A new WHO bottle bioassay method to assess the susceptibility of mosquito vectors to public health insecticides: results from a WHO-coordinated multi- centre study

Abstract

Background: The continued spread of insecticide resistance in mosquito vectors of malaria and arboviral diseases may lead to operational failure of insecticide-based interventions if resistance is not monitored and managed efficiently. This study aimed to develop and validate a new WHO glass bottle bioassay method as an alternative to the WHO standard insecticide tube test to monitor mosquito susceptibility to new public health insecticides with particular modes of action, physical properties or both.Methods: A multi-centre study involving 21 laboratories worldwide generated data on the susceptibility of seven mosquito species (i.e. Aedes aegypti, Ae. albopictus, Anopheles gambiae s.s., An. funestus, An. stephensi, An. minimus and An. albimanus) to seven public health insecticides in five classes, including pyrethroids (metofluthrin, prallethrin, and transfluthrin), neonicotinoids (clothianidin), pyrroles (chlorfenapyr), juvenile hormone mimics (pyriproxyfen) and butenolides (flupyradifurone) in glass bottle assays. The data were analysed using a Bayesian binomial model to determine the concentration–response curves for each insecticide–species combination and to assess the within-bioassay variability in the susceptibility endpoints, killing 50% and 99% of the population (LC50 and LC99) or inhibiting oviposition by 50% and 99% (OI50 and OI99) to measure mortality and the sterilizing effect, respectively. Results: Overall, about 200,000 mosquitoes were tested with the new bottle bioassay, and LC50 / LC99 or OI50 / OI99 values were determined for all insecticides. Variation was seen between laboratories in estimates for some mosquito species/insecticide combinations, while other test results were consistent. The variation was generally greater with transfluthrin and flupyradifurone than with other compounds tested, especially against Anopheles species. Overall, the mean within-bioassay variability in mortality and oviposition inhibition were <10% for most mosquito species-insecticide combinations. Conclusion. Our findings, based on the largest susceptibility dataset ever produced on mosquitoes, showed that the WHO bottle bioassay is adequate for evaluating mosquito susceptibility to new and promising public health insecticides currently deployed for vector control. The datasets presented in this study have been used recently by the WHO to establish 17 new insecticide discriminating concentrations (DCs) for either Aedes spp. or Anopheles spp. The bottle bioassay and DCs can now be widely used to monitor baseline insecticide susceptibility of wild populations of vectors of malaria and Aedes-borne diseases worldwide.

Investigating discriminating concentrations for monitoring susceptibility to broflanilide and cross resistance to other insecticide classes in Anopheles gambiae sensu lato, using the new WHO bottle bioassay method

“…albimanus ), to insecticides that require the addition of an adjuvant in bottle bioassays [ 15 ]. The methods used in our study also followed the new WHO bottle bioassay procedures in terms of bottle coating, drying time, exposure time, endpoints etc [ 17 , 18 ]. This suggests that the lower discriminating concentration of 2.2 μg applied with Mero ® at 800 ppm, as recommended by WHO, could be a more suitable discriminating concentration for monitoring resistance to broflanilide in An .…”

“…The WHO has recently recommended new bottle bioassay procedures and discriminating concentrations for investigating resistance in mosquito vectors to new insecticides that are not suitable for impregnation onto filter papers [ 15 , 17 , 18 ]. The bottle bioassay is a practical method that offers the opportunity to include an adjuvant to help improve the distribution and presentation of insecticides that tend to crystallise on surfaces.…”

Investigating discriminating concentrations for monitoring susceptibility to broflanilide and cross resistance to other insecticide classes in Anopheles gambiae sensu lato, using the new WHO bottle bioassay method

“…albimanus ), to insecticides that require the addition of an adjuvant in bottle bioassays [14]. The methods used in our study also followed the new WHO bottle bioassay procedures in terms of bottle coating, drying time, exposure time, endpoints etc [16, 17]. This suggests that the lower discriminating concentration of 2.2 μg applied with Mero ® at 800 ppm, as recommended by WHO, could be a more suitable discriminating concentration for monitoring resistance to broflanilide in An.…”

“…Laboratory bioassays using insecticide discriminating concentrations have historically provided an efficient method for monitoring insecticide resistance in local malaria vector populations to help guide contextspecific decision making around their deployment for vector control [15]. The WHO has recently recommended new bottle bioassay procedures and discriminating concentrations for investigating resistance in mosquito vectors to new insecticides that are not suitable for impregnation onto filter papers [14,16,17]. The bottle bioassay is a practical method that offers the opportunity to include an adjuvant to help improve the distribution and presentation of insecticides that tend to crystallise on surfaces.…”

“…The WHO has recently recommended new bottle bioassay procedures and discriminating concentrations for investigating resistance in mosquito vectors to new insecticides that are not suitable for impregnation onto filter papers [14, 16, 17]. The bottle bioassay is a practical method that offers the opportunity to include an adjuvant to help improve the distribution and presentation of insecticides that tend to crystallise on surfaces.…”