NE1043: Biology, Ecology & Management of Emerging Disease Vectors

(Multistate Research Project)

Status: Inactive/Terminating

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SAES-422 Reports

Annual/Termination Reports:

[05/04/2010] [05/02/2011] [03/20/2012] [12/22/2014]

Date of Annual Report: 05/04/2010

Report Information

Annual Meeting Dates: 02/25/2010 - 02/26/2010
Period the Report Covers: 10/01/2008 - 09/01/2009

Participants

Allan, Sandra (sandy.allan@ars.usda.gov) USDA-ARS/Florida;
Andreadis, Theodore (theodore.andreadis@ct.gov) Connecticut, Agricultural Experiment Station;
Cortinas, Roberto (rcortinas@unl.edu) Nebraska, University of Nebraska;
Dobson, Stephen (sdobson@uky.edu) Kentucky, University of Kentucky;
Fallon, Ann (fallo002@umn.edu) Minnesota, University of Minnesota;
Federici, Brian (brian.federici@ucr.edu) California, University of California-Riverside;
Fefferman, Nina (fefferman@aesop.rutgers.edu) New Jersey, Rutgers University;
Fonseca, Dina (dinafons@rci.rutgers.edu) New Jersey, Rutgers University;
Gaugler, Randy (gaugler@rutgers.edu) New Jersey, Rutgers University;
Gingrich, Jack (gingrich@udel.edu) Delaware, University of Delaware;
Kaufman, Michael (Kaufma15@msu.edu) Michigan, Michigan State University;
Paskewitz, Susan (paskewit@entomology.wisc.edu) Wisconsin, University of Wisconsin;
Robinson, Mark (mrobinson@nifa.usda.gov) USDA-NIFA;
Ruiz-Moreno, Diego New York, Cornell University;
Stafford, Kirby (kirby.stafford@ct.gov) Connecticut, Agricultural Experiment Station

Brief Summary of Minutes

The annual meeting of Multi-State Project NE1043, "Biology, Ecology & Management of Emerging Disease Vectors" was held on February 25, 2010 at the Rutgers University Center for Vector Biology in New Brunswick, New Jersey.

The following were in attendance: Allan (ARS-Gainesville), Andreadis (CT), Cortines (NE) Dobson (KY), Gingerich (DE), Fallon (MN), Federici (CA), Fefferman (NJ), Fonseca (NJ), Gaugler (NJ), Kaufman (MI), (OK), Paskewitz (WI), Robinson (USDA-NIFA), Ruiz-Moreno (NY), Stafford (CT)

Randy Gaugler (Chair) reported that NE1043 has been formally approved and will run from August 1, 2009 through June 30 2014. Kirby Stafford (CT) is the new Administrative Advisor and Mark Robinson will remain as the USDA/NIFA (formally CSREES) representative. We currently have 22 members representing 11 Agricultural Experiment Stations with representatives from ARS/USDA.

Mark Robinson, (USDA-NIFA representative) discussed the new Farm Bill and the reorganization of CSREES into the National Institute for Food & Agriculture. He noted that there will be more emphasis on the direction of Hatch funding into Multi-State projects from 25% to 30% with a strong emphasis on research. He reviewed the Federal FY2011 budget highlights and announced that $400 million in increased funding will support the following 5 high priority issues with a focus on multi state/agency and interdisciplinary approaches: bioenergy, global climate change, global food security, nutrition and health, and food safety. A discussion ensued as to how the current project may fit into these priority areas.

Nina Fefferman (Rutgers Center for Vector Biology) presented a seminar entitled "Risk assessment models for West Nile virus and eastern equine encephalitis" which stimulated much open discussion on the importance of various factors that should be included in any model.
Brief research and project updates with accompanying discussion were presented by each cooperator.

The following individuals were elected to serve 2 yr terms as officers for the project:

Theodore Andreadis (CT) - chair

Edward Walker (MI) - vice-chair

Sandra Allan (ARS-Gainesville) - secretary

The meeting was adjourned one day early as a result of a snowstorm.

The next meeting is scheduled for late February or early March 2011 at The Connecticut Agricultural Experiment Station, New Haven, CT. The participants agreed to hold a 1.5 day meeting with a similar venue and to include invited research seminar on a topic to be announced. The next meeting should also provide opportunities for 'cooperator reports' by objective, emphasizing presentations that show promise for collaboration.

