistance of An. gambiae s.l. to DDT, deltamethrin, lambdacyholothrin, permethrin, bendiocarb and malathion in four representative sites.ResultsThe L1014F kdr mutation was widely distributed and was predominant in An. gambiae in comparison to An. arabiensis and An. coluzzii. The bioassay tests showed a general trend with a resistance to DDT and pyrethroids and a susceptibility to organophosphate and carbamate according to WHO thresholds. For deltamethrin and permethrin, the two most used insecticides, no significant difference were observed either between the two transects or between mortality rates suggesting no differential selection pressures on malaria vectors. The study of the KD times showed similar trends as comparable levels of resistance were observed, the effect being more pronounced for permethrin.ConclusionsOur study showed a widespread resistance of malaria vectors to DDT and pyrethroids and a widespread distribution of the 1014F kdr allele. These combined observations could suggest the involvement of the kdr mutation. The existence of other resistance mechanisms could not be ruled out as a proportion of mosquitoes did not harbour the kdr allele whereas the populations were fully resistant. The susceptibility to carbamate and organophosphate could be exploited as alternative for insecticide resistance management.

Consolidating strategic planning and operational frameworks for integrated vector management in Eritrea.

BackgroundContemporary malaria vector control relies on the use of insecticide-based, indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs). However, malaria-endemic countries, including Eritrea, have struggled to effectively deploy these tools due technical and operational challenges, including the selection of insecticide resistance in malaria vectors. This manuscript outlines the processes undertaken in consolidating strategic planning and operational frameworks for vector control to expedite malaria elimination in Eritrea.Case descriptionThe effort to strengthen strategic frameworks for vector control in Eritrea was the ‘case’ for this study. The integrated vector management (IVM) strategy was developed in 2010 but was not well executed, resulting in a rise in malaria transmission, prompting a process to redefine and relaunch the IVM strategy with integration of other vector borne diseases (VBDs) as the focus. The information sources for this study included all available data and accessible archived documentary records on malaria vector control in Eritrea. Structured literature searches of published, peer-reviewed sources using online, scientific, bibliographic databases, Google Scholar, PubMed and WHO, and a combination of search terms were utilized to gather data. The literature was reviewed and adapted to the local context and translated into the consolidated strategic framework.DiscussionIn Eritrea, communities are grappling with the challenge of VBDs posing public health concerns, including malaria. The global fund financed the scale-up of IRS and LLIN programmes in 2014. Eritrea is transitioning towards malaria elimination and strategic frameworks for vector control have been consolidated by: developing an integrated vector management (IVM) strategy (2015–2019); updating IRS and larval source management (LSM) guidelines; developing training manuals for IRS and LSM; training of national staff in malaria entomology and vector control, including insecticide resistance monitoring techniques; initiating the global plan for insecticide resistance management; conducting needs’ assessments and developing standard operating procedure for insectaries; developing a guidance document on malaria vector control based on eco-epidemiological strata, a vector surveillance plan and harmonized mapping, data collection and reporting tools.ConclusionEritrea has successfully consolidated strategic frameworks for vector control. Rational decision-making remains critical to ensure that the interventions are effective and their choice is evidence-based, and to optimize the use of resources for vector control. Implementation of effective IVM requires proper collaboration and coordination, consistent technical and financial capacity and support to offer greater benefits.

Rise of multiple insecticide resistance in Anopheles funestus in Malawi: a major concern for malaria vector control.

