Background
Pyrethroids are the only insecticides currently recommended by the World Health Organization for treatment of mosquito nets owing to their strong insecticidal activity at low concentrations and their low mammalian toxicity [
1]. Pyrethroid-treated nets are effective in reducing malaria morbidity and mortality [
2‐
4] and may also provide community protection through mass impact on vector mosquito populations, when used at a high coverage rate [
5,
6].
Pyrethroid resistance in
Anopheles gambiae has become widespread in different regions of Africa [
7‐
9] and may represent a threat for successful and sustainable implementation of insecticide-treated net (ITN) programmes. There is evidence that massive use of DDT against cotton pests in the 1960s and 1970s was responsible for the selection of the
kdr mutation (knock-down resistance) responsible for resistance to pyrethroids [
8]. A point mutation (leucine to phenylalanine) in the S6 transmembrane segment domain II in the sodium channel sequence is associated with the
kdr resistance in West Africa [
10] and is characterized by a reduction in the intrinsic sensitivity of the insect nervous system to DDT and pyrethroids [
11]. A second point mutation (leucine to serine) has also been reported in
An. gambiae from East Africa and is responsible for high level of resistance to permethrin [
12].
The selection of insecticide resistance is a complex evolutionary process [
13] that may depend on genetic, ecological and operational factors [
14]. Since operational factors are the main ones that can, in principle, theoretically be controlled, Georghiou [
15] characterized resistance management tactics according to the intensity of exposure (low
versus high doses), the frequency of applications and their use either in space or time. Because resistant genes are mainly present in a heterozygous state when they first evolve, a strategy to delay the development of resistance would involve doses which preferentially kill heterozygotes (RS) as well as susceptible homozygotes (SS) [
16]. A good understanding of the response to insecticides by mosquitoes heterozygous for resistance is essential to understand how resistance may evolve and to develop tactics for resistance management.
In the present study, investigations were carried out on the behaviour and selection of
An. gambiae of different genotypes for
kdr exposed to nettings treated with a range of permethrin concentrations. Experiments were carried out under laboratory conditions using tunnel tests [
17] and, in the field, using experimental huts [
18].
Discussion
In this tunnel test study, the response of
An. gambiae females heterozygous for
kdr to permethrin treated nets was comparable to that of susceptible ones, confirming that
kdr is recessive in
An. gambiae [
21]. However, a previous study showed a greater efficacy of permethrin treated nets, at the same concentrations, against a pyrethroid-resistant strain of
An. gambiae [
21]. These authors demonstrated that resistant mosquitoes, which could tolerate higher doses of permethrin, stayed longer than susceptible ones on the treated surfaces and took up more insecticide than did susceptibles, resulting in a good efficacy of the pyrethroid treated nettings. Such contrasting results may be explain by larger number of holes in our netting screens of the tunnel tests (9 holes instead of 5), which allowed the resistant mosquitoes to make less contact with the impregnated material and resulted in lower mortality of RR females, even at the highest dose (500 mg/m
2). This dosage, however, was the most efficient in killing preferentially RS and SS females. The experimental hut study showed that nets treated with 1,000 mg/m
2 permethrin were the most effective against pyrethroid-resistant
An. gambiae. However, permethrin concentration as high as 1,000 mg/m
2 should not be recommended without careful consideration of cost and safety issues.
It was also demonstrated that there was little or no correlation between the permethrin concentration and the distribution of
kdr genotypes as far as deterrency, excito-repellency, blood feeding and mortality were concerned. Interestingly, the
kdr allelic frequency was not significantly different between mosquitoes which survived or died after exposure to permethrin treated nets, even at high concentrations. Such results contrasted with findings from other authors [
30] who reported that nets treated either with alpha-cypermethrin or etofenprox in Côte d'lvoire, selected for
kdr in
An. gambiae. This difference could be attributed either to the chemical structure of the insecticide (permethrin being a non α-cyano-pyrethroid) or to differences in the excito-repellent effects of other pyrethroids. Excito-repellency is probably an important factor in the selection of pyrethroid resistance as it affects the duration of exposure of females to impregnated materials.
When
An. gambiae females were exposed to nets treated with permethrin at the concentration recommended by WHO (250 to 500 mg/m
2), the heterozygous ones (RS) were more efficiently killed than the susceptible ones (SS). Since
kdr resistance to the irritant effect appeared to be co-dominant while resistance to lethal effect was recessive [
21], the heterozygotes stayed longer than susceptible ones on the treated netting, picking up more insecticide and being killed in higher proportion. As a consequence, one can expect that permethrin treated nets are unlikely to select for pyrethroid resistance in areas where the
kdr mutation is rare and mostly present in the heterozygous state.
Although other studies in West Africa have consistently shown that pyrethroid treated nets remain effective against
kdr resistant
An. gambiae [
29,
31,
32], it is not possible to predict how long this effectiveness would be maintained should other resistance mechanisms appear. With indoor residual spraying in South Africa, an increased level of mixed function oxidase activity in
Anopheles funestus was associated with severe malaria control failure [
33]. The current spread of pyrethroid resistance in the major malaria vectors
An. gambiae and
An. funestus emphasizes the need to identify alternative insecticides and for the development and implementation of effective and sustainable resistance management strategies. Other experimental huts studies suggested that non-pyrethroid insecticides, such as organophosphates or carbamates, have potential for use on mosquito nets [
34,
35]. These chemicals are less excito-repellent than pyrethroids, and allow for a longer contact between mosquito and insecticide treated netting's, inducing higher mortality [
36]. Unfortunately, a cross resistance to both carbamates and organophospshorous insecticides involving an insensitive acetylcholinesterase has recently been detected in
An. gambiae from Côte d'lvoire [
37,
38] and may, therefore, impede their interest for ITNs in the concerned areas. Furthermore, recent evidence showed that carbosulfan-treated nets (300 mg/m
2) does select for resistance in the
An. gambiae population from the same area [
39], unlike pyrethroids with respect to
kdr. In the absence of new alternative insecticides for treatment of mosquito nets, the possible use of mixtures of insecticides or mosaic treatments [
40‐
42] should be closely investigated, as a possible strategy to maintain the effectiveness of ITNs and prevent the development of resistance. In areas where resistance genes are already present (
Kdr and
Ace.1
R
), ITNs are still the best option. Despite the presence of insensitive acetylcholinesterase at significant level conferring resistance to carbamates and organophosphates [
37], nets treated with these insecticides (carbosulfan and pirimiphos methyl respectively) were very effective in killing resistant mosquitoes and, in the case of carbosulfan, were also effective in preventing blood feeding [
35‐
40]. Recent studies have shown that another organophosphate (chlorpyrifos-methyl) is also effective and potentially safe enough to be considered as a possible alternative, either alone or in combination, for treatment of mosquito nets [
43,
44]. In contrast, in areas where malaria control is based on indoor residual spraying, resistance may lead to failure of control operations, although this is not yet confirmed in areas with
kdr or insensitive AChE resistance.