Persistent spatial clustering and predictors of pediatric La Crosse virus neuroinvasive disease risk in eastern Tennessee and western North Carolina, 2003-2020.

The combined region of eastern Tennessee and western North Carolina has a persistently high risk of pediatric La Crosse virus neuroinvasive disease (LACV-ND). To guide public health intervention in this region, the objectives of this retrospective ecological study were to investigate the geographic clustering and predictors of pediatric LACV-ND risk at the ZIP code tabulation area (ZCTA) level. Data on pediatric cases of LACV-ND reported between 2003 and 2020 were obtained from Tennessee Department of Health and North Carolina Department of Health and Human Services. Purely spatial and space-time scan statistics were used to identify ZCTA-level clusters of confirmed and probable pediatric LACV-ND cases from 2003-2020, and a combination of global and local (i.e., geographically weighted) negative binomial regression models were used to investigate potential predictors of disease risk from 2015-2020. The cluster investigation revealed spatially persistent high-risk and low-risk clusters of LACV-ND, with most cases consistently reported from a few high-risk clusters throughout the entire study period. Temperature and precipitation had positive but antagonistic associations with disease risk from 2015-2020, but the strength of those relationships varied substantially across the study area. Because LACV-ND risk clustering in this region is focally persistent, retroactive case surveillance can be used to guide the implementation of targeted public health intervention to reduce the disease burden in high-risk areas. Additional research on the role of climate in LACV transmission is warranted to support the development of predictive transmission models to guide proactive public health interventions.

LA CROSSE VIRUS VECTOR RESTING BEHAVIORS - FIELD STUDIES WITH PROKOPAK AND RESTING SHELTER COLLECTIONS PROVIDE LOW YIELD.

Resting adult mosquito collections provide opportunities to sample broad physiological conditions (e.g., blood-engorged, gravid, nectar-engorged, and/or parous) that yield important biological information necessary to understand vector and pathogen transmission ecology. In this study, we evaluated Prokopak aspirations of Rhododendron spp. and human-powered pop-up resting shelter collections at 4 residences with historical evidence of proximal La Crosse virus (LACV) transmission from May through September 2022. The goal of this study was to investigate these sampling methods in the context of LACV vector biology-focused principally on Aedes triseriatus (primary LACV vector) and 2 invasive species (Ae. albopictus and Ae. japonicus) that likely serve as secondary LACV vectors. Overall, 304 resting shelters and 80 Prokopak collections yielded a grand total of 33 mosquitoes, of which a third were LACV vectors (Ae. triseriatus [n = 1, 3.0%], Ae. albopictus [n = 4, 12.1%], and Ae. japonicus [n = 6, 18.2%]). Anopheles punctipennis (n = 9, 27.2%) was the most frequently collected species followed by Culex erraticus (n = 7, 21.2%), whereas the least frequently collected species were Ae. triseriatus and Cx. pipiens (n = 1, 3.0%). Despite substantial collection efforts, and concurrent gravid-trap evidence of LACV vectors at the collection sites, Prokopak aspiration of Rhododendron spp. and human-powered pop-up resting shelters did not yield a meaningful number of LACV vectors and thus, as described within, may not be useful adjuncts for the evaluation of LACV ecology and disease risk. Additional approaches to evaluate the resting behavior of these vectors in LACV endemic areas are needed.

La Crosse virus neuroinvasive disease: the kids are not alright.

La Crosse virus (LACV) is the most common cause of neuroinvasive mosquito-borne disease in children within the United States. Despite more than 50 years of recognized endemicity in the United States, the true burden of LACV disease is grossly underappreciated, and there remain severe knowledge gaps that inhibit public health interventions to reduce morbidity and mortality. Long-standing deficiencies in disease surveillance, clinical diagnostics and therapeutics, actionable entomologic and environmental risk indices, case response capacity, public awareness, and availability of community support groups clearly frame LACV disease as neglected. Here we synthesize salient prior research and contextualize our findings as an assessment of current gaps and opportunities to develop a framework to prevent, detect, and respond to LACV disease. The persistent burdens of LACV disease clearly require renewed public health attention, policy, and action.