BackgroundDeciphering the dynamics and evolution of insecticide resistance in malaria vectors is crucial for successful vector control. This study reports an increase of resistance intensity and a rise of multiple insecticide resistance in Anopheles funestus in Malawi leading to reduced bed net efficacy.MethodsAnopheles funestus group mosquitoes were collected in southern Malawi and the species composition, Plasmodium infection rate, susceptibility to insecticides and molecular bases of the resistance were analysed.ResultsMosquito collection revealed a predominance of An. funestus group mosquitoes with a high hybrid rate (12.2 %) suggesting extensive species hybridization. An. funestus sensu stricto was the main Plasmodium vector (4.8 % infection). Consistently high levels of resistance to pyrethroid and carbamate insecticides were recorded and had increased between 2009 and 2014. Furthermore, the 2014 collection exhibited multiple insecticide resistance, notably to DDT, contrary to 2009. Increased pyrethroid resistance correlates with reduced efficacy of bed nets (<5 % mortality by Olyset® net), which can compromise control efforts. This change in resistance dynamics is mirrored by prevalent resistance mechanisms, firstly with increased over-expression of key pyrethroid resistance genes (CYP6Pa/b and CYP6M7) in 2014 and secondly, detection of the A296S-RDL dieldrin resistance mutation for the first time. However, the L119F-GSTe2 and kdr mutations were absent.ConclusionsSuch increased resistance levels and rise of multiple resistance highlight the need to rapidly implement resistance management strategies to preserve the effectiveness of existing insecticide-based control interventions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-015-0877-y) contains supplementary material, which is available to authorized users.

When a discriminating dose assay is not enough: measuring the intensity of insecticide resistance in malaria vectors.

BackgroundGuidelines from the World Health Organization for monitoring insecticide resistance in disease vectors recommend exposing insects to a predetermined discriminating dose of insecticide and recording the percentage mortality in the population. This standardized methodology has been widely adopted for malaria vectors and has provided valuable data on the spread and prevalence of resistance. However, understanding the potential impact of this resistance on malaria control requires a more quantitative measure of the strength or intensity of this resistance.MethodsBioassays were adapted to quantify the level of resistance to permethrin in laboratory colonies and field populations of Anopheles gambiae sensu lato. WHO susceptibility tube assays were used to produce data on mortality versus exposure time and CDC bottle bioassays were used to generate dose response data sets. A modified version of the CDC bottle bioassay, known as the Resistance Intensity Rapid Diagnostic Test (I-RDT), was also used to measure the knockdown and mortality after exposure to different multipliers of the diagnostic dose. Finally cone bioassays were used to assess mortality after exposure to insecticide treated nets.ResultsThe time response assays were simple to perform but not suitable for highly resistant populations. After initial problems with stability of insecticide and bottle washing were resolved, the CDC bottle bioassay provided a reproducible, quantitative measure of resistance but there were challenges performing this under field conditions. The I-RDT was simple to perform and interpret although the end point selected (immediate knockdown versus 24 h mortality) could dramatically affect the interpretation of the data. The utility of the cone bioassays was dependent on net type and thus appropriate controls are needed to interpret the operational significance of these data sets.ConclusionsIncorporating quantitative measures of resistance strength, and utilizing bioassays with field doses of insecticides, will help interpret the possible impact of resistance on vector control activities. Each method tested had different benefits and challenges and agreement on a common methodology would be beneficial so that data are generated in a standardized format. This type of quantitative data are an important prerequisite to linking resistance strength to epidemiological outcomes.

Setting international standards for the management of public health pesticides.

Recent developments have highlighted the urgency of sound management of public health pesticides in vector-borne–disease–endemic countries. Major shortcomings are evident in national-level management practices throughout the pesticide life cycle from production to disposal; these shortcomings will adversely affect the cost-effectiveness and increase the risks of pesticides used. A major thrust has occurred towards developing international standards for improvement of public health pesticide management and towards expanding WHO’s global network on pesticide evaluation. However, to face current and future challenges, such as insecticide resistance in malaria vectors, the global capacity for evaluation of new insecticide products and vector-control tools should be further enhanced. Another area requiring urgent attention is the actual adoption and implementation of the recommended standards, calling for support to strengthen policy, legislation, and capacity.

Scale-up of integrated malaria vector control: lessons from Malawi.

