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Sporadic outbreaks of human cases of West Nile virus (WNV), primarily vectored by Culex quinquefasciatus Say in suburban and urban areas, have been reported since introduction of the virus into Florida in 2001. Miami-Dade County, Florida is part of one of the largest metropolitan areas in the United States, supports Cx. quinquefasciatus year-round, and recently experienced over 60 human cases of WNV during one outbreak. To facilitate more effective integrated vector management and public health protection, we used the Centers for Disease Control and Prevention (CDC) bottle bioassay method to evaluate the susceptibility of adult Cx. quinquefasciatus collected from 29 locations throughout Miami-Dade County to pyrethroid and organophosphate adulticide active ingredients (AIs) used by Miami-Dade County Mosquito Control. We also determined the frequency of the 1014 knockdown resistance (kdr) mutation for Cx. quinquefasciatus from a subset of 17 locations. We detected resistance to two pyrethroid AIs in all tested locations (permethrin: 27 locations, deltamethrin: 28 locations). The 1014F allele was widely distributed throughout all 17 locations sampled; however, 29.4% of these locations lacked 1014F homozygotes even though phenotypic pyrethroid resistance was present. Organophosphate resistance was more variable; 20.7% of the locations tested were susceptible to malathion, and 33.3% of the populations were susceptible to naled. We subsequently conducted a field trial of ReMoa Tri, a recently approved multiple AI adulticide formulation labelled for resistant mosquitoes, against a mixed location field population of Miami-Dade Cx. quinquefasciatus. Average 24-hr mortality was 65.1 ± 7.2% and 48-hr mortality increased to 85.3 ± 9.1%, indicating good control of these resistant Cx. quinquefasciatus. This current study shows that insecticide resistance is common in local Cx. quinquefasciatus but effective options are available to maintain control during active disease transmission in Miami-Dade County.

BackgroundAedes (Stegomyia) albopictus (Skuse) impacts human outdoor activity because of its aggressive biting behavior, and as a major vector of mosquito-borne diseases, it is also of public health importance. Although most mosquito species exhibit crepuscular activity by primarily host seeking at dawn and dusk, Ae. albopictus has been traditionally characterized as a diurnal or day-biting mosquito. With the global expansion and increased involvement of Ae. albopictus in mosquito-borne diseases, it is imperative to elucidate the diel activity of this species, particularly in newly invaded areas.Methodology and principal findingsHuman sweep netting and carbon dioxide-baited rotator traps were used to evaluate the diel activity of Ae. albopictus in two study sites. Both trapping methods were used in New Jersey's Mercer County, USA (temperate/urban), while only human sweep netting was used in Florida's Volusia County, USA (subtropical/suburban). Human sweep netting was performed to determine adult mosquito activity at Sunrise, Solar Noon, Sunset, and Lunar Midnight. Because New Jersey is in a temperate area, diel activity was investigated during the early season (3-19 July), peak season (25 July-19 September), and late season (22 September- 22 October). Aedes albopictus showed the highest activity during peak and late seasons at Solar Noon (P < 0.05). At Sunrise and Sunset during the peak season, Ae. albopictus activity was similar. Lunar Midnight activity was significantly lower than Sunrise and Solar Noon (P < 0.05) but was similar to that of Sunset. In the late season, the highest activity was observed during Solar Noon while the least activity was observed during Sunrise and Lunar Midnight (PConclusion/significanceInvolvement of Ae. albopictus in the transmission of debilitating mosquito-borne pathogens such as chikungunya, dengue, and Zika virus, coupled with its affinity to thrive in human peridomestic environments, substantiates that our findings have global implications in areas where Ae. albopictus populations established. It also highlights the importance of behavioral studies of vector species which will not only help mosquito control professionals plan the timing of their control efforts but also provides empirical evidence against conventional wisdoms that may unjustly persist within public health stewards.

