Vector bionomics and malaria transmission along the Thailand-Myanmar border: a baseline entomological survey.

Baseline entomological surveys were conducted in four sentinel sites along the Thailand-Myanmar border to address vector bionomics and malaria transmission in the context of a study on malaria elimination. Adult Anopheles mosquitoes were collected using human-landing catch and cow-bait collection in four villages during the rainy season from May-June, 2013. Mosquitoes were identified to species level by morphological characters and by AS-PCR. Sporozoite indexes were determined on head/thoraces of primary and secondary malaria vectors using real-time PCR. A total of 4,301 anopheles belonging to 12 anopheline taxa were identified. Anopheles minimus represented >98% of the Minimus Complex members (n=1,683), whereas the An. maculatus group was composed of two dominant species, An. sawadwongporni and An. maculatus. Overall, 25 Plasmodium-positive mosquitoes (of 2,323) were found, representing a sporozoite index of 1.1% [95%CI 0.66-1.50]. The transmission intensity as measured by the EIR strongly varied according to the village (ANOVA, F=17.67, df=3, P<0.0001). Our findings highlight the diversity and complexity of the biting pattern of malaria vectors along the Thailand-Myanmar border that represent a formidable challenge for malaria control and elimination.

Comparison of the Performances of Five Primer Sets for the Detection and Quantification of Plasmodium in Anopheline Vectors by Real-Time PCR.

Quantitative real-time polymerase chain reaction (qrtPCR) has made a significant improvement for the detection of Plasmodium in anopheline vectors. A wide variety of primers has been used in different assays, mostly adapted from molecular diagnosis of malaria in human. However, such an adaptation can impact the sensitivity of the PCR. Therefore we compared the sensitivity of five primer sets with different molecular targets on blood stages, sporozoites and oocysts standards of Plasmodium falciparum (Pf) and P. vivax (Pv). Dilution series of standard DNA were used to discriminate between methods at low concentrations of parasite and to generate standard curves suitable for the absolute quantification of Plasmodium sporozoites. Our results showed that the best primers to detect blood stages were not necessarily the best ones to detect sporozoites. Absolute detection threshold of our qrtPCR assay varied between 3.6 and 360 Pv sporozoites and between 6 and 600 Pf sporozoites per mosquito according to the primer set used in the reaction mix. In this paper, we discuss the general performance of each primer set and highlight the need to use efficient detection methods for transmission studies.

[Influence of plant matter and NPK fertilizer on the biology of Aedes aegypti (Diptera: Culicidae)]. / Influence des matières végétales et des engrais NPK sur la biologie de Aedes aegypti (Diptera : Culicidae).

In this laboratory study, we investigated the attractiveness of gravid Aedes aegypti females for aquatic habitats containing either NPK fertilizer or plant matter or a mixture of both. The development of larvae, adult emergence and weight of emerged adults were measured for each group and physicochemical analysis of water was made for the dosage of minerals and organic compounds. After 23 days experiment, NPK solution remained as attractive as the mixture of NPK + plant matter but did not ensure suitable development of larvae. The plant matter infusion showed less attractiveness for laying eggs than NPK but provided larvae with sufficient organic nutrients for their development. The combination of both NPK + plant matter provided the greatest attractiveness for gravid females and sufficient organic substance allowing mosquito larvae to grow. Physicochemical analyses of water containing NPK showed minerals only (NH4(+)+NO3(-)+P+K) whereas plant matter showed high content of carbon and nitrogen. The NPK + plant matter mixture contained both organic and minerals elements that favoured the proliferation of bacteria and then the development of mosquito larvae. These findings could lead to the development of new traps that could attract females mosquitoes and killed hatched larvae if mix with appropriate larvicides.

[Guidelines for phase III evaluation of vector control methods against malaria]. / Guide pour l'évaluation en Phase III de la lutte antivectorielle contre le paludisme.

