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Studies done in Bouaké (Côte d’Ivoire) about 20-yr ago reported that Anopheles gambiae s.l. Giles was the major malaria vector. The present study aimed to update these data and to identify the main vectors. Mosquitoes were collected in Allokokro and Petessou villages between June 2014 and December 2015 using the human landing catching method. Potential breeding sites of An. gambiae s.l. were identified in August and October 2014 and mapped using GPS. Anopheles species were morphologically and molecularly [polymerase chain reaction (PCR)] identified. Ovaries of female were dissected to determine the parity and infection with Plasmodium was detected in head and thorax by quantitative PCR. In Allokokro, the biting rate of An. gambiae s.s was significantly greater than Anopheles coluzzii, whereas, in Petessou, biting rates of both species were comparable. Plasmodium falciparum (Haemosporida: Plasmodiidae), Plasmodium malariae (Haemosporida: Plasmodiidae), and Plasmodium ovale (Haemosporida: Plasmodiidae) identified in both villages. The infection rates of An. gambiae s.s. and An. coluzzii were not significantly different. The entomological inoculation rate (EIR) of An. gambiae s.s. for P. falciparum was 9-fold greater than that of An. coluzzii in Allokokro; however, in Petessou, the EIRs of both species were comparable. In both village, An. gambiae s.s was responsible for P. falciparum and P. ovale transmission whereas An. coluzzii transmitted all three Plasmodium species.

International audience; Japanese Encephalitis (JE) is the most important cause of human encephalitis in Southeast Asia, and this zoonosis is mainly transmitted from pigs to human by mosquitoes. A better understanding of the host-feeding preference of Japanese encephalitis virus (JEV) major vectors is crucial for identifying risk areas, defining bridge vector species and targeting adapted vector control strategies. To assess host-feeding preference of JE vectors in a rural Cambodian area where JE is known to circulate, in 2017, we implemented four sessions of mosquito trapping (March, June, September, December), during five consecutive nights, collecting four times a night (6 p.m. to 6 a.m.), and using five baited traps simultaneously, i.e., cow, chicken, pig, human, and a blank one for control. In addition, blood meals of 157 engorged females trapped at the same location were opportunistically analyzed with polymerase chain reaction (PCR), using cow, pig, human, and dog blood primers. More than 95% of the 36,709 trapped mosquitoes were potential JE vectors. These vectors were trapped in large numbers throughout the year, including during the dry season, and from 6 p.m. to 6 a.m. Despite the apparent host-feeding preference of Culex vishnui, Cx. gelidus, and Cx. tritaenhyorhincus for cows, statistical analysis suggested that the primary target of these three mosquito species were pigs. Dog blood was detected in eight mosquitoes of the 157 tested, showing that mosquitoes also bite dogs, and suggesting that dogs may be used as proxy of the risk for human to get infected by JE virus.

International audience; Simple Summary: Aedes albopictus is a well-established competent arbovirus vector in Reunion Island, a French overseas territory in the Indian Ocean, occurring in a range of natural to urban environments where it represents a major threat to public health. Following the 2006 Chikungunya outbreak and periodic occurrence of dengue epidemics, the sterile insect technique (SIT) emerged as the most environment-friendly option for integration with the current vector control strategy that relies mainly on the elimination of breeding sites and insecticide applications. This paper describes the trajectory that has been followed in assessing the feasibility of SIT against Ae. albopictus in Reunion Island, and reviews some of the main achievements since 2009. These include essential scientific information so far obtained on the biology and ecology of Ae. albopictus, and the development of the requisite technological capabilities for the production and release of sexually competitive sterile males. Furthermore, it also draws attention to the strategies established to streamline the decision-making process, including an awareness campaign to enhance public understanding, efforts to secure public acceptance and regulatory validation of SIT pilot testing for small-scale suppression of wild Ae. albopictus in selected urban sites on the island. Abstract: The global expansion of Aedes albopictus, together with the absence of specific treatment and vaccines for most of the arboviruses it transmits, has stimulated the development of more sustainable and ecologically acceptable methods for control of disease transmission through the suppression of natural vector populations. The sterile insect technique (SIT) is rapidly evolving as an additional tool for mosquito control, offering an efficient and more environment-friendly alternative to the use of insecticides. Following the devastating chikungunya outbreak, which affected 38% of the population on Reunion Island (a French overseas territory in the southwest of the Indian Ocean), there has been strong interest and political will to develop effective alternatives to the existing vector control strategies. Over the past 10 years, the French Research and Development Institute (IRD) has established an SIT feasibility program against Ae. albopictus on Reunion Island in collaboration with national and international partners. This program aimed to Insects 2020, 11, 770 2 of 25 determine whether the SIT based on the release of radiation-sterilized males is scientifically and technically feasible, and socially acceptable as part of a control strategy targeting the local Ae. albopictus population. This paper provides a review of a multi-year and a particularly broad scoping process of establishing the scientific and technological feasibility of the SIT against Ae. albopictus on Reunion Island. It also draws attention to some prerequisites of the decision-making process, through awareness campaigns to enhance public understanding and support, social adoption, and regulatory validation of the SIT pilot tests.

Aedes aegypti L. (Diptera: Culicidae) is a major vector of yellow fever, dengue, and chikungunya viruses throughout tropical and subtropical areas of the world. Although the southernmost part of Mauritania along the Senegal river has long been recognized at risk of yellow fever transmission, Aedes spp. mosquitoes had never been reported northwards in Mauritania. Here, we report the first observation of Aedes aegypti aegypti (L.) and Aedes (Ochlerotatus) caspius (Pallas, 1771) in the capital city, Nouakchott. We describe the development sites in which larvae of the two species were found, drawing attention to the risk for emergence of arbovirus transmission in the city.

