An entomological review of invasive mosquitoes in Europe.

Among the invasive mosquitoes registered all over the world, Aedes species are particularly frequent and important. As several of them are potential vectors of disease, they present significant health concerns for 21st century Europe. Five species have established in mainland Europe, with two (Aedes albopictus and Aedes japonicus) becoming widespread and two (Ae. albopictus and Aedes aegypti) implicated in disease transmission to humans in Europe. The routes of importation and spread are often enigmatic, the ability to adapt to local environments and climates are rapid, and the biting nuisance and vector potential are both an ecomonic and public health concern. Europeans are used to cases of dengue and chikungunya in travellers returning from the tropics, but the threat to health and tourism in mainland Europe is substantive. Coupled to that are the emerging issues in the European overseas territorities and this paper is the first to consider the impacts in the remoter outposts of Europe. If entomologists and public health authorities are to address the spread of these mosquitoes and mitigate their health risks they must first be prepared to share information to better understand their biology and ecology, and share data on their distribution and control successes. This paper focusses in greater detail on the entomological and ecological aspects of these mosquitoes to assist with the risk assessment process, bringing together a large amount of information gathered through the ECDC VBORNET project.

Diel activity patterns of male Aedes albopictus in the laboratory.

This study was conducted to investigate the flight and dispersal behavior of male Aedes albopictus under laboratory conditions. Two different methods, the Observer and Ethovision software devices, were used to determine the total duration, the mean and maximum flight speed, and the distance covered by these mosquitoes. During 24 h, mosquitoes were more active from 0800 to 1200 h and from 1700 to 2100 h than during the rest of the day. Male Ae. albopictus displayed different activities at different times. The flight activity was 47 min and 57 sec over 24 h. During this period, sugar-fed males flew an average distance of 236.20 m at an average speed of 8.5 cm/sec. The unfed males flew significantly faster than recently fed males, with maximum flight speeds of 44.9 cm/sec and 33.6 cm/sec, respectively. The time used for resting (22 h 2 min +/- 13 min) was significantly higher (P < 0.0001) than walking and flying times (68 min +/- 10 sec and 49 min +/- 5 min, respectively). Overall, both methods allowed observations on flight activity, and the camera recordings allowed these activities to be quantified.

[Updated inventory of mosquitoes (Diptera: Culicidae) of the island of La Réunion, Indian Ocean]. / Inventaire actualisé des moustiques (Diptera : Culicidae) de l'île de La Réunion, océan Indien.

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 Réunion. 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.

Ecological interactions in Aedes species on Reunion Island.

Two invasive, container-breeding mosquito species, Aedes aegypti (Stegomyia aegypti) and Aedes albopictus (Stegomyia albopicta) (Diptera: Culicidae), have different distribution patterns on Reunion Island. Aedes albopictus occurs in all areas and Ae. aegypti colonizes only some restricted areas already occupied by Ae. albopictus. This study investigates the abiotic and biotic ecological mechanisms that determine the distribution of Aedes species on Reunion Island. Life history traits (duration of immature stages, survivorship, fecundity, estimated finite rate of increase) in Ae. aegypti and Ae. albopictus were compared at different temperatures. These fitness measures were characterized in both species in response to competitive interactions among larvae. Aedes aegypti was drastically affected by temperature, performing well only at around 25 °C, at which it achieved its highest survivorship and greatest estimated rate of increase. The narrow distribution of this species in the field on Reunion Island may thus relate to its poor ability to cope with unfavourable temperatures. Aedes aegypti was also more negatively affected by high population densities and to some extent by interactions with Ae. albopictus, particularly in the context of limited food supplies. Aedes albopictus exhibited better population performance across a range of environmental conditions. Its ecological plasticity and its superior competitive ability relative to its congener may further enhance its invasion success on Reunion Island.

An interdisciplinary approach to controlling chikungunya outbreaks on French islands in the south-west Indian ocean.

The outbreak of chikungunya that occurred on French Island territories in the southwest Indian Ocean in 2005 and 2006 caused severe morbidity and mortality. In the aftermath, French authorities set up a scientific task force including experts in epidemiology, public health, entomology, virology, immunology, sociology, animal health, community and hospital medicine. The mission of the task force was to conceive and propose research programs needed to increase understanding of the disease and epidemic and to help public health officials in improving epidemic response measures. The purpose of this article is to describe the findings of the task force at the end of its two-year existence and initial outcomes in the the areas studied. Discussion emphasizes topics requiring further study.

