Multiple invasions, Wolbachia and human-aided transport drive the genetic variability of Aedes albopictus in the Iberian Peninsula.

The Asian tiger mosquito, Aedes albopictus, is one of the most invasive species in the world. Native to the tropical forests of Southeast Asia, over the past 30 years it has rapidly spread throughout tropical and temperate regions of the world. Its dramatic expansion has resulted in public health concerns as a consequence of its vector competence for at least 16 viruses. Previous studies showed that Ae. albopictus spread has been facilitated by human-mediated transportation, but much remains unknown about how this has affected its genetic attributes. Here we examined the factors that contributed to shaping the current genetic constitution of Ae. albopictus in the Iberian Peninsula, where the species was first found in 2004, by combining population genetics and Bayesian modelling. We found that both mitochondrial and nuclear DNA markers showed a lack of genetic structure and the presence of worldwide dominant haplotypes, suggesting regular introductions from abroad. Mitochondrial DNA showed little genetic diversity compared to nuclear DNA, likely explained by infection with maternally transmitted bacteria of the genus Wolbachia. Multilevel models revealed that greater mosquito fluxes (estimated from commuting patterns and tiger mosquito population distribution) and spatial proximity between sampling sites were associated with lower nuclear genetic distance, suggesting that rapid short- and medium-distance dispersal is facilitated by humans through vehicular traffic. This study highlights the significant role of human transportation in shaping the genetic attributes of Ae. albopictus and promoting regional gene flow, and underscores the need for a territorially integrated surveillance across scales of this disease-carrying mosquito.

Susceptibility of Aedes albopictus and Culex quinquefasciatus to Japanese encephalitis virus.

BACKGROUND: Japanese encephalitis virus (JEV) is the principal cause of mosquito-borne encephalitis in human populations within Asia. If introduced into new geographic areas, it could have further implications for public and animal health. However, potential mosquito vectors for virus transmission have not been fully investigated. The Asian tiger mosquito, Aedes albopictus, has emerged in Europe and is now expanding its geographical range into more northerly latitudes. Culex quinquefasciatus, although absent from Europe, has been detected in Turkey, a country with territory in Europe, and could act as a vector for JEV in other regions. To assess the risk of these invasive species acting as vectors for JEV and therefore potentially contributing to its geographical expansion, we have investigated the vector competence of Ae. albopictus and Cx. quinquefasciatus. METHODS: Two colonised lines of Ae. albopictus (Italy and Spain) and a line of Cx. quinquefasciatus (Tanzania) were compared for susceptibility to infection by oral feeding with JEV strain SA-14, genotype III at 106 PFU/ml and maintained at 25 °C. Specimens were processed at 7 and 14 days post-inoculation (dpi). Rates of infection, dissemination and transmission were assessed through detection of viral RNA by real-time polymerase chain reaction (RT-PCR) in mosquito body, legs and saliva, respectively, at each time point. Where possible, infection and dissemination were confirmed by immunohistochemical (IHC) detection of the JEV envelope protein. RESULTS: Aedes albopictus from Italy showed no susceptibility to infection with JEV strain SA-14. Conversely, Ae. albopictus colonised in Spain was susceptible and 100% of infected mosquitoes that were subjected to saliva screening expressed viral RNA at 14 dpi. Culex quinquefasciatus was highly susceptible to infection as early as 7 dpi and 50% of infected mosquitoes that were subjected to saliva screening expressed viral RNA at 14 dpi. Infection and dissemination were confirmed in Cx. quinquefasciatus by IHC detection of JEV envelope protein in both the mid-gut and salivary glands. CONCLUSIONS: Aedes albopictus from two different locations in Europe range from being susceptible to JEV and capable of transmission through to being resistant. Culex quinquefasciatus also appears highly susceptible; therefore, both species could potentially act as vectors for JEV and facilitate the emergence of JEV into new regions.

Confirmed presence of aedes (rusticoidus) refiki Medschid, 1928 in a continental dry Mediterranean peri-urban environment in south-central Spain.

