A CASE REPORT ON PRODUCT ROTATION TO MANAGE SEVERE LYSINIBACILLUS SPHAERICUS RESISTANCE IN CULEX PIPIENS FROM SALT LAKE CITY, UTAH.

The Salt Lake City Mosquito Abatement District (SLCMAD) detected a 20,000-fold resistance to Lysinibacillus sphaericus (Lsph) in Culex pipiens occurring in catch basins of Salt Lake City during 2016. In response, SLCMAD suspended use of Lsph and rotated use of spinosyn and s-methoprene products for the next three years. At the end of the third year, Lsph was evaluated again and efficacy similar to susceptible colony strains. During the second year of Lsph use, technicians observed lack of control of larvae at some urban sites. Bioassays performed during 2021 showed recurrence of some resistance to Lsph to varying degrees across SLCMAD urban areas. The rapidity with which resistant phenotypes reemerged clarifies that SLCMAD cannot in the near future rely on repeated use of Lsph, even after suspending use for three years and using within-season product rotations. Prior reports in other research groups have found long-term selection to Lsph, as is the case at SLCMAD, to not regress in spite of halting use of the products. However, our findings offer some optimism that regression may be relatively quick. More operational review is needed, and future work should characterize resistance alleles in field populations. Collectively, there is a lack of concrete data supporting the prevailing assumptions from adjacent industries that were adopted into mosquito abatement. We provide this short note as additional guidance for mosquito and vector control districts weighing options to remediate Lsph resistance.

Blinded by the light: does heat or light enhance wild mosquito (Diptera: Culicidae) attraction to CO2-baited traps in the Great Salt Lake area?

The New Jersey Light Trap has been among the earliest trap models used for mosquito surveillance in the United States. This trap was modernized in the 1950s to the miniature CDC light trap, with the addition of CO2 following soon after. The incandescent light has the tendency to attract nontarget insects, as well as losing a substantial portion of their energy as heat. Few studies have delineated whether heat or light in isolation make a difference in field collections using the former traps within the United States. Our study focused on isolating heat and light variables by using incandescent bulbs, light emitting diode (LED) bulbs, and electric heating patches affixed to a base model CO2 trap as designed at the Salt Lake City Mosquito Abatement District. Sites were selected in the urban and suburban foothills and canyons of the Wasatch Mountain front, industrial areas near the Salt Lake City International Airport, and rural wetlands in the marshes outlying the Great Salt Lake. Five traps were replicated within each sector during the summer and fall summer seasons. Collections were composed of Aedes dorsalis (Meigen), Culex pipiens L., Culex tarsalis Coquillett, and Culiseta inornata (Williston). Composition changes were a result of seasonal, rather than spatial, shifts. The results showed that LED light traps depressed collections of key species. Otherwise, there were negligible differences in collections among incandescent, heat film, and base model traps. In the Intermountain West, the miniature CDC trap is reliable enough to make programmatic decisions even if light usage varies by district.

Diuresis and α-glucosidase inhibition by erythritol in Aedes aegypti (Diptera: Culicidae) and viability for efficacy against mosquitoes.

BACKGROUND: Sugar alcohols, such as erythritol, are low-impact candidates for attractive toxic sugar baits (ATSB) to kill mosquitoes. To determine whether erythritol has a viable future in ATSB formulations, a suite of assays was conducted to diagnose toxicity mechanisms and starvation effects on mortality in Aedes aegypti (L.) as a model system. METHODS: We measured general carbohydrate load, glucosidase levels, and free glucose in intoxicated adult mosquitoes to observe whether sugar digestion was impaired. We assayed the effects of sugar combinations with erythritol on larvae and adults. To measure erythritol effects when mosquitoes were not resource-deprived, additional assays manipulated the prior starvation status. RESULTS: Up to 50,000 ppm of erythritol in water had no effect on larvae within 72 h, but an ammonia spike indicated diuresis in larvae as early as 4 h (F8,44 = 22.50, P < 0.0001) after sucrose/erythritol combinations were added. Adult consumption of erythritol was diuretic regardless of the sugar pairing, while sucrose and erythritol together generated above 80% mortality (F2,273 = 33.30, P < 0.0001) alongside triple the normal excretion (F5,78 = 26.80, P < 0.0004). Glucose and fructose paired individually with erythritol had less mortality, but still double the fecal excretion. When ingesting erythritol-laced meals, less sugar was detected in mosquitoes as compared to after sucrose meals (χ2 = 12.54, df = 1, P = 0.0004). CONCLUSIONS: Data showed that erythritol is a linear competitive inhibitor of α-glucosidase, marking it as a novel class of insecticide in the current research climate. However, the efficacy on larvae was null and not persistent in adult mosquitoes when compared across various starvation levels. Despite significant diuresis, the combined effects from erythritol are not acute enough for vector control programs considering ATSB against mosquitoes.