The invaders: phylogeography of dengue and chikungunya viruses Aedes vectors, on the South West islands of the Indian Ocean.

Aedes albopictus and Aedes aegypti are the two main worldwide arbovirus vectors that have experienced invasion phases. Aedes aegypti is a pantropical species that spread centuries ago whereas Ae. albopictus started the main wave of invasion in the 1980s. Both species have been at various times on the different islands in Southwestern Indian Ocean (SWIO). This area provides an opportunity to examine the extent to which mosquitoes colonization patterns are influenced by different introductory events likely linked with human settlement and migration between the islands. To explore this hypothesis, we propose a CO1-based phylogeny using a large sampling of fresh Ae. albopictus, Ae. aegypti, and Ae. mascarensis, and 50-year-old dry specimens originating from different Indian Ocean islands. Our data allow us to hypothesize the existence of at least two waves of invasion for Ae. albopictus in the islands of SWIO. The first one most likely occurred several centuries ago with establishments in Madagascar, Mauritius and Reunion Island. The other one that appears to currently still on-going, reached almost all the islands of SWIO during the 1990s or later. The low genetic diversity found between the ancient invasive strain and the contemporary one, indicates with great certainty that Ae. albopictus is not indigenous to the islands of SWIO. Recently, in Madagascar, an invasive lineage of Ae. albopictus has expanded all over the island while Ae. aegypti populations have declined in urban areas. Three clusters of Aedes aegypti have been observed, two fitting with the wild form Ae. aegypti formosus and the other one fitting with the more domestic form Ae. ae. aegypti. Sequence of Ae. mascarensis, endemic to Mauritius suggest that this species might belong to Ae. aegypti species and on this basis we propose to classify it as a sub species or form of Ae. aegypti species. Given the increase of human population flux on these islands, the occurrence of these vectors and their ability to spread quickly are of high importance of arbovirus transmission and the epidemicity of the associated diseases in these islands.

Preliminary study on the galectin molecular diversity in Moroccoan Phlebotomus papatasi sandfly population.

CONTEXT: Galactose binding protein (PpGalec) plays an important role in the specificity of Phlebotomus papatasi sandfly for Leishmania major. The molecular diversity of this ligand is currently unknown but might have some influence on the ability of PpGalec to efficiently recognize L. major in natural sandfly populations. OBJECTIVE: To explore the molecular diversity of the P. papatasi Galectin gene (PpGalec) in natural sandfly population of Morocco. RESULTS & CONCLUSIONS: Sequence variations of PpGalec was analyzed in 31 P. papatasi specimens collected from endemic and non-endemic zoonotic cutaneous leishmaniasis foci of Morocco. Among the 211 amino acid positions analyzed, 11 are subjected to mutation. Interestingly, we observe that one mutation directly affect an amino acid known to be involved in the substrate recognition by galectin. The repercussion of this polymorphism on the capacity of the galectin to efficiently bind the L. major Lipophosphoglycane (LPG) awaits further investigations.

Sexual performance of male mosquito Aedes albopictus.

Issues of male fertility must be addressed to support the development of a sterile insect technique (SIT) programme for the control of Aedes albopictus Skuse (Diptera: Culicidae) populations on Reunion Island in the Indian Ocean. The mating ability of a local strain of Ae. albopictus was tested using several batches of females and different cage sizes under laboratory conditions. Individual males were able to inseminate up to 14 females at an average of 9.5 females per male when exposed to 20 females over 7 days. Males filled between three and 27 spermathecal capsules at an average of 15.5 capsules per male. The average number of females inseminated per male was 5.3 when two virgin females were introduced to one male and replaced every day for 12 days, and 8.6 when 10 virgin females were introduced to one male and replaced every day for 14 days. A continuous decrease in the number of both inseminated females and filled spermathecal capsules was observed over time, until no mating occurred after 14 days. The high number of females inseminated by one male and the duration of male activity may have strong implications for SIT control of mosquitoes.

Bacterial flora as indicated by PCR-temperature gradient gel electrophoresis (TGGE) of 16S rDNA gene fragments from isolated guts of phlebotomine sand flies (Diptera: Psychodidae).