Most new vector control methods against malaria involve the use of pesticides. Prior to release of these products for general use, their efficacy, persistence, and cross-resistance must be tested on mosquito colonies raised in the laboratory (phase I) then on wild mosquitoes in the field (small-scale), individual dwellings, or experimental huts (phase II). The goal of phase III studies is to evaluate the efficacy and effectiveness of the vector-control product or method against malaria in a population at regular risk for transmission. The main objective of phase III tests is to measure the epidemiologic impact, e.g. on the incidence or prevalence of malaria in humans. This article presents guidelines for carrying out phase III tests of vector-control methods against malaria (e.g. home insecticide spraying or insecticide-impregnated bednet use). It was written by participants in a workgroup formed to define recommendations for the WHOPES (WHO Pesticide Evaluation Scheme).

Aedes albopictus, an arbovirus vector: from the darkness to the light.

The Asian tiger mosquito, Aedes albopictus (Skuse, 1894), is an invasive species that can be found on all continents. The species, originally considered a secondary vector of viruses such as Dengue viruses, has recently been suggested to play a role in the transmission of Chikungunya virus in several countries bordering the Indian Ocean, Central Africa and Europe. Here we review the current geographic range and the relevant biological traits of A. albopictus in order to explain its rapid spread. We examine and discuss recent changes in its role as a vector, particularly in the transmission of arboviruses, and its importance in the current and future emergence of pathogens. Finally, we report conventional and innovative ways to control A. albopictus.

[Aedes aegypti oviposition in response to NPK fertilizers]. / Influence des engrais de type NPK sur l'oviposition d'Aedes aegypti.

Fertilizers are mineral associations intended to bring to the plants nutritive complements necessary to their growth. Modern fertilizers (NPK) combine the three basic elements which are the nitrogen (N), phosphorus (P) and potassium (K). In this study, we investigated in tunnel apparatus the influence of aqueous solutions containing low, moderate and high concentrations of NPK on the oviposition of Aedes aegypti. The results showed that the solutions containing moderate concentrations (NK = 17-33 mg/l and P = 23-47 mg/l) attracted significantly more gravid females than distilled water (P < 0.001). Conversely, the solutions containing either low or high concentrations of NPK (NK = 8 mg/l and P = 12 mg/l; NK = 50 mg/I and P = 70 mg/l) did not induce significant attraction (P > 0.05). These findings suggest that NPK fertilizers may influence the egg-laying behaviour of Ae. aegypti in field situations.

Multiple insecticide resistance mechanisms in Anopheles gambiae and Culex quinquefasciatus from Benin, West Africa.

Because free-insecticide treated net distribution is planned in Benin (West Africa) during the next few years, we investigated the type, frequency and distribution of insecticide resistance mechanisms in Anopheles gambiae and Culex quinquefasciatus mosquitoes in four localities selected on the basis of contrasting agricultural practices, use of insecticides and environment. Bioassays with WHO diagnostic test kits were carried out using pyrethroid, carbamate, organophosphate and organochlorine insecticides. An. gambiae mosquitoes were identified to species and to M or S molecular forms using PCR techniques. Molecular and biochemical assays were carried out to identify kdr and Ace.1 mutations in individual mosquitoes and to detect any increase in the activity of enzymes typically involved in insecticide metabolism (oxidase, esterase and glutathion-S-transférases). WHO diagnostic tests showed high frequency of resistance in An. gambiae and Cx. quinquefasciatus to permethrin and DDT in three areas. This was consistent with the presence of target site insensitivity due to kdr mutation and to increased metabolism through enzymatic activity. Kdr was expressed in both M and S forms. However, less than 1% of An. gambiae or Cx. quiqnuefasciatus showed the presence of the Ace.1(R) mutation. Carbamate/OP resistance was present at higher frequency in Culex than in An. gambiae. Dieldrin resistance was present in both species at all four localities. A higher frequency of pyrethroid-resistance was found in An. gambiae mosquitoes collected in urban areas compared to those collected in rice growing areas. The expansion of vegetable growing within urban areas probably contributed to selection pressure on mosquitoes. The detection of multiple resistance mechanisms in both An. gambiae and Cx. quinquefasciatus in Benin may represent a threat for the efficacy of ITNs and other forms of vector control such as indoor residual spraying in the future.

