Scented Sugar Baits Enhance Detection of St. Louis Encephalitis and West Nile Viruses in Mosquitoes in Suburban California.

Scented sugar baits deployed in California deserts detected early West Nile virus (WNV) transmission by mosquitoes, representing a potential improvement to conventional arbovirus surveillance that relies heavily on infection rates in mosquito pools. In this study, we expanded deployment of scented sugar baits into suburban Sacramento and Yolo (2015, 2016) and Riverside Counties (2016), California. The goal of the study was to determine whether scented sugar baits detect WNV and St. Louis encephalitis virus (SLEV) concurrent with mosquito infections in trapped pools in areas of high human density. Between 8 and 10% of sugar baits were WNV RNA positive in both study years across the three counties. In Riverside County, where SLEV re-emerged in 2015, 1% of sugar baits were SLEV positive in 2016. Rates of sugar bait positives were at least 100 times higher than infection rates in trapped mosquitoes in the same districts. The prevalence of sugar bait positives varied temporally and did not coincide with infections in mosquitoes collected at the same sites each week. WNV RNA positive sugar baits were detected up to 2 wk before and after concurrent surveillance detected infection in mosquito pools at the same sites. Sugar baits also detected WNV in Riverside County at locations where no WNV activity was detected in mosquito pools. Sugar baits generated between 0.8 and 1.2 WNV positives per $1,000 and can be more economical than carbon dioxide baited traps that produce 0.8 positives per $1,000. These results indicate that the sugar bait approach enhances conventional arbovirus surveillance in mosquitoes in suburban California.

Host selection patterns of Culex tarsalis (Diptera: Culicidae) at wetlands near the Salton Sea, Coachella Valley, California, 1998-2002.

The bloodmeal hosts used by Culex tarsalis Coquillett collected along the Salton Sea in Coachella Valley, CA, during 1998-2002 were identified using sequences of the cytochrome c oxidase I gene identified from Barcode of Life database. Overall, 265 (83.3%) of 318 bloodmeals were identified, of which 76.6% fed on birds, 18.1% on mammals, and 5.3% on reptiles. Forty-seven different hosts were identified, none of which comprised > 12.5% of the total. Although Cx. tarsalis exhibits specific host-seeking flight patterns, bloodmeals seemed to be acquired opportunistically, thereby limiting potential arbovirus transmission efficiency in species-rich environments.

Use of scented sugar bait stations to track mosquito-borne arbovirus transmission in California.

Laboratory and field research was conducted to determine if Culex tarsalis Coquillett expectorated West Nile virus (WNV) during sugar feeding and if a lure or bait station could be developed to exploit this behavior for WNV surveillance. Experimentally infected Cx. tarsalis repeatedly expectorated WNV onto filter paper strips and into vials with wicks containing sucrose that was readily detectable by a quantitative reverse transcriptase-polymerase chain reaction assay. Few females (33%, n = 27) became infected by imbibing sugar solutions spiked with high concentrations (10(7) plaque forming units/ml) of WNV, indicating sugar feeding stations probably would not be a source of WNV infection. In nature, sugar bait stations scented with the floral attractant phenyl acetaldehyde tracked WNV transmission activity in desert but not urban or agricultural landscapes in California. When deployed in areas of the Coachella Valley with WNV activity during the summer of 2011, 27 of 400 weekly sugar samples (6.8%) tested positive for WNV RNA by reverse transcriptase-polymerase chain reaction. Prevalence of positives varied spatially, but positive sugar stations were detected before concurrent surveillance measures of infection (mosquito pools) or transmission (sentinel chicken seroconversions). In contrast, sugar bait stations deployed in urban settings in Los Angeles or agricultural habits near Bakersfield in Kern County supporting WNV activity produced 1 of 90 and 0 of 60 positive weekly sugar samples, respectively. These results with sugar bait stations will require additional research to enhance bait attractancy and to understand the relationship between positive sugar stations and standard metrics of arbovirus surveillance.

Residual mosquito barrier treatments on U.S. military camouflage netting in a southern California desert environment.

Treating perimeters of vegetation with residual insecticides for protection from mosquito vectors has potential for U.S. military force health protection. However, for current U.S. military operations in hot-arid environments with little or no vegetation, residual applications on portable artificial materials may be a viable alternative. We evaluated bifenthrin residual treatments of U.S. military camouflage netting under hot-arid field conditions in a desert area in southern California exposed to abundant wild Culex tarsalis mosquitoes. We assessed the ability of the treatment to reduce the numbers of mosquitoes penetrating perimeters of netting and reaching CO2-baited mosquito traps. Treated camouflage netting barriers reduced mosquitoes by > or = 50% for 7-14 days and by 20-35% for 21-28 days compared to untreated barriers. Although reductions may be translated into reductions in risk of exposure to mosquito-borne diseases, we emphasize that barrier treatments should be a component in a suite of insect control measures to be effective.

