Improving the efficiency of aerosolized insecticide testing against mosquitoes.

Developing robust and standardised approaches for testing mosquito populations against insecticides is vital for understanding the effectiveness of new active ingredients or formulations. Methods for testing mosquito susceptibility against contact insecticides or products, such as those delivered through public health programmes, are well-established and standardised. Nevertheless, approaches for testing volatile or aerosolized insecticides used in household products can be challenging to implement efficiently. We adapted WHO guidelines for household insecticides to develop a standardised and higher-throughput methodology for testing aerosolized products in a Peet Grady test chamber (PG-chamber) using caged mosquitoes and an efficient decontamination method. The new approach was validated using insecticide resistant and susceptible Aedes and Anopheles mosquito colonies. An added feature is the inclusion of cage-facing cameras to allow real-time quantification of knockdown following insecticide exposure. The wipe-based decontamination method was highly effective for removing pyrethroids' aerosolized oil-based residues from chamber surfaces, with < 2% mortality recorded for susceptible mosquitoes tested directly on the surfaces. There was no spatial heterogeneity for knockdown or mortality of caged mosquitoes within the PG chamber. The dual-cage approach we implement yields eight-times the throughput compared to a free-flight protocol, allows simultaneous testing of different mosquito strains and effectively discriminates susceptible and resistant mosquito colonies tested side-by-side.

Development and application of a tri-allelic PCR assay for screening Vgsc-L1014F kdr mutations associated with pyrethroid and organochlorine resistance in the mosquito Culex quinquefasciatus.

BACKGROUND: Culex quinquefasciatus has a widespread distribution across tropical and sub-tropical regions, and plays an important role in the transmission of vector-borne diseases of public health importance, including lymphatic filariasis (LF) and multiple arboviruses. Increased resistance to insecticides threatens the efficacy and sustainability of insecticide-based anti-vector interventions which mitigate the burden of mosquito transmitted diseases in endemic regions. In C. quinquefasciatus two non-synonymous voltage gated sodium channel (Vgsc) variants, both resulting in a leucine to phenylalanine change at codon 1014, are associated with resistance to pyrethroids and DDT. This tri-allelic variation has compromised the ability to perform high-throughput single-assay screening. To facilitate the detection and monitoring of the Vgsc-1014 locus in field-caught mosquitoes, an Engineered-Tail Allele-Specific-PCR (ETAS-PCR) diagnostic assay was developed and applied to wild mosquitoes from Brazil, Tanzania and Uganda. RESULTS: This new cost-effective, single-tube assay was compared to two, well-established, genotyping approaches, pyrosequencing and TaqMan. The ETAS-PCR assay showed high specificity for discriminating the three alleles at Vgsc-L1014F, with genotyping results strongly correlated with pyrosequencing and TaqMan results (98.64% and 100% respectively). CONCLUSIONS: Our results support the utility of the ETAS-PCR/Vgsc-1014 diagnostic assay, which stands as an effective alternative for genotyping tri-allelic variants.

Entomological Monitoring and Evaluation: Diverse Transmission Settings of ICEMR Projects Will Require Local and Regional Malaria Elimination Strategies.

The unprecedented global efforts for malaria elimination in the past decade have resulted in altered vectorial systems, vector behaviors, and bionomics. These changes combined with increasingly evident heterogeneities in malaria transmission require innovative vector control strategies in addition to the established practices of long-lasting insecticidal nets and indoor residual spraying. Integrated vector management will require focal and tailored vector control to achieve malaria elimination. This switch of emphasis from universal coverage to universal coverage plus additional interventions will be reliant on improved entomological monitoring and evaluation. In 2010, the National Institutes for Allergies and Infectious Diseases (NIAID) established a network of malaria research centers termed ICEMRs (International Centers for Excellence in Malaria Research) expressly to develop this evidence base in diverse malaria endemic settings. In this article, we contrast the differing ecology and transmission settings across the ICEMR study locations. In South America, Africa, and Asia, vector biologists are already dealing with many of the issues of pushing to elimination such as highly focal transmission, proportionate increase in the importance of outdoor and crepuscular biting, vector species complexity, and "sub patent" vector transmission.

Nested clade and phylogeographic analyses of the Chagas disease vector Triatoma brasiliensis in Northeast Brazil.

Triatoma brasiliensis (Hemiptera: Reduviidae: Triatominae) is the most important Chagas disease vector in the semiarid areas of Northeast Brazil. We analyzed mitochondrial cytochrome b sequence variation among 136 individuals representing 16 populations from across the species' distribution. Neighbor-joining and parsimony tree-building methods were used in conjunction with nested clade analysis to describe the systematics and phylogeography of this species. Our results indicate that T. brasiliensis is composed of four genetically distinct chromatic forms (referred to as brasiliensis, macromelasoma, juazeiro, and melanica) that present inter-population divergence values (0.027-0.119, corrected K2-p) and a pattern of haplotype geographic distribution compatible with the existence of a species complex. As a consequence, such forms can be treated as isolated targets in vector control programs. We were unable to infer what is shaping the population structure of the brasiliensis form as we obtained mutually exclusive causes of structure, namely a barrier to gene flow caused by past population fragmentation, and isolation by distance between populations (which would permit gene flow). We found indication of mitochondrial DNA introgression occurring among forms in putative hybrid zones.