Dancing in the purple rain: color affinity and oviposition choices in Aedes sierrensis (Diptera: Culicidae).

The western tree hole mosquito, Aedes sierrensis (Ludlow) (Diptera: Clucidae), is a pestiferous mosquito with a range extending over the entire pacific seaboard and into portions of the intermountain west. As a peridomestic heartworm vector, it demands at least some level of surveillance to understand its abundance. However, the species is refractory to a majority of conventional vector surveillance approaches for tracking mosquitoes. To find more options for Aedes sierrensis surveillance, a variety of oviposition attractants were evaluated in arena-style choice assays using colony reared adults. A range of infusion treatments (e.g., alfalfa, oak, and beetroot) were examined and then combined with investigations of liquid color as well as ovicup color and entryway position. These studies revealed that Ae. sierrensis have an affinity for purple coloration, plain water, and larger entryway sizes for oviposition cups. A prototype ovicup was 3D-printed using purple filament and multiple types of entryways, and used to re-test infusion waters. No particular attraction differences were detected after normalizing for purple color. Comparisons to black 3D-printed cups yielded surprising observations that male mosquitoes also aggregated on purple cups while females sheltered, but not necessarily oviposited, in black cups. Although this was only a laboratory-based assessment, these studies provide useful information for future field trials of potential oviposition traps for surveillance of Ae. sierrensis.

Do it yourself: 3D-printed miniature CDC trap for adult mosquito (Diptera: Culicidae) surveillance.

The central component of mosquito and vector surveillance programs globally is the adult mosquito trap, which is intended to collect host-seeking mosquitoes. The miniature CDC trap is a widely distributed trap style in part due to its relative affordability and compact nature. Despite already being a simple trap, in-house production methods, such as 3D printing, could improve the accessibility of the CDC trap by eliminating some of the supply chain variables. We present here several trials with the Salt Lake City (SLC) trap, a three-dimensional (3D) printed trap design. Functional assessments were made on secondary components and found no statistically significant differences when comparing CO2 line height (above vs. below fan), battery types (sealed lead acid vs. USB battery pack), and trap body collection shape (funnel body vs. simple/straight body). The SLC trap was compared directly to a commercial equivalent, the ABC trap, with comparative assessment on species diversity and evenness in collections and found to be statistically equivalent on all metrics. Methods also detail an accompanying optional transport system for a pressurized CO2/regulator set-up, should a practitioner elect not to use dry ice. Our final design is presented here with the publicly published stereolithography (STL) files and a detailed outline of the transport container system. Alternative models are available for in-house manufacture of mosquito traps, and we contribute these designs in an effort to stimulate further growth in vector surveillance.

Do It Yourself: A Modernized Gravid Trap Design for Mosquito Surveillance.