BACKGROUND: The 'snow-melt mosquito' aedes (rusticoidus) refiki is a rare species with a wide distribution in Europe that is usually defined as an aggressive mosquito for mammals, including humans. During a mosquito survey in a peri-urban area in south-central mainland Spain, adult Ae. refiki females were captured and identified by morphological traits. The presence of this species of mosquito has never been molecularly confirmed under continental dry Mediterranean climatic influence with scarce number of days with snow on soil. The aim of this study was to confirm by amplification and sequencing of mitochondrial cytochrome c oxidase subunit I (COI) and internal transcribed spacer 2 (ITS2) region. RESULTS: We also successfully amplified and typed the species molecularly by COI and ITS2 regions. The peri-urban area where Ae. refiki was found contrasts with the reported cold, humid and snowy environments required by the species to breed. CONCLUSIONS: This finding suggests that the species is already adapted to continental dry Mediterranean environments, questioning whether it is a truly stenotopic species of cold snowy environments.

AIMSurv: First pan-European harmonized surveillance of Aedes invasive mosquito species of relevance for human vector-borne diseases.

Human and animal vector-borne diseases, particularly mosquito-borne diseases, are emerging or re-emerging worldwide. Six Aedes invasive mosquito (AIM) species were introduced to Europe since the 1970s: Aedes aegypti, Ae. albopictus, Ae. japonicus, Ae. koreicus, Ae. atropalpus and Ae. triseriatus. Here, we report the results of AIMSurv2020, the first pan-European surveillance effort for AIMs. Implemented by 42 volunteer teams from 24 countries. And presented in the form of a dataset named "AIMSurv Aedes Invasive Mosquito species harmonized surveillance in Europe. AIM-COST Action. Project ID: CA17108". AIMSurv2020 harmonizes field surveillance methodologies for sampling different AIMs life stages, frequency and minimum length of sampling period, and data reporting. Data include minimum requirements for sample types and recommended requirements for those teams with more resources. Data are published as a Darwin Core archive in the Global Biodiversity Information Facility- Spain, comprising a core file with 19,130 records (EventID) and an occurrences file with 19,743 records (OccurrenceID). AIM species recorded in AIMSurv2020 were Ae. albopictus, Ae. japonicus and Ae. koreicus, as well as native mosquito species.

Progressive Invasion of Aedes albopictus in Northern Spain in The Period 2013-2018 and A Possible Association with the Increase in Insect Bites.

1) Background: Aedes albopictus has rapidly expanded throughout Europe, becoming a public health concern in the Mediterranean Basin. 2) Methods: Following the detection of Ae. albopictus in the southwestern French region of Aquitaine in 2012, an entomological surveillance programme was implemented in the Basque Country (Northern Spain) in 2013. 3) Results: Ae. albopictus eggs were first detected in 2014 in a transited parking area in the northeastern sampling point, 22 km away from the nearest French site with recorded presence of tiger mosquito. At this site, eggs were found throughout the study (2014-2018). Other western and southern municipalities became positive in 2017 and 2018. Ae. albopictus adults were first captured in 2018 by aspiration of the vegetation in an area where eggs had been detected since 2015, suggesting a progressive establishment of a self-sustained population. Incidence of insect bites in humans was roughly constant over the study period except for a significant increase in 2018 in the Health County where eggs had been detected since 2014. Densities of Ae. albopictus eggs in positive areas remained at similar levels over the years. 4) Conclusion: Multiple approaches and standardized methods are necessary to successfully control this vector.

A randomized, blinded, controlled trial to assess sand fly mortality of fluralaner administered orally in dogs.