Accomplishments

Objective 1. Strengthen basic and applied research on the mosquito, pathogen, hosts, and environmental factors that influence disease emergence. An introduction of Aedes albopictus was documented at a commercial tire recycling plant in northeastern CT. Adult females were collected from late Jul Oct at the plant. Host-seeking females attempting to alight on human subjects and larvae hatching from eggs collected in ovitraps placed in the woodlands surrounding the tire plant, were detected from mid-Aug - Oct denoting seasonal establishment. However, no larvae were recovered from eggs collected in ovitraps placed in the surrounding woodlands or in traps placed 1.6 km away, nor were females detected by human subjects the following season, indicating that the species did not survive winter conditions. The failure of Ae. albopictus to overwinter was likely due to winter egg mortality and/or interspecific competition from Ae. triseriatus and Ae. japonicus. (CT) Blood-engorged female Ae. j. japonicus were collected and blood meals identified by sequencing portions of the cytochrome b gene of mitochondrial DNA. Mammalian hosts included white-tailed deer (53%), humans (36%), fallow deer (5%), horse (3%), and opossum (3%). No avian, amphibian, reptilian or mixed blood meals were identified. (CT, NJ) Engorged An. quadrimaculatus and An. punctipennis mosquitoes were collected from EEE virus foci in central NY, and throughout NJ and their blood meals were identified. Analysis of An. quadrimaculatus and An. punctipennis from NY revealed 97.7% and 97.2% acquired blood from mammalian hosts, respectively. An. quadrimaculatus and An. punctipennis from NJ revealed 100% and 96% mammalian-derived blood meals. Individual mosquitoes containing mixed-blood meals from both avian and mammalian hosts were detected in 1.6% of An. quadrimaculatus in NY, and 2.8% and 4.0% of An. punctipennis from NY and NJ, respectively. White-tailed deer constituted the most common vertebrate host, comprising 85.8% to 97.7% of all blood meals. One horse- and two human-derived blood meals were detected from An. quadrimaculatus collected in NJ. EEE virus was isolated from one An. punctipennis collected in NY. Limited avian-derived blood meals were detected from mourning dove, sharp-shinned hawk, and house finch. Occasional feeding on avian hosts suggests that these mosquitoes may participate as epidemic/epizootic bridge vectors of EEE virus from viremic birds to horses and humans. Predominance of white-tailed deer as a source of blood meals supports enzootic amplification of deer-associated arboviruses in this region, including Jamestown Canyon, Cache Valley, and Potosi viruses. (CT, NJ, NY). We examined the genetic variations of Cx. pipiens mosquitoes from Chicago, IL that were determined to be principally ornithophilic but exhibited a higher inclination for mammalian hosts including humans. Microsatellite analysis of 10 polymorphic markers was performed on Cx. pipiens specimens with identified avian or mammalian blood meals. There were no significant differences in allelic richness, the pattern of conformity to Hardy-Weinberg equilibrium and linkage disequilibrium, nor was there overall genetic differentiation between specimens with avian- and mammalian-derived blood meals. However, Cx. pipiens form pipiens with mammalian- (including human-) derived blood meals had significantly higher ancestry and proportion of hybrids from the Cx. pipiens form molestus (population from NYC) than did those with avian-derived blood meals. By contrast, there were no significant differences in the ancestry and the proportion of hybrids from Cx. quinquefasciatus (TX). No temporal genetic variation was detected in accordance with the observation that there was no shift in blood feeding from birds to mammals. The results suggest that the probability of genetic ancestry from Cx. pipiens f. molestus may predispose mosquitoes to feed more readily on mammals, however the genetic mechanisms are unknown. (CT, MI) We have completed initial investigations of dispersal and survival of marked male Ae. albopictus in the field using MRR experiments. Additional effort will be devoted to comparing marking methods for persistence in the field and potential impacts on male survivorship and dispersal. Comparisons of longevity, dispersal rate and survivorship will be used to determine the advantages/disadvantages with the differing approaches. The experimental design (e.g. collection times, types of habitat) and mathematical interpretation of results will be coordinated with other participants. Pending appropriate regulatory approval, males infected with differing Wolbachia infection types may also be compared using the developed approaches. A large greenhouse mesocosm has been used to examine a Wolbachia-based microbial biopesticide approach against Ae. albopictus. Additional tests will compare the percent 'incompatibly mated' females with the incompatible males and allow assessment of male competitiveness for mates. (KY) Operational research was conducted in preparation for a large scale study of Culex pipiens dispersal from larval habitats (catch basins in urban Chicago). The research focused on the development and application of 13C and 15N stable isotope labeling methods of larval foods, in particular either in solution or incorporated into algae. The concept is to amend the larval habitats with these materials which results in 'marked' adults at a ca. 10-fold higher ratio of 13C/12C or 15N/14N than natural occurs. Mass spectroscopy is then used to detect the isotopic signature in individual, or in pooled, mosquitoes captured at the study site. Additional research was continued on the distribution and abundance of Oc. japonicus in Michigan, and new studies on the significance of algae to larval nutrition of this species. (MI) High resolution spatial population genetics has revealed two independent introductions of the invasive species Ae. japonicus, that are now mixing. We have obtained specimens from multiple locations in Japan for this analysis. In a study on sibling analysis of Cx pipiens populations, we have finished analysis of the proportion of egg masses fertilized by multiple males by examining offspring from successive egg masses from females mated in captivity. We have also dissected the spermatheca of females caught in the wild and will examined these next. We have completed inheritance analyses on real-time rates of dispersal of insect populations. A panel of 17 microsatellites were developed for Cx restuans. We have confirmed that, unlike Cx pipiens, this species over winters in large numbers even in forested areas with few underground human-made structures. Also, bioassays to examine Cx. pipiens and Ae. albopictus resistance to organophosphates and pyrethroids have been developed. (NJ) While examining hybrid populations in the Culex pipiens complex in Asia we identified and characterized a system where males are genetically distinguishable from females at the DNA level using a simple PCR- based assay. This unusual but very useful condition resulted from an old hybridization event coupled with suppressed recombination on chromosome 1, as ascertained using Single Nucleotide Polymorphisms (SNPs). Since suppressed recombination is often associated with critical sex determining genes, and the MDL has been linked to several genetic markers we have co-located in this region, we assert that the elusive MDL is located within this region of about 1,000,000 bp. An initial scan of the genes present has identified the testis development protein (TDP), a candidate gene in the sex determination pathway. (NJ) A knowledge, attitudes, and practices (KAP) questionnaire combined with entomological surveys of residential mosquito breeding sites were conducted in two neighborhoods. We tested the hypothesis that "correct" West Nile virus (WNV) knowledge and perceptions correspond with the use of practices that prevent mosquitoes from breeding and biting. Our results demonstrate that perceptions of WNV relate to the number of positive containers in yards and the use of mosquito preventive measures. In contrast, WNV knowledge was not related. Culex pipiens and Cx. restuans were common species found breeding in containers. Ochlerotatus japonicus was the most abundant species in 77% of positive containers (buckets, flower pots and birdbaths). This new invasive mosquito together with the Culex species identified in this study represents significant potential as disease vectors. This is the first study to directly investigate the relationship between KAP and breeding of WNV vectors in residential yards. (NY) Behavioral analysis of oviposition behavior by Aedes albopictus and Aedes aegypti revealed differences between the two species and provide the basis for subsequent evaluations of potential oviposition repellents or toxicants. (ARS-Gainesville) Objective 2. Use knowledge of mosquito, pathogen, vertebrate reservoir, and environment interactions to enhance ability to predict conditions leading to disease. Field studies suggest that Cx. pipiens feeds preferentially on American robins. To determine the contribution of innate preferences to observed preference patterns in the field, we conducted host preference trials with adult female Cx. pipiens in outdoor cages comparing the relative attractiveness of American robins with two sympatric bird species, European starling and house sparrow. Host seeking Cx. pipiens were 3x more likely to enter robin-baited traps when paired with a starling and almost twice more likely when paired with a house sparrow. There was no difference in the probability of trap entry when two robins were offered. Logistic regression analysis determined that the age, sex and weight of the birds, the date of the trial, starting-time, temperature, humidity, wind-speed and age of the mosquitoes had no effect on the probability of a choosing a robin over an alternate bird. Findings indicate that preferential feeding by Cx. pipiens mosquitoes on certain avian hosts is likely to be inherent. (CT) The optimal method for early prediction of West Nile virus (WNV) infection risk remains controversial. We analyzed the predictive utility of risk factor data for human WNV over a six-year period in CT. Using multiple logistic regression, the 30-day risk of human WNV infection by town was modeled using environmental variables as well as mosquito and wild bird surveillance. Using only environmental variables or animal sentinel data was less predictive than a model that considered all variables. In the final parsimonious model, population density, growing degree-days, temperature, WNV positive mosquitoes, dead birds and WNV positive birds were significant predictors of human infection risk, with an ROC value of 0.75. A real-time model using climate, land use, and animal surveillance data to predict WNV risk appears feasible. The dynamic patterns of WNV infection suggest a need to periodically refine such prediction systems (CT) WNV infection rate in C. pipiens is highly seasonal, being virtually zero in early summer, rising rapidly in mid-summer, then declining. The rise in infection rate mid-summer is strongly correlated with the appearance of fledgling birds, suggesting that age structure of the bird populations being fed upon influenced viral infection dynamics. However, results of a field study did not uphold the hypothesis that variation in avian community species diversity influenced the intensity of transmission, nor were nestling birds found to have higher exposure to infectious mosquito bites. In the latter case, nestling birds were not concluded to be the major age class promoting virus transmission; rather, post-nestling birds (fledglings) are likely the important age class whose appearance seasonally promotes annual, mid-summer epizootics. Blood meal analysis of C. pipiens, the primary vector in the study area, revealed that few bird species contribute to most of the estimated amplification fraction based on an ecological index estimator, namely, American robin, house finch, and blue jay. Further, C. pipiens fed at a higher than anticipated frequency on mammals, in particular humans (~16%). The tendency to feed on avians versus mammals was found to have an association with genetic ancestry within this mosquito population, where those individuals that had fed on mammals had a significantly higher ancestry for the unusual molestus form, whilst the avian feeding individuals lacked these genetic markers. Additional research on molecular genetic diversity of WNV over three year epidemic period showed that the virus is slowly evolving under a model of neutral, stabilizing selection with an increase of approximately 0.1% per yr in nucleotide-level genetic diversity. Analysis of mosquito infection among sites and years showed a strong relationship to temperature; cooler temperatures resulted in lower and seasonally delayed infection rates. (MI) A multivariate model was developed using both climatic and biotic variables in predicting the number of weekly New Jersey human cases using data from 2002-2006 data. We were able to account for greater than 70 percent of the variability. The model featured variables that were lagged to include time from being bitten by an infected mosquito to showing symptoms (i.e., incubation time up to 14 days). (NJ) We are developing a mathematical model that explores how behavioral influences of feeding preferences could cause disease amplification