ProblemIndoor residual spraying and long-lasting insecticidal nets (LLINs) are key tools for malaria vector control. Malawi has struggled to scale up indoor residual spraying and to improve LLIN coverage and usage.ApproachIn 2002, the Malawian National Malaria Control Programme developed guidelines for insecticide treated net distribution to reach the strategic target of at least 60% coverage of households with an LLIN. By 2005, the target coverage was 80% of households and the Global Fund financed the scale-up. The US President’s Malaria Initiative funded the indoor residual spraying intervention.Local settingMalawi’s entire population is considered to be at risk of malaria. Poor vector control, insecticide resistance in malaria vectors and insufficient technical and financial support have exacerbated the malaria burden.Relevant changesBetween 2002 and 2012, 18 248 206 LLINs had been distributed. The coverage of at least one LLIN per household increased from 27% (3689/13 664) to 58% (1974/3404). Indoor residual spraying coverage increased from 28 227 to 653 592 structures between 2007 and 2011. However, vector resistance prompted a switch from pyrethroids to organophosphates for indoor residual spraying, which increased the cost and operations needed to be cut back from seven to one district. Malaria cases increased from 2 853 315 in 2002 to 6 748 535 in 2010, and thereafter dropped to 4 922 596 in 2012.Lessons learntA single intervention-based approach for vector control may have suboptimal impact. Well-coordinated integrated vector management may offer greater benefits. A resistance management plan is essential for effective and sustainable vector control.

A genetic model of the effects of insecticide-treated bed nets on the evolution of insecticide-resistance.

The evolution of insecticide-resistance in malaria vectors is emerging as a serious challenge for the control of malaria. Modelling the spread of insecticide-resistance is an essential tool to understand the evolutionary pressures and dynamics caused by the application of insecticides. We developed a population-genetic model of the spread of insecticide-resistance in a population of Anopheles vectors in response to insecticides used either as adulticides (focussing on insecticide-treated bed nets (ITNs)) or as larvicides (either for the control of malaria or, as an inadvertent side-product, in agriculture). We show that indoor use of insecticides leads to considerably less selection pressure than their use as larvicides, supporting the idea that most resistance of malaria vectors is due to the agricultural use of the insecticides that are also used for malaria control. The reasons for the relatively low selection pressure posed by adulticides are (i) that males are not affected by the ITNs and, in particular, (ii) that the insecticides are also repellents, keeping mosquitoes at bay from contacting the insecticide but also driving them to bite either people who do not use the insecticide or alternative hosts. We conclude by discussing the opposing public health benefits of high repellency at an epidemiological and an evolutionary timescale: whereas repellency is beneficial to delay the evolution of resistance, other models have shown that it decreases the population-level protection of the insecticide.

Impact of agriculture on the selection of insecticide resistance in the malaria vector Anopheles gambiae: a multigenerational study in controlled conditions.

BackgroundResistance of mosquitoes to insecticides is mainly attributed to their adaptation to vector control interventions. Although pesticides used in agriculture have been frequently mentioned as an additional force driving the selection of resistance, only a few studies were dedicated to validate this hypothesis and characterise the underlying mechanisms. While insecticide resistance is rising dramatically in Africa, deciphering how agriculture affects resistance is crucial for improving resistance management strategies. In this context, the multigenerational effect of agricultural pollutants on the selection of insecticide resistance was examined in Anopheles gambiae.MethodsAn urban Tanzanian An. gambiae population displaying a low resistance level was used as a parental strain for a selection experiment across 20 generations. At each generation larvae were selected with a mixture containing pesticides and herbicides classically used in agriculture in Africa. The resistance levels of adults to deltamethrin, DDT and bendiocarb were compared between the selected and non-selected strains across the selection process together with the frequency of kdr mutations. A microarray approach was used for pinpointing transcription level variations selected by the agricultural pesticide mixture at the adult stage.ResultsA gradual increase of adult resistance to all insecticides was observed across the selection process. The frequency of the L1014S kdr mutation rose from 1.6% to 12.5% after 20 generations of selection. Microarray analysis identified 90 transcripts over-transcribed in the selected strain as compared to the parental and the non-selected strains. Genes encoding cuticle proteins, detoxification enzymes, proteins linked to neurotransmitter activity and transcription regulators were mainly affected. RT-qPCR transcription profiling of candidate genes across multiple generations supported their link with insecticide resistance.ConclusionsThis study confirms the potency of agriculture in selecting for insecticide resistance in malaria vectors. We demonstrated that the recurrent exposure of larvae to agricultural pollutants can select for resistance mechanisms to vector control insecticides at the adult stage. Our data suggest that in addition to selected target-site resistance mutations, agricultural pollutants may also favor cuticle, metabolic and synaptic transmission-based resistance mechanisms. These results emphasize the need for integrated resistance management strategies taking into account agriculture activities.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-014-0480-z) contains supplementary material, which is available to authorized users.

