MAJOR ARTICLE
Interplay Between Plasmodium Infection and
Resistance to Insecticides in Vector Mosquitoes
Haoues Alout,1,2,a Bienvenue Yameogo,2 Luc Salako Djogbénou,3 Fabrice Chandre,1 Roch Kounbobr Dabiré,2
Vincent Corbel,1,4 and Anna Cohuet1,2
1
Institut de Recherche pour le Développement, Maladies Infectieuses et Vecteurs, Ecologie, Génétique, Evolution et Contrôle, UM1-UM2-CNRS5290-IRD
224, Montpellier, France; 2Institut de Recherche en Sciences de la Santé, Bobo-Dioulasso, Burkina Faso; 3Institut Régional de Santé Publique/Université
d’Abomey-Calavi, Cotonou, Bénin; and 4Department of Entomology, Faculty of Agriculture, Kasetsart University, Bangkok, Thailand
Despite its epidemiological importance, the impact of insecticide resistance on vector-parasite interactions and
malaria transmission is poorly understood. Here, we explored the impact of Plasmodium infection on the level
of insecticide resistance to dichlorodiphenyltrichloroethane (DDT) in field-caught Anopheles gambiae sensu
stricto homozygous for the kdr mutation. Results showed that kdr homozygous mosquitoes that fed on infectious blood were more susceptible to DDT than mosquitoes that fed on noninfectious blood during both
ookinete development (day 1 after the blood meal) and oocyst maturation (day 7 after the blood meal) but not
during sporozoite invasion of the salivary glands. Plasmodium falciparum infection seemed to impose a fitness
cost on mosquitoes by reducing the ability of kdr homozygous A. gambiae sensu stricto to survive exposure to
DDT. These results suggest an interaction between Plasmodium infection and the insecticide susceptibility of
mosquitoes carrying insecticide-resistant alleles. We discuss this finding in relation to vector control efficacy.
Keywords. malaria; cost of infection; Anopheles gambiae; insecticide exposure; kdr; insecticide resistance; Plasmodium falciparum.
Human malaria is caused by a parasitic protist of the
genus Plasmodium, which is transmitted through the
bites of infected mosquitoes of the Anopheles genus.
Plasmodium parasites are dependent on completing a
complex life cycle in the Anopheles mosquito vector
for transmission to occur. Thus, reducing the mosquito
abundance or interfering with its ability to support the
parasite cycle can interrupt malaria transmission. Consequently, malaria control mainly relies on targeting the
mosquito vector by using insecticides, but its efficacy
has been challenged by the emergence of insecticide resistance [1]. Insecticide resistance in the main malaria
Received 25 February 2014; accepted 28 April 2014; electronically published 14
May 2014.
a
Present affiliation: Arthropod-Borne Infectious Diseases Laboratory, Department
of Microbiology, Immunology, and Parasitology, Colorado State University, Fort
Collins.
Correspondence: Haoues Alout, PhD, Colorado State University, AIDL, 1692 Campus delivery, Fort Collins, CO 80523-1692 ([email protected]).
The Journal of Infectious Diseases® 2014;210:1464–70
© The Author 2014. Published by Oxford University Press on behalf of the Infectious
Diseases Society of America. All rights reserved. For Permissions, please e-mail:
[email protected].
DOI: 10.1093/infdis/jiu276
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vector species, such as Anopheles gambiae, is spreading
dramatically in Africa [2]. Insecticides impose intense
selection pressure on mosquito populations as a result
of crop protection practices and increasing coverage
of disease vector controls required for public health
purposes [3]. Two main mechanisms have been described for resistance: target site mutations and enhanced metabolic detoxification. The molecular basis
of target site insensitivity has been characterized in
many insect species [3] and has demonstrated conserved resistant mutations across insect vectors. Point
mutations in the gene coding for the voltage-gated sodium channel, named kdr for knockdown resistance,
confer resistance to pyrethroids and dichlorodiphenyltrichloroethane (DDT) insecticides [4, 5]; and mutations in
the ace-1 gene, which encodes the acetylcholinesterase
enzyme, confer cross-resistance to carbamate and organophosphate insecticides [6]. Metabolic resistance is
the result of elevated levels of detoxifying enzymes
such as cytochrome P450 monooxygenases, esterases,
or glutathione-S-transferases [3]. Currently, insecticide
resistance is widespread, and multiple mechanisms are
selected together as a result of increasing insecticide
selective pressure. Because resistant mosquitoes are the only
mosquitoes able to survive in the presence of lethal dose of insecticides, pathogen transmission is ensured mostly by resistant
vectors in areas where vector control is implemented.
