Challenges and opportunities associated with the introduction of next generation long lasting insecticidal nets for malaria control: a case study from Burkina Faso.

BACKGROUND Reductions in malaria incidence in Africa can largely be attributed to increases in malaria vector control activities; predominately the use of long lasting insecticidal nets (LLINs). With insecticide resistance affecting an increasing number of malaria endemic countries and threatening the effectiveness of conventional LLINs, there is an increasing urgency to implement alternative tools that control these resistant populations. The aim of this study was to identify potential challenges and opportunities for accelerating access to next generation LLINs in Burkina Faso, a country with areas of high levels of insecticide resistance. METHODS An analytical framework was used to guide the selection of interviewees, data collection and analysis. Semi structured interviews were carried out with key informants in April 2014 in Burkina Faso. Interviews were conducted in French and English, audio recorded, transcribed and entered into NVivo10 for data management and analysis. Data were coded according to the framework themes and then analysed to provide a description of the key points and explain patterns in the data. RESULTS Interviewees reported that the policy architecture in Burkina Faso is characterised by a strong framework of actors that contribute to policymaking and strong national research capacity which indirectly contributes to national policy change via collaboration with internationally led research. Financing significantly impacts the potential adoption, availability and affordability of next generation LLINs. This confers significant power on international donors that fund vector control. National decisions around which LLINs to procure were restricted to quantity and delivery dates; the potential to tackle insecticide resistance was not part of the decision-making process. Furthermore, at the time of the study there was no World Health Organisation (WHO) guidance on where and when next generation LLINs might positively impact on malaria transmission, severely limiting their adoption, availability and affordability. CONCLUSIONS This study shows that access to next generation LLINs was severely compromised by the lack of global guidance. In a country like Burkina Faso where WHO recommendations are relatively quickly adopted, a clear WHO recommendation and adequate financing will be key to accelerate access to next generation LLINs. Key Words: Malaria, Next generation long lasting insecticidal nets, LLINs, PBO nets, Policy Analysis, Vector Control, Burkina Faso

Effective population size of Anopheles funestus chromosomal forms in Burkina Faso

BACKGROUND: As Anopheles funestus is one of the principal Afro-tropical malaria vectors, a more complete understanding of its population structure is desirable. In West and Central Africa, An. funestus population structure is complicated by the coexistence of two assortatively mating chromosomal forms. Effective population size (N(e)) is a key parameter in understanding patterns and levels of intraspecific variation, as it reflects the role of genetic drift. Here, N(e )was estimated from both chromosomal forms, Kiribina and Folonzo, in Burkina Faso. METHODS: Short-term N(e )was estimated by evaluating variation at 16 microsatellite loci across temporal samples collected annually from 2000–2002. Estimates were based on standardized variance in allele frequencies or a maximum likelihood method. Long-term N(e )was estimated from genetic diversity estimates using mtDNA sequences and microsatellites. RESULTS: For both forms, short-term and long-term N(e )estimates were on the order of 10(3 )and 10(5), respectively. Long-term N(e )estimates were larger when based on loci from chromosome 3R (both inside and outside of inversions) than loci outside of this arm. CONCLUSION: N(e )values indicate that An. funestus is not subject to seasonal bottlenecks. Though not statistically different because of large and overlapping confidence intervals, short-term N(e )estimates were consistently smaller for Kiribina than Folonzo, possibly due to exploitation of different breeding sites: permanent for Folonzo and intermittent for Kiribina. The higher long-term N(e )estimates on 3R, the arm carrying the two inversions mainly responsible for defining the chromosomal forms, give natural selection broader scope and merit further study