International audience; Background: Malaria is endemic in the southernmost Sahelian zone of Mauritania where the major known mosquito vector is Anopheles arabiensis. Understanding seasonal population dynamics, feeding preferences and insecticide resistance status of these vectors in the area is essential to improve vector control measures implemented at a local scale. Here, malaria vector populations' bionomics is described in two sentinel sites located in the Sahelian zone of Mauritania. Methods: Between September 2014 and December 2016, longitudinal entomological surveys were conducted in Kobeni (15 degrees 49'N, 09 degrees 24'W) and Rosso (16 degrees 30'N; 15 degrees 48'W), two localities in the southern Sahelian zone of Mauritania. Adult mosquitoes were collected using indoor pyrethrum spray catch (PSC). Morphological and PCR-based methods were used to identify the species, detect Plasmodium parasites and analyze blood meals in individual mosquitoes. WHO insecticide susceptibility tests were performed with malathion (5%), bendiocarb (0.1%), permethrin (0.75%) and deltamethrin (0.05%) using female An. gambiae (s.l.) reared from larval and pupal collections from natural breeding sites. Results: A total of 2702 Anopheles mosquitoes were collected by PSC during the study period comprising 2291 Anopheles gambiae (s.l.), 376 Anopheles rufipes and 35 Anopheles pharoensis. In Rosso, all mosquitoes from the An. gambiae (s.l.) complex were molecularly identified as An. arabiensis (n = 455/455, 100%). Anopheles pharoensis represented 2.5% (n = 35/1420) of the specimens collected by PSC in Rosso. In Kobeni, An. arabiensis was dominant (n = 278/301, 92.3%) and occurred together with Anopheles coluzzii (n = 18/301, 6%) and An. gambiae (s.s.) (n = 3/301, 1%). Two An. coluzzii x An. arabiensis hybrids were also detected (0.7%) in Kobeni, and An. rufipes was the only other Anopheles species found resting indoors (n = 376/1277, 29.4%). There was an average of 5.6 and 3.6 indoor resting female An. gambiae (s.l.) per room in Kobeni and Rosso, respectively. Indoor resting female An. gambiae (s.l.) mosquitoes in both sites fed most frequently on bovine blood (35.5% in Rosso and 37% in Kobeni). The proportion of An. gambiae (s.l.) mosquitoes that took human blood was significantly higher in Kobeni (HBI = 37%) than in Rosso (HBI = 5.6%) and 32% of An. gambiae (s.l.) mosquitoes contained blood from more than one host species. None of the 1414 tested mosquitoes in both sites were found positive for Plasmodium spp. sporozoites. WHO insecticide resistance tests revealed resistance to permethrin in the An. arabiensis population from Rosso (mortality = 64%) as well as reduced mortality to deltamethrin (mortality = 97%). Conclusion: This study provides updated information on the composition and dynamics of the malaria vector system in southern Mauritania where malaria is endemic. Such data are a necessary prerequisite to devise and implement tailored malaria elimination strategies in areas of low residual transmission.

International audience; Internal transcribed spacer regions of ribosomal DNA were sequenced, and species-specific primerswere designed to simplify the identification of two morphologically similar species of the Detrituscomplex,Ochlerotatus detritusandOchlerotatus coluzzii. Each newly designed primer was able toamplify a species-specific fragment with a different size. Samples from France and Germany were suc-cessfully tested. This new tool prompts for bio-ecological studies to refine basic knowledge on thebionomics of this species complex, towards a better control and prevention of ensuing nuisances.Moreover, ITS2 sequencing revealed the existence of (1) two distinct haplotypes ofOc. detritusthatare sympatric and widely distributed along the French Atlantic and Mediterranean littorals and (2)a specific haplotype in mosquitoes sampled from Tunisia, raising the question of the taxonomic statusof this North-African population.

In the Phlebotomine sandflies, a few molecular studies related on the genus Sergentomyia have been published. The present study explored the genetic variability within Sergentomyia (Sintonius) clydei (Diptera, Psychodidae). The sampling included 15 populations originating from 12 countries. A morphological approach was coupled to the sequencing of two molecular markers (cytochrome b mtDNA and cacophony nuclear DNA). The most variable morphological characters resided in the cibarium of the females, especially (i) the pigment patch pattern and (ii) the number of cibarial teeth and denticles in the armature. However this morphological approach was unable to individualize any population within S. clydei. The NJ trees based on both molecular markers individualized the specimens from the Aldabra group of islands in the Seychelles. Surprisingly, cyt b variability was not compatible with the known data about the complete submersion of Aldabra occurring relatively recently some 125,000 years ago. The settlement of these islands by S. clydei from continental Africa, the Middle East or Asia, and the value of mtDNA markers are discussed.

A literature analysis coupled with new entomological surveys conducted between 2009 and 2012 led to changes in the list of mosquito species present on the island of La Reunion. Using morphological criteria, Orthopodomyia arboricollis is replaced by Or. reunionensis. On the basis of morphometrical and genetic criteria, Culex univittatus is replaced by Cx. neavei. Cx. poicilipes, which was already reported missing 40 years ago, has not been found again. Anopheles arabiensis is confirmed as the only species of the Gambiae complex present on the island. Thus, twelve species are currently known. For each of them, elements of taxonomic, biological and medical interest are listed. An. arabiensis is a major vector of human Plasmodium (last case of indigenous malaria in 1967). In the Indian Ocean, Aedes albopictus and Ae. aegypti both are competent for transmitting dengue and chikungunya viruses. In Africa, Cx. quinquefasciatus transmits Wuchereria bancrofti and Cx. neavei transmits the Sindbis virus; both species also transmit the West Nile virus. Cx. tritaeniorhynchus is the major vector of Japanese Encephalitis virus in Asia. Two species are endemic (Ae. dufouri and Or. reunionensis), the ten other ones are also found in Madagascar and on the African continent (An. coustani, An. arabiensis, Ae. fowleri, Cx. tritaeniorhynchus, Cx. neavei, Cx. insignis, Lutzia tigripes), with three of them having also a cosmopolitan distribution (Ae. aegypti, Ae. albopictus and Cx. quinquefasciatus). Among the twelve recorded taxa, eight species are anthropophilic, three are supposedly zoophilic and one is a predatory species. No new invasive anthropophilic species did settle on the island. Updated identification keys of larval and adult stages are proposed.