The selection of insecticide resistance in mosquito vectors is
thought to interfere with the development and transmission of
parasites because of pleiotropic effects on vector longevity, vector competence, and vector feeding behavior [7]. In agreement
with this, we recently demonstrated the impact of insecticide resistance on parasite infection in mosquitoes, using the natural
system A. gambiae–Plasmodium falciparum; target-site mutations responsible for insecticide resistance (ace-1 G119S and
kdr L1014F) increased the prevalence of P. falciparum infection
in A. gambiae sensu stricto and probably its transmission
through increased sporozoite prevalence [8]. However, no investigations have been made on the impact of infection on the
susceptibility to insecticide in this epidemiologically relevant
vectorial system.
P. falciparum sets up complex interactions and undergoes
various developmental stages before the sporozoites reach the
salivary glands of the mosquito vector and can be transmitted
to human hosts. Although the cost of infection in natural vector-parasite associations is still in debate, development of the
Plasmodium parasite is thought to reduce the overall fitness of
its arthropod vector [9] because of mosquito cell damage when
parasites cross the midgut epithelium [10, 11] or the cost of immune system activation [12]. Indeed, Plasmodium parasites are
associated with increased oxidative stress in the mosquito midgut because of the production of reactive oxygen species (ROS)
[13]. These ROS enhance immunity but are also detrimental to
the mosquitoes, which respond by modifying expression of
detoxifying enzymes [14, 15]. Insecticides, similar to other xenobiotics, induce a detoxification response through the overexpression of specific enzymes involved in binding, transport, and
breakdown of exogenous compounds, resulting in an increase in
tolerance [16]. This detoxification response is also observed in
mosquitoes carrying mutations responsible for insecticide resistance [17]. Because enzymatic detoxification is implicated in
both response to infection and insecticide exposure, we hypothesized that Plasmodium infection could then impact the susceptibility to insecticides. Previous works have demonstrated
the interaction of insecticide exposure and infection on host
life history traits [18, 19]. For example, infection with entomopathogenic fungi increases insecticide-induced mortality
among resistant mosquitoes [18].
Therefore, in this study, we aimed to determine whether P.
falciparum infection influences the level of insecticide resistance
in wild-caught A. gambiae. Female mosquitoes were allowed to
feed on control or gametocyte-infected blood, and mortality
due to 4% DDT was tested at various times following blood
feeding, corresponding to different parasite stages of the Plasmodium sporogony.
MATERIALS AND METHODS
Ethical Statement
Ethical approval was obtained from the Centre Muraz Institutional Ethics Committee under the ethical clearance number
003-2009/cE-cM. All human volunteers were enrolled after receipt of written informed consent from the participant and/or
their legal guardians.
Mosquito Sampling
Anopheline larvae were sampled in Soumousso village, located
40 km southeast of Bobo-Dioulasso, Burkina Faso, during the
rainy season (from August to September) in 2011 and 2012.
This site was selected because anopheline mosquitoes are highly
resistant to DDT. During the rainy season, this site is composed
predominantly of Anopheles gambiae S molecular form, now
called A. gambiae sensu stricto [40], with a high frequency of
the kdr allele ( fkdr = 0.77), a low frequency of the ace-1R allele
( face-1R = 0.15), and absence or low levels of amplified detoxification enzymes [21]. Larvae were collected from typical A. gambiae sensu lato breeding sites, including gutters, swallow wells,
and pools of standing water. Larvae were brought back to the
insectary and reared to adults. Emerging adults were morphologically identified using standard identification keys as A. gambiae sensu lato females and were further used for blood feeding.