within a vector species without associated amplification being a necessary consequence in any of the host species. Additionally, a predictive model is being developed to explore hotspots for the potential evolution of newly arising insecticide resistance. This work will set the foundation for the construction of strategies for pesticide use which will enable long-term disease control rather than merely short-term effects which can lose efficacy as vector populations develop resistance. (NJ) Due to clear differences in rates of human infections between high/low latitude and mid latitude eastern seaboard states, in collaboration with Dr. Kramer we are analyzing the effect of underlying genetic make-up on the WNV vector competence of the northern Cx. pipiens, tropical/southern Cx quinquefasciatus, and hybrids between the two. We have optimized diapause markers for Cx pipiens in our lab to examine how the interplay of temperature and genetics affects vector competence in the Culex pipiens complex across the United States. (NJ) The behavioral role of acoustics in mosquito mating was explored, with male and female Aedes converging flight tone harmonics to match each other. We further demonstrated that this interaction occurs most frequently when the potential mate represents the optimal physical conditions to maximize reproductive success. Consequently, this acoustic behavior may be used in mate assessment. Further studies are investigating potential approaches for acoustic disruption of this essential pre-copulatory behavior. (NY) Studies been aimed at identifying male reproductive proteins that are transferred to females and may have significant impacts on female physiology. These proteins represent potential and novel targets for mosquito control. A novel method of reverse isotope labeling followed by proteomic analysis was used to examine proteins transferred to females. Sperm enriched proteins were also identified and analyzed. A total of 50 male seminal fluid proteins and 38 sperm proteins have been identified using this approach. The seminal fluid protein classes suggested roles in sperm storage and protection from oxidative stress, ecdysteroidogenesis, and protein activation. Many of the sperm proteins were homologous to Drosophila sperm proteins, suggesting conservation of sperm-related function across Diptera. (NY) Objective 3. Develop strategies to control mosquito vectors. Previous studies have shown that several species of anophelines are sensitive to the Bacillus sphaericus (Bs) binary toxin (Bin toxin). Thus, we initiated studies on the efficacy and resistance management properties of our two best recombinant larvicidal stains of B. thuringiensis subsp. israelensis (Bti) against Anopheles gambiae. Both of these strains have been genetically engineered to produce large amounts of the Bs Bin toxin. The best strain, referred to here as Bti/BsBin, produces the Bs Bin in combination with the four major endotoxins of Bti, which are Cry4A, Cry4B, Cry11A and Cyt1A. When tested against fourth instars of An. gambiae, this strain was nine-fold more effective at the LC95 level than the strains of Bti and Bs used in current commercial products. Interestingly and importantly, preliminary selection studies with the Bti/BsBin recombinant show no resistance development in An. gambiae after nine generations of selection. These results are similar to what we have observed in selection studies with the same Bti/BsBin strain against Cx. quinquefasciatus, which is known to develop high levels of resistance quickly under field conditions when only treated with B. sphaericus formulations. (CA) Factors that directly impact horizontal transmission of the microsporidium Amblyospora albifasciati to its intermediate copepod host, Mesocyclops annulatus were examined in laboratory bioassays in relation to life history strategies that facilitate persistence of the parasite in natural populations of its definitive mosquito host, Oc. albifasciatus. A moderately high quantity of meiospores from mosquito larvae was required to infect adult female copepods (IC50 = 3.6 x 10 4 meiospores/ml). Meiospore infectivity following storage at 25C was detected up to 30 days, while meiospores stored at 4C remained infectious to copepods for 17 months. The pathological impact of A. albifasciati infection on M. annulatus resulted in a 30% reduction in survivorship after 7 days followed by gradual progressive mortality up to 40 days. Infected female copepods survived in sediment under desiccation up to 30 days. The susceptibility of late stage copepodid M. annulatus to meiospores of A. albifasciati and transstadial transmission of infection to adult females was established. (CT) Our goal is to establish research collaborations related to mosquito reproduction, mosquito cell cultures, and maintenance of the bacterium Wolbachia pipientis, for eventual modification as an agent for manipulation of mosquito populations. (MN) In an Asian tiger mosquito area-wide management project, we achieved 50%-60% reductions in intervention sites in Mercer and Monmouth counties. Ae. albopictus populations in the 3 Monmouth sites increased very slowly (compared to both the 2008 season and Mercer) and therefore significant reduction occurred only in late August and September when the populations peaked. Education efforts resulted in a significant reduction in Ae. albopictus populations in Mercer but not in Monmouth. Dramatic socioeconomic differences between the two counties had unexpected results. That is, it was considerably easier to access teachers and their students in inner-city Mercer than in suburban Monmouth. Overall, Mercer sites were easier to work in, both for the education and the control teams. The number and nature of Ae. albopictus larval sources in the two counties was also different and impacted both the type and outcome of education and management measures. (NJ, ARS-Gainesville) We are examining the potential of biological control to supplement current mosquito control efforts in southern Wisconsin. We determined that fathead minnows serve as good predators of Culex spp. in the lab and as excellent biocontrol tools in a pilot study of three field sites. In 2009, we expanded field introductions to nine locations and measured ecological parameters that affected fish persistence. In 6 of the 9 sites, control of Culex spp. was nearly absolute for most of the summer. In three sites, fish were not retained. Depth and energy dissipaters were key factors in persistence. Further, we are examining the effectiveness of copepods as predators. These crustaceans are more effective against mosquitoes than any other invertebrate predator and they can be mass produced inexpensively. We collected copepods from catch basins in Fall of 2009, identified three species, and began culturing them in the lab. Preliminary trials using Macrocyclops albidus indicated that local isolates are capable of feeding on early instar Culex spp. Current work focuses on optimizing culture conditions in the lab for scaled up production and releases into catch basins. (WI) Emphasis has been on examination of host odors for attractant discovery, use of structure-activity models to develop new repellents and toxicants, and characterization of barrier sprays. Odors from chickens were identified by gas chromatography/mass spectrometry and bioassays of these compounds are in progress. Research with repellents and toxicants is drawn from data generated by the USDA screening program from 1942-1977. From a set of structurally similar acylpiperdine compounds, a successful model was developed, and novel efficacious repellents were predicted, synthesized and verified by bioassay. Some compounds from this set are being evaluated against other arthropods. The current emphasis of modeling work is on the development of new toxicants. Collaborative projects with barrier sprays involved comparison of sprayers and comparison of efficacy duration for three pyrethroid active ingredients. (ARS-Gainesville) Landing responses of mosquitoes (Aedes albopictus) and sand flies (Lutzomyia shannoni, Phlebotomus papatsi) on wax myrtle leaves treated with commercial residual pesticides (maximum label rate) were video-taped and analyzed to determine effect on landing frequency and duration. Significant differences existed in landing and resting responses between mosquito species (Aedes aegypti, Anopheles quadrimaculatus and Culex quinquefasciatus) exposed to sub-lethal rates of pyrethroids and better knowledge of the differences helps optimize efficacy of treatment against targeted species. Lab evaluation of components of Duet® ULV spray using Culex quinquefasciatus and Lutzomyia shannoni indicated that prallethrin enhanced flight activity resulting in more droplets and greater mortality. Species of plant, application method, rain exposure and sunlight significantly decreased efficacy of bifenthrin-treated leaves for eliciting mortality of Ae. aegypti determined in laboratory assays. (ARS-Gainesville) In addition to studies on structure activity relationships of compounds for new toxicant discovery, we are using gene silencing using RNAi technology as a method to knock down critical proteins in mosquitoes with possible applications for control. An inhibitor of apoptosis protein 1 gene in Aedes aegypti (AaeIAP1) was chosen as a target for the development of molecular pesticides and when topically applied was shown to cause mortality in adult Aedes aegypti. Other potential targets and new constructs are being evaluated. (ARS-Gainesville) Semi-field and field studies were conducted on the use of barrier sprays to reduce host-vector contact. Barriers were constructed of either treated vegetation or camouflage fabric. These barrier materials were sprayed with one of several candidate synthetic pyrethroid compounds. Traps baited with carbon dioxide and 1-octen-3-ol served as surrogate hosts. The traps were placed in the center of the treated barrier plots ( either 5 x 5 or 7 x 7). Preliminary results indicate effective protection from mosquitoes for 3-6 weeks depending on the mosquito species, type of insecticide treatment, barrier material and/or environmental conditions. (ARS-Gainesville) Objective 4. Enhance surveillance technologies for mosquitoes and mosquito-borne pathogens. Statewide mosquito trapping and testing for mosquito-borne arboviruses was conducted from Jun - Oct in CT. A total of 291,641 mosquitoes (16,895 pools) representing 38 species were trapped and tested. 35 isolations of WNV were made from: Cx. pipiens (25), Cx. restuans (3), Cx. salinarius (1) and Oc. stimulans (1), collected at 19 sites in 5 counties (Fairfield, Hartford, Middlesex, New Haven, and New London) from Jul 15 - Oct 8. Widespread eastern equine encephalitis (EEE) activity was detected. 123 isolations of EEE were made from: Cs. melanura (84), Oc. canadensis (10), Ae. cinereus (6), Ur. sapphirina (4), An. punctipennis (4), Cx. salinarius (3), Ae. vexans (2), An. quadrimaculatus (2), An. walkeri (2), Oc. trivittatus (2), Cx. restuans (1), Oc. cantator (1), Oc. triseriatus (1), and Ps. ferox (1) collected at 25 sites in 7 counties from Aug 17 - Oct 27. One horse case and infections in 2 commercial pheasant flocks were reported. Other mosquito-borne viruses isolated included: Jamestown Canyon (43), Cache Valley (1), Highlands J (60), Trivittatus (29), and Flanders (1). (CT) New Jersey surveillance for EEE indicated high activity levels in the enzootic vector Culiseta melanura with 57 positives of 371 pools submitted from the traditional resting box sites -- a five-fold increase from the previous year. Dissemination of the virus was widespread and began earlier (15 July 2009 versus early August of 2008). Non-melanura species were also positive for the first time in several years, including first time positive pools for Culex erraticus and Aedes japonicus. Six horses and two alpaca were EEE positive. Surveillance for WNV found 322 positive pools from the 9,965 pools submitted, mostly in Culex pipiens (primary enzootic vector), Culex restuans or mixed Culex pools. One horse case and three human cases developed. Positive mosquitoes appeared well before the first positive dead bird, indicating that the value of dead bird reports have decreased. Surveillance for SLE and LAC viruses was negative. (NJ) We compare the efficiency and sensitivity of the Rapid Analyte Measurement Platform (RAMP) to reverse transcriptase polymerase chain reaction (RT-PCR) from 2005 to 2008 to detect West Nile virus (WNV). Overall, 316 pools tested negative and 115 pools tested positive for WNV. Eighty-nine pools tested positive using RAMP and all were confirmed by RT-PCR; 26 pools were WNV-negative using RAMP but positive using RT-PCR. False-positives from RAMP were not detected. (NJ) More Ae. albopictus females were collected at 1 m compared to 6 m above ground with significantly more collected in suburban compared to sylvatic habitats. Optimal placement of traps will increase surveillance sensitivity. Propane powered commercial traps producing CO2 were as effective as CDC traps that required a separate CO2 source for Ae. albopictus surveillance efforts and these types of traps may provide additional surveillance tools. (ARS-Gainesville) Objective 5. Develop strategies for sustainable mosquito control by including training at all levels. No specific reports submitted; however, training of students and postdocs is an integral component of nearly all of the above reports.

