Comparative study of the effect of solvents on the efficacy of neonicotinoid insecticides against malaria vector populations across Africa

Background:New insecticides with a novel mode of action such as neonicotinoids have recently been recommended for public health by WHO.Resistance monitoring of such novel insecticides requires a robust protocol to monitor the development of resistance in natural populations.In this study,we comparatively used three different solvents to assess the susceptibility of malaria vectors to neonicotinoids across Africa.Methods:Mosquitoes were collected from May to July 2021 from three agricultural settings in Cameroon(Njombe-Penja,Nkolondom,and Mangoum),the Democratic Republic of Congo(Ndjili-Brasserie),Ghana(Obuasi),and Uganda(Mayuge).Using the CDC bottle test,we compared the effect of three different solvents(ethanol,acetone,MERO)on the efficacy of neonicotinoids againstAnopheles gambiae s.l.In addition,TaqMan assays were used to genotype key pyrethroid-resistant markers inAn.gambiae and odds ratio based on Fisher exact test were used to evaluate potential cross-resistance between pyrethroids and clothianidin.Results:Lower mortality was observed when using absolute ethanol or acetone alone as solvent for clothianidin(11.4-51.9% mortality in Nkolondom,31.7-48.2% in Mangoum,34.6-56.1% in Mayuge,39.4-45.6% in Obuasi,83.7-89.3% in Congo and 71.1-95.9% in Njombe pendja)compared to acetone+MERO for which 100% mortality were observed for all the populations.Similar observations were done for imidacloprid and acetamiprid.Synergist assays(PBO,DEM and DEF)with clothianidin revealed a significant increase of mortality suggesting that metabolic resistance mechanisms are contributing to the reduced susceptibility.A negative association was observed between the L1014F-kdr mutation and clothianidin resistance with a greater frequency of homozygote resistant mosquitoes among the dead than among survivors(OR=0.5;P=0.02).However,the I114T-GSTe2 was in contrast significantly associated with a greater ability to survive clothianidin with a higher frequency of homozygote resistant among survivors than other genotypes(OR=2.10;P=0.013).Conclusions:This study revealed a contrasted susceptibility pattern depending on the solvents with ethanol/acetone resulting to lower mortality,thus possibly overestimating resistance,whereas the MERO consistently showed a greater efficacy of neonicotinoids but it could prevent to detect early resistance development.Therefore,we recommend monitoring the susceptibility using both acetone alone and acetone+MERO(4μg/ml for clothianidin)to capture the accurate resistance profile of the mosquito populations.

First detection of F1534C knockdown resistance mutation in Aedes aegypti(Diptera:Culicidae)from Cameroon

Background: Aedes borne viral diseases,notably dengue,are increasingly reported in Cameroon with Aedes aegypti being a major vector.Data on insecticide resistance of this vector and underlying mechanisms needed for outbreak preparedness remain scarce in Cameroon.Here,we present the nationwide distribution of insecticide resistance in Ae.aegypti and investigate the potential resistance mechanisms involved.Methods:: Immature stages of Ae.aegypti were collected between March and July 2017 in 13 locations across Cameroon and reared until G1/G2/G3 generation.Larval,adult bioassays,and piperonyl butoxide(PBO)synergist assays were carried out according to World Health Organization guidelines.F1534C mutation was genotyped using allele specific polymerase chain reaction in field collected adults(Go)and the polymorphism of the sodium channel gene was assessed.Theχ2 test was used to compare the mortality rate between bioassays with insecticides only and bioassays after preexposure to PBO synergist.Results: Larval bioassay revealed that all the three populations tested with temephos were susceptible.Adult bioassays showed a good level of susceptibility toward both pyrethroids tested,0.25%permethrin and 0.05%deltamethrin,with six out of 10 populations susceptible.However,two populations(Douala and Edéa)were resistant(deltamethrin[73.2–92.5%mortality],permethrin[2.6–76.3%mortality]).The resistance to 4%dichlorodiphenyltrichloroethane was observed in four out of 10 populations tested(16.8–87.1%mortality).Resistance was also reported to carbamates including 0.1%propoxur(60.8–87.1%mortality)and to 0.1%bendiocarb(82.9%mortality).All populations tested were fully susceptible to 1%fenitrothion.A partial recovery of susceptibility was observed in the pyrethroid resistant population of Douala after pre-exposed to PBO suggesting the implication of cytochrome P450 monoxygenases permethrin resistance.Genotyping and sequencing detected the F1534C kdr mutation in the two pyrethroid resistant locations of Edéa and Douala,with allelic frequency of 3.3%and 33.3%respectively.However,the high genetic diversity of the sodium channel gene supports the recent introduction of this mutation in Cameroon.Conclusions: This study revealed the contrasting resistance profiles to insecticides of Ae.aegypti populations in Cameroon suggesting that,instead of a unique nationwide control approach,a regionally adapted strategy will be needed to control this vector.The localised distribution of the F1534C kdr mutation supports this region-specific control strategy.