P. falciparum Experimental Infection by Membrane
Feeding Assay
Membrane feeding assays were performed as described by Alout
et al [8]. Briefly, P. falciparum gametocyte carriers were selected
by examining thick blood smears of blood samples from children aged 5–11 years from 2 villages in southwestern Burkina
Faso (Dandé and Soumousso, located 60 km north and 40 km
southeast, respectively, of Bobo-Dioulasso). Children with a gametocyte density of >20 gametocytes/µL of blood were selected,
and a venous blood sample (8 mL) was taken after a second
measure of the gametocyte density for confirmation. Blood
serum was replaced with European naive AB serum to limit
the potential effect of human transmission blocking immunity
[39]. Reconstituted blood samples were divided in 2 batches,
from which one was used directly as infectious blood, and the
other was heated at 42°C for 15 minutes as noninfectious control blood. This treatment results in the inactivation of gametocytes [41, 42] and does not affect the fitness of mosquitoes [43].
Membrane feeders were filled with 500 µL of reconstituted
blood and maintained at 37°C by water jackets. Female mosquitoes aged 3–5 days were allowed to feed separately on infected or
heat-inactivated (control) blood through a Parafilm membrane
for up to 30 minutes. A total of 2–3 feeders were used for each
blood type (infectious or control), to limit potential feeder effect. Unfed female mosquitoes were discarded, and only fully
fed mosquitoes were maintained in a large cage (area,
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30 × 30 × 30 cm) under standard insectary conditions on a 5%
sucrose solution. This procedure was repeated 9 times, with
each feeding assay using a different gametocyte-infected blood.
Insecticide Exposure
Insecticide exposure was performed on female mosquitoes, using
the World Health Organization standard tube protocol [44].
Adult females were exposed for 1 hour to 4% DDT-impregnated
paper, a discriminating concentration that kills all individuals
that do not carry alleles conferring insecticide resistance [44].
Exposure to insecticides was performed on days 1, 7, and 14
after the blood meal. For each replicate, a batch of adult females
was exposed to a nonimpregnated paper to ensure that natural
mortality was <10%. Mortality was recorded 24 hours after insecticide exposure. Mosquitoes that died and those that survived
after insecticide exposure were then stored separately on silica
gel for subsequent DNA extraction and polymerase chain reaction (PCR) analysis.
Molecular Analysis
Genomic DNA was extracted using the CTAB protocol [45]
from all individual A. gambiae sensu lato mosquitoes that survived or died after DDT exposure. They were identified to the
species and molecular level by PCR, as described elsewhere
[46]. The presence of the L1014F kdr-west mutation, responsible for resistance to DDT, was determined using the diagnostic
test of Martinez-Torres et al [4]. P. falciparum infection was determined by PCR with Pf1/Pf2 primers [47] for females that
were exposed to DDT on day 7 and 14 after the blood meal.
Statistical Analysis
To analyze the effect of P. falciparum infection on resistance
phenotype, the mortality rate 24 hours after insecticide exposure was used as the response variable. Here, only mosquitoes
belonging to A. gambiae sensu stricto and identified as homozygous for the kdr mutation ( predominant in the studied sample) were included, to avoid confounding effects (n = 839
among 9 feeding assays). We used 4 explanatory variables:
blood meal (a 2-level categorical variable, defined as infectious
or control), donor (a categorical variable, with each gametocyte
carrier representing a level), gametocyte density of the blood
donor (a numerical variable), and day (a numerical variable, defined as the day after the blood meal when mosquitoes were exposed to insecticide). To explore the impact of P. falciparum
infection in detail, mosquitoes that were not infected following
an infectious blood meal (ie, P. falciparum–negative PCR result)
were removed, and only those positive for P. falciparum infection were compared to the control group. Therefore, only those
that were exposed to DDT on day 7 and day 14 after the blood
meal were included (n = 556 among 7 feeding assays) because
we could not determine by PCR on day 1 after the blood
meal whether mosquitoes were infected, because of the potential
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Alout et al
presence of parasite DNA in the remaining blood in the midgut.