Seasonal distribution of Anopheles funestus chromosomal forms from Burkina Faso

Abstract Background Previous studies of Anopheles funestus chromosomal inversion polymorphisms in Burkina Faso showed large departures from Hardy-Weinberg equilibrium and linkage disequilibrium among inversions located on different chromosomes, implying the existence of two taxonomic units ("chromosomal forms") with limited genetic flow. One chromosomal form, named Folonzo, is highly polymorphic for alternative rearrangements of 3R a, 3R b, 2R a, and 3L a; the other, Kiribina, is predominantly characterized by the standard arrangement of these inversions. To investigate the temporal distribution of these chromosomal forms, further collections were carried out in two villages near Ouagadougou where they are found in sympatry. Methods Chromosomal karyotypes were determined from indoor-resting, half-gravid females sampled within and across six breeding seasons, from December 1998 to April 2007. Results As expected, the pattern of chromosomal polymorphism in An. funestus was consistent with assortatively mating Folonzo and Kiribina forms. When samples were assigned to each chromosomal form, their relative abundance varied within successive breeding seasons in a repeating pattern of temporal variability. Relative abundance of the Folonzo form was correlated with climatic variables related to temperature and rainfall. Conclusion The relative abundance of Folonzo and Kiribina forms of An. funestus likely reflects different larval ecologies that are linked to varying climatic conditions. Further analysis of the bionomics of these vectors is recommended in light of its relevance to vector control.http://deepblue.lib.umich.edu/bitstream/2027.42/112459/1/12936_2009_Article_987.pd

Behavioural plasticity of Anopheles coluzzii and Anopheles arabiensis undermines LLIN community protective effect in a Sudanese-savannah village in Burkina Faso

Background Despite the overall major impact of long-lasting insecticide treated nets (LLINs) in eliciting individual and collective protection to malaria infections, some sub-Saharan countries, including Burkina Faso, still carry a disproportionately high share of the global malaria burden. This study aims to analyse the possible entomological bases of LLIN limited impact, focusing on a LLIN-protected village in the Plateau Central region of Burkina Faso. Methods Human landing catches (HLCs) were carried out in 2015 for 12 nights both indoors and outdoors at different time windows during the highest biting activity phase for Anopheles gambiae (s.l.). Collected specimens were morphologically and molecularly identified and processed for Plasmodium detection and L1014F insecticide-resistance allele genotyping. Results Almost 2000 unfed An. gambiae (s.l.) (54% Anopheles coluzzii and 44% Anopheles arabiensis) females landing on human volunteers were collected, corresponding to a median number of 23.5 females/person/hour. No significant differences were observed in median numbers of mosquitoes collected indoors and outdoors, nor between sporozoite rates in An. coluzzii (6.1%) and An. arabiensis (5.5%). The estimated median hourly entomological inoculation rate (EIR) on volunteers was 1.4 infective bites/person/hour. Results do not show evidence of the biting peak during night hours typical for An. gambiae (s.l.) in the absence of bednet protection. The frequency of the L1014F resistant allele (n = 285) was 66% in An. coluzzii and 38% in An. arabiensis. Conclusions The observed biting rate and sporozoite rates are in line with the literature data available for An. gambiae (s.l.) in the same geographical area before LLIN implementation and highlight high levels of malaria transmission in the study village. Homogeneous biting rate throughout the night and lack of preference for indoor-biting activity, suggest the capacity of both An. coluzzii and An. arabiensis to adjust their host-seeking behaviour to bite humans despite bednet protection, accounting for the maintenance of high rates of mosquito infectivity and malaria transmission. These results, despite being limited to a local situation in Burkina Faso, represent a paradigmatic example of how high densities and behavioural plasticity in the vector populations may contribute to explaining the limited impact of LLINs on malaria transmission in holo-endemic Sudanese savannah areas in West Africa

Efficacy of Olyset Duo, a bednet containing pyriproxyfen and permethrin, versus a permethrin-only net against clinical malaria in an area with highly pyrethroid-resistant vectors in rural Burkina Faso: a cluster-randomised controlled trial

Background Substantial reductions in malaria incidence in sub-Saharan Africa have been achieved with massive deployment of long-lasting insecticidal nets (LLINs), but pyrethroid resistance threatens control. Burkina Faso is an area with intense malaria transmission and highly pyrethroid-resistant vectors. We assessed the effectiveness of bednets containing permethrin, a pyrethroid, and pyriproxyfen, an insect growth regulator, versus permethrin-only (standard) LLINs against clinical malaria in children younger than 5 years in Banfora, Burkina Faso. Methods In this two-group, step-wedge, cluster-randomised, controlled, superiority trial, standard LLINs were incrementally replaced with LLINs treated with permethrin plus pyriproxyfen (PPF) in 40 rural clusters in Burkina Faso. In each cluster, 50 children (aged 6 months to 5 years) were followed up by passive case detection for clinical malaria. Cross-sectional surveys were done at the start and the end of the transmission seasons in 2014 and 2015. We did monthly collections from indoor light traps to estimate vector densities. Primary endpoints were the incidence of clinical malaria, measured by passive case detection, and the entomological inoculation rate. Analyses were adjusted for clustering and for month and health centre. This trial is registered as ISRCTN21853394. Findings 1980 children were enrolled in the cohort in 2014 and 2157 in 2015. At the end of the study, more than 99% of children slept under a bednet. The incidence of clinical malaria was 2·0 episodes per child-year in the standard LLIN group and 1·5 episodes per child-year in the PPF-treated LLIN group (incidence rate ratio 0·88 [95% CI 0·77–0·99; p=0·04]). The entomological inoculation rate was 85 (95% CI 63–108) infective bites per transmission season in the standard LLIN group versus 42 (32–52) infective bites per transmission season in the PPF-treated LLIN group (rate ratio 0·49, 95% CI 0·32–0·66; p<0·0001). Interpretation PPF-treated LLINs provide greater protection against clinical malaria than do standard LLINs and could be used as an alternative to standard LLINs in areas with intense transmission of Plasmodium falciparum malaria and highly pyrethroid-resistant vectors. Funding EU Seventh Framework Programme