Publications

Allan, S. A., D. L. Kline and T. Walker. 2009. Environmental factors affecting efficacy of bifenthrin-treated vegetation for mosquito control. J. Amer. Mosq. Control Assoc. 25: 338-346. Alphey, L., M. Benedict, R. Bellini, G. G. Clark, D. A. Dame, M. W. Service, and S. L. Dobson. 2009. Sterile-Insect Methods for Control of Mosquito-Borne Diseases: An Analysis. Vector Borne Zoonotic Dis. Andreadis, T. G. 2009. Failure of Aedes albopictus (Diptera: Culicidae) to overwinter following introduction and seasonal establishment at a tire recycling plant in the northeastern USA. J. Amer. Mosq. Control Assoc. 25: 25-31. Andreadis, T. G. and Wolfe, R. J. Evidence for reduction of native mosquitoes with increased expansion of the invasive Ochlerotatus japonicus japonicus (Diptera: Culicidae) in the northeastern United States. J. Med. Entomol. (in press). Arthur, BJ, Wyttenbach, RA, Harrington, LC and R R. Hoy. Neural Responses to One- and Two-Tone Stimuli in the Hearing Organ of the Dengue Vector Mosquito. J. Exp. Biol. (in press). Bartlett-Healy, K, W Crans & R Gaugler. 2009. Vertebrate hosts and phylogenetic relationships of amphibian trypanosomes from a potential invertebrate vector, Culex territans. J Parasitol 95:381-7. Bataille A, Cunningham AA, Cedeño V, Cruz M, Eastwood G, Fonseca DM, Causton CE, Azuero R, Loayza J, Cruz Martinez JD, and Goodman SJ. 2009. Evidence for regular on-going introductions of mosquito disease vectors into the Galápagos Islands. Proc Roy Soc Lond, Ser B. 276:3769-75. Bentley, M.T., Kaufman, P.E., Kline, D.L. & Hogsette, J.A. 2009. Response of adult mosquitoes to light-emitting diodes placed in resting boxes and in the field. J. Am. Mosq. Control Assoc. 25: 285-291. Brelsfoard, C. L., W. St Clair, and S. L. Dobson. 2009. Integration of irradiation with cytoplasmic incompatibility to facilitate a lymphatic filariasis vector elimination approach. Parasites & Vectors 2: 8. Cameron E, RC Wilkerson, M Mogi, Toma T, Myiagi I, Kim H-C, Fonseca DM. 2010. Molecular phylogenetics of Aedes japonicus, a disease vector that recently invaded Western Europe, North America, and the Hawaiian Islands. J Med Entomol (in press). Cantrell, C., Klun, J., Pridgeon, J. W., & Becnel, J. J. 2009. Structure-activity relationship studies on the mosquito toxicity and biting deterrency of callicarpenal derivatives. Chem. Biodiversity. 6: 447-458. Cator, LJ, Arthur, BJ, Harrington, LC and RR Hoy. 2009. Harmonic convergence in the love songs of the dengue vector mosquito. Science. 323(5917):1077-9. Chaves, L.F., Harrington, L.C., Keogh, C.L, Nguyen, A.M. and U. D. Kitron. Blood feeding patterns of mosquitoes: random or reflective of preferences? Frontiers in Zool.. (in press). Condon, GC, SP Healy & A Farajollahi. 2009. Sentinel chicken coop modification for canopy-level mosquito-borne disease surveillance. J Am Mosq Control Assoc. 25:390-3. Condon, GC, PJ Clayson, E Williges & A Farajollahi. 2009. A simplified tripod support for use with carbon dioxide-baited vector surveillance traps. J Am Mosq Control Assoc. 25:221-3. Cooperband, M. F. and S. A. Allan. 2009. Effects of different pyrethroids on landing behavior of female Aedes aegypti, Anopheles quadrimaculatus, and Culex quinquefasciatus mosquitoes (Diptera: Culicidae) J. Med. Entomol. 46: 292-306. Diuk-Wasser, M. A., Molaei, G., Simpson, J. E., Folsom-OKeefe, C. M., Armstrong, P. M., and Andreadis, T. G. Avian communal roosts as amplification foci for West Nile virus in urban areas in northeastern United States. Am. J. Trop. Med Hyg. (in press). Doyle, M. A., D. L. Kline, S. A. Allan and P. E. Kaufman. 2009. Efficacy of residual bifenthrin applied to landscape vegetation against Aedes albopictus. J. Amer. Mosq. Control Assoc. 25: 179-183. Fallon AM and Witthuhn BA, 2009. Proteasome activity in a naive mosquito cell line infected with Wolbachia pipientis wAlbB. In vitro cellular and developmental biology-Animal 45:460-6. Farajollahi, A, B Kesavaraju, M Nelder, S Crans & R Gaugler. 2009. A unique larval collection and survival of Orthopodomyia signifera in the presence of the predator Toxorhynchites rutilus septentrionalis. J Am Mosq Contr Assoc 25:370-3. Farajollahi, A, B Kesavaraju, D Price, G Williams, S Healy, R Gaugler & M Nelder. 2009. Field evaluation of BG-Sentinel and industry-standard traps for Aedes albopictus and West Nile virus surveillance. J Med Entomol 46:919-25. Farajollahi, A & MP Nelder. 2009. Changes in Aedes albopictus (Diptera: Culicidae) populations in New Jersey and implications for arbovirus transmission. J Med Entomol. 46:1220-4. Federici, B. A. 2010. Recombinant bacterial larvicides for control of important mosquito vectors of disease. Pages 163-176, In Vector Biology, Ecology and Control, (P. W. Atkinson, Editor). Springer: Dordrecht, Heidelberg, London, New York. Fonseca D.M, Harrison B, Faybyshev M, Kramer LD. Selection on phenotypic traits in the US Culex pipiens complex hybrid swarm. Genes (in press) Fonseca DM, Widdel A, Spichiger S-E, Hutchinson M, Kramer LD. Fine-scale spatial and temporal population genetics of a new US mosquito, reveal multiple introductions. Mol Ecol (in press) Fonseca DM, Smith JL, Kim H-C, Mogi M. 2009. Population genetics of the mosquito Culex pipiens pallens reveals sex-linked asymmetric introgression by Culex quinquefasciatus Infection. Genet Evol 9:1197-203. Gavotte, L., D. R. Mercer, R. Vandyke, J. W. Mains, and S. L. Dobson. 2009. Wolbachia Infection and Resource Competition Effects on Immature Aedes albopictus (Diptera: Culicidae). J. Med. Entomol. 46: 451-459. Haddow, A, J Moulton, R Gerhardt, L McCuiston & C Jones. 2009. Description of the egg of Ochlerotatus japonicus japonicus (Diptera: Culicidae) using variable pressure scanning electron microscopy. J. Med. Entomol. 46:9-14. Hamer G, Kitron U, Goldberg TL, Brawn JD, Loss SR, Ruiz MO, Hayes DB, Walker ED. 2009. Host selection by Culex pipiens indicts birds responsible for West Nile virus amplification. Am. J. Trop. Med. Hyg. 80: 268-278. Harvell,D, Altizer,S, Cattadori, IM, Harrington, LC, and E Weil. 2009. Climate Change and Wildlife Disease: When Does the Host Matter the Most? Ecology. 90: 912-920. Hoel, D. F., D. L. Kline and S. A. Allan. 2009. Evaluation of six mosquito traps for collection of Aedes albopictus and associated mosquito species in a suburban setting in north central Florida. J. Amer. Mosq. Control Assoc. 25: 47-57. Hoffmann, W.C., Farooq, M., Walker, T.W., Fritz, B., Szumlas, D., Quinn, B., Bernier, U., Hogsette, J., Lan, Y., and Huang, Y. 2009. Canopy Penetration and Deposition of Barrier Sprays from Electrostatic and Conventional Sprayers, J. Am. Mosq. Control Assoc. 25: 323-331. Huang, S., Hamer, G., L., Molaei, G., Walker, E., D., Goldberg, T., L., Kitron, U. D., and Andreadis, T. G. 2009. Genetic variation associated with mammalian feeding in Culex pipiens from a West Nile virus epidemic region in Chicago, Illinois. Vector-Borne Zoonotic Dis. 9:637-642. Irwin, P. and S. Paskewitz. 2009. Investigation of fathead minnows (Pimephales promelas) as a biological control agent of Culex mosquitoes under laboratory and field conditions. J. Amer. Mosq. Conro. Assoc. 25: 301-309. Kesavaraju, B, A Afify & R Gaugler. Growth and survival of the invasive Aedes albopictus larvae on Diospyrus virginiana (American persimmon) leaves. J Med Entomol (in press). Liu, A., Lee, V., Galusha, D., Slade, M., Diuk-Wasser, M., Andreadis, T., Scotch, M., and Rabinowitz, P. 2009. Risk factors for human infection with West Nile virus in Connecticut: a multi-year analysis. Int. J. Health Geographics 8:67 doi:10.1186/1476-072X-8-67. Loss SR, Hamer GL, Walker ED, Ruiz MO, Goldberg TL, Kitron UD, Brawn JD. 2009. Avian host community structure and prevalence of West Nile virus in Chicago, Illinois. Oecologia 159: 415-425. Loss, SR, Loss SR, Hamer GL, Walker ED, Ruiz MO, Goldberg TL, Kitron UD, Brawn JD. 2009. Nestling passerines are not important hosts for amplification of West Nile virus in Chicago, Illinois. Vector-Borne Zoonotic Dis. 9: 13-17. McCann, S., J. F. Day, S. A. Allan and C. C. Lord. 2009. Age modifies effect of body size on fecundity in Culex quinquefasciatus Say. J. Vector Ecol. 34: 174-181. Micieli, M. V., García, J. J. and Andreadis, T. G. 2009. Factors affecting horizontal transmission of the microsporidium Amblyospora albifasciati to its intermediate host, Mesocyclops annulatus. J. Invertebr. Pathol. 101:228-233. Molaei, G., Farajollahi, A., Scott, J. J., Gaugler, R., and Andreadis, T. G. 2009. Human blood feeding by the recently introduced mosquito, Aedes japonicus japonicus and public health implications. J. Amer. Mosq. Control Assoc. 25:210-214. Molaei, G., Farajollahi, A., Armstrong, P. M., Oliver, J., Howard, J. J., and Andreadis, T. G. 2009. Identification of blood meals in Anopheles quadrimaculatus and Anopheles punctipennis from eastern equine encephalitis virus foci in northeastern USA. Med. Vet. Entomol. 23:350-356. Nelder, M, A Farajollahi, S Healy, B Kesavaraju, I Unlu, T. Crepeau, D. Fonseca & R Gaugler. A novel combination of monomolecular film and (S)-methoprene to combat immatures of the Asian tiger mosquito Aedes albopictus. Amer J Trop Med & Hyg (in press). Obenauer, P.J., Kaufman, P.E., Allan, S.A. & Kline, D.L. 2009. Host-seeking height preferences of Aedes albopictus (Diptera: Culicidae) in North Central Florida suburban and sylvatic locales. J. Med. Entomol. 46: 900-908. Obenauer, P.J., Kaufman, P.E., Allan, S.A. & Kline, D.L. 2009. Infusion-baited ovitraps to survey ovipositional height preferences of container-inhabiting mosquitoes in two Florida habitats. J. Med. Entomol. 46: 1507-1513. Pang YP, Ekstrom F, Polsinelli GA, Gao Y, Rana S, Hua DH, Andersson B, Andersson PO, Peng L, Singh SK, Mishra RK, Zhu KY, Fallon AM, Ragsdale DW, and Brimijoin, 2009. Selective and irreversible inhibitors of mosquito acetylcholinesterases for controlling malaria and other mosquito-borne diseases. PLoS ONE 4:e6851, 2009; PubMed ID: 19714254 Park, H.-W., M. Tang, Y. Sakano, and B. A. Federici. 2009. I nsertion of a 1.1 kb downstream region from Bacillus sphaericus 2362 into B. sphaericus 2297 decreases Bin toxin synthesis and mosquitocidal activity. Applied and Environmental Microbiology 75, 878-881. Ponlawat, A., and L.C.Harrington. 2009. Factors associated with male mating success of the dengue vector mosquito, Aedes aegypti. Amer. Soc. Trop. Med. Hyg. 80: 312-8. Porcar, M., A.-M. Grenier, B. A. Federici, and Yvan Rahbe. 2009. Effect of Bacillus thuringiensis-endotoxins on the pea aphid, Acrythrosiphon pisum. Appl. Environ. Microbiol. 75, 4897-4900. Pridgeon, J.W., Bernier, U.R., and Becnel, J.J. 2009. Toxicity comparison of eight repellents against four species of female mosquitoes (Diptera: Culicidae). J. Am. Mosq. Control Assoc. 25: 168-173. Pridgeon, J. W., Becnel, J. J., Clark, G. G., & Linthicum, K. J. 2009. A High throughput screening method to identify potential pesticides for mosquito control. J. Med. Entomol. 46: 335-341. Reed, L, M Johansson, N Panella, R McLean, T Creekmore, R Puelle & N Komar. 2009. Declining mortality in american crow (Corvus brachyrhynchos) following natural West Nile virus infection. Avian Dis 53:458-46 Shively, J. M., G. C. Cannon, S. Heinhorst, J. A. Fuerst, D. A. Bryant, E. Gantt, J. A. Maupin-Furlow, D. Schuler, F. Pfeifer, R. Docampo, C. Dahl, J. Preiss, A. Steinbuchel, and B. A. Federici. 2009. Intracellular structures of Prokaryotes: Inclusions, compartments, and assemblages. Encyclopedia of Microbiology, pp. 404-424. Elsevier Sciences, Amsterdam. Simpson, J. E., Folsom-OKeefe, C. M., Childs, J. E., Simons, L. E., Andreadis, T. G. Diuk-Wasser, M. A. 2009. Avian host-selection by Culex pipiens Say (Diptera: Culicidae) in experimental trials. PLOS One 4(11): e7861. doi:10.1371/journal.pone.0007861. Suh, E., D. R. Mercer, Y. Q. Fu, and S. L. Dobson. 2009. Pathogenicity of Life- Shortening Wolbachia in Aedes albopictus after Transfer from Drosophila melanogaster. Appl. Environ. Microbiol 75: 7783-7788. Tuiten, W. Koenraadt, C.J.M, McComas, K. and L.C. Harrington. 2009. The effect of West Nile virus perceptions and knowledge on protective behavior and presence of vector breeding habitats in residential yards in New York State. Eco Health. DOI 10.1007/s10393-009-0219-z. Williges, E, A Farajollahi, M Nelder & R Gaugler. 2009. Comparative field analysis of rapid analyte measurement platform and reverse transcriptase polymerase chain reaction assays for West Nile virus surveillance. J Vect Ecol 34:324-8. Wirth, M. C., W. E. Walton, and B. A. Federici. Resistance to the Bacillus sphaericus Bin toxin is delayed significantly through combination with the mosquitocidal proteins of Bacillus thuringiensis subspecies israelensis. Environ. Microbiol. (in press). Wirth, M. C., W. E. Walton, and B. A. Federici. Inheritance patterns, dominance, stability and allelism of resistance and cross-resistance in two colonies of Culex quinqufasciatus (Diptera: Culicidae) selected with Cry endotoxins of Bacillus thuringiensis subsp. Israelensis. (in press.) Xue, R., Pridgeon, J. W., Becnel, J. J., & Ali, A. 2009. Fipronil as a larvicide against container-inhabiting mosquito, Aedes albopictus. J. Am. Mosq. Control Assoc. 25: 224-227. Zhao, L., Pridgeon, J. W., Becnel, J. J., Clark, G. G., & Linthicum, K. J. 2009. Identification of genes differentially expressed during heat shock treatment in Aedes aegypti. J. Med. Entomol. 46: 490-495. Zhao, L., Pridgeon, J. W., Becnel, J. J., Clark, G. G., & Linthicum, K. J. 2009. Mitochondrial Gene Cytochrome b Developmental and Environmental Expression in Aedes aegypti (Diptera: Culicidae). J. Med. Entomol. 46:1361-1369.