A second analysis was then performed, in which the blood meal
variable was replaced by the infection variable (a 2-level categorical variable, defined as infected or control). Both maximal models included the variables blood meal or infection, gametocyte
density, and day and their interactions as fixed effects. The
donor variable was used as a random variable to account for
the correlation between individuals from the same feeding
assay. Statistical analyses were performed with Statistical Analysis
Software (SAS Institute, Cary, NC). For each analysis, the relevance of the random structure was compared with a generalized
linear model with no random effect, based on the lowest Akaike
information criterion. The mortality rate was analyzed using the
GLIMMIX procedure with a binomial error structure.
RESULTS
Species Identification and kdr Genotyping
Of the 888 blood-fed female mosquitoes exposed to DDT over 9
replicates, 872 were identified as A. gambiae sensu stricto, and
16 were identified as Anopheles arabiensis. Molecular diagnosis
of the kdr mutation revealed that 839 A. gambiae sensu stricto
mosquitoes (>96%) were homozygous for the L1014F mutation;
13 were heterozygous and 10 were homozygous for the susceptible allele. Only 1 of 16 A. arabiensis specimens was heterozygous for the kdr mutation; all others were homozygous for the
susceptible allele (Table 1). Regardless of species, all mosquitoes
heterozygous or homozygous for the susceptible allele died after
insecticide exposure. Thus, we analyzed only individuals homozygous for the kdr mutation, which included only A. gambiae
sensu stricto (n = 839 females) and no A. arabiensis mosquitoes.
Impact of Plasmodium Infection on Insecticide Resistance
Our first analysis looked at the influence of the presence of infectious gametocytes in the blood but not at the outcome of infection in mosquitoes (Table 2). The type of blood meal
(infectious or not), day of exposure, and the interaction of
both variables had a significant effect on DDT-associated
Table 1. Frequency of kdr Genotypes in Anopheles gambiae
Sensu Stricto and Anopheles arabiensis Mosquitoes Collected
in Soumousso, Burkina Faso
kdr Genotype,a No. (f b )
Anopheles Species
RR
RS
SS
Total, No.
(f b)
A. gambiae sensu
stricto
A. arabiensis
839 (0.96)
23 (0.03)
10 (0.01)
872 (1)
1 (0.06)
15 (0.94)
16 (1)
a
0
RR and SS indicate mosquitoes homozygous for the resistant allele and the
susceptible allele, respectively, and RS indicates mosquitoes heterozygous for
the kdr mutation.
b
f: frequency
Table 2. Insecticide Bioassay Results for Anopheles gambiae
Sensu Stricto Mosquitoes Exposed to an Infectious or
Noninfectious (Control) Blood Meal
Day
After
Blood
Meal
Infectious
Blood
Meal
Sample
Size,
No.
Death,
No.
Mortality Rate,a
% ± SEM
P
Value a
1
No
94
12
12.8 ± 3.44
1
Yes
120
38
31.7 ± 4.24
.002
7
7
No
Yes
149
145
13
41
8.7 ± 2.31
28.3 ± 3.74
<.001
14
No
185
85
45.9 ± 3.66
14
Yes
179
90
50.3 ± 3.74
.470
Abbreviation: SEM, standard error of the mean.
a
Measured 24 hours after a 1-hour exposure to 4% dichlorodiphenyltrichloroethane (DDT). Mosquitoes were exposed to DDT at different times after the
blood meal.
By the χ2 test, for comparisons of mortality among mosquitoes exposed to
infectious blood meal vs those exposed to a control blood meal.
b
mortality (Table 3). Regardless of the blood meal type, DDTassociated mortality changed depending on the day of exposure.
