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

Background: Broflanilide is a new insecticide being developed for malaria vector control. As new insecticide chemistries become available, strategies to preserve the susceptibility of local malaria vectors and extend their useful life need to be considered before large scale deployment. This requires the development of appropriate testing procedures and identification of suitable discriminating concentrations for monitoring susceptibility in wild vector populations to facilitate decision making by control programmes.

Methods: Dose-response WHO bottle bioassays were conducted using the insecticide-susceptible Anopheles gambiae s.s. Kisumu strain to determine a discriminating concentration of broflanilide. Bioassays were performed without the adjuvant Mero^® and with two concentrations of Mero^® (500 ppm and 800 ppm) to investigate its impact on the discriminating concentration of the insecticide. Probit analysis was used to determine the lethal doses at 50% (LC₅₀) and 99% (LC₉₉) at 24-, 48- and 72-hours post-exposure. Cross-resistance to broflanilide and pyrethroids, DDT, dieldrin and carbamates, was investigated using An. gambiae s.l. Covè and An. coluzzii Akron strains. The susceptibility of wild pyrethroid-resistant mosquitoes from communities in Southern Benin to broflanilide was assessed using the estimated discriminating concentrations.

Results: Broflanilide induced a dose-dependent and delayed mortality effect. Mortality rates in bottles treated without Mero^® were <80% using the range of broflanilide doses tested (0-100 µg/bottle) leading to high and unreliable estimates of LC₉₉ values. The discriminating concentrations defined as 2XLC₉₉ at 72 hours post exposure were estimated to be 2.2 µg/bottle with 800 ppm of Mero^® and 6.0 µg/bottle with 500 ppm of Mero^®. Very low resistance ratios (0.6-1.2) were determined with the insecticide resistant An. gambiae s.l. Covè and An. coluzzii Akron strains suggesting the absence of cross-resistance via the mechanisms of resistance to pyrethroids, DDT, dieldrin and carbamates they possess. Bottle bioassays performed with broflanilide at both discriminating concentrations of 6 µg/bottle with 500 ppm of Mero^® and 2.2 µg/bottle with 800 ppm of Mero^®, showed susceptibility of wild highly pyrethroid-resistant An. gambiae s.l. from villages in Southern Benin.

Conclusion: Here we determined discriminating concentrations for monitoring susceptibility to broflanilide in bottle bioassays, using susceptible An. gambiae vectors. Using the estimated discriminating concentrations, we showed that wild pyrethroid-resistant populations of An. gambiae s.l. from southern Benin were fully susceptible to the insecticide. Broflanilide also shows potential to be highly effective against An. gambiae s.l. vector populations that have developed resistance to other public health insecticides.