International audience; Although mosquitoes (Diptera: Culicidae) are important disease vectors, information on their biodiversity in Mauritania is scarce and very dispersed in the literature. Data from the scientific literature gathered in the country from 1948 to 2016 were collected and analyzed. Overall 51 culicid species comprising 17 Anopheles spp., 14 Aedes spp., 18 Culex spp. and two Mansonia spp. have been described in Mauritania among which Anopheles arabiensis, Aedes vexans, Culex poicilipes and Culex antennatus are of epidemiological significance. Anopheles arabiensis is widely distributed throughout the country and its geographic distribution has increased northwards in recent years, shifting its northern limit form 17 degrees 32'N in the 1960s to 18 degrees 47'N today. Its presence in the central region of Tagant highlights the great ecological plasticity of the species. Conversely, the distribution of Anopheles gambiae (s.s.) and Anopheles melas has shrunk compared to that of the 1960s. Anopheles rhodesiensis and An. d'thali are mainly confined in the mountainous areas (alt. 200-700 m), whereas Anopheles pharoensis is widely distributed in the Senegal River basin. Culex poicilipes and Cx. antenattus were naturally found infected with Rift valley fever virus in central and northern Mauritania following the Rift valley outbreaks of 1998 and 2012. Recently, Ae. aegypti emerged in Nouakchott and is probably responsible for dengue fever episodes of 2015. This paper provides a concise and up-to-date overview of the existing literature on mosquito species known to occur in Mauritania and highlights areas where future studies should fill a gap in knowledge about vector biodiversity. It aims to help ongoing and future research on mosquitoes particularly in the field of medical entomology to inform evidence-based decision-making for vector control and management strategies.

J. Med. Entomol. 45(3): 391Ð400 (2008) ABSTRACT Polymorphisms at eight microsatellite loci and a fragment of the mitochondrial DNA (mtDNA)-ND4 gene were surveyed in Aedes aegypti (L.) (Diptera: Culicidae) populations collected from six localities in North Cameroon, with emphasis on comparing domestic versus sylvan popula- tions. The microsatellites revealed signiÞcant genetic differentiation among sylvan populations, with meanFST 0.066. Domestic collections were genetically homogeneous (meanFST 0.012). No pattern of isolation by distance was detected, and one of highest levels of genetic differentiation was estimated between populations sampled a few kilometers apart, each in a distinctly different ecological envi- ronment (FST 0.076). Analyses of mtDNA-ND4 polymorphisms and divergence between the two neighboring populations revealed increased genetic diversity within the domestic population, with molecular signatures suggesting recent demographic expansion, whereas a single haplotype was observed in the sylvan sample. These data suggest reduced gene sow between sylvan and domestic Ae. aegypti populations in North Cameroon, reminiscent of the situation for Ae. aegypti in Kenya in East Africa.

Background Aedes aegypti and Ae. albopictus are the main epidemic vectors of dengue, chikungunya and Zika viruses worldwide. Their control during epidemics relies mainly on control of larvae and adults with insecticides. Unfortunately, loss of susceptibility of both species to several insecticide classes limits the efficacy of interventions. In Africa, where Aedes-borne viruses are of growing concern, few data are available on resistance to insecticides. To fill this gap, we assessed the susceptibility to insecticides of Ae. aegypti and Ae. albopictus populations in the Central African Republic (CAR) and studied the mechanisms of resistance. Methods Immature stages were sampled between June and September 2014 in six locations in Bangui (the capital of CAR) for larval and adult bioassays according to WHO standard procedures. We also characterized DDT- and pyrethroid-resistant mosquitoes molecularly and biochemically, including tests for the activities of nonspecific esterases (α and β), mixed-function oxidases, insensitive acetylcholinesterase and glutathione S-transferases. Results Larval bioassays, carried out to determine the lethal concentrations (LC50 and LC95) and resistance ratios (RR50 and RR95), suggested that both vector species were susceptible to Bacillus thuringiensis var. israeliensis and to temephos. Bioassays of adults showed susceptibility to propoxur and fenitrothion, except for one Ae. albopictus population that was suspected to be resistant to fenithrothion. None of the Ae. aegypti populations was fully susceptible to DDT. Ae. albopictus presented a similar profile to Ae. aegypti but with a lower mortality rate (41%). Possible resistance to deltamethrin was observed among Ae. aegypti and Ae. albopictus, although some were susceptible. No kdr mutations were detected in either species; however, the activity of detoxifying enzymes was higher in most populations than in the susceptible Ae. aegypti strain, confirming decreased susceptibility to DDT and deltamethrin. Conclusion These findings suggested that regular, continuous monitoring of resistance is necessary in order to select the most effective adulticides for arbovirus control in Bangui. Electronic supplementary material The online version of this article (doi:10.1186/s13071-016-1887-5) contains supplementary material, which is available to authorized users.

International audience; In Tunisia, malaria transmission has been interrupted since 1980. However, the growing number of imported cases and the persistence of putative vectors stress the need for additional studies to assess the risk of malaria resurgence in the country. In this context, our aim was to update entomological data concerning Anopheles mosquitoes in Tunisia. From May to October of 2012, mosquito larval specimens were captured in 60 breeding sites throughout the country and identified at the species level using morphological keys. Environmental parameters of the larval habitats were recorded. Specimens belonging to the An. maculipennis complex were further identified to sibling species by the ribosomal deoxyribonucleic acid (rDNA)-internal transcribed spacer 2 (ITS2) polymerase chain reaction (PCR) technique. In total, 647 Anopheles larvae were collected from 25 habitats. Four species, including An. labranchiae, An. multicolor, An. sergentii, and An. algeriensis, were morphologically identified. rDNA-ITS2 PCR confirmed that An. labranchiae is the sole member of the An. maculipennis complex in Tunisia. An. labranchiae was collected throughout northern and central Tunisia, and it was highly associated with rural habitat, clear water, and sunlight areas. Larvae of An. multicolor and An. sergentii existed separately or together and were collected in southern Tunisia in similar types of breeding places.