Mortality increased significantly between day 7 and day 14 after
the blood meal (mean mortality [±standard error of the mean
{SEM}], 13.02% ± 7.98% and 60.09% ± 14.88%, respectively;
P = .0143) but not between day 1 and day 7 after the blood
meal (16.09% ± 9.53% and 13.02% ± 7.98%, respectively;
P = .804). Across all days of exposure, ingestion of an infectious
blood meal increased DDT-associated mortality (mean mortality [±SEM], 17.17% ± 5.89% to 37.27% ± 9.27%; P < .001). Mortality among young noninfected mosquitoes (12.8% on day 1
after the blood meal) is consistent with previously described
DDT-associated mortality among kdr A. gambiae sensu stricto
mosquitoes from the same sample site in 2010 [20, 21]. On day 1
and day 7 after the blood meal, DDT-associated mortality was
greater among female mosquitoes that took an infectious blood
meal, compared with those that fed on heat-inactivated blood
(mean mortality [±SEM], 31.7% ± 4.24% for the infectious
group and 12.8% ± 3.44% for the control group on day 1
[P = .004] and 28.3% ± 3.74% for the infectious group and
8.7% ± 2.31% for the control group on day 7 [P < .001]).
Table 3. Statistical Analysis of DichlorodiphenyltrichloroethaneInduced Mortality in Relation to the Blood Meal Type
Source
Infectious blood meal
Day
Infectious blood
meal × day
df,
df,
Numerator Denominator
1
2
2
823
823
823
F
When mosquitoes were exposed to insecticide on day 14 after
the blood meal, there was no difference in DDT-associated
mortality between blood meal types (mean mortality [±SEM],
50.3% ± 3.74% for the infectious group and 45.9% ± 3.66% for
the control group [P = .2536]; Figure 1).
We ran a second analysis in which only females positive for
P. falciparum infection were compared to females that fed on a
noninfectious blood (infected vs control). The data consisted
only of females that were exposed on day 7 and day 14 after the
blood meal (n = 556), as we were not able to confirm the success
of infection on day 1 after the blood meal. The results of this
second analysis agreed with those of the first analysis (Table 4).
Overall, the DDT-associated mortality among infected mosquitoes was greater than that among noninfected mosquitoes (mean
mortality [±SEM], 50.14% ± 14.0% and 22.55% ± 9.88%, respectively; P < .001) and was greater on day 14 after the blood meal
than on day 7 after the blood meal (mean mortality [±SEM],
64.55% ± 16.70% and 13.86% ± 9.79%, respectively; P = .0277).
When mosquitoes were exposed on day 7 after the blood meal,
DDT-associated mortality among infected mosquitoes was greater than that among control mosquitoes (mean mortality [±SEM],
Table 4. Statistical Analysis of DichlorodiphenyltrichloroethaneInduced Mortality in Relation Plasmodium falciparum Infection
df,
Numerator
df,
Denominator
F
P Value
Plasmodium infection
1
545
24.11
<.001
Day
Infection × day
2
2
545
545
4.87
6.29
P Value
24.55 <.001
3.79
4.23
Figure 1. Dichlorodiphenyltrichloroethane (DDT)–induced mortality
among blood-fed female mosquitoes. The mortality rate among Anopheles
gambiae sensu stricto females that fed on gametocyte-infected blood (red)
or heat-inactivated blood (blue) was recorded 24 hours after DDT exposure
at different times after the blood meal. Bars above and below the means
represent standard errors of the mean. **P < .01, ***P < .001. Abbreviation: NS, nonsignificant.
.023
.0148
Significance of variables obtained after selection of the minimal mixed effect
model is presented.
Source
.0227
.0125
Significance of variables obtained after selection of the minimal mixed effect
model is presented.
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Figure 2. Dichlorodiphenyltrichloroethane (DDT)–induced mortality
among Plasmodium falciparum–infected and control female mosquitoes.
The mortality rate among P. falciparum–positive (red) and noninfected control (blue) Anopheles gambiae sensu stricto females was recorded 24 hours
after DDT exposure at different times after the blood meal. Bars above and
below the means represent the standard errors of the mean. ***P < .001.
Abbreviation: NS, nonsignificant.