[Vectors of malaria: biology, diversity, prevention, and individual protection]. / Vecteurs du paludisme: biologie, diversité, contrôle et protection individuelle.

Only the Anopheles mosquitoes are implicated in the transmission of malaria. Among the numerous species of anopheles, around fifty are currently involved in the transmission. 20 are responsible for most of the transmission in the world. The diversity of behavior between species and in a single species of anopheles as well as climatic and geographical conditions along with the action of man on the environment condition the man vector contact level and the various epidemiological aspects of malaria. The anopheles are primarily rural mosquitoes and are less likely to be found in city surroundings in theory. But actually, the adaptation of some species to urban surroundings and the common habit of market gardening in big cities or in the suburbs is responsible for the de persistence of Anopheles populations in town. Except for South-East Asia, urban malaria has become a reality. The transmission risk of malaria is heterogeneous and varies with time. There is a great variation of risk within a same country, a same zone, and even within a few kilometers. The transmission varies in time according to seasons but also according to years and to the level of climatic events. For the traveler, prevention at any time relies on the strict application of individual protection, as well in rural than in urban surroundings.

Lethal and behavioural effects of three synthetic repellents (DEET, IR3535 and KBR 3023) on Aedes aegypti mosquitoes in laboratory assays.

The knock-down, mortality and 'irritancy' effects of three synthetic repellents (DEET, IR3535 and KBR 3023) on Aedes aegypti (L) (Diptera: Culicidae) were evaluated in the laboratory in the absence of animal bait. Filter paper tests were carried out to assess the knock-down effect (KDt(50) and KDt(95)) and mortality (LC(50) and LC(95)) induced by each repellent. 'Irritancy' tests were carried out to compare the flight response (time to first take-off, or FT) to increasing concentrations of repellents (2-7%) and at five distances from the treated surface (0-40 mm). DEET had an insecticidal effect (KDt(50) = 9.7 min at 7%; CL(50)= 1165 mg/m(2)), whereas IR3535 and KBR 3023 did not. Relative to an untreated control, IR3535 was an irritant (relative irritancy or RI > 1) at doses of 5% and 7% (RI = 17.7 and 9.9, respectively), whereas DEET was an irritant at lower concentrations (RI = 12.3 at 2% DEET). KBR 3023 was the weakest irritant over the same range of concentrations (RI(max) = 3.6 at 6%). DEET was more of an irritant (RI(20) = 9.4) than IR3535 (RI(20) = 2.9) over a range of distances (0-20 mm), and KBR 3023 was not an irritant unless mosquitoes made contact with the treated surface. All three repellents had a significant effect on mosquitoes, but DEET exhibited a more complex mode of action than the others due to its insecticidal properties. The repellents do not behave as a single class of compounds with a common mode of action, but most probably affect different physiological systems in insects. The physiological and molecular mechanisms of repellents, especially DEET, should be investigated to ensure a better use of these molecules for skin applications and/or for treating materials against mosquitoes.

[Aedes albopictus: chronical of a spreading vector]. / Aedes albopictus: chronique d'un vecteur expansionniste.

Over the last 50 years the Asian tiger mosquito Aedes (stegomyia) albopictus (Skuse) has spread to all continents in the old and new world. This anthropophilous species is able to adapt to most climates. Although long considered as a secondary disease vector, it has been shown to be competent for arbovirus transmission under laboratory conditions. In several locations that it has invaded, the tiger mosquito has played a major role in arbovirus transmission (dengue fever and chikungunya). A recent example is the outbreak of chikungunya on the Indian Ocean island of Reunion

[Effect of pyrethoid and organophosphate bi-treated nets on the behaviour of Kdr+ Anopheles gambiae mosquitoes]. / Comportement d'Anopheles gambiae Kdr+ face a des moustiquaires bi-imprégnées d'insecticides pyréthri- noïde et organophosphoré.