Widespread pyrethroid and DDT resistance in the major malaria vector Anopheles funestus in East Africa is driven by metabolic resistance mechanisms.

BackgroundEstablishing the extent, geographical distribution and mechanisms of insecticide resistance in malaria vectors is a prerequisite for resistance management. Here, we report a widespread distribution of insecticide resistance in the major malaria vector An. funestus across Uganda and western Kenya under the control of metabolic resistance mechanisms.Methodology/Principal FindingsFemale An. funestus collected throughout Uganda and western Kenya exhibited a Plasmodium infection rate between 4.2 to 10.4%. Widespread resistance against both type I (permethrin) and II (deltamethrin) pyrethroids and DDT was observed across Uganda and western Kenya. All populations remain highly susceptible to carbamate, organophosphate and dieldrin insecticides. Knockdown resistance plays no role in the pyrethroid and DDT resistance as no kdr mutation associated with resistance was detected despite the presence of a F1021C replacement. Additionally, no signature of selection was observed on the sodium channel gene. Synergist assays and qRT-PCR indicated that metabolic resistance plays a major role notably through elevated expression of cytochrome P450s. DDT resistance mechanisms differ from West Africa as the L119F-GSTe2 mutation only explains a small proportion of the genetic variance to DDT resistance.ConclusionThe extensive distribution of pyrethroid and DDT resistance in East African An. funestus populations represents a challenge to the control of this vector. However, the observed carbamate and organophosphate susceptibility offers alternative solutions for resistance management.

Increased pyrethroid resistance in malaria vectors and decreased bed net effectiveness, Burkina Faso.

Malaria control is dependent on insecticides. Increases in prevalence of insecticide resistance in malaria vectors across Africa are well-documented. However, few attempts have been made to quantify the strength of this resistance and link it to the effectiveness of control tools. Using quantitative bioassays, we show that in Burkina Faso pyrethroid resistance in Anopheles gambiae mosquitoes has increased in intensity in recent years and now exceeds 1,000-fold. In laboratory assays, this level of resistance renders insecticides used to impregnate bed nets ineffective. Thus, the level of personal and community protection afforded by long-lasting insecticide-treated net campaigns will probably be reduced. Standardized methods are needed to quantify resistance levels in malaria vectors and link these levels to failure of vector control methods.

Molecular population genetics of the NADPH cytochrome P450 reductase (CPR) gene in Anopheles minimus.

Development of insecticide resistance (IR) in mosquito vectors is a primary huddle to malaria control program. Since IR has genetic basis, and genes constantly evolve with response to environment for adaptation to organisms, it is important to know evolutionary pattern of genes conferring IR in malaria vectors. The mosquito Anopheles minimus is a major malaria vector of the Southeast (SE) Asia and India and is susceptible to all insecticides, and thus of interest to know if natural selection has shaped variations in the gene conferring IR. If not, the DNA fragment of such a genecould be used to infer population structure and demography of this species of malaria vector. We have therefore sequenced a ~569bp DNA segment of the NADPH cytochrome P450 reductase (CPR) gene (widely known to confer IR)in 123 individuals of An. minimus collected in 10 different locations (eight Indian, one Thai and one Vietnamese). Two Indian population samples were completely mono-morphic in the CPR gene. In general, low genetic diversity was found with no evidence of natural selection in this gene. The data were therefore analyzed to infer population structure and demography of this species. The10 populations could be genetically differentiated into four different groups; the samples from Thailand and Vietnam contained high nucleotide diversity. All the 10 populations conform to demographic equilibrium model with signature of past population expansion in four populations. The results in general indicate that the An. minimus mosquitoes sampled in the two SE Asian localities contain several genetic characteristics of being parts of the ancestral population.

An online tool for mapping insecticide resistance in major Anopheles vectors of human malaria parasites and review of resistance status for the Afrotropical region.