Retrospective Assessment of Preparedness for Mosquito Control Post-Hurricane Florence in North Carolina.

OBJECTIVES: Ideally, mosquito control programs (MCPs) use surveillance to target control measures to potentially dangerous mosquito populations. In North Carolina (NC), where there is limited financial support for mosquito control, communities may suffer from mosquito-related issues post-hurricane due to lack of existing MCPs. Here, study objectives were to (1) investigate the emergency response of a subset of NC counties post-Hurricane Florence and (2) develop guidelines and policy recommendations to assist MCPs in post-hurricane mosquito control response. METHODS: A survey was administered to a subset of eastern NC counties (an area previously impacted by hurricanes) with various levels of MCPs (from none to well-developed). RESULTS: All respondents indicated that having Federal Emergency Management Agency (FEMA) training would be helpful in developing a post-hurricane emergency response plan for mosquito control. There was concern related to a lack of knowledge of emergency control methods (eg, aerial/ground, adulticiding/larviciding) post-hurricane. MCP structure (eg, infrastructure, resources, operational plans/policies) could facilitate response activities and help ensure necessary emergency financial support from agencies such as FEMA. CONCLUSIONS: Mosquito control post-hurricane protects public health. Public health and other agencies can be networking resources for MCPs. Policy recommendations include implementation of routine FEMA assistance training workshops to improve an understanding of processes involved in assistance and reimbursement.

Phylogenomics reveals the history of host use in mosquitoes.

Mosquitoes have profoundly affected human history and continue to threaten human health through the transmission of a diverse array of pathogens. The phylogeny of mosquitoes has remained poorly characterized due to difficulty in taxonomic sampling and limited availability of genomic data beyond the most important vector species. Here, we used phylogenomic analysis of 709 single copy ortholog groups from 256 mosquito species to produce a strongly supported phylogeny that resolves the position of the major disease vector species and the major mosquito lineages. Our analyses support an origin of mosquitoes in the early Triassic (217 MYA [highest posterior density region: 188-250 MYA]), considerably older than previous estimates. Moreover, we utilize an extensive database of host associations for mosquitoes to show that mosquitoes have shifted to feeding upon the blood of mammals numerous times, and that mosquito diversification and host-use patterns within major lineages appear to coincide in earth history both with major continental drift events and with the diversification of vertebrate classes.

A Comparison of Oak Leaf and Fescue Hay Infusion-Baited Gravid Trap Collections-An Analysis Steeped in the Context of La Crosse Virus Vector Surveillance Effectiveness.

Neuroinvasive La Crosse virus disease remains the primary cause of pediatric arboviral encephalitis in the USA. In spite of the persistent public health burden, there are limited entomologic surveillance options that target both native and invasive La Crosse virus (LACV) vectors. In this study we used Reiter/Cummings tacklebox gravid traps to compare white oak (Quercus alba) and hay (predominately Festuca arundinacea) infusions within a LACV-endemic area of western North Carolina. Paired gravid traps (approximately 1,728 total trap-hours for each infusion) yielded 485 mosquitoes, with 3 species (Aedes japonicus [n = 265], Ae. triseriatus [n = 156], and Culex restuans [n = 45]) accounting for 96.1% of the total collection. The hay-infusion traps collected 2.5 times more Ae. triseriatus and 1.3 times more Ae. japonicus than the oak-infusion traps. The sum differences in overall collections for these 2 species by infusion type were statistically significant (χ2 = 9.61, df = 1, P = 0.0019). Poisson ratio tests to compare capture rates suggest that hay infusions were more effective for capturing Ae. triseriatus, but that hay and white oak leaf infusions had equivocal capture rates for Ae. japonicus (an invasive LACV accessory vector) and Cx. restuans (an enzootic West Nile virus vector). These results are discussed in the context of operational considerations for LACV vector surveillance.

Survey of Invasive Mosquito Surveillance and Control Capacity in Southeastern USA Reveals Training and Resource Needs.