Evaluation of ULV and thermal fog mosquito control applications in temperate and desert environments.

Ultra-low-volume (ULV) and thermal fog aerosol dispersals of pesticides have been used against mosquitoes and other insects for half a century. Although each spray technology has advantages and disadvantages, only 7 studies have been identified that directly compare their performance in the field. US military personnel currently operating in hot-arid environments are impacted by perpetual nuisance and disease vector insect problems, despite adulticide operations using modern pesticide-delivery equipment such as ULV. None of the identified comparative studies has looked at the relative feasibility and efficacy of ULV and thermal fog equipment against mosquitoes in hot-arid environments. In this study we examine the impact of ULV and thermal fog applications of malathion against caged sentinel mosquitoes in the field in a warm temperate area of Florida, followed by a similar test in a hot-dry desert area of southern California. Patterns of mortality throughout 150 m x 150 m grids of sentinel mosquitoes indicate greater efficacy from the thermal fog application in both environments under suboptimal ambient weather conditions. We discuss the implications of these findings for future military preventive medicine activities and encourage further investigations into the relative merits of the 2 technologies for force health protection.

Effects of warm winter temperature on the abundance and gonotrophic activity of Culex (Diptera: Culicidae) in California.

Culex tarsalis Coquillett, Cx. quinquefasciatus Say, and Cx. pipiens L. were collected during the warm winter of 2009 using dry ice-baited and gravid traps and walk-in red boxes positioned in desert, urban, and agricultural habitats in Riverside, Los Angeles, Kern, and Yolo Counties. Temperatures exceeded the preceding 50 yr averages in all locations for most of January, whereas rainfall was absent or below average. Abundance of Culex species in traps during January ranged from 83 to 671% of the prior 5 yr average in all locations. Few females collected resting were in diapause during January based on follicular measurements. Evidence for early season gonotrophic activity included the detection of freshly bloodfed, gravid, and parous females in resting collections, gravid oviposition site-seeking females in gravid female traps, and nulliparous and parous host-seeking females at dry ice-baited traps. Female Culex seemed to employ multiple overwintering strategies in California, including larval and adult quiescence, adult female diapause, and an intermediate situation with adult females collected with enlarged follicles, but without evident vitellogenesis. West Nile, St. Louis, or western equine encephalitis viruses were not detected in 198 pools of adults or 56 pools of adults reared from field-collected immatures collected during January and February 2009. Our preliminary data may provide insight into how climate change may extend the mosquito season in California.

Evaluation of barrier treatments on native vegetation in a southern California desert habitat.

Treating perimeters with residual insecticides for protection from mosquito vectors has shown promise. These barrier treatments are typically evaluated in temperate or tropical areas using abundant vegetation as a substrate. However, there is an emerging interest to develop this technology to protect deployed US troops in extreme desert environments with sparse vegetation. We used a remote desert area in the Coachella Valley, California, to 1) evaluate bifenthrin barrier treatments on native xeric vegetation and 2) compare treatments applied with electrostatic and conventional spray technologies. Through a combination of laboratory bioassays on treated and control vegetation sampled at specific intervals over 63 days, synchronized with field surveillance of mosquitoes, we measured the temporal pattern of bioactivity of bifenthrin barriers under natural hot, dry, and dusty desert conditions. Regardless of spray technology, mosquito catch in treated plots was about 80% lower than the catch in control plots 1 day after treatment. This reduction in mosquito numbers in treated plots declined each week after treatment but remained at about 40% lower than control plots after 28 days. Field data were corroborated by results from bioassays that showed significantly higher mosquito mortality on treated vegetation over controls out to 28 days postspray. We concluded that barrier treatments in desert environments, when implemented as part of a suite of integrated control measures, may offer a significant level of protection from mosquitoes for deployed troops. Given the comparable performance of the tested spray technologies, we discuss considerations for choosing a barrier treatment sprayer for military scenarios.

Persistent West Nile virus transmission and the apparent displacement St. Louis encephalitis virus in southeastern California, 2003-2006.