We constructed an electric multi-rotor autonomous unmanned aerial system (UAS) to perform mosquito control activities. The UAS can be equipped with any of four modules for spraying larvicides, dropping larvicide tablets, spreading larvicide granules, and ultra-low volume spraying of adulticides. The larvicide module sprayed 124 μm drops at 591 mL/min over a 14 m swath for a total application rate of 1.6 L/ha. The tablet module was able to repeatedly deliver 40-gram larvicide tablets within 1.1 m of the target site. The granular spreader covered a 6 m swath and treated 0.76 ha in 13 min at an average rate of 1.8 kg/ha. The adulticide module produced 16 μm drops with an average deposition of 2.6 drops/mm2. UAS pesticide applications were made at rates prescribed for conventional aircraft, limited only by the payload capacity and flight time. Despite those limitations, this system can deliver pesticides with much greater precision than conventional aircraft, potentially reducing pesticide use. In smaller, congested environments or in programs with limited resources, UAS may be a preferable alternative to conventional aircraft.

We constructed an electric multi-rotor autonomous unmanned aerial system (UAS) to perform mosquito control activities. The UAS can be equipped with any of four modules for spraying larvicides, dropping larvicide tablets, spreading larvicide granules, and ultra-low volume spraying of adulticides. The larvicide module sprayed 124 μm drops at 591 mL/min over a 14 m swath for a total application rate of 1.6 L/ha. The tablet module was able to repeatedly deliver 40-gram larvicide tablets within 1.1 m of the target site. The granular spreader covered a 6 m swath and treated 0.76 ha in 13 min at an average rate of 1.8 kg/ha. The adulticide module produced 16 μm drops with an average deposition of 2.6 drops/mm2. UAS pesticide applications were made at rates prescribed for conventional aircraft, limited only by the payload capacity and flight time. Despite those limitations, this system can deliver pesticides with much greater precision than conventional aircraft, potentially reducing pesticide use. In smaller, congested environments or in programs with limited resources, UAS may be a preferable alternative to conventional aircraft.

BACKGROUND:Usutu virus (USUV) is a member of the Japanese encephalitis virus (JEV) serocomplex in the Flaviviridae family. Emergence of USUV in Europe has led to disease burdens in birds and created increasing concern for the potential zoonotic transmission to humans. Whilst USUV has not been detected in the New World, the identification of competent vector species in North America is critical in the assessment of the likelihood of its dispersal and establishment of enzootic transmission cycles. The objective of this study was to determine vector competence of potential mosquito vectors in North America for USUV. Three medically important mosquito species were selected for testing because of their involvement in the transmission of West Nile virus and St. Louis encephalitis virus, two related JEV serocomplex flaviviruses in the New World. METHODOLOGY/PRINCIPAL FINDINGS:Oral challenge of Culex pipiens, Culex quinquefasciatus, and Aedes albopictus was performed to determine the susceptibility and vector competence of North American mosquitoes for USUV. Infection status was monitored by the isolation of virus from homogenized mosquito tissues. The disseminated form of infection was demonstrated by the detection of infectious virus in the head, wings, and legs of infected mosquitoes. The presence of viral RNA in saliva of infected Cx. pipiens and Cx. quinquefasciatus indicated that both species are competent for transmission of USUV. CONCLUSIONS/SIGNIFICANCE:Results indicate that members of the Cx. pipiens complex are susceptible to USUV and competent for its transmission potential in North America in the event of its introduction. In contrast, Ae. albopictus were highly refractory to USUV infection, suggesting that this species is unlikely to contribute to USUV transmission in North America.

Aedes albopictus is a major nuisance pest and also a public health concern because of the role it plays in the transmission of arboviruses. There is a continuing demand for effective surveillance tools for this species. The first generation of Biogents Sentinel (BGS1) traps have proven to be an effective tool for surveillance of Ae. albopictus throughout its range, however, some defects in construction led to the eventual development of the next generation. We compared the performance of the new generation prototype trap (BGS2P) to the original. Studies were conducted in suburban and urban areas in Florida, Louisiana, New Jersey, and Virginia, USA in the summer of 2014 (July-October). BGS1 traps collected significantly more Ae. albopictus when compared to the BGS2P with or without CO2 in all locations (P