Impact Statements

Work carried out in 2009 as part of the Northeast Regional Research Project, NE-507, provided valuable theoretical and practical knowledge to help manage mosquito-borne disease. For example, the detection of a comparatively high prevalence of human blood feeding in Ae. j. japonicus in association with its local abundance, vector competence and repeated detection of West Nile virus illustrates the potential for this invasive mosquito to serve as a bridge vector.
The RAMP system was validated as a fast and reliable tool to augment existing RT-PCR-based methods for West Nile virus surveillance programs.
Research on new chemical insecticides is moving novel technologies from the bench to operational use.
Statewide mosquito and arbovirus surveillance programs provide an early warning system for intervention activities by local mosquito control agencies.
The feasibility of initiating large field cage experiments on the sterile insect technique was demonstrated.
A risk model was developed and made available via website to end-users (local mosquito control agencies) for predicting eastern equine encephalitis.
The groundwork was prepared for analysis of proteins to target for disrupting mosquito reproduction. Cutting-edge molecular methods are being deployed to identify gene function in proteins with potential for mosquito control.
Research led to an enhanced understanding of mosquito biology and host-mosquito interactions across an array of ecosystems. Unraveling these ecological relationships is essential to developing economical and effective integrated pest management programs for mosquitoes.
Genetic engineering research is resulting in new bacterial toxins with enhanced virulence against mosquito larvae with potential for commercial development as new, low risk biological insecticides.
Provided for and encouraged environmentally sound, scientifically based, and professional control by local mosquito control agencies.

Date of Annual Report: 05/02/2011

Report Information

Annual Meeting Dates: 03/01/2011 - 03/02/2011
Period the Report Covers: 10/01/2009 - 09/01/2010

Participants

Allan, Sandra (sandy.allan@ars.usda.gov) USDA-ARS/Florida;
Anderson, John (john.f.anderson@ct.gov) Connecticut, Agricultural Experiment Station;
Andreadis, Theodore (theodore.andreadis@ct.gov) Connecticut, Agricultural Experiment Station;
Armstrong, Philip (philip.armstrong@ct,gov) Connecticut, Agricultural Experiment Station;
Becnel, James (james.becnel@ars.usda.gov) USDA ARS Gainesville Florida;
Cortinas, Roberto (rcortinas@unl.edu) Nebraska, University of Nebraska;
Fallon, Ann (fallo002@umn.edu) Minnesota, University of Minnesota;
Federici, Brian (brian.federici@ucr.edu) California, University of California-Riverside;
Fonseca, Dina (dinafons@rci.rutgers.edu) New Jersey, Rutgers University;
Hardstone, Melissa (melissa.hardstone@ct.gov) Connecticut, Agricultural Experiment Station;
Kaufman, Michael (Kaufma15@msu.edu) Michigan, Michigan State University;
Daniel, Kline (dan.kline@ars.usda.gov) USDA ARS Gainesville Florida;
Molaei, Goudarz (goudarz.molaei@ct.gov) Connecticut, Agricultural Experiment Station;
Shepard, John (john.shepard@ct.gov) Connecticut, Agricultural Experiment Station;
Thomas, Michael (michael.c.thomas@ct.gov) Connecticut, Agricultural Experiment Station;
Stafford, Kirby (kirby.stafford@ct.gov) Connecticut, Agricultural Experiment Station;
Vossbrinck, Charles (charles.vossbrinck@ct.gov)Connecticut, Agricultural Experiment Station;

Brief Summary of Minutes

The annual meeting of the Multi-State Project NE1043, Biology, Ecology and Management of Emerging Disease Vectors was held on March 1-2, 2011 at The Connecticut Agriculture Experiment Station, New Haven, CT.

The following were in attendance: Allan (ARS-Gainesville), Anderson (CT), Andreadis (CT), Armstrong (CT), Becnel (ARS-Gainesville), Cortinas (NB), Fallon (MN), Federici (CA), Fonseca (NJ), Hardstone (CT), Kaufman (MI), Kline (ARS-Gainesville), Molaei (CT), Shepard (CT), Stafford (CT), Thomas (CT), and Vossbrinck (CT).

Dr. Kirby Stafford presented an administrative report and indicated that the project had 24 members representing 12 states with additional representation from ARS/USDA. Mark Robinson (USDA-NIFA representative) was unable to attend due to travel restrictions. Future funding opportunities through NIFA remain uncertain pending budget reductions.