West Nile virus (family Flaviviridae, genus Flavivirus, WNV) invaded the Colorado Desert biome of southern California during summer 2003 and seemed to displace previously endemic St. Louis encephalitis virus (family Flaviviridae, genus Flavivirus, SLEV, an antigenically similar Flavivirus in the Japanese encephalitis virus serocomplex). Western equine encephalomyelitis virus (family Togaviridae, genus Alphavirus, WEEV), an antigenically distinct Alphavirus, was detected during 2005 and 2006, indicating that conditions were suitable for encephalitis virus introduction and detection. Cross-protective "avian herd immunity" due to WNV infection possibly may have prevented SLEV reintroduction and/or amplification to detectable levels. During 2003-2006, WNV was consistently active at wetlands and agricultural habitats surrounding the Salton Sea where Culex tarsalis Coquillett served as the primary enzootic maintenance and amplification vector. Based on published laboratory infection studies and the current seroprevalence estimates, house sparrows, house finches, and several Ardeidae may have been important avian amplifying hosts in this region. Transmission efficiency may have been dampened by high infection rates in incompetent avian hosts, including Gamble's quail, mourning doves, common ground doves, and domestic pigeons. Early season WNV amplification and dispersal from North Shore in the southeastern portion of the Coachella Valley resulted in sporadic WNV incursions into the urbanized Upper Valley near Palm Springs, where Culex pipiens quinquefasciatus Say was the primary enzootic and bridge vector. Although relatively few human cases were detected during the 2003-2006 period, all were concentrated in the Upper Valley and were associated with high human population density and WNV infection in peridomestic populations of Cx. p. quinquefasciatus. Intensive early mosquito control during 2006 seemed to interrupt and delay transmission, perhaps setting the stage for the future reintroduction of SLEV.

Intensive early season adulticide applications decrease arbovirus transmission throughout the Coachella Valley, Riverside County, California.

In the Coachella Valley of California the seasonal onset of St. Louis encephalitis virus (SLEV), western equine encephalomyelitis virus (WEEV), and West Nile virus (WNV) has been detected consistently at the shoreline of the Salton Sea near the community of North Shore. The timing and intensity of initial amplification in the Culex tarsalis Coquillett/wild bird cycle at this focus seemed closely linked to the subsequent dispersal of virus to the rest of the Coachella Valley and perhaps southern California. In 2004, an attempt was made to interrupt the amplification and dispersal of WNV using ground ultra-low volume (ULV) applications of Pyrenone 25-5. Although these localized treatments were started 1 month after the initial detection in April, surveillance indicated no dispersal from this focus at this time. However, these treatments appeared to have little effect, and WNV eventually was detected throughout the valley, with seven human cases reported in the urbanized upper valley near Palm Springs. In 2005, the initial detection of WNV at North Shore at the end of May was followed rapidly by dispersal throughout the valley precluding efforts at containment. Evaluation of ground and aerial applications at North Shore during May and June 2005, respectively, indicated variable kill of sentinel mosquitoes (overall mortality: ground, 43%; air, 34%) and limited control of the target Cx. tarsalis population. In 2006, aerial ULV applications with the same chemical were begun immediately following the first detection of virus in mid-April, resulting in an apparent reduction of Cx. tarsalis abundance and delay of WNV activity in the rural lower valley and a marked decline in transmission by Culex quinquefasciatus Say populations in the densely populated upper northwestern valley with no human cases reported.

Overwintering of West Nile virus in Southern California.

West Nile virus (family Flaviviridae, genus Flavivirus, WNV) invaded southern California during 2003, successfully overwintered, amplified to epidemic levels, and then dispersed to every county in the state. Although surveillance programs successfully tracked and measured these events, mechanisms that allowed the efficient overwintering and subsequent amplification of WNV have not been elucidated. Our current research provided evidence for three mechanisms whereby WNV may have persisted in southern California during the winters of 2003-2004 and 2004-2005: 1) continued enzootic transmission, 2) vertical transmission by Culex mosquitoes, and 3) chronic infection in birds. WNV was detected in 140 dead birds comprising 32 species, including 60 dead American crows, thereby verifying transmission during the November-March winter period. Dead American crows provide evidence of recent transmission because this species always succumbs rapidly after infection. However, WNV RNA was not detected concurrently in 43,043 reproductively active female mosquitoes comprising 11 species and tested in 1,258 pools or antibody in sera from 190 sentinel chickens maintained in 19 flocks. Although efficient vertical transmission by WNV was demonstrated experimentally for Culex tarsalis Coquillett infected per os, 369 females collected diapausing in Kern County and tested in 32 pools were negative for WNV. Vertical transmission was detected in Culex pipiens quinquefasciatus Say adults reared from field-collected immatures collected from Kern County and Los Angeles during the summer transmission period. Chronic infection was detected by finding WNV RNA in 34 of 82 birds that were inoculated with WNV experimentally, held for >6 wk after infection, and then necropsied. Frequent detection of WNV RNA in kidney tissue in experimentally infected birds >6 wk postinfection may explain, in part, the repeated detection of WNV RNA in dead birds recovered during winter, especially in species such as mourning doves that typically do not die after experimental infection. In summary, our study provides limited evidence to support multiple modes of WNV persistence i n southern California. Continued transmission andvertical transmission by Culex p. quinquefasciatus Say seem likely candidates for further study.