Gravid traps have become a common and frequently essential surveillance tool for parous Culex spp. vectors of West Nile virus and other encephalitis-causing pathogens. The recent closing of BioQuip Products Inc., an entomological supply company, has jeopardized the commercial availability of gravid traps. The Salt Lake City Mosquito Abatement District presents herein a template for making your own gravid trap, but with some modernizations for quieter fans and longer lasting, light weight, lithium battery packs. At the time of writing, the materials cost for the fan ($14 USD), toolbox ($13), cables ($9), ABS pipe ($2.50), aluminum brackets ($10), catch container with lid ($9), trap net ($10), USB battery pack ($35) and the negligible amount of 3D-printed filament ($2), is approximately half the cost (not including labor) of the formerly available commercial model. Additionally, performance validation in the laboratory (t4,9 = 0.1191, P < 0.9109) and within two field sites (χ2 = 0.107, P < 0.744) demonstrated no significant differences in collections of gravid Culex pipiens. We do not present an overhaul of the previous gravid trap blueprint, but the quality-of-life updates to the trap design, the feasibility of in-house manufacture, and the mirrored collection efficacy to the commercial model can allow improved maintenance of gravid trap surveillance networks without a commercial supplier.

Evaluation of An Aerial Application of Duet Hd® Against Aedes Dorsalis and Culex Tarsalis in Rural Habitats of the Great Salt Lake, Utah.

The Salt Lake City Mosquito Abatement District (SLCMAD) has been conducting aerial applications using an organophosphate insecticide against adult mosquitoes for several decades. In order to evaluate a potential rotation product, aerial applications of Duet HD™, a pyrethroid, were conducted under operational conditions against wild populations of Aedes dorsalis and Culex tarsalis and against colony strains of Cx. pipiens and Cx. quinquefasciatus. The erratic wind patterns of the greater Salt Lake area did not prevent sufficient droplet deposition flux at 9 monitoring locations spread across a 5,120-acre (2,072 ha) spray block within rural habitats. Three separate aerial application trials showed great efficacy against Ae. dorsalis. In contrast, Cx. tarsalis exhibited inconsistent treatment-associated mortalities, suggesting the presence of less susceptible or resistant field populations as a result of spillover from agricultural or residential pyrethroid usage. Bottle bioassays to diagnose pyrethroid resistance using field-collected Cx. tarsalis indicated that some populations of this species, especially those closest to urban edges, failed to show adequate mortality in resistance assays. Despite challenging weather conditions, Duet HD worked reasonably well against susceptible mosquito species, and it may provide a crucial role as an alternative for organophosphate applications within specific and sensitive areas. However, its area-wide adoption into control applications by the SLCMAD could be problematic due to reduced impacts on the most important arboviral vector species, Cx. tarsalis, in this area. This study demonstrates the importance of testing mosquito control products under different operational environments and against potentially resistant mosquito populations by municipal mosquito control districts.

Do it yourself: fabricating and evaluating a mosquito (Diptera: Culicidae) blood-feeding device to replace a commercial option.

Tools for rearing hematophagous insects, such as mosquitoes (Diptera: Culicidae), in an insectary are essential for research and operational evaluations in vector biology and control. There is an abundance of low-cost options for practitioners without conventional infrastructure. However, few midrange options exist that provide a balance of efficiency and low material waste. We present here a reproducible design for an electrically powered blood-feeding device that offers long-term reusability, low material waste, and customizability for different species or experiments. The limitation is the requirement for electricity, but the gain is a simple, low-skill device that can be modified as needed. To validate the design, assessments of feeding angle and blood-feeding success were compared between the Salt Lake City Mosquito Abatement District artificial membrane feeder (SLAM) and a commercial system (Hemotek). Engorgement in Aedes aegypti (80-90%), Culex pipiens (50-80%), and Culex tarsalis (30-75%) was similar between the 2 units, resulting in nearly identical fecundity outcomes between devices. Additionally, 45° angles were more successful, generally, than presenting the feeders flat or vertical to the mosquitoes (df3,48, P = 1.014 × 10-15). This angle is simple to present with the SLAM device. Materials for in-house reproduction of the SLAM system are now widely available in regions with access to e-commerce and shipped goods. This results in a device schematic that should fit well into a relatively modular, do-it-yourself paradigm where the practitioner needs only to assemble some materials without complex engineering. This article provides schematics, cost comparison, and validation of the in-house-made SLAM system.