The M and S molecular forms of Anopheles gambiae sensu stricto Giles are thought to be reproductively isolated through premating barriers. However, the exact mechanisms of recognition of conspecific partners are unknown. Because mating in An. gambiae occurs in swarms, one might expect swarming behavior between the M and S forms to be different and that this probably reduces the risk of contact between males and females of the different forms in areas where they are sympatric. We report the occurrence of four mixed swarms, containing males of M and S forms, out of a total of 26 swarms sampled in Soumousso, a typical savannah village of Burkina Faso, West Africa. However, the frequency of mixed swarms was lower than that expected by chance. This observation suggests partial segregation between the swarms of the molecular forms, which may contribute to their isolation. Because the frequency of mixed swarms seems too high to explain the low frequency of cross-mating and hybrids, we suggest that mate recognition in a swarm is more important than swarm segregation.

We have characterized the spliced transcripts of nef and envelope genes of SIVagm from African green monkey of the sabaeus subspecies. Most of the transcripts we have studied, representing the most abundant mRNA species in our assay, have undergone a specific splicing event that removes a part of the trans-activation response (TAR) element. This region is predicted to form a stable secondary structure (four stem–loop elements in SIVagm-sab) that affects the trans-activation of viral gene expression by Tat and the translation of the viral transcripts. Contrary to what is observed in other viruses, in which this R-region splicing has also been described (e.g., HIV-2), the LTR splicing in SIVagm-sab removes part of the first stem–loop and the following ones, nearly completely disrupting the TAR element secondary structure. Because LTR splicing seems to be a conserved feature among the strains we have characterized, these results suggest that this phenomenon could have important consequences for viru...

International audience; Four mosquito species, including a new species of the genus Stegomyia, are reported from Mayotte in the western Indian Ocean. The most abundant species were Stegomyia aegypti and St. albopicta. Only one species of the St. simpsoni group was observed, St. bromeliae. The fourth species is Stegomyia pia Le Goff & Robert n. sp. of which the larva, pupa, male and female are here described. The larval stages of St. pia n. sp. are morphologically similar to St. aegypti but differ in the number of branches of the seta 1-X; the adult is morphologically distinct for a number of characters, for instance the scutal ornamentation. Stegomyia pia n. sp. is uncommon but not rare, and largely distributed across Mayotte. Its larval habitats are natural and diverse including rock pools, tree holes, and cut and severed bamboos. The biology of adults remains unknown, especially female biting behaviour. Both morphological characters and nucleotide sequences of the ITS2 and COI genes indicate that this species is best placed in the genus Stegomyia. Dichotomous keys to the four species of Mayotte Stegomyia are presented for adults and fourth-instar larvae. The potential vector role of these mosquitoes is hypothesised. This paper underlines advances in knowledge of the biodiversity in the French overseas departments and territories.; Quatre espèces de moustiques du genre Stegomyia sont rapportées de Mayotte, Océan Indien Occidental, dont une nouvelle espèce pour la science. Les espèces les plus abondantes étaient Stegomyia aegypti et St. albopicta. Une seule espèce du groupe St. simpsoni a été observée, St. bromeliae. La quatrième espèce est Stegomyia pia Le Goff & Robert n. sp., dont la larve, la nymphe, et les adultes mâles et femelles sont ici décrits. Les stades larvaires de St. pia n. sp. sont morphologiquement proches de ceux de St. aegypti mais en diffèrent par le nombre de branche des soies 1-X; l’adulte est aisément reconnaissable grâce à de nombreux caractères, par exemple l’ornementation du scutum. Stegomyia pia n. sp. est une espèce peu commune sans être rare et largement distribuée sur l’ensemble de Mayotte. Ses gîtes préimaginaux sont naturels et variés, trous de rocher, trous d’arbres, bambous coupés et fendus. On ignore encore tout de la biologie des adultes, en particulier des comportements trophiques des femelles. Les caractères morphologiques ainsi que les séquences nucléotidiques des gènes ITS2 et COI indiquent que cette espèce est placée dans le genre Stegomyia. Des clés dichotomiques de détermination des quatre espèces de Stegomyia de Mayotte sont proposées pour les adultes et pour les larves de stade 4. Le rôle de vecteur potentiel de ces moustiques est envisagé. Cet article contribue à la connaissance de la biodiversité de l’outre-mer français.

International audience; Plant-derived sugar is the only source of dietary carbohydrate for males of most mosquito species. Male resource acquisition and utilization remain an under-researched area of behavior in vectors of human diseases. However, the renewed interest in the use of sterile males against disease vector mosquitoes reinforces the urgent need for studies on the behavioral and ecological processes that underpin male fitness and reproductive success. Here an attempt was made first to characterize the conditions and modes of resource acquisition (plant derived sugar meals) early in the life of An. arabiensis males, and second to test the hypothesis that the plants chosen for their sugar meals are those which maximize their fitness in terms of energy gains (i.e. amount of lipids, proteins, glycogen and glucose). Olfactometry assays demonstrated the ability of An. arabiensis males to discriminate among a sample of ten abundant flowering plants present in their natural habitats. In further experiments, we observed significant variations in the sugar intake rates that matched their olfactory preferences, with the most attractive plants eliciting significantly higher sugar intake rates. Consistent with our expectations, analyses of the whole-body free sugars, lipids and glycogen unequivocally showed that the energy reserve accumulated post-feeding is dependent on the diet of the adult males, with the preferred plants providing more energy reserves than the less preferred ones, despite mosquitoes actively feeding on both. Taken together, these results show that An. Arabiensis males are able to discern between food sources, preferentially feeding on those species of plant that provide the highest metabolic payoff. Ensuring or somehow heightening the ability to detect and obtain rewarding sugar meals by male mosquitoes reared for field release could enhance their competitive ability in the field.