Several invasive mosquito species that are nuisances or of medical and veterinary importance have been introduced into the Southeastern region of the USA, posing a threat to other species and the local ecosystems and/or increasing the risk of pathogen transmission to people, livestock, and domestic pets. Prompt and effective monitoring and control of invasive species is essential to prevent them from spreading and causing harmful effects. However, the capacity for invasive mosquito species surveillance is highly variable among mosquito control programs in the Southeast, depending on a combination of factors such as regional geography and climate, access to resources, and the ability to interact with other programs. To facilitate the development of invasive mosquito surveillance in the region, we, the Mosquito BEACONS (Biodiversity Enhancement and Control of Non-native Species) working group, conducted a survey on the capacities of various public health agencies and pest control agencies engaged in mosquito surveillance and control in seven Southeastern states (Alabama, Florida, Georgia, Louisiana, Mississippi, North Carolina, and South Carolina). Ninety control programs completed the survey, representing an overall response rate of 25.8%. We report key findings from our survey, emphasizing the training and resource needs, and discuss their implications for future invasive mosquito surveillance and control capacity building. By increasing communication and collaboration opportunities (e.g., real-time sharing of collection records, coordinated multistate programs), the establishment of Mosquito BEACONS and the implementation of this survey can accelerate knowledge transfer and improve decision support capacity in response to or in preparation for invasive mosquito surveillance and can establish infrastructure that can be used to inform programs around the world.

Survey of United States Mosquito Control Programs Reveals Opportunities to Improve the Operational Value of Centers for Disease Control and Prevention Bottle Bioassays.

Mosquito control programs have increasingly used Centers for Disease Control and Prevention bottle bioassays (BB) to evaluate insecticide resistance (IR). The reported utility, benefits, and limitations of BB and other methods were assessed via electronic survey of U.S. mosquito control professionals to identify potential areas for improvement, future study, and professional training. Opportunities were identified to improve BB operational value and IR detection.

A Cross-Sectional Household Survey in the US Virgin Islands (2019) Reveals Cisterns as Challenging Peridomestic Aedes aegypti Habitats.

Most residences in the United States Virgin Islands (USVI) rely on household rainwater-catchment systems and subterranean cisterns for long-term water storage that may provide suitable habitats for mosquitoes of public health relevance. We conducted a household cistern survey (n = 164) on the islands of St. Croix, St. John, and St. Thomas in 2019. The survey revealed that 45.7% (95% CI: 38.3-53.4%) of cisterns contained mosquitoes (adult and/or immature mosquitoes). Aedes aegypti, a vector of chikungunya, dengue, and Zika viruses in the USVI, was found in 27.4% (95% CI: 21.2-34.7%) of cisterns and accounted for 83.3% of the total mosquitoes identified in the study. The odds of detecting mosquitoes in a cistern were 5.45 times higher at locations where the residents reported that they had observed adult mosquitoes coming out of their cisterns (95% CI: 2.25-14.21), suggesting that vector control personnel should consider resident complaints about mosquitoes in their cistern as valid and likely reliable self-assessments. Resident mosquito management practices in cisterns did not correspond with decreased odds of mosquito detection. We conclude that cisterns in the USVI commonly provide habitat for immature and adult Ae. aegypti, which may decrease the effectiveness of area-wide mosquito control strategies. Additional studies are necessary to evaluate the importance of these cisterns as they relate to mosquito production and arbovirus transmission risk, and to assess physical and chemical control methods.

Operational Insights Into Mosquito Control Disaster Response in Coastal North Carolina: Experiences With the Federal Emergency Management Agency After Hurricane Florence.

Preparation for post-hurricane mosquito control is essential for an effective emergency response to protect public health and promote recovery efforts. Effective pre-hurricane planning includes laying the groundwork for a successful reimbursement application to the Federal Emergency Management Agency. The critical and overlapping need to sustain funding for mosquito control programs is highlighted here in the context of both normal and emergency responses. Community support is an integral component of an effective integrated pest management program and is established over time with appropriate communication and engagement. Experienced mosquito control operators who are familiar with treatment areas are an essential component of successful operations. Here, practical advice is provided to plan, prepare, and implement a successful ground- and aerial-based mosquito control response.