Aging partially restores the efficacy of malaria vector control in insecticide-resistant populations of Anopheles gambiae s.l. from Burkina Faso

&lt;br&gt;Background: The operational impact of insecticide resistance on the effectiveness of long-lasting insecticide nets (LLINs) and indoor residual spraying (IRS) is poorly understood. One factor which may prolong the effectiveness of these tools in the field is the increase in insecticide susceptibility with mosquito age. In this study, LLINs and IRS were tested against young (three to five days) and old (17-19 days) pyrethroid resistant Anopheles gambiae s.l. from Burkina Faso.&lt;/br&gt; &lt;br&gt;Methods: Blood-fed adult Anopheles gambiae s.l. were collected from south-west Burkina Faso and identified to species/form level. Cohorts of the F1 progeny of An. gambiae s.s. S-forms were exposed to deltamethrin (0.05%) at three to five or 17-19 days post-emergence and tested for the frequency of the resistance allele 1014F. Isofemale lines of the M, S- form of An. gambiae s.s. and Anopheles arabiensis were exposed in WHO cone tests to either a) LLINs deployed in households for two years or (b) bendiocarb sprayed walls.&lt;/br&gt; &lt;br&gt;Results: Mortality rates in response to deltamethrin (0.05%) increased from levels indicative of strong resistance in three to five day old F1 mosquitoes, to near full susceptibility in the 17-19 day old cohort. On exposure to LLINs sampled from the field, the mortality rate in isofemale lines was higher in older mosquitoes than young (OR = 5.28, CI 95% = 2.81-9.92), although the mortality estimates were affected by the LLIN tested. In general, the LLINs sampled from the field performed poorly in WHO cone bioassays using either laboratory susceptible or field caught mosquito populations. Finally, there was a clear relationship between mortality and age on exposure to bendiocarb-sprayed walls, with older mosquitoes again proving more susceptible (OR = 3.39, CI 95% = 2.35-4.90).&lt;/br&gt; &lt;br&gt;Conclusions: Age is a key factor determining the susceptibility of mosquitoes to insecticides, not only in laboratory studies, but in response to field-based vector control interventions. This has important implications for understanding the epidemiological impact of resistance. If mosquitoes old enough to transmit malaria are still being suppressed with available insecticides, is resistance potentially having less of an impact than often assumed? However, the poor performance of LLINs used in this study in Burkina Faso, is a cause for concern and requires urgent investigation.&lt;/br&gt

De Novo Transcriptome Sequencing in Anopheles funestus Using Illumina RNA-Seq Technology