In this study, we tested the capacity of Temperature Gradient Gel Electrophoresis (TGGE)-based fingerprinting of 16S rDNA PCR fragments to assess bacterial composition in a single isolated sand fly gut. Bacterial content was studied in different life stages of a laboratory-reared colony of Phlebotomus duboscqi and in a wild-caught Phlebotomus papatasi population. Our study demonstrates that a major reorganization in the gut bacterial community occurs during metamorphosis of sand flies. Chloroflexi spp. was dominant in the guts of pre-imaginal stages, although Microbacterium spp. and another as yet unidentified bacteria were detected in the gut of the adult specimen. Interestingly, Microbacterium spp. was also found in all the adult guts of both species. We demonstrate that the analysis of bacterial diversity in an individualized sand fly gut is possible with fingerprinting of 16S rDNA. The use of such methodology, in conjunction with other culture-based methods, will be of great help in investigating the behavior of the Leishmania-bacterial community in an ecological context.

Contribution of Anopheles funestus, An. gambiae and An. nili (Diptera: Culicidae) to the perennial malaria transmission in the southern and western forest areas of Côte d'Ivoire.

The involvement of members of the Anopheles gambiae complex Giles and An. funestus Giles and An. nili Theobald groups in the transmission of Plasmodium falciparum was recently investigated in the villages of Gbatta and Kpéhiri, which lie, respectively, in forest areas in the west and south of Côte d'Ivoire. Adult female mosquitoes were collected, using human landing catches, inside and outside dwellings. After identification and dissection, the heads and thoraces of all the anopheline mosquitoes were tested, in an ELISA, for circumsporozoite protein (CSP). All the female anopheline mosquitoes collected and identified to species using PCR were found to be An. gambiae s.s., An. nili s.s. or An. funestus s.s., with An. gambiae s.s. and An. funestus s.s. predominant in Gbatta but An. nili s.s. the most common species in Kpéhiri. In Gbatta, 3·1% of the female An. gambiae collected, 5·0% of the female An. funestus and 1·8% of the female An. nili were found CSP-positive. The corresponding values in Kpéhiri were even higher, at 5·9%, 6·2% and 2·4%, respectively. The estimated entomological inoculation rates (EIR) were very high: 302 infected bites (139 from An. gambiae, 146 from An. funestus and 17 from An. nili)/person-year in Gbatta and 484 infected bites (204 from An. gambiae, 70 from An. funestus and 210 from An. nili)/person-year in Kpéhiri. In Gbatta, An. gambiae s.s. was responsible for most of the rainy-season transmission while An. funestus became the main malaria vector in the dry seasons. In Kpéhiri, however, An. nili appeared to be the main vector throughout the year, with An. gambiae of secondary importance and An. funestus only becoming a significant vector during the rainy season. Although, in both study sites, intense transmission was therefore occurring and the same three species of anopheline mosquito were present, the relative importance of each mosquito species in the epidemiology of the human malaria at each site differed markedly.

Optimization of breeding output for larval stage of Anopheles gambiae (Diptera: Culicidae): prospects for the creation and maintenance of laboratory colony from wild isolates.

Domesticating anopheline species from wild isolates provides an important laboratory tool but requires detailed knowledge of their natural biology and ecology, especially the natural breeding habitats of immature stages. The aim of this study was to determine the optimal values of some parameters of Anopheles gambiae larval development, so as to design a standard rearing protocol of highland isolates, which would ensure: the biggest fourth instars, the highest pupae productivity, the shortest duration of the larval stage and the best synchronization of pupation. The density of larvae, the size of breeding water and the quantity of food supplied were tested for their effect on larval growth. Moreover, three cheap foodstuffs were selected and tested for their capability to improve the breeding yield versus TetraMin® as the standard control. The larval density was a very sensitive parameter. Its optimal value, which was found to be ≈1 cm-2 surface area, yielded a daily pupation peak of 38.7% on day 8 post-oviposition, and a global pupae productivity of 78.7% over a duration range of three days. Anopheles gambiae's larval growth, survival and developmental synchronization were density-dependent, and this species responded to overcrowding by producing smaller fourth instars and fewer pupae, over elongated immature lifetime and duration range of pupae occurrence, as a consequence of intraspecific competition. While shallow breeding waters (<3 cm) produced a higher number of pupae than deeper ones, no effect of the breeding habitat's absolute surface area on larval development was observed. Increasing the daily food supply improved the pupae productivity but also boosted the water pollution level (which was assessed by the biological oxygen demand (BOD) and the chemical oxygen demand (COD)) up to a limit depending on the food quality, above which a rapid increase in larval mortality was recorded. The food quality that could substitute the manufactured baby fish food was obtained with weighed mixture of 1 wheat+1 shrimp+2 fish. On establishing an anopheline mosquito colony in the laboratory, special care should be taken to design and maintain the appropriate optimal values of larval density, water depth, daily diet quantity and nutritional quality.