Enhancing toxic sugar meals against Aedes aegypti (Diptera: Culicidae) by adulterating with erythritol in combination with other active ingredients.

Attractive toxic sugar baits (ATSBs) are an underexploited method for mosquito control. For ATSBs to be more widely accepted, demonstrably effective ingredients need to be verified. We investigated erythritol as a toxic additive in sugar meals against Aedes aegypti (L.) for potential future use in ATSBs. Erythritol is a sugar alcohol that is commonly used as a sugar substitute, while also being toxic to mosquitoes. Our studies tested formulations of erythritol, sucrose, and blends of both. Secondary investigations included combinations with the active ingredients Bacillus thuringiensis israelensis, spinosyn, and boric acid. Adult Ae. aegypti were separated into test groups and provided various combinations. Formulations containing erythritol, with or without another toxicant, exhibited 90% mortality within 72 h of observation (P = 0.03192). Additionally, erythritol appeared more effective when combined with sucrose in a 1:1 ratio (5% concentration each). This combination showed a 24% and 85% increase in mortality when combined with boric acid and Bti, respectively, at 48 h compared with equivalent groups containing only 10% sucrose. Erythritol appears to kill adult mosquitoes, even in relatively low concentrations, without another toxicant being required. However, erythritol also effectively enhances kill of main ingredient toxicants such as boric acid and Bti, showing a supporting role. The low concentration of erythritol needed to provide significant kill, its ability to fill in as both a sugar base and toxicant, and its ability to be safely handled by humans makes erythritol a strong candidate for use as a supporting ingredient in future bait formulations.

Mosquito Surveillance for West Nile Virus.

Identifying the mosquitoes responsible for transmitting human disease-causing pathogens is of critical importance for effective control of mosquito-borne outbreaks. West Nile virus is often transferred by adult female mosquitoes in the genus Culex, which deposit eggs in a variety of aquatic habitats throughout the world. Herein we describe several methodological approaches to monitor these species in nature, as well as offering details for data collection and analysis.

Laboratory evaluation of sugar alcohols for control of mosquitoes and other medically important flies.

Insecticide application for vector control is the most controversial component of a public health program due to concerns about environmental and human health safety. One approach to overcome this challenge is the use of environmentally benign active ingredients. Among the most promising emerging strategies are attractive toxic sugar baits. Sugar alcohols-naturally occurring molecules safe for human consumption but potentially toxic to insects when ingested, have received increased attention for use with this approach. For this study, we screened the toxicity of four different sugar alcohols on several mosquito species, a biting midge, and a filth fly. Sugar alcohol mortalities exceeded those in the sucrose (positive control) only group. However, only erythritol and highly concentrated xylitol induced mortalities exceeding those in the water only (negative control) treatment ranging from approximately 40-75%. Formulations containing erythritol and xylitol should be further investigated under field conditions for efficacy in reducing populations of biting flies and for assessing potential non-target impacts.

Origins of high latitude introductions of Aedes aegypti to Nebraska and Utah during 2019.

Aedes aegypti (L.), the yellow fever mosquito, is also an important vector of dengue and Zika viruses, and an invasive species in North America. Aedes aegypti inhabits tropical and sub-tropical areas of the world and in North America is primarily distributed throughout the southern US states and Mexico. The northern range of Ae. aegypti is limited by cold winter months and establishment in these areas has been mostly unsuccessful. However, frequent introductions of Ae. aegypti to temperate, non-endemic areas during the warmer months can lead to seasonal activity and disease outbreaks. Two Ae. aegypti incursions were reported in the late summer of 2019 into York, Nebraska and Moab, Utah. These states had no history of established populations of this mosquito and no evidence of previous seasonal activity. We genotyped a subset of individuals from each location at 12 microsatellite loci and ~ 14,000 single nucleotide polymorphic markers to determine their genetic affinities to other populations worldwide and investigate their potential source of introduction. Our results support a single origin for each of the introductions from different sources. Aedes aegypti from Utah likely derived from Tucson, Arizona, or a nearby location. Nebraska specimen results were not as conclusive, but point to an origin from southcentral or southeastern US. In addition to an effective, efficient, and sustainable control of invasive mosquitoes, such as Ae. aegypti, identifying the potential routes of introduction will be key to prevent future incursions and assess their potential health threat based on the ability of the source population to transmit a particular virus and its insecticide resistance profile, which may complicate vector control.