Author(s): Ya-Umphan, Phubeth; Cerqueira, Dominique; Parker, Daniel M; Cottrell, Gilles; Poinsignon, Anne; Remoue, Franck; Brengues, Cecile; Chareonviriyaphap, Theeraphap; Nosten, Francois; Corbel, Vincent | Abstract: BackgroundThe modalities of malaria transmission along the Thailand-Myanmar border are poorly understood. Here we address the relevance of using a specific Anopheles salivary biomarker to measure the risk among humans of exposure to Anopheles bites.MethodsSerologic surveys were conducted from May 2013 to December 2014 in 4 sentinel villages. More than 9400 blood specimens were collected in filter papers from all inhabitants at baseline and then every 3 months thereafter, for up to 18 months, for analysis by enzyme-linked immunosorbent assay. The relationship between the intensity of the human antibody response and entomological indicators of transmission (human biting rates and entomological inoculation rates [EIRs]) was studied using a multivariate 3-level mixed model analysis. Heat maps for human immunoglobulin G (IgG) responses for each village and survey time point were created using QGIS 2.4.ResultsThe levels of IgG response among participants varied significantly according to village, season, and age (Pl.001) and were positively associated with the abundance of total Anopheles species and primary malaria vectors and the EIR (Pl.001). Spatial clusters of high-IgG responders were identified across space and time within study villages.ConclusionsThe gSG6-P1 biomarker has great potential to address the risk of transmission along the Thailand-Myanmar border and represents a promising tool to guide malaria interventions.

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.

In West Africa, M and S molecular forms of Anopheles gambiae sensu stricto (Diptera: Culicidae) Giles, frequently occur together, although with different population bionomics. The S form typically breeds in rain-dependant water collections and is present during the rainy season only whereas the M form can thrive all year long in areas with permanent breeding opportunities. In the present study, we explored physiological and developmental trade-offs at play in laboratory colonies and field populations of the M and S forms that originated from an area of sympatry in Burkina Faso, where M and S larvae exhibit such habitat segregation. In the laboratory, larvae of the M form developed slower than the S form (mean values 9.51 and 8.85 days, respectively, Wilcoxon's test, P < 0.001). Although wing length and dry weight at emergence showed large variations, M females were on average 8% heavier than S females of similar wing length. Higher nutritional reserves (proteins and lipids) in teneral adults explained part of this weight difference, reflecting a better ability of the M form to garner resources at the larval stage. Furthermore, a higher rate of ovarian maturation was observed in the M form after a single bloodmeal. The relevance of these findings for parasite transmission is discussed.

No Phlebotomine sandflies had ever been reported in the Comoros Archipelago, including the three islands of the Republic of the Union of Comoros (Grande Comore, Moheli and Anjouan) and the French oversea department of Mayotte. During three field surveys carried out in 2003, 2007 and 2011, we provided the first record of Phlebotomine sandflies in this area. A total of 85 specimens belonging to three species were caught: a new species S. (Vattieromyia) pessoni n. sp. (two females from Grande Comore), a new subspecies of Sergentomyia (Rondanomyia) goodmani (80 specimens from Grande Comore and one from Anjouan) and Grassomyia sp. (two females from Moheli). The individualisation of these taxa was inferred both from morphological criteria and sequencing of a part of the cytochrome b of the mitochondrial DNA. These taxa are closely related to Malagasy sandflies.

Background During recent periods, the islands of the Republic of Seychelles experienced many diseases such as dengue, chikungunya, Bancroft’s filaria and malaria. Mosquitoes transmit the agents that cause these diseases. Published information on mosquitoes in the Seychelles is notably dispersed in the literature. The maximum number of species obtained on a single field survey does not exceed 14 species. Methods We performed a comprehensive bibliographic review using mosquito and Seychelles as the key words, as well as conducted a mosquito field survey for larval and adult stages during the rainy season in December 2008. Sixteen sites were sampled on four granitic islands (Mahé, Praslin, La Digue and Aride) and six sites on coralline atolls in the extreme southwest of the country (Aldabra group). Results We found published references to 21 mosquito species identified at least on one occasion in the Seychelles. Our collections comprised 18 species of mosquitoes, all of them from the subfamily Culicinae; no Anophelinae was found. We also confirm that Aedes seychellensis is a junior synonym of Ae. (Aedimorphus) albocephalus. The first records for Culex antennatus and Cx. sunyaniensis are presented from the country, specifically from Aldabra and Praslin, respectively. Based on a comparison of the taxa occurring on the granitic versus coralline islands, only three species, Ae. albocephalus, Cx. scottii and Cx. simpsoni are shared. Aedes albopictus appeared to exclude largely Ae. aegypti on the granitic islands; however, Ae. aegypti was common on Aldabra, where Ae. albopictus has not been recorded. The notable aggressiveness of mosquitoes towards humans on coralline islands was mainly due to two species, the females of which are difficult to distinguish: Ae. fryeri and Ae. (Aedimorphus) sp. A. The number of mosquito species collected at least once in the Seychelles is now 22, among which five species (Ae. (Adm) sp. A, Cx. stellatus, Uranotaenia browni. Ur. nepenthes and Ur. pandani) and one subspecies (Ae. vigilax vansomerenae) are considered as endemic. Two illustrated identification keys, one for adult females and the other for larval stages, are presented. Conclusions The knowledge of the culicidian fauna in the Seychelles has been notably updated. The number of mosquito species is relatively large with regards to land surface and distances to continental Africa, although the anophelines are totally lacking. The complex natural history of mosquitoes in the Seychelles provides examples of both vicariance- and dispersal-mediated divergences. They present superb examples for theoretical and applied island biology.