Mapping malaria transmission risk in northern morocco using entomological and environmental data.

Malaria resurgence risk in Morocco depends, among other factors, on environmental changes as well as the introduction of parasite carriers. The aim of this paper is to analyze the receptivity of the Loukkos area, large wetlands in Northern Morocco, to quantify and to map malaria transmission risk in this region using biological and environmental data. This risk was assessed on entomological risk basis and was mapped using environmental markers derived from satellite imagery. Maps showing spatial and temporal variations of entomological risk for Plasmodium vivax and P. falciparum were produced. Results showed this risk to be highly seasonal and much higher in rice fields than in swamps. This risk is lower for Afrotropical P. falciparum strains because of the low infectivity of Anopheles labranchiae, principal malaria vector in Morocco. However, it is very high for P. vivax mainly during summer corresponding to the rice cultivation period. Although the entomological risk is high in Loukkos region, malaria resurgence risk remains very low, because of the low vulnerability of the area.

[Comparison of PCR, ELISA-CSP and direct microscopic observation methods for the detection of Plasmodium falciparum sporozoites in Anopheles gambiae M in Senegal]. / Comparaison des méthodes de la PCR, d'ELISA-CSP et d'observation microscopique directe pour la détection des sporozoïtes de Plasmodium falciparum chez Anopheles gambiae M au Sénégal.

A comparative study between the Enzyme-Linked Immuno Sorbent Assay (ELISA-CSP) for circumsporozoitic antigen detection method, the direct observation after dissection and the polymerase chain reaction (PCR) technique used to identify Plasmodium falciparum genomic DNA markers was carried out. This to evaluate the sensibility and the specificity of the PCR, for the determination of both sporozoitic index (ICSP) and the entomological inoculation rate (EIR). The study is conducted in laboratory on eighty six specimens of Anopheles gambiae M infected after being fed with the blood of a gametocytes carrier from Dielmo (Senegal). Salivary glands of forty-eight specimens randomly selected (test A) among the infected eighty six are microscopically observed after manual dissection for the sporozoites detection. The content of these salivary glands and the crushed head/thorax of the remaining 38 specimens (test B) are tested in ELISA-CSP and PCR. The positive and negative results obtained were recorded and summarized for each method. A pair-comparison of the results obtained with each method generally revealed a good sensibility and an excellent specificity The kappa coefficient (K) of test A indicated a "moderate" to "excellent" concordance between the three different methods performed. By using the crushed head/thorax sample, generally used to determine the transmission parameters (ICSP and EIR), the PCR/ELISA-CSP concordance was excellent. In the light of the values of sensibility and specificity obtained, this PCR is comparable to the other methods for the assessment of sporozoitic index and entomological inoculation rate.

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.

Influence of temperature on immature development, survival, longevity, fecundity, and gonotrophic cycles of Aedes albopictus, vector of chikungunya and dengue in the Indian Ocean.

Aedes albopictus is a mosquito originating from Asia, which has extended its range worldwide the last decades. It is a competent vector for several arboviruses. It was first described in La Réunion (an island of the South West part of the Indian Ocean) in 1913. Since then, it has become the dominant Aedes species and a serious threat to public health, especially during the two last arboviruses outbreaks of dengue (1977) and chikungunya (2005-2006). Despite its pest status, data on the biology of this vector are scarce, especially the population present in the Indian Ocean (IO), which has never been studied in detail. Therefore, the immature development, survival, longevity, fecundity, and gonotrophic cycles of Ae. albopictus were studied for an F2 population of the IO. These biological parameters were studied in controlled conditions at eight constant temperatures (5, 10, 15, 20, 25, 30, 35, and 40 degrees C). The minimal threshold of immature stages development was found at 10.4 degrees C and its optimum at 29.7 degrees C. The shortest periods for immature development were found at 30 degrees C, with in average of 8.8 d. The optimum intrinsic rate of growth (r) was observed between 25 and 30 degrees C. The gonotrophic cycles were also evaluated, and the shortest cycles were found at 30 degrees C (mean, 3.5 d). Those results are according to the field repartition of this species in La Réunion, allowing Ae. albopictus survival at a large range of temperatures.