A presentation was given by Dr. Goudarz Molaei (CAES) entitled Discriminating blood-feeding mosquitoes, their catholic counterparts, and transmission dynamics of West Nile virus in the US. Discussion on blood meal sources, role of nestlings and effect of virus infection on host choice followed.

Updates were provided by each cooperator with active discussion ensuing. The group photo was taken.

Several in depth presentations were made providing stimulus for further discussion and collaboration. Two discrete efforts that emerged were to develop databases on maintained mosquito colonies and cell culture lines with efforts headed by Allan (colonies) and Fallon (cell cultures).

Possible expansion of the group was discussed to include representatives from other Experiment Stations with scientists involved in mosquito research and the US Department of Defense Armed Forces Pest Management Board.

Elected officers remain in position for two years with no vacancies until next year. The next meeting will be at The Connecticut Agricultural Experiment Station at the same time of year. The format will remain the same with a 1.5 day meeting starting on Tuesday and featuring an invited speaker.

Accomplishments

Objective 1. Strengthen basic and applied research on the mosquito, pathogen, hosts, and environmental factors that influence disease emergence. In an effort to evaluate the invasion success and impact of Aedes japonicus japonicus on populations of native container dwelling species, thirteen waste tire disposal sites and four natural rock pool habitats were sampled for mosquito larvae throughout Connecticut in 2005, and data were compared with results from prior surveys of similar sites made in 1987 and 1999. Ae. j. japonicus was the predominant species collected at the waste tire disposal sites regardless of surrounding landscape features, accounting for 55.9% of all larvae. A comparison with collections from prior surveys revealed a 90% reduction in the relative abundance of larval populations of Aedes triseriatus and significant reductions among larval populations of Aedes atropalpus and Culex restuans. Ae.. j. japonicus was also the most abundant mosquito collected in rock pool habitats, accounting for nearly 80% of all collected larvae, except where water temperatures exceeded 30oC. This was concomitant with significant declines in the relative abundance of Oc. atropalpus and Cx. restuans. We conclude that Oc. j. japonicus is a potentially effective competitor in rock pool and tire environments and may be responsible for reducing populations of several native species occupying these habitats through interspecific competition for limited resources. The exclusion of Oc. j. japonicus from warm water pools further suggests that a temperature barrier may exist for Oc. j. japonicus and that populations may not be able to colonize effectively regions of the United States with relatively high summer temperatures. (CT) A 3-yr study was undertaken to examine the parity status, survival and prevalence of West Nile virus (WNV) in overwintering populations of Culex pipiens collected from a hibernaculum located in a WNV endemic region in New York City. Nearly 6,000 females were collected from December through April. Parity rates were highest among females collected in December and January, ranging from 12.3% to 21.9% depending on the year. In each year of the study, the proportion of parous females declined significantly during the course of the winter with the percentage of parous females found in April ranging from 0.9% to 10%. Results provide unequivocal evidence that parous female Cx. pipiens from this region of the northeastern US enter hibernacula in the fall in comparatively high proportions not previously recognized for this species, and while these females experience significant mortality during the winter some survive to April to emerge in the spring. The absence of any detectible blood remnants in overwintering females reaffirm that blood feeding does not occur among diapusing females during the winter. The possibility that a portion of the diapausing population may be autogenous as a result of hybridization with sympatric below-ground populations of Cx. p. pipiens form molestus is discussed. A single isolation of WNV was obtained in Vero cell culture from a pool of 50 females collected on January 11, 2007 representing an infection prevalence of 0.07% in the overwintering population in 2007. No isolations of WNV were made from mosquitoes collected in 2008 or 2009). Findings provide further evidence for local overwintering of WNV in diapausing Cx. p. pipiens, albeit at very low rates, consistent with the paucity of WNV positive mosquitoes detected in June and early July despite the emergence of females from hibernacula in early May in this region. (CT) Eastern equine encephalitis virus (EEEV) is maintained in an enzootic cycle involving Culiseta melanura mosquitoes and avian hosts, whereas other mosquito species that feed opportunistically on mammals have been incriminated as bridge vectors to humans and horses. To evaluate the capacity of these mosquitoes to acquire, replicate, and possibly transmit EEEV, we estimated the infection prevalence and virus titers in mosquitoes collected in CT by cell culture, plaque titration, and quantitative RT-PCR. Cs. melanura was the most important source of EEEV (68% of 122 virus isolations) and the only species to support consistently high virus titers required for efficient transmission. Our findings suggest that Cs. melanura serves as the primary enzootic and epidemic vector of EEEV in this region, which may explain the relative paucity of human cases. More generally, this study emphasizes the importance of evaluating virus titers from field-collected mosquitoes to help assess their potential role as vectors. (CT) Southern California remains an important focus of WNV virus activity, with persistently elevated incidence after invasion by the virus in 2003 and subsequent amplification to epidemic levels in 2004. Ecoepidemiological studies of vectors-hosts-pathogen interactions are of paramount importance for better understanding of the transmission dynamics of WN virus and other emerging mosquito-borne arboviruses. We investigated vector-host interactions and blood feeding behavior of 4 competent mosquito vectors by using a PCR method targeting mitochondrial DNA to identify vertebrate hosts of blood-fed mosquitoes. Diagnostic testing by cell culture, real-time RT-PCR, and immunoassays were additionally used to examine WNV infection in blood-fed mosquitoes, mosquito pools, dead birds, and mammals. Analyses of engorged Culex quinquefasciatus revealed that this mosquito species acquired 88% of the blood meals from avian and 11.6% from mammalian hosts, including humans. Similarly, Cx. tarsalis fed 82% on birds and 18% on mammals. Culex erythrothorax fed opportunistically on both birds (59%) and mammals (41%). In contrast, Cx. stigmatosoma acquired all blood meals from avian hosts. House finches and a few other mostly passeriform birds served as the main hosts for the blood-seeking mosquitoes. Evidence of WN virus infection was detected in mosquito pools, wild birds, dead birds, and mammals, including humans with cases of fatalities during the study. These findings emphasize the important role of house finches and several other passeriform birds in the maintenance and amplification of WNV in southern CA, with Cx. quinquefasciatus acting as both the principal enzootic and bridge vector responsible for the spillover of WNV to humans. Cx. tarsalis and Cx. stigmatosoma, are important but less widely distributed (CT) Approvals have been obtained, allowing for the open release of Aedes albopictus males in Kentucky. Initial field trials have tested mark-release-recapture methods for assessing Ae. albopictus male dispersal and survival following a point release. Methods developed in Project Year 1 were trialed, including classical fluorescent dyes and molecular methods. One of the molecular methods was observed to show good persistence in the field with little fitness impact on treated males. A deficiency observed is the current inability to capture Ae. albopictus males. Collections were typically female biased (e.g. 61%, 56%) and less than 1% of the released males were recaptured, which complicates interpretation of the results. In an effort to increase male recapture rates, future MRR experiments will include additional collection methods and tools. (KY) Research revolved around analysis of the tree-hole ecosystem as a model for studies on larval mosquito nutritional relationships and resultant production of adult mosquitoes, and in particular larval mosquito/microbial interactions. Larval mosquitoes were shown to be strong top-down predators on the microbial community in tree holes, such that a trophic cascade was evident in the protozoan and bacterial compartments of the community. The cascade was most intense on flagellate and ciliate populations and dampened at the bacterial trophic level. However, as larval mosquito feeding pressure was experimentally released and protozoan populations rebounded, predation pressure on bacteria increased due to protozoans, such that a group of predation-resistant bacteria emerged as elongated filamentous forms. In related research, stable isotope analysis showed that larval Ae. triseriatus mosquitoes utilized primarily plant based detritus early in the development season but that animal detritus became relatively more important later in the season. One key finding, through stable isotope analysis, was the important of summer inputs of tree flower parts into the tree hole system as a nutrient pulse. (MI) Aedes japonicus was collected during summer 2010 in eastern Nebraska as part of the Nebraska Department of Health and Human Services WNV surveillance program. Specimens were collected using CDC-light traps baited with carbon dioxide. Specimens were confirmed by Harry Savage (CDC/OID/NCEZID). The mosquito was collected in at least four independent trapping visits to separate sites. These are the first recorded collections of Ae. japonicus in Nebraska. The importance of Ae. japonicus to vector disease ecology and epidemiology in eastern Nebraska is not known. (NE) We found that low concentrations of malathion (0.11 ppm) that are often detected in aquatic systems affect competition between two invasive mosquito species Ae. albopictus and Ae. japonicus. There were no survivors of Ae. japonicus larvae in malathion. There was a significant negative effect of Ae. japonicus density on Ae. albopictus survival but this effect was absent in the presence of malathion. These findings indicate that pesticide mediated alterations in competition and species-specific differences in susceptibility to pesticides could play a role in enhancing invasive Ae. albopictus potential. (NJ) Accomplishments have been made toward strengthening our understanding of how pathogens, hosts and environmental factors influence mosquito-borne disease. Our laboratory has examined how invasive plants affect container breeding mosquitoes in Florida, and has extended this work to invasive plant dominated ecosystems in the southern Midwest. In collaboration with colleagues from the University of Florida, we have examined how abiotic factors influence the distribution of vectors of dengue fever virus and chikungunya virus, how chikungunya virus affects adult longevity in Ae. aegypti and Ae. albopictus, and how abiotic conditions in larval environments impact vector competence. In addition, we completed one study on how combinations of leaf species affect competition in Ae. aegypti and Ae. albopictus (OK). Objective 2. Use knowledge of mosquito, pathogen, vertebrate reservoir, and environment interactions to enhance ability to predict conditions leading to disease. The emergence of arthropod vectors of human disease, and the pathogens and pathogen systems associated with them, is of primary concern in our modern era of rapid population movement, trade, and environmental change. The risk of human infection with West Nile virus (WNV) in urban areas of the upper midwestern US remains one of the distinct study topics in this project. Models of the effects of environmental factors on WNV disease risk have yielded conflicting outcomes, in part because of the temporal and spatial scales of investigation. We used spatial and statistical modeling techniques to analyze and forecast fine scale spatial (2000 m grid) and temporal (weekly) patterns of WNV mosquito infection relative to changing weather conditions in the urban landscape of the greater Chicago, Illinois. Increased air temperature was the strongest temporal predictor of increased infection in Culex pipiens and Culex restuans mosquitoes, with differences in cumulative high temperature differences being a key factor distinguishing years with higher mosquito infection and higher human illness rate from those with lower rates. Spatially, precipitation was the most important variable predicting mosquito infection; precipitation and temperature alone could explain the pattern of spatial variability better than could other environmental variables. Other spatial environmental factors that tended to be important, such as impervious surfaces and elevation, mediate the effect of rainfall on soils and in urban catch basins. Evolutionary studies of WNV from 2005 suggest moderate evolutionary change but significant local scale processes. We analyzed viral nucleotide sequences from mosquitoes collected in 2005, 2006, and 2007 from a known transmission hot spot in suburban Chicago. Within this small area, the viral envelope gene has increased approximately 0.1% per year in nucleotide-level genetic diversity. In each year, viral diversity was higher in residential sites characterized by dense housing than in urban green space sites such as cemeteries and parks. Phylodynamic analyses show an increase in viral effective population size around 2005, consistent with a higher-than-average peak in mosquito and human infection rates that year. Analyses of times to most recent common ancestor suggest that WNV in 2005 and 2006 may have arisen predominantly from viruses present during 2004 and 2005, respectively, but that WNV in 2007 had an older common ancestor, perhaps indicating a predominantly mixed or exogenous origin. (MI) From the environmental models developed, we conclude that finely grained temporal and spatial patterns of precipitation and air temperature have consistent and significant impacts