BackgroundAedes albopictus (Skuse, 1884) (Diptera: Culicidae), a mosquito native to Asia, has recently invaded all five continents. In Central Africa it was first reported in the early 2000s, and has since been implicated in the emergence of arboviruses such as dengue and chikungunya in this region. Recent genetic studies of invasive species have shown that multiple introductions are a key factor for successful expansion in new areas. As a result, phenotypic characters such as vector competence and insecticide susceptibility may vary within invasive pest species, potentially affecting vector efficiency and pest management. Here we assessed the genetic variability and population genetics of Ae. albopictus isolates in Cameroon (Central Africa), thereby deducing their likely geographic origin.Methods and resultsMosquitoes were sampled in 2007 in 12 localities in southern Cameroon and analyzed for polymorphism at six microsatellite loci and in two mitochondrial DNA regions (ND5 and COI). All the microsatellite markers were successfully amplified and were polymorphic, showing moderate genetic structureamong geographic populations (F(ST) = 0.068, P < 0.0001). Analysis of mtDNA sequences revealed four haplotypes each for the COI and ND5 genes, with a dominant haplotype shared by all Cameroonian samples. The weak genetic variation estimated from the mtDNA genes is consistent with the recent arrival of Ae. albopictus in Cameroon. Phylogeographic analysis based on COI polymorphism indicated that Ae. albopictus populations from Cameroon are related to tropical rather than temperate or subtropical outgroups.ConclusionThe moderate genetic diversity observed among Cameroonian Ae. albopictus isolates is in keeping with recent introduction and spread in this country. The genetic structure of natural populations points to multiple introductions from tropical regions.

Aedes aegypti (Linné, 1762) is a major vector of arboviruses such as Yellow Fever, Dengue and Chikungunya. In Africa, where the species exhibits major variations in morphology, ecology, behavior and vector competence, two subspecies have been described: a light form, named Ae. aegypti aegypti (Aaa) with highly domestic and anthropophilic habits and a cosmotropical distribution; and a dark form, referred to as Ae. aegypti formosus (Aaf), which is endemic to Africa and thrives in sylvan environments. In East Africa, both forms were described to occur in sympatry whereas only Aaf was reported from Central/West Africa. However, recent findings suggest Aaa was also common in Senegal. Here, we report on a longitudinal survey of morphological and genetic variability of Ae. aegypti sampled in the rural environment of Niakhar, Senegal. In agreement with recent findings, most of specimens we analyzed were classified as Aaa suggesting typical Aaf was scarce in the studied area. Among Aaa, significant temporal variations in abdominal pale scales pattern were detected. Depending on the season and the nature of larval breeding places, the specimens (particularly females) tend to segregate in two main morphological groups. Microsatellite-based estimates of genetic differentiation did not provide any clear evidence that the two groups were genetically distinct. Overall, these results improve our understanding of the diversity of Ae. aegypti in West Africa, where data are crucially lacking.

The Camargue is an extensive wetland in the southeast of France, which is highly influenced by human activities. Large ponds, marshes and irrigated fields pro- vide abundant potential breeding sites for mosquitoes. Mosquitoes, which are important in terms of the nuisance they cause to people and animals, the limitations they impose on tourism and their potential threat to human health. Several of the mosquito species present are potential vectors of malaria and West Nile virus. Therefore, the population dynamics of these species were monitored over an entire breeding season during March - October 2005. Mosquito populations were sampled in two study areas once every 2 weeks, using CDC light traps baited with CO 2 . Sixteen species were collected. The majority (98.7%) of the catch were Aedes caspius (Pallas) (Diptera: Culicidae), Culex modestus (Ficalbi), Culex pipiens L. and Anopheles hyrcanus (Pallas). The population dynamics of these species varied considerably in relation to the species ' biology, climatic conditions (rainfall, temperature and season), water management, implementa- tion of mosquito control campaigns and landscape use.

Background Malaria is a former endemic problem in the Camargue, South East France, an area from where very few recent data concerning Anopheles are available. A study was undertaken in 2005 to establish potential malaria vector biology and dynamics and evaluate the risk of malaria re-emergence. Methods Mosquitoes were collected in two study areas, from March to October 2005, one week every two weeks, using light traps+CO2, horse bait traps, human bait catch, and by collecting females in resting sites. Results Anopheles hyrcanus was the most abundant Anopheles species. Anopheles melanoon was less abundant, and Anopheles atroparvus and Anopheles algeriensis were rare. Anopheles hyrcanus and An. melanoon were present in summer, whereas An. atroparvus was present in autumn and winter. A large number of An. hyrcanus females was collected on humans, whereas almost exclusively animals attracted An. melanoon. Based on an enzyme-linked immunosorbent assay, almost 90% of An. melanoon blood meals analysed had been taken on horse or bovine. Anopheles hyrcanus and An. melanoon parity rates showed huge variations according to the date and the trapping method. Conclusion Anopheles hyrcanus seems to be the only Culicidae likely to play a role in malaria transmission in the Camargue, as it is abundant and anthropophilic.

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, carbarnate, organophosphate and organochlorime 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-transferases). 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. quiclnuefasciatus 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.

International audience; We report the first finding of the knockdown Leu-Phe and Leu-Ser mutations associated with resistance to pyrethroids and DDT insecticides in the malaria mosquito Anopheles gambiae from Cameroon. The Leu-Phe mutation was found in both the M and S molecular forms of An. gambiae. Importantly, two specimens of the S molecular form were found to carry both mutations in a heterozygous state.