Entomological investigations in the region of the last malaria focus in Morocco.

To evaluate the risk of malaria transmission resumption in Morocco, we have studied the current level of receptivity of the region of the last malaria focus in the country. Anopheles (Anopheles) maculipennis labranchiae and Anopheles (Cellia) sergentii, the major vectors of malaria in Morocco, are still presents but their anthropopathic index was low and no parasite positive samples were detected. An. labranchiae was very rare; only 34 females were caught over all the study period. The human biting rate was nil and none of its blood meal was human. An. sergenti was more abundant but its low human aggressiveness and its zoophilic behaviour would not attribute to this species an important vectorial capacity. Thus, the receptivity of Chefchaouen province, the region of the last malaria focus in Morocco, under the current vector control measures undertaken by Public Health services, is low and despite the likely presence of Plasmodium vivax gametocyte carriers, the malariogenic potential appears to be low and the risk of malaria resumption is, at this time, unimportant.

La presence des bovins comme hotes alternatifs peut-elle modifier le comportement trophique des vecteurs du paludisme en zone de foret?

Des enquetes entomologiques longitudinales ont ete effectuees de novembre 1998 a novembre 2000 en vue de determiner le comportement trophique des anopheles en presence de bovins comme hotes alternatifs a l`homme dans deux localites de la zone de foret du sud Cameroun; Obala et Nkolbisson. Les collectes des moustiques ont eu lieu a l`interieur des habitations par pulverisation intra domiciliaire et a l`exterieur a l`aide de captures sous moustiquaire piege avec comme appats l`homme et le veau. L`origine du repas de sang et l`infectivite des moustiques ont ete identifiees par la technique ELISA (Enzyme Linked Immunosorbent Asay). Au total; 992 anopheles appartenant a 8 especes ont ete collectees au cours de cette etude. An. funestus etait l`espece la plus abondante a Obala tandis qu`An. nili predominait a Nkolbisson. Le taux d`anthropophilie etait plus eleve chez les moustiques collectes a l`interieur des habitations que chez ceux collectes a l`exterieur sous moustiquaire piege. An. gambiae et An. funestus ont montre une nette preference pour l`homme tandis qu`An. nili a pique indifferemment l`homme et le boeuf. An. gambiae etait de loin le vecteur principal dans les deux sites d`etude avec 24 cas d`infections sur les 42 trouves. Ces resultats montrent que les vecteurs en zone de foret peuvent diversifier la source de leur repas de sang en presence d`hotes alternatifs tels que les bovins. Toutefois; il serait important de comprendre si l`introduction de l`elevage de bovins dans cette region pourrait influencer le cycle de transmission du paludisme

Epidemiological Importance of the Anopheles Nili Group of Malaria Vectors in Equatorial Villages of Cameroon; Central Africa

From March 1999 to March 2007; entomological surveys were conducted in eight riverside locations of the equatorial domain in Cameroon; to assess the contribution of Anopheles nili group to malaria transmission. Results showed that the anthropophilic Anopheles nili; Anopheles carnevalei and Anopheles ovengensis were involved in 5-68of the Plasmodium falciparum entomological inoculation rates (EIRs) in all the study sites. Alongside; vectors such as Anopheles Gambiae; Anopheles moucheti; Anopheles funestus; Anopheles marshallii and Anopheles hancocki also took a substantial part to these transmission rates in sympatric conditions. Anopheles nili s.s. was the major vector in villages situated near the Sanaga river; with more than 100 infective bites/person/year. Its true Plasmodium falciparum infections rates ranged from 1.4to 6.1. Anopheles carnevalei and Anopheles ovengensis were mainly distributed in the equatorial cross-bordering areas; where they contributed for less than 30of the Plasmodium falciparum EIRs in villages located along the Nten river and Congo river networks. These findings confirm Anopheles nili s.s. as a major malaria vector mosquito; and also provided more data regarding the epidemiological role of Anopheles carnevalei and Anopheles ovengensis througout the central African region