Do tigers hunt during the day? Diel activity of the Asian tiger mosquito, Aedes albopictus (Diptera: Culicidae), in urban and suburban habitats of North America.

BACKGROUND: Aedes (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 FINDINGS: Human 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 (P<0.05). Bottle rotator traps used in conjunction with the human sweep net technique exhibited similar results. Seasonal activity was not differentiated in Florida due to the consistent subtropical climate. The highest adult activity was observed at Sunrise using human sweep netting, but it was not significantly different from Solar Noon and Sunset. The lowest adult activity was observed at Lunar Midnight; however, it was not significantly different from Solar Noon and Sunset. These results provide evidence that the diel activity of Ae. albopictus, contrary to the common perception of its diurnal activity, is much more varied. CONCLUSION/SIGNIFICANCE: Involvement 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.

Heterodissemination: precision insecticide delivery to mosquito larval habitats by cohabiting vertebrates.

Conventional larvicide delivery strategies originally developed for permanent and floodwater mosquitoes have proved suboptimal in the small, scattered, and cryptic larval habitats preferred by container-inhabiting Aedes mosquitoes. New methods such as autodissemination, wherein adult mosquitoes spread insecticides to their own larval habitats, have been under study. Another novel delivery method termed heterodissemination, i.e. larvicide delivery by other species sharing the same habitats, has also been proposed. We conducted a proof-of-concept study with four independent experiments using American bullfrogs (Lithobates catesbeianus) and green frogs Lithobates clamitans as carriers of pyriproxyfen, an insect growth regulator, under semi-field conditions in three different locations, two in New Jersey, and one in Utah. Frogs with attached slow-release pyriproxyfen tablets were introduced into outdoor enclosures with water containers. Water samples from the containers were periodically tested using larval Aedes albopictus and Culex pipiens mosquitoes to assess mortality and percent eclosure inhibition. Overall pupal mortality [95% credible intervals] estimated by Bayesian analysis for the treatment group was 73.4% [71.3-75.2] compared to 4.1% [2.9-5.5] for the control group. Mortality within treatment groups in four different experiments ranged from 41 to 100%, whereas control mortalities ranged from 0.5% to 11%. We conclude that heterodissemination is a promising and effective approach deserving of further study.

Toys or Tools? Utilization of Unmanned Aerial Systems in Mosquito and Vector Control Programs.

Organized mosquito control programs (MCP) in the United States have been protecting public health since the early 1900s. These programs utilize integrated mosquito management for surveillance and control measures to enhance quality of life and protect the public from mosquito-borne diseases. Because much of the equipment and insecticides are developed for agriculture, MCP are left to innovate and adapt what is available to accomplish their core missions. Unmanned aerial systems (UAS) are one such innovation that are quickly being adopted by MCP. The advantages of UAS are no longer conjectural. In addition to locating mosquito larval habitats, UAS affords MCP real-time imagery, improved accuracy of aerial insecticide applications, mosquito larval detection and sampling. UAS are also leveraged for applying larvicides to water in habitats that range in size from multi-acre wetlands to small containers in urban settings. Employing UAS can reduce staff exposure to hazards and the impact associated with the use of heavy equipment in sensitive habitats. UAS are utilized by MCP nationally and their use will continue to increase as technology advances and regulations change. Current impediments include a dearth of major UAS manufacturers of equipment that is tailor-made for mosquito control, pesticides that are optimized for application via UAS and regulations that limit the access of UAS to national airspace. This manuscript highlights the strengths and weaknesses of UAS within MCP, provides an update on systems and methods used, and charts the future direction of UAS technology within MCP tasked with public health protection.

No Evidence That Salt Water Ingestion Kills Adult Mosquitoes (Diptera: Culicidae).