BACKGROUND:The Asian tiger mosquito, Aedes albopictus, is a vector of dengue, Chikungunya, and Zika viruses. This mosquito inhabits a wide range of artificial water-holding containers in urban and suburban areas making it difficult to control. We tested the hypothesis that female-driven autodissemination of an insect growth regulator could penetrate cryptic oviposition habitats difficult to treat with conventional insecticidal sprays. METHODOLOGY:Oviposition preferences of Ae. albopictus females for open and cryptic cups were tested in semi-field experiments. Two conventional larvicidal sprayers were tested to determine droplet penetration and larvicidal efficacy in open and cryptic habitats using Bacillus thuringiensis var. israelensis (Bti) in the field. Finally, the efficacy of pyriproxyfen autodissemination stations was assessed in cryptic and open cups in residential areas during 2013 and 2014. PRINCIPAL FINDINGS:Gravid females strongly preferred cryptic (53.1±12.9 eggs/cup) over open (10.3±4.3 eggs/cup) cups for oviposition. Cryptic cups showed limited droplet penetration and produced 0.1-0.3% larval mortality with a conventional backpack and low-volume sprays of Bti. The autodissemination stations effectively contaminated these cryptic cups (59.3-84.6%) and produced 29.7-40.8% pupal mortality during 2013-2014. Significant pupal mortality was also observed in open cups. CONCLUSIONS:The autodissemination station effectively exploits the oviposition behavior of wild gravid females to deliver pyriproxyfen to targeted oviposition habitats. Although the pupal mortality in cryptic cups was relatively lower than expected for the effective vector control. Autodissemination approach may be a suitable supporting tool to manage Ae. albopictus immatures in the cryptic habitats those are less accessible to conventional larvicidal sprays.

Adult control of Aedes albopictus via ultra-low volume is difficult because this species occurs primarily in peridomestic habitats where obstacles such as buildings and vegetation can disrupt spray plumes and droplet dispersion. We determined droplet penetration and characterization of a pyrethroid adulticide applied from the ground at mid (46.77 ml/ha) and maximum (93.53 ml/ha) label rates within cryptic habitats of urban and suburban environments. Droplets were collected from all habitats, with no significant differences detected between locations within the same application rate or collection method. No differences were detected in droplet densities (drops per mm2) between rates within urban environments, but more droplets were collected in urban (149.93 ± 11.07 SE) than suburban sites (114.37 ± 11.32) at the maximum label rate (P = 0.003). The excellent penetration of aerosols into cryptic habitats of an urban site was likely due to the shorter spray paths afforded by our network of roads and alleys. Mid label rates displayed similar droplet density values as maximum label rates in urban areas, indicating that lower rates may be used effectively to reduce costs, lessen non-target effects, and increase environmental stewardship. Advances in formulations and technology are driving changes in adulticide applications, leading to use of the minimum effective dose for maximum efficacy, precision, and accountability.

Aedes albopictus is an invasive species which continues expanding its geographic range and involvement in mosquito-borne diseases such as chikungunya and dengue. Host selection patterns by invasive mosquitoes are critically important because they increase endemic disease transmission and drive outbreaks of exotic pathogens. Traditionally, Ae. albopictus has been characterized as an opportunistic feeder, primarily feeding on mammalian hosts but occasionally acquiring blood from avian sources as well. However, limited information is available on their feeding patterns in temperate regions of their expanded range. Because of the increasing expansion and abundance of Ae. albopictus and the escalating diagnoses of exotic pathogens in travelers returning from endemic areas, we investigated the host feeding patterns of this species in newly invaded areas to further shed light on its role in disease ecology and assess the public health threat of an exotic arbovirus outbreak.We identified the vertebrate source of 165 blood meals in Ae. albopictus collected between 2008 and 2011 from urban and suburban areas in northeastern USA. We used a network of Biogents Sentinel traps, which enhance Ae. albopictus capture counts, to conduct our collections of blooded mosquitoes. We also analyzed blooded Culex mosquitoes collected alongside Ae. albopictus in order to examine the composition of the community of blood sources. We found no evidence of bias since as expected Culex blood meals were predominantly from birds (n = 149, 93.7%) with only a small proportion feeding on mammals (n = 10, 6.3%). In contrast, Aedes albopictus fed exclusively on mammalian hosts with over 90% of their blood meals derived from humans (n = 96, 58.2%) and domesticated pets (n = 38, 23.0% cats; and n = 24, 14.6% dogs). Aedes albopictus fed from humans significantly more often in suburban than in urban areas (χ(2), p = 0.004) and cat-derived blood meals were greater in urban habitats (χ(2), p = 0.022). Avian-derived blood meals were not detected in any of the Ae. albopictus tested.The high mammalian affinity of Ae. albopictus suggests that this species will be an efficient vector of mammal- and human-driven zoonoses such as La Crosse, dengue, and chikungunya viruses. The lack of blood meals obtained from birds by Ae. albopictus suggest that this species may have limited exposure to endemic avian zoonoses such as St. Louis encephalitis and West Nile virus, which already circulate in the USA. However, growing populations of Ae. albopictus in major metropolitan urban and suburban centers, make a large autochthonous outbreak of an arbovirus such as chikungunya or dengue viruses a clear and present danger. Given the difficulties of Ae. albopictus suppression, we recommend that public health practitioners and policy makers install proactive measures for the imminent mitigation of an exotic pathogen outbreak.