During extensive sampling in Burkina Faso and other African countries, the Leu-Phe mutation producing the kdr pyrethroid resistance phenotype was reported in both Anopheles gambiae ss and A. arabiensis. This mutation was widely distributed at high frequency in the molecular S form of A. gambiae while it has been observed at a very low frequency in both the molecular M form and A. arabiensis in Burkina Faso. While the mutation in the M form is inherited through an introgression from the S form, its occurrence is a new and independent mutation event in A. arabiensis. Three nucleotides in the upstream intron of the kdr mutation differentiated A. arabiensis from A. gambiae ss and these specific nucleotides were associated with kdr mutation in A. arabiensis. Ecological divergences which facilitated the spread of the kdr mutation within the complex of A. gambiae ss in West Africa, are discussed.

Samples of the dengue vector mosquito Aedes aegypti (L.) (Diptera: Culicidae) were collected from 13 localities between 1995 and 1998. Two laboratory strains, Bora (French Polynesia) and AEAE, were both susceptible to DDT and permethrin; all other strains, except Larentuka (Indonesia) and Bouake (Ivory Coast), contained individual fourth-instar larvae resistant to permethrin. Ten strains were subjected to a range of biochemical assays. Many strains had elevated carboxylesterase activity compared to the Bora strain; this was particularly high in the Indonesian strains Salatiga and Semarang, and in the Guyane strain (Cayenne). Monooxygenase levels were increased in the Salatiga and Paea (Polynesia) strains, and reduced in the two Thai strains (Mae Kaza, Mae Kud) and the Larentuka strain. Glutathione S-transferase activity was elevated in the Guyane strain. All other enzyme profiles were similar to the susceptible strain. The presence of both DDT and pyrethroid resistance in the Semarang, Belem (Brazil) and Long Hoa (Vietnam) strains suggested the presence of a knock-down resistant (kdr)-type resistance mechanism. Part of the S6 hydrophobic segment of domain II of the voltage-gated sodium channel gene was obtained by RT-PCR and sequenced from several insects from all 13 field strains. Four novel mutations were identified. Three strains contained identical amino acid substitutions at two positions, two strains shared a different substitution, and one strain was homozygous for a fourth alteration. The leucine to phenylalanine substitution that confers nerve insensitivity to pyrethroids in a range of other resistant insects was absent. Direct neurophysiological assays on individual larvae from three strains with these mutations demonstrated reduced nerve sensitivity to permethrin or lambda cyhalothrin inhibition compared to the susceptible strains.

Background Olfaction plays a significant role in insect behavior during critical steps of their life-cycle, such as host-seeking during foraging or the search for a mate. Here, we explored genetic polymorphism within and divergence between sibling species of the African malaria mosquito, Anopheles gambiae sensu lato in the gene sequence and encoded peptides of an odorant receptor, Or39. This study included sympatric specimens of An. gambiae sensu stricto, An. coluzzii and An. arabiensis sampled together in the village of Dielmo, Senegal. Results A 1,601 bp genomic sequence composed of 6 exons and 5 introns was obtained for Or39 from 6–8 mosquitoes in each of the 3 species. DNA sequence analysis revealed a high level of molecular polymorphism (π = 0.0154; Haplotype diversity = 0.867) and high overall genetic differentiation between taxa (Fst > 0.92, P

International audience; Anopheles gambiae s.s. is a complex of sibling taxa characterized by various paracentric inversions. In west and central Africa, where several taxa are sympatric, a kdr mutation responsible for pyrethroid resistance has been described in only one (the S taxon), suggesting an absence of gene flow between them. Following a thorough sampling, we have found a kdr mutation in another taxon (M). To establish whether this mutation is the same event or not, the large intron upstream of the kdr mutation was sequenced to find polymorphic sites in susceptible/resistant and M/S mosquitoes. The low genetic diversity found in this DNA region indicates that a local genetic sweep has recently occurred. However, some polymorphic sites were found, and it is therefore concluded that the kdr mutation in the M taxon is not an independent mutation event, and is best explained by an introgression from the S taxon. These results are discussed within the context of possible gene flow between members of An. gambiae s.s. taxa, and with the possible spread of the kdr mutation in other closely related malaria vectors of the An. gambiae complex.

We investigated HIV-1 diversity by means of heteroduplex mobility assay (HMA) genotyping. We studied 199 samples from patients originating from 26 countries and living in France. The HMA successfully genotyped 182 (91 %) of these samples, as follows : 77 (42 %) subtype A, 57 (31 %) subtype B, 5 (3 %) subtype C, 5 (3 %) subtype D, 8 (4 %) subtype E, 22 (12 %) subtype F, 5 (3 %) subtype G, and 3 (2 %) subtype H. We were not able to genotype 12 samples by means of the HMA. These latter strains were sequenced, and phylogenetic analyses revealed that they were highly divergent subtype A-, D-, or G-related strains. Eight (of 12) subtype D strains were indeterminate by HMA, owing to the broad intrasubtype diversity, suggesting that new reference subtype D plasmids are required, as previously proposed. Thirty-seven strains belonging to the different subtypes were sequenced, and the results showed perfect concordance with the HMA results. Interlaboratory quality controls confirmed the reliability of the HMA for HIV-1 subtyping, despite the extensive viral variability. However, plasmid selection must be continuously revised to cover viral diversification. (Résumé d'auteur)