29.1% ± 14.3% for the infected group and 5.9% ± 5.2% for the
control group; P < .001), but the difference was not significant
on day 14 after the blood meal (mean mortality [±SEM],
71.2% ± 14.6% for the infected group and 57.3% ± 17.5% for
the control group; P = .304) as illustrated in Figure 2.
DISCUSSION
In this study, we investigated the impact of Plasmodium infection on the insecticide resistance phenotype of wild-caught A.
gambiae. We determined the impact of different stages of Plasmodium development on the ability of kdr homozygous mosquitoes to survive DDT exposure. The results clearly showed
that kdr mosquitoes that fed on infectious blood were less
able to survive after DDT exposure during ookinete development (day 1 after the blood meal) and oocyst maturation
(days 7 after the blood meal), compared with those that fed
on control blood. These results were confirmed when comparing only P. falciparum–positive individuals on day 7 after the
blood meal to noninfected controls. However, DDT-induced
mortality was not different between sporozoite-infected and
control mosquitoes on day 14 after the blood meal. As reported
in other studies, the level of insecticide resistance decreases with
age in field-collected and laboratory-colonized Anopheles mosquitoes [22, 23]. This suggests that the age of mosquitoes has a
greater impact on insecticide susceptibility than sporozoite infection (ie, day 14 after the blood meal).
The results presented here contrast with that of Anopheles
stephensi infected by Plasmodium chabaudi, for which no increase of mortality was observed after permethrin exposure
[24]. However, several experimental differences may account
for this discrepancy, such as the use of different vector-parasite
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Alout et al
combination, the use of an insecticide-susceptible strain with
no history of insecticide exposure, or the use of a different
insecticide ( permethrin vs DDT). Additionally, an important
difference is the presence of insecticide-resistant alleles
(L1014F) in our sampled mosquitoes. Also, these collected samples may have experienced insecticide exposure in the field as
larvae or in previous generations, which could affect various
life history traits, particularly vector competence [25, 26]. Previous exposures of kdr homozygous mosquitoes to insecticides
may have induced a detoxification response resulting in a variable insecticide resistance phenotype. Although we did not test
for metabolic resistance mechanisms in the sampled mosquitoes, glutathione-S-transferases may be present in mosquitoes
from the village where they were collected [21].
Our results show that Plasmodium infection increases DDTassociated mortality of kdr-resistant mosquitoes during the first
week after infection (ie, from day 1 to day 7 after the blood
meal) but not later. This suggests a trade-off between mounting
an effective immune response and surviving DDT exposure.
About 18–24 hours after the infectious blood meal, ookinetes
cross the midgut epithelium and trigger the activation of the
mosquito immune system [48], which is mediated in part by
the production and detoxification of ROS in the mosquito midgut [13, 14]. Plasmodium infection has been shown to decrease
the expression of a large number of genes coding detoxification
enzymes at the ookinete stage, particularly cytochrome P450
and the glutathione-S-transferases [15]. This may explain the
increased susceptibility to DDT in our experiment. Thus, we
hypothesize that the change in expression of detoxification
genes induced by infection would lead to a trade-off between
the control of ROS levels and the elimination of insecticides.
This would trigger immune defenses against P. falciparum that
may induce a cost on the ability of kdr-homozygous A. gambiae
to survive insecticide exposure. In line with this hypothesis, Farenhorst et al [18] have demonstrated that fungal infection increases
insecticide susceptibility of resistant mosquitoes that is probably
due to a reallocation of insecticide-detoxifying enzymes toward
fungal toxins. An alternative hypothesis might be that insecticide
exposure affects the susceptibility to infection, leading to a greater
Plasmodium-induced mortality rate, which is consistent with the
immunotoxic effect of insecticides in vertebrates [27, 28].