on the timing and location of increased mosquito infection with WNV. Changes in weather patterns with global climate change make it especially important to improve our ability to predict how inter-related local weather and environmental factors affect vectors and vector-borne disease risk. However, we now have a distinctive set of models that predict with a high degree of accuracy the variation in WNV infection in vector mosquito populations with solely two variables (temperature and precipitation), providing simplicity to a complex system. Interestingly, viral evolutionary analyses show that the population of WNV in suburban Chicago is an admixture of viruses that are both locally derived and introduced from elsewhere, containing evolutionary information aggregated across a breadth of spatial and temporal scales. Our results suggest that the population of WNV in suburban Chicago is broadly representative of that on coarser spatial scales, but also that it contains population genetic and phylogenetic signatures of local ecological and epidemiological processes, such as urban landscape type and climate. Moreover, WNV in suburban Chicago appears to show substantial temporal variability, both in its propensity to increase in diversity across years and in its variable evolutionary dynamics from year to year. WNV in our small hot spot study area thus appears to represent an admixture of viruses that are both locally derived and introduced from elsewhere and that reflect transmission dynamics aggregated across a breadth of spatial and temporal scales. We suggest that further studies of WNV on fine spatial and temporal scales in other regions might help clarify the importance of micro-scale processes to the transmission and evolution of this and other emerging arboviral pathogens. (MI) Related research examined the application of sequential sampling methodology to the process of seasonal amplification of WNV in Cx. pipiens vector populations. The goal is to develop a system of early warning as infection rate in the vector population rises, so as to anticipate the timing and intensity of human cases. This was accomplished by analyzing through time series cross correlation analysis the time-lagged relationships between vector infection rate, scored as the number of positive pools found per week of season, with the number of human cases scored by date of onset. Neither percentage of positive pools nor classic MIR were as useful as sheer number of positive pools per week as the predictor variable. Lags were in the range of 3-4 weeks, suggesting that the rise in vector infection rate in most summers precedes the rise of the human epidemic curve. From these relationships, sampling and outbreak threshold lines were developed, incorporating data on sampling characteristics of the mosquito population. Trends were examined at the local, regional (county), and state level using long term data from metropolitan Chicago and Illinois, our main study sites. Results showed that a sequential sampling system of mosquito infection, based on the number of positive pools found per week, is indeed useful as a tool for anticipating the beginning of the human epidemic curve, and it was most sensitive at the local vs. regional or state spatial scales. (MI) We have made a number of accomplishments in predicting and preventing disease transmission in Oklahoma through two projects. First, a project funded by the Oklahoma Center for the Advancement of Science and Technology (OCAST) has examined the landscape ecology of mosquito assemblages in the Cross Timbers region of Oklahoma. We have determined a high degree of habitat fidelity in mosquito assemblages, suggesting targeting certain areas for control would be more efficient and effective for certain diseases. Also funded through OCAST is a project on response to temperature of Ae. albopictus. This study has found significant differences in response to temperature depending upon nutrient level and population origin. In addition, we found an apparent change in the allometric relationship between measurements of mosquito total growth as a function of temperature induced changes. We also have a project examining the potential transmission of dog heartworm in urban versus rural areas in OK, data from which are currently being analyzed. (OK) Objective 3. Develop strategies to control mosquito vectors. An aposymbiotic laboratory strain of Ae. albopictus has been compared with a recently established, wild caught strain. No differences were observed in male performance, including competition for mates. Experimental values for percent hatching broods did not differ statistically with predicted values through three replications. In anticipation of Wolbachia-based biopesticide experiments, a closed greenhouse mesocosm system has been developed for maintaining and monitoring closed populations of Ae. albopictus. Replicate populations were maintained and monitored for 50 days, recording eclosion rates, sex ratio, egg production and estimating hatch rates. The results support downstream large-scale experiments assessing proposed control strategies. (KY) We evaluated pellet formulations of a monomolecular film (Agnique®) and (S)-methoprene (Altosid®) against Ae. albopictus larvae and pupae. In the lab, Agnique® provided 80% control for 20 d, whereas Altosid® in combination with Agnique® provided 80% control for > 60 d. During field trials, the 1:1 pellet ratio of products provided > 95% control for at least 32 d and 50% control for at least 50 d. Altosid® remained effective after a 107 d lab-induced drought, suggesting the product may serve as a possible means of control during drought conditions. Agnique® and Altosid®, when used in tandem for difficult-to-treat locations, can provide long-term control and reduce the number of treatments. (NJ) A broad based approach to new toxicant discovery includes screening of conventional chemical libraries and structure activity relationship analysis, evaluation of registered compounds for mosquitocidal activity and bioassays of compounds derived from natural products. In addition, we are exploring gene silencing using RNAi technology as a method to knock down critical proteins in mosquitoes with possible applications for control. A new research agreement with an industry partner has been initiated that will focus on identification of effective gene targets, production of large quantities of dsRNA and evaluation of delivery methods. (ARS-Gainesville, FL). Host odor analysis to discover new attractants and attraction-inhibitors was continued in 2010. Odors from horses were analyzed by gas chromatography/mass spectrometry and further evaluations of the identified compounds are in progress. Research progress was also made in the development of novel repellents and insecticides by structure-activity modeling of compounds. The data for model development are taken from the USDA data archive of more than 30,000 compound activities. A model of carboxamide repellents was developed and these have been evaluated against Ae. aegypti, An. albimanus, and An. gambiae. In collaboration with ARS-Oxford researchers, natural product repellents have been discovered and reported. The emphasis of the ARS-Gainesville modeling work has shifted towards development of novel insecticides. Several lead compounds have been found, but models to develop novel compounds have been unsatisfactory. In conjunction with the Natick Soldier Center, the bite protection perfomance of permethrin-treated US Army combat uniforms was evaluated. Results from these studies indicated that Fire-Resistant Army Combat Uniforms (FRACUs) treated with permethrin at the factory level were highly efficacious at preventing bites of Ae. aegypti and An. albimanus through 50 standardized wash cycles. The US Army has transitioned to a FRACU stock supply that is 100% factory-level permethrin treated as of September 1, 2010. The US Marine Corps used this same procedure to transition to factory treated combat uniforms in 2007. (ARS-Gainesville, FL) The role of prallethrin as a component of the formulated product, Duet", was evaluated in a wind tunnel against female Cx. quinquefasciatus. By using commercially formulated product without each of the active ingredients (prallethrin, sumethrin, piperonyl butoxide), the impact of prallethrin on resting mosquitoes was evaluated. Prallethrin enhanced flight during the spray period and resulted in a larger number of droplets detected on the treated mosquitoes with higher subsequent mortality. Similar results were obtained with sand flies (Lutzomyia shannoni). On-going studies are examining the effects of prallethrin exposure on activation and flight of unfed and blood-fed Ae. aegypti and Ae. albopictus. (ARS-Gainesville, FL) Towards development of a mosquito control strategy based on the nectar feeding behavior of mosquitoes, a series of effective oral toxicants with different modes of action were identified against three genera of mosquitoes in laboratory assays. Further examination on the role of sugar and amino acid ingestion by Culex has underscored the critical need for sugar-feeding by male and female mosquitoes. Flowering plants attractive to Culex spp. in north Florida have been identified and studies are on-going to identify attractant chemicals from these plants that could be used for surveillance or with an attract-and-kill approach. (ARS-Gainesville, FL) Semi-field and field studies were conducted on the use of spatial repellents/barriers to reduce host-vector contact. Barriers were constructed of camouflage material. These barrier materials were sprayed with lambda-cyhalothrin. Traps baited with carbon dioxide and 1-octen-3-ol served as surrogate hosts. The traps were placed in the center of the treated barrier plots (either 5 x 5 ). Spatial repellents tested included S.C Johnsons Clip-On, ThermaCell and Conceal (linalool). Preliminary results indicate some protection from mosquitoes by the physical barrier alone; the addition of a spatial repellent usually resulted in increased protection, but not always. The effectiveness of the spatial repellent varied with species. (ARS-Gainesville, FL) Objective 4. Enhance surveillance technologies for mosquitoes and mosquito-borne pathogens. Statewide trapping and testing for mosquito-borne arboviruses was conducted in CT from June through October at 91 fixed collection sites. A total of 115,725 mosquitoes (10,654 pools) representing 35 species were trapped and tested. 220 isolations of WNV were made from 9 mosquito species: Cx. pipiens = 170, Cx. restuans = 25, Cx. salinarius = 15, Ae. japonicus = 4, Ae. vexans = 2, Cs. melanura = 1, Oc. taeniorhynchus = 1, Oc. triseriatus = 1, Ur. sapphirina = 1. The first positive mosquitoes were collected on Jun. 14, and the last on Oct. 7. The majority of virus activity was detected in densely populated urban and suburban regions in southwestern and central CT. Eight human cases of WNV were locally acquired with no fatalities (age range 50 - 81 yrs., date of onset Jun. 10 Sept. 18). Four isolations of EEE were made from Cs. melanura at a single location (Aug. 20 Sept. 30). Other mosquito-borne viruses isolated included: Cache Valley = 6 isolates from 4 species (Sept. 1 28); Highlands J = 1 isolate from 1 species (Oct. 26); Jamestown Canyon = 22 isolates from 11 species (Jun. 3 Aug. 17) (CT). The Delaware Dept. of Natural Resources and Environmental Control (DNREC) asked us to address this question can NJ light trap mosquito collections be used for both population estimation and WNV detection purposes? The problem in past years has often been the overwhelming number of mosquitoes and also non-target species that need to be sorted with these trap collections. We selected 10 sites from which to keep DNREC-collected mosquitoes for an 8-week period from July 5 to September 3, 2010. Of the ten, only five had good numbers of Culex spp. mosquitoes. Because of the extreme drought, numbers were low throughout the period. However, we found one WNV-positive pool during the week of 15021 August at a site that has been endemic for WNV over the past five years. The MFIR was relatively high (5.94) compared to past years, most likely because of the drought and reductions in mosquito emergence during August. The problem is that although detections can be made, species identity is very hard to determine in these trap collections. (DE). In rural areas of Nebraska, dry ice availability can be a problem. Octenol has a stable shelf life (if sealed), does not need to be refrigerated and is commercially available. The Nebraska Department of Health and Human Services was interested in finding out if a widely-available octenol gel product (Nosquito octenol lure 24%) could be used in lieu of dry ice in CDC light traps. The study was conducted at ten different sites within Lancaster County, NE during the summer of 2010. At each site, an octenol-baited light trap, a dry ice-baited trap, and an unbaited trap (control) were used. Overall mosquito and Culex spp. capture using octenol was not statistically significant compared with the control. Culex spp. and total mosquito captures were significantly higher using dry ice compared to octenol and the control. These findings are in line with studies conducted in other regions of the US. (NE) The oviposition behavior of Ae. albopictus was examined to help engineer a gravid trap specifcally for this important vector. This OCAST funded project has shown that Ae. albopictus prefer containers with small surface area but greater depth, over containers of the same volume but larger. This experimental work will continue this summer with a focus on oviposition media, and the results will be applied to field collections at the end of the summer. (OK) Lactic acid, as a component of human skin and an effective attractant for Ae. aegypti in the presence of carbon dioxide, was examined as an attractant for Culex. Olfactometer studies indicated that it was attractive for Cx. quinquefasciatus (although less so than for Ae. aegypti) but decreased attraction of both Cx. tarsalis and Cx. nigripalpus. A blend of lactic acid, acetone and dimethyl disulfide was effective to Ae. aegypti but less so for Culex. This research is continuing with field evaluations and additional host odors. (ARS-Gainesville, FL) Objective 5. Develop strategies for sustainable mosquito control by including training at all levels. No specific reports submitted; however, training of students and postdocs is an integral component of nearly all of the above reports.