In Africa the highest HIV infection rate has been reported among female commercial sex workers (CSWs) who are at increasing risk of acquiring and transmitting HIV infection. In October 1995, 176 CSWs were studied in Bamako, the capital city of Mali. The ages of the CSWs ranged from 15 to 50 years old (mean, 28.8 years). Only 20.45% of the 176 CSWs were Malian; the majority were from Nigeria (32.9%) and Ghana (31.8%), and the remaining were from other African countries. Forty-one percent were active for less than 1 year as a commercial sex worker, and the length of prostitution for the remaining women ranged from 1 to 15 years (mean, 2.76). A total of 81 (46.02%) of the 176 CSWs were positive for HIV antibodies; 63 (35.8%) were HIV-1 positive, (3.9%) were HIV-2 positive, 11 (6.2%) had antibodies to HIV-1 and HIV-2, and none of them had antibodies to group O viruses. For all HIV antibody-positive samples, PBMCs were separated and genetic subtypes of HIV-1 were determined using the heteroduplex mobility assay (HMA), with ED5-ED12 as outer and ES7-ES8 as inner primers. Among the 66 HIV-1 strains characterized, 53 (80.3%) were subtype A, 2 (3.1%) belonged to subtype C, 1 (1.5%) belonged to subtype D, and 10 (15.1%) were identified as subtype G. Among the 10 subtype G strains, 8 were obtained from women who were very recent CSWs, with an activity of 1 year or less, assuming that there is a high probability that these infections occurred recently. Genetic subtypes of five HIV-2 viruses were determined by sequencing of the env and/or gag genes followed by phylogenetic analysis, and all of them belonged to subtype A. Comparison of HIV-1 and HIV-2 seroprevalence data from our study with previous data from Mali shows a significant rise in HIV-1 prevalence and a significant decrease in HIV-2 prevalence and confirms similar trends observed in neighboring countries. We have found four different genetic subtypes of HIV-1; however, subtype A is predominant and accounts for 80% of the cases and 15% of the HIV-1 infections were subtype G. It is important to continue the surveillance of subtypes on a systematic basis in order to see to what extent the proportions of the different subtypes will change over time.The genetic variability of HIV-1 was investigated in a 1995 study of 176 commercial sex workers (CSWs) recruited in different areas in Bamako, Mali. 36 CSWs (20.45%) were born in Mali; 58 (32.9%) were from Nigeria and 56 (31.8%) were from Ghana. They ranged in age from 15-50 years (mean, 28.8 years). 41% of sex workers had been active for less than 12 months; the remaining women had been CSWs for 1-15 years (mean, 2.76 years). Of the 81 CSWs (46.02%) who were HIV-positive, 63 (35.8%) were infected with HIV-1, 7 (3.9%) with HIV-2, and the remaining 11 (6.2%) had antibodies to both HIV-1 and HIV-2. In contrast to other studies conducted among CSWs in Africa, none of these sex workers had antibodies to group O viruses. HIV-1 prevalence increased with age and length of time in prostitution and was higher among women with a history of sexually transmitted diseases. Among the 66 HIV-1 strains characterized, 53 (80.3%) were subtype A, 2 (3.1%) belonged to subtype C, 1 (1.5%) belonged to subtype D, and 10 (15.1%) were identified as subtype G. These results indicate a significant rise in HIV-1 prevalence and significant decreases in HIV-2 and combined HIV-1 and HIV-2 prevalence (10%, 15%, and 13%, respectively, in 1985). Ongoing surveillance of HIV-1 subtypes in Africa is important to identify shifts in the proportions of different subtypes over time. The genetic diversity of HIV has important implications for vaccine development.

The African malaria vector Anopheles gambiae is polymorphic for alternative arrangements on the left arm of chromosome 2 (2La and 2L+ a ) that are non-randomly distributed with respect to degree of aridity. Detailed studies on the ecological role of inversion 2La have been hindered by the technical demands of traditional karyotype analysis and by sex- and stage-specific limitations on the availability of polytene chromosomes favorable for analysis. Recent molecular characterization of both inversion breakpoints presented the opportunity to develop a polymerase chain reaction (PCR)-based method for karyotype analysis. Here we report the development of this molecular diagnostic assay and the results of extensive field validation. When tested on 765 An. gambiae specimens sampled across Africa, the molecular approach compared favorably with traditional cytologic methods, correctly scoring > 94% of these specimens. By providing ready access to the 2La karyotype, this tool lays groundwork for future studies of the ecological genomics of this medically important species.

Background The yellow fever mosquito Aedes aegypti is a major vector of dengue and hemorrhagic fevers, causing up to 100 million dengue infections every year. As there is still no medicine and efficient vaccine available, vector control largely based on insecticide treatments remains the only method to reduce dengue virus transmission. Unfortunately, vector control programs are facing operational challenges with mosquitoes becoming resistant to commonly used insecticides. Resistance of Ae. aegypti to chemical insecticides has been reported worldwide and the underlying molecular mechanisms, including the identification of enzymes involved in insecticide detoxification are not completely understood. Results The present paper investigates the molecular basis of insecticide resistance in a population of Ae. aegypti collected in Martinique (French West Indies). Bioassays with insecticides on adults and larvae revealed high levels of resistance to organophosphate and pyrethroid insecticides. Molecular screening for common insecticide target-site mutations showed a high frequency (71%) of the sodium channel 'knock down resistance' (kdr) mutation. Exposing mosquitoes to detoxification enzymes inhibitors prior to bioassays induced a significant increased susceptibility of mosquitoes to insecticides, revealing the presence of metabolic-based resistance mechanisms. This trend was biochemically confirmed by significant elevated activities of cytochrome P450 monooxygenases, glutathione S-transferases and carboxylesterases at both larval and adult stages. Utilization of the microarray Aedes Detox Chip containing probes for all members of detoxification and other insecticide resistance-related enzymes revealed the significant constitutive over-transcription of multiple detoxification genes at both larval and adult stages. The over-transcription of detoxification genes in the resistant strain was confirmed by using real-time quantitative RT-PCR. Conclusion These results suggest that the high level of insecticide resistance found in Ae. aegypti mosquitoes from Martinique island is the consequence of both target-site and metabolic based resistance mechanisms. Insecticide resistance levels and associated mechanisms are discussed in relation with the environmental context of Martinique Island. These finding have important implications for dengue vector control in Martinique and emphasizes the need to develop new tools and strategies for maintaining an effective control of Aedes mosquito populations worldwide.