The main method to control malaria transmission is through
insecticide, as it reduces the longevity of the vector, the most important parameter of the vectorial capacity for a mosquito population [29]. Insecticide resistance limits the efficacy of vector
control, as mosquitoes are able to survive high doses of insecticide. Alleles responsible for insecticide resistance may also impact
other life history traits of the vectors, including vector competence and other parameters of the vectorial capacity, such as biting frequency, and vector density [7]. We recently showed that
target-site resistant mutations affect vector competence, particularly the kdr mutation, which increases the probability of
P. falciparum infection and decreases the intensity of infection in
A. gambiae [8]. Therefore, insecticide resistance would lead to an
increase of malaria transmission. However, because of the widespread application of insecticides for agricultural and public
health purposes, insecticide-resistant mosquitoes are likely to
be in contact with insecticides several times during their feeding
cycles. Mosquitoes carrying insecticide-resistant alleles will be
more susceptible to insecticides as they get older [22, 23], and
Plasmodium infection may even reduce their insecticide resistance level. This might keep malaria vector control efficient, as
older and infected females would be killed more easily even in
insecticide-resistant populations. Consistently, by modeling the
effect of late-life-acting insecticides, Read et al showed that an
age-dependent biocide (ie, an insecticide that kills disproportionally older mosquitoes) would keep vector control efficient while
slowing the evolution of resistance [30]. Moreover, if this insecticide is more effective against Plasmodium-infected mosquitoes,
the evolution of resistance would be further slowed.
While insecticides affect directly the vectorial capacity of a
mosquito population and, probably, pathogen transmission
through the longevity of mosquitoes and their vector competence
(ie, the ability to support parasite development) [31], they also
have indirect effects through the selection of insecticide resistance
mechanisms [8]. Indeed, declined insecticide concentrations over
time on insecticide-treated surfaces would lead to multiple exposures to sublethal doses among resistant populations. This could
induce the production of detoxification enzymes in the mosquito,
affecting vector competence [7, 14]. Last, other ecological/environmental factors may influence the insecticide resistance phenotype, such as humidity, temperature [32], larval diet [33], or
urban pollutant [34]. For example, susceptibility to organophosphate insecticides in natural populations of A. aegypti increases
with increasing larval rearing temperature [32]. Taken together,
these observations reveal the complex ecological consequence of
insecticide application on the selection of resistance and on malaria transmission. A better understanding of ecological and environmental factors affecting mosquito resistance to pyrethroids
may thus represent a way to better manage vector population and
insecticide resistance.
In conclusion, we provide evidence that P. falciparum infection
can reduce the level of resistance to DDT among field-caught
mosquitoes homozygous for the kdr mutation. Insecticides that
are no longer active on younger kdr homozygous mosquitoes
may be still partially efficient in controlling disease transmission
because infected mosquitoes carrying insecticide-resistant alleles
have less chance to survive after insecticide exposure. This
might explain why selection of insecticide resistance in A. gambiae
by operational control was not associated with an increase in malaria prevalence. Indeed, only 1 case of control failure due to
pyrethroid resistance, in South Africa during 2000, has been reported [35]. Data from other studies suggest that insecticide resistance may have a smaller effect than expected on malaria control,
such as in Bioko Island, where the frequency of kdr increases but
transmission index and malaria cases decrease [2]. In Burundi and
Cote d’Ivoire, distribution of insecticide-treated nets led to a reduction in malaria incidence despite a high kdr frequency
(>80%) [36, 37]. Future studies in various ecological settings
should help to determine the impact of both insecticide resistance
and vector control programs on malaria incidence. These results
support the importance of environmental stresses (ie, insecticides)
on the ability of mosquitoes to transmit Plasmodium parasites
[38] and emphasizes the need to better understand the relationships between insecticide selection, insecticide resistance, and
Plasmodium infection, which is critical for effective implementation of vector control strategies.
Notes
Acknowledgments. We thank the participating children and their parents, for their involvement in this study; the local authorities, for their support; the IRSS staff in Burkina Faso, for technical assistance; and the
Laboratoire Mixte International LAMIVECT in Bobo-Dioulasso, Burkina
Faso, for technical support.
Financial support. This work was supported by the European Community’s Seventh Framework Program FP7/2007-2013 (grants 242095
and 223736) and the Institut de recherche pour le developpement (financial
support and fellowship to H. A.).
Potential conflicts of interest. All authors: No reported conflicts.
All authors have submitted the ICMJE Form for Disclosure of Potential
Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
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