Mosquito species that utilize peridomestic containers for immature development are commonly aggressive human biters, and because they often reach high abundance, create significant nuisance. One of these species, the Asian tiger mosquito Aedes albopictus, is an important vector of emerging infectious diseases, such as dengue, chikungunya, and Zika fevers. Integrated mosquito management (IMM) of Ae. albopictus is particularly difficult because it requires access to private yards in urban and suburban residences. It has become apparent that in the event of a public health concern due to this species, homeowners will have to be active participants in the control process by reducing mosquito habitats in their properties, an activity known as source reduction. However, limited attempts at quantifying the effect of source reduction by homeowners have had mixed results. Of note, many mosquito control programs in the US have some form of education outreach, however the primary approach is often passive focusing on the distribution of education materials as flyers. In 2010, we evaluated the use of active community peer education in a source reduction program, using AmeriCorps volunteers. The volunteers were mobilized over a 4-week period, in two areas with approximately 1,000 residences each in urban Mercer and suburban Monmouth counties in New Jersey, USA. The volunteers were first provided training on peridomestic mosquitoes and on basic approaches to reducing the number of container habitats for mosquito larvae in backyards. Within the two treatment areas the volunteers successfully engaged 758 separate homes. Repeated measures analysis of variance showed a significant reduction in container habitats in the sites where the volunteers actively engaged the community compared to untreated control areas in both counties. Our results suggest that active education using community peer educators can be an effective means of source reduction, and a critical tool in the arsenal against peridomestic mosquitoes.

The increasing range of Aedes albopictus, the Asian tiger mosquito, in the USA and the threat of chikungunya and dengue outbreaks vectored by this species have necessitated novel approaches to control this peridomestic mosquito. Conventional methods such as adulticiding provide temporary relief, but fail to manage this pest on a sustained basis. We explored the use of cold aerosol foggers and misting machines for area-wide applications of Bacillus thuringiensis var. israelensis (VectoBac WDG) as a larvicide targeting Aedes albopictus. During 2010-2013 we performed initially open field trials and then 19 operational area-wide applications in urban and suburban residential areas in northeastern USA to test three truck-mounted sprayers at two application rates. Area-wide applications of WDG in open field conditions at 400 and 800 g/ha killed on average 87% of tested larvae. Once techniques were optimized in residential areas, applications with a Buffalo Turbine Mist Sprayer at a rate of 800 g/ha, the best combination, consistently provided over 90% mortality. Importantly, there was no significant decrease in efficacy with distance from the spray line even in blocks of row homes with trees and bushes in the backyards. Under laboratory conditions Bti deposition in bioassay cups during the operational trials resulted in over 6 weeks of residual control. Our results demonstrate that area-wide truck mounted applications of WDG can effectively suppress Ae. albopictus larvae and should be used in integrated mosquito management approaches to control this nuisance pest and disease vector.