The efficacy of mosquito nets treated with bifenthrin (pyrethroid) and chlorpyrifos-methyl (organophosphate), alone and in mixture, was evaluated against the pyrethroid-resistant strain (VKPR) of Anopheles gambiae using the tunnel test technique. Results first showed that bifenthrin and chlorpyrifos-methyl kept a good excito-repellency effect against resistant mosquitoes despite the low doses applied on nets. With the mixture, irritancy significantly increased, which consequently limited the duration of exposure of females to impregnated surface. This phenomenon most probably explained the absence of synergism between bifenthrin and chlorpyrifos-methyl concerning the insecticidal activity of this mixture. This study demonstrated the importance of the mosquito behaviour in the global effectiveness of insecticide treated materials and in the development of alternative strategies for vector control. Concerning the prospective insecticide mixtures, not only will we have to consider the insecticidal properties of each insecticide selected, but also their excito-repellency which affects the time of duration of mosquitoes on the treated surface.

Synergism between permethrin and propoxur against Culex quinquefasciatus mosquito larvae.

To see if synergism occurs between carbamate and pyrethroid insecticides, we tested permethrin and propoxur as representatives of these two classes of compounds used for mosquito control. Larvicidal activity of both insecticides was assessed separately and together on a susceptible strain of the mosquito Culex quinquefasciatus (Diptera: Culicidae) by two methods. When mixed at a constant ratio (permethrin : propoxur 1 : 60 based on LC50) and tested at serial concentrations to plot dose/mortality regression, significant synergy occurred between them (co-toxicity coefficient = 2.2), not just an additive effect. For example, when the mixture gave 50% mortality, the same concentrations of permethrin and propoxur alone would have given merely 2 x 1% mortality. When a sublethal dose (LC0) of permethrin or propoxur was added to the other (range LC10-LC95), synergism occurred up to the LC80 level. Synergistic effects were attributed to the complementary modes of action by these two insecticide classes acting on different components of nerve impulse transmission. Apart from raising new possibilities for Culex control, it seems appropriate to consider using such mixtures or combinations for insecticide-treated mosquito nets in situations with insecticide-resistant Anopheles malaria vectors.

Resistance to carbosulfan in Anopheles gambiae from Ivory Coast, based on reduced sensitivity of acetylcholinesterase.

Resistance to carbosulfan, a carbamate insecticide, was detected in field populations of the malaria vector mosquito Anopheles gambiae Giles (Diptera: Culicidae) from two ecologically contrasted localities near Bouaké, Ivory Coast: rural M'bé with predominantly M form of An. gambiae susceptible to pyrethroids; suburban Yaokoffikro with predominantly S form of An. gambiae highly resistant to pyrethroids (96% kdr). The discriminating concentration of 0.4% carbosulfan (i.e. double the LC100) was determined from bioassays with the susceptible An. gambiae Kisumu strain. Following exposure to the diagnostic dosage (0.4% carbosulfan for 1 h), mortality rates of female An. gambiae adults (reared from larvae collected from ricefields) were 62% and 29% of those from M'bé and Yaokoffikro, respectively, 24 h post-exposure. Exposure for 3 min to netting impregnated with the operational dosage of carbosulfan 200 mg/m2 gave mortality rates of 88% of those from M'bé and only 12.2% for Yaokoffikro. In each case the control untreated mortality rate was insignificant. Biochemical assays to detect possible resistance mechanism(s) revealed the presence of insensitive AChE in populations of An. gambiae at both localities, more prevalent in the S form at Yaokoffikro than in M form at M'bé, as expected from bioassays results. Our study demonstrates the need to monitor carbamate resistance among populations of the An. gambiae complex in Africa, to determine its spread and anticipate vector control failure if these insecticides are employed.

