Background
Malaria vectors control mainly relies on the use of insecticide-treated nets (ITN) and indoor residual spraying (IRS). Pyrethroids are the only group of insecticides currently recommended for net treatment [
1]. Although pyrethroid resistance in the most important malaria vector
Anopheles gambiae s.s. has become widespread in several African countries [
2‐
5], field studies in experimental huts and at community level using malaria indicators have shown that pyrethroid-treated bed nets remain usually effective against pyrethroid resistant mosquitoes [
6‐
9]. However, the evolution of pyrethroid resistance in
An. gambiae s.s. represent a threat for malaria control. To prevent any significant decline of the efficiency of pyrethroids, harmful to the malaria control, management strategies of pyrethroid resistance are envisaged through the exploration of news tools or combination of existing ones.
One of these strategies, used in agriculture as well as in public health, consists to associate in the same treatment, several molecules having different modes of action. Although developed initially for agricultural use and for indoor residual spraying, carbamates and organophosphates constitute a new prospect to circumvent pyrethroid resistance in An. gambiae s.s.
In area of high prevalence of
kdr in Côte d'Ivoire, experimental hut trials of carbamates or organophosphates alone and in combination with pyrethroids on mosquito nets showed very promising results [
8,
10‐
13]. However, little is known about the susceptibility status of pyrethroid resistance populations of
An. gambiae to organophosphates and carbamates in Côte d'Ivoire, as well as potential resistance mechanisms.
Acetylcholinesterase (AChE) is a common target for carbamates and organophosphates. These insecticides blocks transmission of nerve impulses by irreversible inhibition of AChE at cholinergic synapses, causing insect death. Cross-resistance to carbamates and organophosphates can arise by insensitive AChE mechanism due to the glycine to serine substitution (G119S mutation) resulting from a single point mutation in the
ace-1 gene [
14]. The G119S mutation was selected independently in several mosquitoes species including
An. gambiae s.s., the major malaria vector in Africa [
11,
14‐
17]. This mutation was found in both M and S molecular forms of
An. gambiae from Côte d'Ivoire [
16,
17].
In the current study, the geographic extent of insensitive AChE mechanism in An. gambiae s.s. populations from Côte d'Ivoire according to molecular forms, as well as their susceptibility status to carbamates and organophosphates were investigated.
Discussion
The distribution of M and S molecular forms of
An. gambiae s.s. in the study agrees with previous findings that reported both M and S forms in Guinea savannah areas and only the M form in the forest areas [
27‐
30]. This geographic distribution seems to follow more the global environment than the breeding sites nature. Both forms are involved in carbamate and organophosphate resistance, although at different level according to insecticides. Indeed, in this study,
An. gambiae s.s. displayed large variations in resistance level to carbamates and organophosphates. Although the wild populations were all resistant to carbamates, resistance was less marked to propoxur than to carbosulfan at WHO diagnostic concentrations.
All these populations were as resistant to carbosulfan as the population of Yaokoffikro in surrounding area of Bouaké [
11]. The resistance reported in Bouaké was attributed to agricultural or domestic hygiene or public health use of carbamates. In Burkina-Faso, Diabaté
et al[
31] attributed
An. gambiae s.s. pyrethroid resistance in cotton field areas to their use in agriculture.
The observed cross-resistance to organophosphates and carbamates in Tiassalé and Toumodi highlights implication of their common target site: the AChE-1. Although the mutation
ace-1 G119S provided cross-resistance to organophosphates and carbamates, the resistance level greatly varied between both insecticide families. This difference observed in resistance level could be the consequence of their difference observed in dominance level relied on insecticide specificity. According to Djogbénou
et al[
32], dominance status of
ace-1 G119S varied between semirecessivity with fenitrothion and chlorpyrifos methyl to semidominance with propoxur and carbamates. The fact that low cross-resistance was observed in the other populations, suggests and confirms potential involvement of metabolic resistance mechanisms and/or alternative mutation associated to G119S. This may explain why mortality rates to organophosphates among samples from Nieky, Abidjan and Yamoussoukro were so strong despite confirmed resistance level to carbamates in bioassays.
Such result could also be explained by possible cross-resistance between organophosphates and pyrethroids based on an increased detoxification mechanism were as suggested for other anopheline species selected for pyrethroid resistance [
33].
Moreover an alternative mutation in
ace-1 gene was described in the
Culex pipiens strain originating from Cyprus. This mutation is F290V substitution and it confers cross-resistance to OP and carbamate insecticides [
34]. Because
C. pipiens and
An. gambiae s.s. share G119S, it is possible that they share also this other mutation. Asidi
et al[
13] had noted that G119S mutation did not confer effective resistance to chlorpyrifos-methyl. Yet, the G119S mutation involved certainly a high resistance to carbamate but could enhance organophosphate hydrolysis. Similar mutations in a homologous position to G119S are known to alter substrate specificity in
Drosophila melanogaster and enhanced hydrolysis of some organophosphates [
35].
The presence of G119S mutation in both M and S forms of
An. gambiae s.s. has already been reported by Weill
et al[
16] and Djogbénou
et al[
17] and was suggested to result from introgression between forms. The wide distribution of
ace-1
R
reported here could result from an unique event that then spread as reported in
C. pipiens amplified esterase B2 genes through the world [
36].
The absence of homozygous resistant individuals might be related to high fitness cost of the G119S mutation, resulting on death of the homozygous resistant [
13,
16,
17]. Indeed, greater mortality of resistant individuals during pupation relative to their sensitive counterparts was reported. There was also evidence for costs to adult fitness as resistant individuals were smaller than sensitive adults [
37]. Consequently, in area where the resistant allele
ace-1
R
is present, resistant mosquitoes will mainly at heterozygote state (
ace-1
RS
). Because of this fitness cost, at least one duplication combining resistant and susceptible alleles of the
ace-1 locus has recently appeared, started to spread and replace
ace-1
R
in treated areas [
17,
38‐
41]. Duplications lead to an excess of heterozygotes in natural populations because that heterozygotes involving either
ace-1
S
or
ace-1
R
alleles do not exhibit deleterious side effects. To date, no specific test is available for detecting specifically
ace-1 duplications as mosquitoes carrying duplications appear as heterozygous for
ace-1
R
mutation.
Further investigation is needed to tackle the origin of the difference of resistance between carbamates and organophosphates.
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
LPAA, AAK designed the study, conducted the field work, genotyping, summarized the data and drafted the manuscript. MAA, ET jointly carried out PCR assays, and interpreted the results. PKK and MK supervised LPAA and AAK and contributed to the manuscript. FC contributed to design of the study and manuscript drafting. All authors read and approved the final manuscript.