BackgroundMalaria control programmes across Africa and beyond are facing increasing insecticide resistance in the major anopheline vectors. In order to preserve or prolong the effectiveness of the main malaria vector interventions, up-to-date and easily accessible insecticide resistance data that are interpretable at operationally-relevant scales are critical. Herein we introduce and demonstrate the usefulness of an online mapping tool, IR Mapper.MethodsA systematic search of published, peer-reviewed literature was performed and Anopheles insecticide susceptibility and resistance mechanisms data were extracted and added to a database after a two-level verification process. IR Mapper ( http://www.irmapper.com) was developed using the ArcGIS for JavaScript Application Programming Interface and ArcGIS Online platform for exploration and projection of these data.ResultsLiterature searches yielded a total of 4,084 susceptibility data points for 1,505 populations, and 2,097 resistance mechanisms data points for 1,000 populations of Anopheles spp. tested via recommended WHO methods from 54 countries between 1954 and 2012. For the Afrotropical region, data were most abundant for populations of An. gambiae, and pyrethroids and DDT were more often used in susceptibility assays (51.1 and 26.8% of all reports, respectively) than carbamates and organophosphates. Between 2001 and 2012, there was a clear increase in prevalence and distribution of confirmed resistance of An. gambiae s.l. to pyrethroids (from 41 to 87% of the mosquito populations tested) and DDT (from 64 to 91%) throughout the Afrotropical region. Metabolic resistance mechanisms were detected in western and eastern African populations and the two kdr mutations (L1014S and L1014F) were widespread. For An. funestus s.l., relatively few populations were tested, although in 2010–2012 resistance was reported in 50% of 10 populations tested. Maps are provided to illustrate the use of IR Mapper and the distribution of insecticide resistance in malaria vectors in Africa.ConclusionsThe increasing pyrethroid and DDT resistance in Anopheles in the Afrotropical region is alarming. Urgent attention should be afforded to testing An. funestus populations especially for metabolic resistance mechanisms. IR Mapper is a useful tool for investigating temporal and spatial trends in Anopheles resistance to support the pragmatic use of insecticidal interventions.

Impact of agriculture on the selection of insecticide resistance in the malaria vector Anopheles gambiae : a multigenerational study in controlled conditions

Resistance of mosquitoes to insecticides is mainly attributed to their adaptation to vector control interventions. Although pesticides used in agriculture have been frequently mentioned as an additional force driving the selection of resistance, only a few studies were dedicated to validate this hypothesis and characterise the underlying mechanisms. While insecticide resistance is rising dramatically in Africa, deciphering how agriculture affects resistance is crucial for improving resistance management strategies. In this context, the multigenerational effect of agricultural pollutants on the selection of insecticide resistance was examined in Anopheles gambiae. An urban Tanzanian An. gambiae population displaying a low resistance level was used as a parental strain for a selection experiment across 20 generations. At each generation larvae were selected with a mixture containing pesticides and herbicides classically used in agriculture in Africa. The resistance levels of adults to deltamethrin, DDT and bendiocarb were compared between the selected and non-selected strains across the selection process together with the frequency of kdr mutations. A microarray approach was used for pinpointing transcription level variations selected by the agricultural pesticide mixture at the adult stage. A gradual increase of adult resistance to all insecticides was observed across the selection process. The frequency of the L1014S kdr mutation rose from 1.6% to 12.5% after 20 generations of selection. Microarray analysis identified 90 transcripts over-transcribed in the selected strain as compared to the parental and the non-selected strains. Genes encoding cuticle proteins, detoxification enzymes, proteins linked to neurotransmitter activity and transcription regulators were mainly affected. RT-qPCR transcription profiling of candidate genes across multiple generations supported their link with insecticide resistance. This study confirms the potency of agriculture in selecting for insecticide resistance in malaria vectors. We demonstrated that the recurrent exposure of larvae to agricultural pollutants can select for resistance mechanisms to vector control insecticides at the adult stage. Our data suggest that in addition to selected target-site resistance mutations, agricultural pollutants may also favor cuticle, metabolic and synaptic transmission-based resistance mechanisms. These results emphasize the need for integrated resistance management strategies taking into account agriculture activities.