Our ultimate objective is to design cost-effective control strategies for Aedes albopictus, the Asian tiger mosquito, an important urban nuisance and disease vector that expanded worldwide during the last 40 years. We conducted mosquito larval surveys from May through October 2009 in the City of Trenton, New Jersey, USA, while performing intensive monthly source-reduction campaigns that involved removing, emptying, or treating all accessible containers with larvicides and pupicides. We examined patterns of occurrence of Ae. albopictus and Culex pipiens, another urban mosquito, among different container types by comparing observed and expected number of positive containers of each type. Expected use was based on the relative frequency of each container type in the environment. Aedes albopictus larvae and pupae were found significantly more often than expected in medium volumes of water in buckets and plant saucers but were rarely collected in small volumes of water found in trash items such as discarded cups and cans. They were also absent from large volumes of water such as in abandoned swimming pools and catch basins, although we consistently collected Cx. pipiens from those habitats. The frequency of Ae. albopictus in tires indicated rapid and extensive use of these ubiquitous urban containers. Standard larval-based indices did not correlate with adult catches in BG-Sentinel traps, but when based only on Ae. albopictus key containers (buckets, plant saucers, equipment with pockets of water, and tires) they did. Although we found that only 1.2% of the 20,039 water-holding containers examined contained immature Ae. albopictus (5.3% if only key containers were counted), adult populations were still above nuisance action thresholds six times during the 2009 mosquito season. We conclude that in urban New Jersey, effective source reduction for Ae. albopictus control will require scrupulous and repeated cleaning or treatment of everyday use containers and extensive homeowner collaboration.

The Biogents® Sentinel (BGS) trap is the standard tool to monitor adult Aedes (Stegomyia) albopictus (Skuse) (Diptera: Culicidae), the Asian tiger mosquito. BGS traps are commonly placed in residential properties during surveillance operations, but locations within properties may have significant differences in ambient light, temperature, and humidity (e.g. between a sunlit lawn and shady underbrush). We examined the effect of BGS trap placement on Ae. albopictus capture rates in three residential properties in Monmouth County, New Jersey, USA. In each property we visually selected locations as shade, partial shade, and sun. Traps in "partial shade" locations were under vegetation and were exposed to filtered sunlight during some parts of the day while "shaded" locations were never exposed to direct sunlight. Locations defined as "sun" were exposed to direct sunlight for large parts of the day. We placed a BGS trap in each of the three location types and used small data loggers to measure temperature, relative humidity, and light exposure at each trap during a 24-hour deployment. To address temporal variability, we made seven separate measurements from 31 August to 22 September 2010. We found that "partial shade" and "full shade" locations did not differ but that "full sun" locations had significantly higher light exposure, higher temperature, and lower humidity. Importantly, Ae. albopictus catches (males, females, or both) were consistently and significantly over 3 times higher in traps located in shaded locations. To further investigate the effects of local temperature and humidity on surveillance we examined Ae. albopictus collections from 37 BGS traps fitted with data loggers and deployed weekly from August through mid October, during the 2009 season, in three urban sites in Mercer County, NJ. We confirmed that local climate influences capture rates and that Ae. albopictus surveillance projects need to monitor trap placement carefully for maximum efficiency.

Aedes albopictus, the Asian tiger mosquito, continues expanding its geographic range and involvement in mosquito-borne diseases such as chikungunya and dengue. Vector control programs rarely attempt to suppress this diurnal species with an ultra-low volume (ULV) adulticide because for maximum efficacy applications are conducted at night. During 2009-2011 we performed experimental nighttime applications of a novel adulticide (DUET®) against field populations of Ae. albopictus within an urban site composed of approximately 1,000 parcels (home and yard) in northeastern USA. Dual applications at mid label rate of the adulticide spaced one or two days apart accomplished significantly higher control (85.0 ± 5.4% average reduction) than single full rate applications (73.0 ± 5.4%). Our results demonstrate that nighttime ULV adulticiding is effective in reducing Ae. albopictus abundance and highlight its potential for use as part of integrated pest management programs and during disease epidemics when reducing human illness is of paramount importance.