Efficacy of mosquito nets treated with a pyrethroid-organophosphorous mixture against Kdr- and Kdr+ malaria vectors (Anopheles gambiae).

In order to prevent the resistance of Anopheles gambiae s.l. to pyrethroids from spreading too quickly and to lengthen the effectiveness of insecticide impregnated mosquito nets, it has recently been suggested to use mixtures of insecticides that have different modes of action. This study presents the results obtained with tulle mosquito nets treated with bifenthrin (a pyrethroid) and chlorpyrifos-methyl (an organophosphorous) both separately and in mixture on two strains of An. gambiae, one sensitive to all insecticides, and the other resistant to pyrethroids. The values of KDt50 and KDt95 and the mortality induced with the mixture of bifenthrin (25 mg/m2) and chlorpyrifos-methyl (4.5 mg/m2) show a significant synergistic effect on the strain of An. gambiae susceptible to insecticides. However, the tested combination does not induce any synergistic effect on the VKPR strain selected with permethrin, but only enhances the effectiveness of the two insecticides taken separately.

Efficacy of mosquito nets treated with insecticide mixtures or mosaics against insecticide resistant Anopheles gambiae and Culex quinquefasciatus (Diptera: Culicidae) in Côte d'Ivoire.

Only pyrethroid insecticides have so far been recommended for the treatment of mosquito nets for malaria control. Increasing resistance of malaria vectors to pyrethroids threatens to reduce the potency of this important method of vector control. Among the strategies proposed for resistance management is to use a pyrethroid and a non-pyrethroid insecticide in combination on the same mosquito net, either separately or as a mixture. Mixtures are particularly promising if there is potentiation between the two insecticides as this would make it possible to lower the dosage of each, as has been demonstrated under laboratory conditions for a mixture of bifenthrin (pyrethroid) and carbosulfan (carbamate). The effect of these types of treatment were compared in experimental huts on wild populations of Anopheles gambiae Giles and the nuisance mosquito Culex quinquefasciatus Say, both of which are multi-resistant. Four treatments were evaluated in experimental huts over six months: the recommended dosage of 50 mg m(-2) bifenthrin, 300 mg m(-2) carbosulfan, a mosaic of 300 mg m(-2) carbosulfan on the ceiling and 50 mg m(-2) bifenthrin on the sides, and a mixture of 6.25 mg m(-2) carbosulfan and 25 mg m(-2) bifenthrin. The mixture and mosaic treatments did not differ significantly in effectiveness from carbosulfan and bifenthrin alone against anophelines in terms of deterrency, induced exophily, blood feeding inhibition and overall mortality, but were more effective than in earlier tests with deltamethrin. These results are considered encouraging, as the combination of different classes of insecticides might be a potential tool for resistance management. The mixture might have an advantage in terms of lower cost and toxicity.

Insecticide mixtures for mosquito net impregnation against malaria vectors.

Insecticides belonging to the pyrethroid family are the only compounds currently available for the treatment of mosquito nets. Unfortunately, some malaria vector species have developed resistance to pyrethroids and the lack of alternative chemical categories is a great concern. One strategy for resistance management would be to treat mosquito nets with a mixture associating two insecticides having different modes of action. This study presents the results obtained with insecticide mixtures containing several proportions of bifenthrin (a pyrethroid insecticide) and carbosulfan (a carbamate insecticide). The mixtures were sprayed on mosquito net samples and their efficacy were tested against a susceptible strain of Anopheles gambiae, the major malaria vector in Africa. A significant synergism was observed with a mixture containing 25 mg/m2 of bifenthrin (half the recommended dosage for treated nets) and 6.25 mg/m2 of carbosulfan (about 2% of the recommended dosage). The observed mortality was significantly more than expected in the absence of any interaction (80% vs 41%) and the knock-down effect was maintained, providing an effective barrier against susceptible mosquitoes.