BACKGROUND: Leishmania infantum is the parasite responsible for the disease in humans known as zoonotic visceral leishmaniasis (ZVL). Dogs are considered the main domestic reservoir of ZVL and sand flies are the proven vectors. The use of systemic insecticides in dogs has been studied as an alternative strategy to control ZVL in endemic areas. One systemic insecticide in dogs, fluralaner, has a proven anti-sand fly effect in membrane-fed studies. However, the efficacy and duration on sand flies directly feeding from dogs treated with fluralaner remains unknown. METHODS: Direct feeding bioassays were performed on 10 beagle dogs that had been randomly assigned to two groups: one with five dogs orally treated with Bravecto® (fluralaner) and other five as a control. About 30 females of Phlebotomus papatasi were allowed to directly feed from dogs at seven days before the administration of the treatment and Days 3, 17, 31, 45 and 73 post-treatment. Sand fly mortality after feeding was observed every 24 h for 5 days. The Kaplan-Meyer method, Henderson-Tilton formula and a negative binomial mixed model were used to respectively calculate: (i) mortality and its 95% confidence interval (CI); (ii) efficacy of the insecticide at killing sand flies in 24 h; and (iii) differences in the risk of sand fly death at 24 h after feeding. RESULTS: Control sand fly mortality 24 h after feeding was always ≤ 20% and mortality in the fluralaner group ranged from 2% (95% CI: 0-4%) 7 days before treatment to 100% at 3 days post-treatment. Fluralaner efficacy was 100, 93, 94 and 75% at Days 3, 17, 31 and 45, respectively (P < 0.0001). The increase in the risk of sand fly death was 32.9 (95% CI: 4-263), 76 (95% CI: 8-705), 95.8 (95% CI: 9-1029) and 10.6 times (95% CI: 1.43-79) on Days 3, 17, 31 and 45, respectively CONCLUSIONS: The efficacy of fluralaner, orally administered to dogs, against sand-flies was above 90% for 31 days. Fluralaner administered to dogs should be further evaluated as a control strategy in ZVL endemic areas.

Citizen science provides a reliable and scalable tool to track disease-carrying mosquitoes.

Recent outbreaks of Zika, chikungunya and dengue highlight the importance of better understanding the spread of disease-carrying mosquitoes across multiple spatio-temporal scales. Traditional surveillance tools are limited by jurisdictional boundaries and cost constraints. Here we show how a scalable citizen science system can solve this problem by combining citizen scientists' observations with expert validation and correcting for sampling effort. Our system provides accurate early warning information about the Asian tiger mosquito (Aedes albopictus) invasion in Spain, well beyond that available from traditional methods, and vital for public health services. It also provides estimates of tiger mosquito risk comparable to those from traditional methods but more directly related to the human-mosquito encounters that are relevant for epidemiological modelling and scalable enough to cover the entire country. These results illustrate how powerful public participation in science can be and suggest citizen science is positioned to revolutionize mosquito-borne disease surveillance worldwide.

First Data on Resistance to Pyrethroids in Wild Populations of Aedes albopictus from Spain.

The invasive mosquito, Aedes albopictus, found in Spain since 2004, is a competent vector of yellow fever, Zika, dengue, and chikungunya viruses among other diseases. Although controversial, the use of adulticiding is a relevant tool for vector control and could be crucial for the management of any possible outbreak of imported diseases. We present the 1st study in Spain on the susceptibility of field populations from Barcelona, Peñíscola, Castellón, and Mallorca of Ae. albopictus to several pyrethroids using either bioassays under the World Health Organization methodology and biochemical tests. In the bioassays, the discriminating concentrations were calculated using a local, susceptible laboratory strain. Different susceptibility levels were found for some combinations of populations and products. The biochemical tests carried out by enzymatic analysis supported these results, showing an overexpression of glutathione S-transferase activity in 1 population.

Review of ten-years presence of Aedes albopictus in Spain 2004-2014: known distribution and public health concerns.

Ten years have gone by since the Asian tiger mosquito Aedes albopictus was recorded for the first time in Spain. In this paper, all relevant published information about this vector in Spain for the period 2004-2014 is reviewed. The known distribution for 2014 is provided, including all historical records (published and unpublished data) and the results from samplings of the last year. The consequences on public health about the presence of the Asian tiger mosquito in Spain are also highlighted. Further, legal aspects and control plans related to the management and diseases transmitted by this invasive vector species are also discussed.

Characterization of Aedes albopictus akirin for the control of mosquito and sand fly infestations.

The control of arthropod vectors of pathogens that affect human and animal health is important for the eradication of vector-borne diseases. Recent evidences showed a reduction in the survival and/or fertility of mosquitoes, sand flies and poultry red mites fed in vitro with antibodies against the recombinant Aedes albopictus akirin. These experiments were the first step toward the development of a multi-target arthropod vaccine. In this study, we showed that the oviposition of A. albopictus and Phlebotomus perniciosus fed on mice vaccinated with recombinant A. albopictus akirin was reduced by 17% and 31%, respectively when compared to controls. However, Aedes aegypti mosquitoes were not affected after feeding on vaccinated mice. These results showed that recombinant A. albopictus akirin could be used to vaccinate hosts for the control of mosquito and sand fly infestations and suggested new experiments to develop improved vaccine formulations.