Determination of the discriminating concentration of chlorfenapyr (pyrrole) and Anopheles gambiae sensu lato susceptibility testing in preparation for distribution of Interceptor® G2 insecticide-treated nets

Abstract

Background: Following agricultural use and large-scale distribution of insecticide-treated nets (ITNs), malaria vector resistance to pyrethroids is widespread in sub-Saharan Africa. Interceptor® G2 is a new dual active ingredient (AI) ITN treated with alpha-cypermethrin and chlorfenapyr for the control of pyrethroid-resistant malaria vectors. In anticipation of these new nets being more widely distributed, testing was conducted to develop a chlorfenapyr susceptibility bioassay protocol and gather susceptibility information.

Methods: Bottle bioassay tests were conducted using five concentrations of chlorfenapyr at 12.5, 25, 50, 100, and 200 µg AI/bottle in 10 countries in sub-Saharan Africa using 13,639 wild-collected Anopheles gambiae sensu lato (s.l.) (56 vector populations per dose) and 4,494 pyrethroid-susceptible insectary mosquitoes from 8 colonized strains. In parallel, susceptibility tests were conducted using a provisional discriminating concentration of 100 µg AI/bottle in 16 countries using 23,422 wild-collected, pyrethroid-resistant An. gambiae s.l. (259 vector populations). Exposure time was 60 min, with mortality recorded at 24, 48 and 72 h after exposure.

Results: Median mortality rates (up to 72 h after exposure) of insectary colony mosquitoes was 100% at all five concentrations tested, but the lowest dose to kill all mosquitoes tested was 50 µg AI/bottle. The median 72-h mortality of wild An. gambiae s.l. in 10 countries was 71.5, 90.5, 96.5, 100, and 100% at concentrations of 12.5, 25, 50, 100, and 200 µg AI/bottle, respectively. Log-probit analysis of the five concentrations tested determined that the LC₉₅ of wild An. gambiae s.l. was 67.9 µg AI/bottle (95% CI: 48.8-119.5). The discriminating concentration of 203.8 µg AI/bottle (95% CI: 146-359) was calculated by multiplying the LC₉₅ by three. However, the difference in mortality between 100 and 200 µg AI/bottle was minimal and large-scale testing using 100 µg AI/bottle with wild An. gambiae s.l. in 16 countries showed that this concentration was generally suitable, with a median mortality rate of 100% at 72 h.

Conclusions: This study determined that 100 or 200 µg AI/bottle chlorfenapyr in bottle bioassays are suitable discriminating concentrations for monitoring susceptibility of wild An. gambiae s.l., using mortality recorded up to 72 h. Testing in 16 countries in sub-Saharan Africa demonstrated vector susceptibility to chlorfenapyr, including mosquitoes with multiple resistance mechanisms to pyrethroids.

Keywords: Anopheles gambiae; CDC bottle bioassay; Chlorfenapyr; Discriminating concentration; Insecticide resistance; Insecticide-treated net; Interceptor G2; Pyrrole.

Fig. 1

Locations of insecticide susceptibility testing sites where mortality of wild Anopheles gambiae s.l. was measured in bioassays following exposure to a full (yellow) or limited (blue) range of concentrations of chlorfenapyr. *denotes countries where a susceptible insectary strain was also tested with the full range of concentrations

Fig. 2

Percentage mortality (72 h) after exposure to each of five concentrations (12.5, 25, 50, 100, 200 µg AI/bottle) of chlorfenapyr in bottle bioassays using pyrethroid susceptible colony mosquito strains in eight countries

Fig. 3

Median percentage mortality of wild Anopheles gambiae s.l. 72 h after exposure to chlorfenapyr at concentrations of 12.5, 25, 50, 100, and 200 µg AI/bottle in bottle bioassay in 10 countries

Fig. 4

Percentage mortality of wild Anopheles gambiae s.l. 60 min, 24 h, 48 h, and 72 h after exposure to the provisional discriminating concentration of chlorfenapyr at 100 µg AI/bottle in 16 countries

Fig. 5

Percentage mortality of wild Anopheles gambiae s.l. 60 min, 24 h, 48 h, and 72 h after exposure to chlorfenapyr at 200 µg AI/bottle in 10 countries