BACKGROUND: Anopheles funestus is one of the primary vectors of human malaria, which causes a million deaths each year in sub-Saharan Africa. Few scientific resources are available to facilitate studies of this mosquito species and relatively little is known about its basic biology and evolution, making development and implementation of novel disease control efforts more difficult. The An. funestus genome has not been sequenced, so in order to facilitate genome-scale experimental biology, we have sequenced the adult female transcriptome of An. funestus from a newly founded colony in Burkina Faso, West Africa, using the Illumina GAIIx next generation sequencing platform. METHODOLOGY/PRINCIPAL FINDINGS: We assembled short Illumina reads de novo using a novel approach involving iterative de novo assemblies and "target-based" contig clustering. We then selected a conservative set of 15,527 contigs through comparisons to four Dipteran transcriptomes as well as multiple functional and conserved protein domain databases. Comparison to the Anopheles gambiae immune system identified 339 contigs as putative immune genes, thus identifying a large portion of the immune system that can form the basis for subsequent studies of this important malaria vector. We identified 5,434 1:1 orthologues between An. funestus and An. gambiae and found that among these 1:1 orthologues, the protein sequence of those with putative immune function were significantly more diverged than the transcriptome as a whole. Short read alignments to the contig set revealed almost 367,000 genetic polymorphisms segregating in the An. funestus colony and demonstrated the utility of the assembled transcriptome for use in RNA-seq based measurements of gene expression. CONCLUSIONS/SIGNIFICANCE: We developed a pipeline that makes de novo transcriptome sequencing possible in virtually any organism at a very reasonable cost ($6,300 in sequencing costs in our case). We anticipate that our approach could be used to develop genomic resources in a diversity of systems for which full genome sequence is currently unavailable. Our An. funestus contig set and analytical results provide a valuable resource for future studies in this non-model, but epidemiologically critical, vector insect

Combining organophosphate treated wall linings and long-lasting insecticidal nets for improved control of pyrethroid resistant Anopheles gambiae.

BACKGROUND: New approaches to delivering insecticides need to be developed to improve malaria vector control. Insecticidal durable wall lining (DL) and net wall hangings (NWH) are novel alternatives to indoor residual spraying which can be produced in a long-lasting format. Non-pyrethroid versions could be used in combination with long-lasting insecticidal nets for improved control and management of insecticide resistant vector populations. METHODS: Experimental hut trials were carried out in Valley du Kou, Burkina Faso to evaluate the efficacy of pirimiphos methyl treated DL and NWH either alone or in combination with LLINs against pyrethroid resistant Anopheles gambiae ss. Comparison was made with pyrethroid DL. Mosquitoes were genotyped for kdr and ace-1R resistant genes to investigate the insecticide resistance management potential of the combination. RESULTS: The overall kdr and ace-1(R) allele frequencies were 0.95 and 0.01 respectively. Mortality with p-methyl DL and NWH alone was higher than with pyrethroid DL alone (>95% vs 40%; P0.1). Combining p-methyl DL or NWH with LLINs reduced biting rates significantly (8-9%) compared to p-methyl DL and NWH alone (>40%) and killed all An gambiae that entered the huts. Mosquitoes bearing the ace-1(R) gene were more likely to survive in huts with p-methyl DL alone (p<0.03) whereas all resistant and susceptible genotypes were killed by the combination. CONCLUSION: P-methyl DL and NWH outperformed pyrethroid DL. Combining p-methyl DL and NWH with LLINs could provide significant epidemiological benefits against a vector population which is resistant to pyrethroids but susceptible to organophosphates. There was evidence that the single intervention would select kdr and ace-1(R) resistance genes and the combination intervention might select less strongly. Technology to bind organophosphates to plastic wall lining would be worth developing

Do bednets including piperonyl butoxide offer additional protection against populations of Anopheles gambiae s.l. that are highly resistant to pyrethroids? An experimental hut evaluation in Burkina Fasov

Malaria control is dependent on the use of Long Lasting Insecticidal Nets (LLINs) containing pyrethroids. A new generation of LLINs containing both pyrethroids and the synergist piperonyl butoxide (PBO LLINs) have been developed in response to increasing pyrethroid resistance in African malaria vectors but questions remain about the performance of these nets in areas where pyrethroid resistance is at very high levels. This study was conducted in two settings in Southwest Burkina Faso, Vallée du Kou 5 and Tengrela where Anopheles gambiae s.l. mortality rates in WHO discriminating dose assays were < 14 % for permethrin < 33 % for deltamethrin (< 33 %). When mosquitoes were pre-exposed to PBO in WHO tube assays mortality rates increased substantially but full susceptibility was not restored. Molecular characterisation revealed high levels of kdr alleles and elevated levels of P450s previously implicated in pyrethroid resistance. In cone bioassays and experimental huts the PBO LLINs outperformed the pyrethroid only equivalents from the same manufacturers. Blood feeding rates were 1.6 - 2.2 fold lower, and mortality rates were increased by 1.69 – 1.78 -fold in huts with PBO LLINs versus non PBO LLINs. This study indicates that PBO LLINs provide greater personal and community level protection than standard LLINs against highly pyrethroid resistant mosquito populations