Role of corvids in epidemiology of west Nile virus in southern California.

The invasion of different southern California landscapes by West Nile virus (WNV) and its subsequent amplification to epidemic levels during 2004 enabled us to study the impact of differing corvid populations in three biomes: the hot Colorado desert with few corvids (Coachella Valley), the southern San Joaquin Valley (Kern County) with large western scrub-jay but small American crow populations, and the cool maritime coast (Los Angeles) with a large clustered American crow population. Similar surveillance programs in all three areas monitored infection rates in mosquitoes, seroconversion rates in sentinel chickens, seroprevalence in wild birds, numbers of dead birds reported by the public, and the occurrence of human cases. Infection rates in Culex tarsalis Coquillett and sentinel chicken seroconversion rates were statistically similar among all three areas, indicating that highly competent mosquito hosts were capable of maintaining enzootic WNV transmission among less competent and widely distributed avian hosts, most likely house sparrows and house finches. In contrast, infection rates in Culex pipiens quinquefasciatus Say were statistically higher in Kern and Los Angeles counties with elevated corvid populations than in Coachella Valley with few corvids. Spatial analyses of dead corvids showed significant clusters near known American crow roosts in Los Angeles that were congruent with clusters of human cases. In this area, the incidence of human and Cx. p. quinquefasciatus infection was significantly greater within corvid clusters than without, indicating their importance in virus amplification and as a risk factor for human infection. In contrast the uniform dispersion by territorial western scrub-jays resulted in a high, but evenly distributed, incidence of human disease in Kern County.

Factors influencing the outcome of mark-release-recapture studies with Culex tarsalis (Diptera: Culicidae).

Three potentially important aspects of mark-release-recapture experimentation were addressed: 1) source of mosquitoes for release, 2) time of release, and 3) weather during recapture. Culex tarsalis Coquillett mosquitoes collected as adult host-seeking females from dry ice-baited traps (CO2 traps) operated within the study area (local) were recaptured more frequently than females collected from traps operated outside the study area (foreign) or reared from field-collected immatures (reared). These results supported published studies on Anopheles and Ochlerotatus that indicated mosquitoes may "memorize" flight paths within their environment. Releasing gravid females provided a potentially useful replacement for reared females, because these gravids oviposited at wetlands and then dispersed to upland traps. Releasing local, foreign, or reared mosquitoes just after sunrise or just before sunset did not alter recapture success or the distance dispersed. Elevated wind speeds inhibited dispersal from protected microhabitats with citrus orchards and resulted in most recaptures being found at the leeward portion of the orchard.

Nocturnal microhabitat distribution of adult Culex tarsalis (Diptera: Culicidae) impacts control effectiveness.

Suction traps (30 cm diameter) were more effective for non-attractant sampling of flying adult Culex tarsalis Coquillett than were smaller CDC (5.5 cm diameter), Malaise or ramp traps. Comparative catch in suction traps operated in a variety of vegetation types indicated that females congregated along elevated ecotones and were significantly less abundant flying over low vegetation or under and over elevated vegetation. Most females taken at upland orchards or Tamarisk tree lines were unfed (97%, n = 5,278) and similar in reproductive condition to host-seeking females. Blood fed and gravid females and males were only abundant near emergence sites. Pyrocide 7396 (Pyrethrin 5%, PBO 25%) was applied at the label rate of 5 oz/min by truck mounted Pro-Mist ultra low volume (ULV) equipment and particle drift measured by bioassay. ULV particles dispersed well downwind over low vegetation, between citrus orchard rows, and under date orchard canopy, but did not penetrate citrus orchards or vineyards when rows were perpendicular to wind direction. Particles did move up and over vegetation contacting sentinel mosquitoes placed above the canopy. The congregation of adult mosquitoes at vegetative ecotones and within orchard vegetation may afford protection from ground applied ULV particles, negatively impacting control. These data may explain why repeated applications often fail to interrupt encephalitis virus transmission.