Population Growth Rates of Aedes atropalpus (Diptera: Culicidae) Are Depressed at Lower Temperatures Where Aedes japonicus japonicus (Diptera: Culicidae) Are Naturally Abundant in Rock Pools.

Recent studies report extensive reductions in the abundance of the North American rock pool mosquito, Aedes atropalpus (Diptera: Culicidae), following the invasion of Ae. japonicus japonicus in the United States. Although developmental temperature is recognized as an important component of the invasion biology of Ae. j. japonicus, its impacts on the population growth and fitness of Ae. atropalpus remain largely undefined. In this study we reared Ae. atropalpus larvae at three temperature ranges reflecting ecologically important temperatures in natural rock pools: a low temperature range (mean: 19°C) where Ae. j. japonicus is common and Ae. atropalpus is often rare, a middle temperature range (mean: 25°C) where both species are naturally found in similar relative abundances, and a higher temperature range (mean: 31°C) where Ae. atropalpus is the dominant species. We measured survival, development time, wing length, and fecundity to calculate a finite population growth rate at each temperature. Our results indicate that Ae. atropalpus population growth suffers in colder rock pools, which informs the perceived displacement of the species in temperate habitats. The population growth rate was highest in the middle temperature range, but not significantly higher than in the highest temperature range used in this study. The developmental success of Ae. atropalpus at the intermediate temperature range suggests that competition with Ae. j. japonicus in rock pools within that range may significantly impact natural Ae. atropalpus populations.

Peridomestic conditions affect La Crosse virus entomological risk by modifying the habitat use patterns of its mosquito vectors.

Anthropogenic land-use change may affect the transmission risk for endemic vector-borne diseases such as La Crosse encephalitis. In this study, we applied a comparative ecological approach to evaluate differences in vector species abundance, gonotrophic status, and environmental variables among six residential habitats (historical case houses) and six paired adjacent forest patches in a La Crosse virus endemic area of North Carolina. A total of 93,158 container Aedes spp. eggs were obtained by ovistrips and 1,040 resting mosquito adults were collected by large-bore aspiration from 10 June through 8 October, 2010. At sites characterized by high densities of artificial containers, the totals of eggs and adult mosquitoes were higher in the peridomestic plots. At sites characterized by lower densities of artificial containers, the totals of eggs and adult mosquitoes were higher in the forested plots. Although Aedes triseriatus, the primary La Crosse virus vector, was more commonly found in forested habitat overall, in sites characterized by high density of artificial containers, it was found in higher abundance in the peridomestic habitat. Similarly, the proportion of gonotrophically active (parous and gravid) mosquitoes was higher in the peridomestic habitat for sites with highartificial container density. Our study suggests that La Crosse virus transmission risk may be higher at peridomestic habitats with higher densities of artificial containers and thus reinforces the importance of public health measures to improve source reduction efforts.

Diapause influenced oviposition behavior and physical egg hatch cues of Aedes atropalpus (Diptera: Culicidae): traits that may influence successful colonization of riverine rock pools.

Mosquitoes have developed specialized oviposition strategies that allow them to develop in a wide variety of aquatic habitats. Environmentally cued hatching traits may also play an important role in the successful colonization of some larval habitats, but this subject has remained largely unexplored in Culicidae. Aedes atropalpus (Coquillett) is an autogenous rock pool specialist that may maintain unique adaptations for oviposition and egg hatching. We investigated the egg-laying strategies of Ae. atropalpus exposed to standard (non-diapausing) rearing conditions and diapause-inducing conditions and tested the impact of physical agitation on egg hatch rates by exposing floating and submerged eggs to physical agitation treatments. The results of the oviposition experiment indicate that Ae. atropalpus females primarily lay non-diapausing eggs directly onto the water surface and lay diapausing eggs directly on solid surfaces. The egg-hatching experiment demonstrated that physical agitation significantly increases Ae. atropalpus hatch rates. Floating and submerged eggs responded similarly to the agitation treatment. These data suggest that oviposition behaviors based on both egg diapause status and environmentally-cued hatching strategies may be important adaptations for Ae. atropalpus in riverine rock pools.