Various products and insecticides are available that purport to reduce wild populations of adult mosquitoes. Recently, several manufacturers and general public comments on the internet have promoted devices that claim that ingestion of salt will significantly reduce populations of wild mosquitoes to near zero; there are no known scientific efficacy data that support these claims. We tested the survival of nine mosquito species of pest and public health importance across four adult diets: Water Only, Sugar Water Only (8.00%), Salt Water Only (1.03%), and Sugar + Salt Water. Species included the following: Aedes aegypti (L.), Aedes albopictus (Skuse), Aedes dorsalis (Meigen), Aedes notoscriptus (Skuse), Aedes vigilax (Skuse), Anopheles quadrimaculatus (Say), Culex pipiens (L.), Culex quinquefasciatus (Say), and Culex tarsalis (Coquillett). Male and female mosquitoes were placed in cages and allowed to feed on liquid diets under controlled environmental conditions for 1 wk. For seven of the nine species, adult survival was significantly higher in the presence (Sugar Water, Sugar + Salt Water) versus the absence (Water Only, Salt Only) of sugar, with no indication that salt had any effect on survival. Anopheles quadrimaculatus showed intermediate survival in Sugar + Salt to either Sugar Only or no sugar diets, whereas Aedes dorsalis showed low survival in Salt Only versus other diets. Based on our data and coupled with the fact that mosquitoes have physiological and behavioral adaptations that allow them to avoid or process excess salt (as found in blood meals), we conclude that there is no scientific foundation for salt-based control methods of mosquitoes.

Trap Comparison for Surveillance of the Western Tree Hole Mosquito (Diptera: Culicidae).

The western tree hole mosquito, Aedes sierrensis (Ludlow), is a common nuisance mosquito and vector of Dirofilaria immitis (Leidy), the etiologic agent of dog heartworm, in western North America. Here, we compare weekly mosquito collections made with Mosquito Magnet (MM) traps, Biogents Sentinel (BGS) traps, and Biogents Bowl (BGS Bowl) traps set in Salt Lake City, UT, from the start of June to mid-August 2017. We found the number of mosquitoes decreased with rainfall and temperature independently of trap type. The highest number of mosquitoes were caught by BGS traps baited with carbon dioxide (CO2) and BG lure, which collected 62% (n = 422) of all mosquitoes, followed by the MM at 31% (n = 213), and both the BGS and BG Bowl with BG lure had 3.5% (n = 24) each. Aedes sierrensis females were caught weekly at similar densities (mean ±â€…SD) in BGS with CO2 and lure (1.17 ±â€…2.93) and the MM (1.17 ±â€…2.66) traps during the study period. Given that BGS with CO2 and lure traps have several operational advantages over MM traps, including a quicker setup, smaller size, and lower cost, we consider BGS with CO2 and lure traps as the best suited surveillance tool to detect and remove Ae. sierrensis in the western United States and similar settings throughout North America.

West Nile Virus Mosquito Vectors in North America.

In North America, the geographic distribution, ecology, and vectorial capacity of a diverse assemblage of mosquito species belonging to the genus Culex determine patterns of West Nile virus transmission and disease risk. East of the Mississippi River, mostly ornithophagic Culex pipiens L. complex mosquitoes drive intense enzootic transmission with relatively small numbers of human cases. Westward, the presence of highly competent Culex tarsalis (Coquillett) under arid climate and hot summers defines the regions with the highest human risk. West Nile virus human risk distribution is not uniform geographically or temporally within all regions. Notable geographic 'hotspots' persist with occasional severe outbreaks. Despite two decades of comprehensive research, several questions remain unresolved, such as the role of non-Culex bridge vectors, which are not involved in the enzootic cycle, but may be involved in virus transmission to humans. The absence of bridge vectors also may help to explain the frequent lack of West Nile virus 'spillover' into human populations despite very intense enzootic amplification in the eastern United States. This article examines vectorial capacity and the eco-epidemiology of West Nile virus mosquito vectors in four geographic regions of North America and presents some of the unresolved questions.