Short-Term, Large-Area Survey of Container Aedes spp. (Diptera: Culicidae): Presence and Abundance is Associated with Fine-scale Landscape Factors in North Carolina, USA.

Container Aedes mosquitoes are responsible for the transmission of anthroponotic and zoonotic viruses to people. The surveillance and control of these mosquitoes is an important part of public health protection and prevention of mosquito-borne disease. In this study, we surveyed 327 sites over 2 weeks in late June and early July in 2017 in North Carolina, USA for the presence and abundance of Aedes spp. eggs in an effort to better target potential Ae. aegypti collections. We examined the ability of 2 types of landscape data, Light Detection And Ranging (LIDAR) and National Land Cover Database (NLCD) to explain the presence and abundance of eggs using principal component analysis to deal with collinearity, followed by generalized linear regression. We explained variation of both egg presence and abundance for Aedes albopictus (Skuse) and Aedes triseriatus (Say) using both NLCD and LIDAR data. However, the ability to make robust predictions was limited by variation in the data. Increased sampling time and better landscape data would likely improve the predictive ability of our models, as would a better understanding of oviposition behavior.

Assessing Insecticide Resistance in Adult Mosquitoes: Perspectives on Current Methods.

Mosquito insecticide resistance (IR) is a growing global issue that must be addressed to protect public health. Vector control programs (VCPs) should regularly monitor local mosquito populations for IR and plan control measures accordingly. In some cases, state/federal resources financially support this testing with expertise and/or training programs. Standardization of methods (eg, Centers for Disease Control and Prevention bottle bioassay, World Health Organization tube testing, dose-mortality bioassay) for monitoring IR must be prioritized. One solution is regional hubs of IR monitoring at the state or other level. Training programs on methodology and interpretation of results should be developed and routinely offered to local VCPs conducting IR testing in mosquitoes. Here, current methods for assessing mosquito IR are discussed and insights into a variety of questions from VCPs are considered. It is critical that methods for IR monitoring and data interpretation are standardized through routine training, with the goal of evidence-driven decision making to improve control of mosquitoes and mosquito-borne disease.

Classification of Mosquitoes with Infrared Spectroscopy and Partial Least Squares-Discriminant Analysis.

Mosquito-borne diseases are responsible for considerable morbidity and mortality globally. Given the absence of effective vaccines for most arthropod-borne viruses, mosquito control efforts remain the dominant method of disease prevention. Ideal control efforts begin with entomologic surveillance in order to determine the abundance, identity, and infection status of pathogen-vectoring mosquito populations. Traditionally, much of the surveillance work involves morphological species identification by trained entomologists. Limited operational funding and lack of specialized training is a known barrier to surveillance and effective control efforts for many operational mosquito control personnel. Therefore, there is a need for surveillance workflow improvements and rapid mosquito identification methods. Herein, is presented a proof of concept study in which infrared spectroscopy coupled with partial least squares-discriminant analysis was explored as a means of automatically classifying mosquitoes at the species level. The developed method resulted in greater than 94% accuracy for four mosquitoes of public health relevance: Aedes aegypti, Aedes albopictus, Aedes japonicus, and Aedes triseriatus.

Horizontal distribution affects the vertical distribution of native and invasive container-inhabiting Aedes mosquitoes within an urban landscape.

The vertical dimension constitutes an important niche axis along which mosquitoes may adjust their distribution. Here, we evaluated whether the vertical distribution of container-inhabiting Aedes mosquitoes differs along a gradient of anthropogenic land-use intensity within an urban landscape. Using a pulley system, we hung oviposition cups at three heights (ground level, 4.5, and 9 m) and in three habitats: forest, park, and a built environment. We hypothesized that mosquito abundance and diversity would be highest in the least disturbed forest habitat, decrease in the park, and be lowest at the UNC-Greensboro campus. We also expected Aedes albopictus (Skuse) and Ae. triseriatus (Say) to mainly oviposit at ground level and Ae. hendersoni (Cockerell) at canopy height. Aedes albopictus was the most common species (68.8%) collected in all three habitat types and was the only species found in the built environment. In that habitat, Ae. albopictus exhibited a bimodal distribution with the lowest activity at the intermediate height (4.5 m). Aedes triseriatus (28.9%) did not differ in egg abundance between the forest and park habitats but did exhibit diverse vertical habitat use while avoiding the canopy in the park habitat. Aedes hendersoni (2.3%) was the most sylvatic species and oviposited only at ground level. Our results indicate that the vertical distribution of mosquitoes is affected by the type of habitat in which they occur, and that this variation could be driven via local-scale modification of microclimatic factors.

Phenology of Rock Pool Mosquitoes in the Southern Appalachian Mountains: Surveys Reveal Apparent Winter Hatching of Aedes japonicus and the Potential For Asymmetrical Stage-Specific Interactions.

The North American rock pool mosquito, Aedes atropalpus, has reportedly decreased in abundance following the introduction of Ae. japonicus japonicus to the USA, but the specific mechanisms responsible for the reduction remain unclear. Thus, there is a need for field studies to improve our knowledge of natural rock pool systems where both species co-occur. We sampled rock pool invertebrates over a 12-month period along the Chattooga River at a high-elevation site (728 m) near Cashiers, NC, and at a lower-elevation site (361 m) near Clayton, GA. We identified 12 orders of macroinvertebrates representing at least 19 families and 5 mosquito species. Aedes j. japonicus was present year-round at both sites. We observed overwintering Ae. j. japonicus larvae in pools with water temperatures as cold as 3°C and detected apparent winter egg hatching in water below 10°C. Aedes atropalpus was rarely encountered at the high-elevation site but was highly abundant in the summer months at the low-elevation site. Late-stage Ae. j. japonicus larvae inhabited pools in March 2019 when Ae. atropalpus first appeared in the same pools, creating the potential for asymmetrical stage-specific interactions. Our observations provide evidence of overwintering and early hatching of Ae. j. japonicus in the southeastern climate. Further study of the importance of stage-dependent competition and winter egg hatching of diapausing Ae. j. japonicus eggs is warranted.

Context-Dependent Accuracy of the BG-Counter Remote Mosquito Surveillance Device in North Carolina.

There is a clear need for improved vector surveillance approaches that are affordable, labor efficient, and safer than traditional methods. The BG-Counter (Biogents USA, Moorefield, WV) is a device for remotely monitoring mosquito activity in combination with the BG-Sentinel (Biogents USA), a widely used trap for the collection of host-seeking mosquitoes. The BG-Counter uses a wireless connection to provide real-time counts of mosquitoes captured by the BG-Sentinel, allowing users to remotely monitor mosquito populations. This study tested the effectiveness of the BG-Counter in 5 North Carolina counties. A total of 96 trap-days resulted in the collection of >45,000 individual mosquitoes representing 35 species. Aedes albopictus was the most common species collected in all counties, except for New Hanover County where Culex nigripalpus was the most common. The mean daily accuracy ranged from 80.1% (New Hanover County) to 9.4% (Jackson County). There was a significant linear relationship between the actual number of mosquitoes collected and the device counts at all sites except Jackson County, the site with the lowest relative mosquito abundance compared with nontarget organisms. A linear regression of daily BG-Counter accuracy and the daily proportion of mosquitoes to the total number of arthropods collected revealed a significant positive linear relationship, supporting the premise that the BG-Counter is less effective when the relative abundance of mosquitoes is low. Mosquito surveillance programs using the BG-Counter should recognize its context-dependent accuracy and routinely evaluate the accuracy of the device based on local conditions.

First Record of Mansonia dyari From Saint Croix, United States Virgin Islands.

The first report of Mansonia dyari on Saint Croix, United States Virgin Islands (USVI), is confirmed. Adult and larval specimens were collected in 2018 and 2019 through adult surveillance and larval collections. Specimens were identified by microscopic methods, and a representative specimen was confirmed by DNA sequencing (mitochondrial cytochrome c oxidase subunit I). Morphological features are reviewed and compared with Mansonia flaveola, a species previously reported in the USVI. Notes are provided on the locations, collection methods, and mosquito associates found with Ma. dyari in the USVI.