Publications

Allan SA. Susceptibility of adult mosquitoes to insecticides in aqueous sucrose baits. J Vector Ecol (in press). Allan SA, Bernier UR, Kline DL. 2010. Laboratory evaluation of human-associated odors on attraction of Culex spp. (Diptera: Culicidae). J Vector Ecol 35:318-324. Amin E, El-Hawary SS, Fathy MM, Mohammed R, Ali Z, Tabanca N, Wedge DE, Becnel JJ, Khan IA. 2010. Triterpenoidal saponins: Bioactive secondary metabolites from Zygophyllum coccineum. Planta Medica DOI: 10.1055/s-0030-1250463. Amore G, Bertolotti L, Hamer GL, Kitron UD, Walker ED, Ruiz MO, Brawn JD, Goldberg TL. 2010. Multi-year evolutionary dynamics of West Nile virus in suburban Chicago, USA, 2005-2007. Phil Trans Roy Soc B 365:1871-1878. PMID: 20478882. Anderson JF, Ferrandino FJ, Dingman DW, Main AJ, Andreadis TG, Becnel JJ. Control of West Nile virus vectors in catch basins in Connecticut with Bacillus thuringiensis israelensis, Bacillus sphearicus and spinosid. J Am Mosq Control Assoc (in press). Andreadis TG., Armstrong PA, Bajwa WJ. 2010. Studies on hibernating populations of Culex pipiens (Diptera: Culicidae) from a West Nile virus endemic focus in New York City: parity rates and isolation of West Nile virus. J Am Mosq Control Assoc 26:257-264. Andreadis TG, Wolfe RJ. 2010. Evidence for reduction of native mosquitoes with increased expansion of the invasive Ochlerotatus japonicus japonicus (Diptera: Culicidae) in the northeastern United States. J Med Entomol 47:43-52. Armstrong PA, Andreadis TG. 2010. Eastern equine encephalitis virus in mosquitoes and their role as bridge vectors. Emerging Inf Dis 16:1869-1874. Barnard DR, Bernier, UR, Clark GG, Nathan M, Zaim M. 2009. Guidelines for efficacy testing of mosquito repellents for human skin. WHO Pesticide Evaluation Scheme. 30pp. Brey C, Farajollahi A, Gaugler R, Evans H, Kesavaraju B. 2011. Effect of malathion on larval competition between Aedes albopictus and Aedes atropalpus (Diptera: Culicidae). J Med Entomol (in press). Britch SC, Linthicum KJ, Wynn WW, Walker TW, Farooq M, Smith VL, Robinson CA., Lothrop BB, Snelling M, Gutierrez A, Lothrop HD, Kerce JD, Becnel JJ, Bernier UR, Pridgeon JW. 2010. Evaluation of ULV and thermal fog mosquito control applications in temperate and desert environments. J Am Mosq Control Assoc. 26:183-97. Calvitt M, Moretti R, Lampazzi RE, Bellini R, Dobson SL. 2010. Characterization of a new Aedes albopictus (Diptera: Culicidae): Wolbachia pipientis (Rickettsiales: Rickettsiaceae) symbiotic association generated by artificial transfer of the wPip strain from Culex pipiens (Diptera: Culicidae). J Med Entomol 47:179-187. Cantrell CL, Pridgeon JW, Fronczek FR, Becnel JJ. 2010. Structure-activity relationship studies on derivatives of eudesmanolides from Inula helenium as toxicants against Aedes aegypti larvae and adults. Chem Biodivers. 7:1681-97. Chaves LF, Harrington LC, Keogh CL, Nguyen AM, Kitron UD. 2010. Blood feeding patterns of mosquitoes: random or reflective of preferences? Frontiers in Zool 7:3. Highly Accessed Article http://www.frontiersinzoology.com/content/7/1/3/abstract Cheng GJ, Cox J, Wang P, Krishnan MN, Dai J, Quan F, Anderson JF, Fikrig E. 2010. A C-type lectin collaborates with a CD45 phosphatase homolog to facilitate West Nile virus infection of mosquitoes. Cell 142:714-725. Cooperband MF, Ellison FV, Clark GG, Jany W, Allan SA. 2010. Prallethrin-induced excitation increases contact between sprayed ultra-low-volume droplets and flying mosquitoes (Diptera: Culicidae) in a wind tunnel. Med Entomol 47:1099-1106. Diuk-Wasser MA., Molaei G, Simpson JE, Folsom-OKeefe, CM, Armstrong PM, Andreadis TG. 2010. Avian communal roosts as amplification foci for West Nile virus in urban areas in northeastern United States. Am J Trop Med Hyg 82:337-343. Fu Y, L. Gavotte L, Mercer DR, Dobson SL. 2010. Artificial triple Wolbachia Infection in Aedes albopictus yields a new pattern of unidirectional cytoplasmic incompatibility. Appl Environ Microbiol 76:5887-5891. Gavotte LD, Mercer R, Stoeckle JJ, Dobson SL. 2010. Costs and benefits of Wolbachia infection in immature Aedes albopictus depend upon sex and competition level. J Invert Pathol 105:341-346. Gingrich JB, OConnor L-L, Meredith WH, Pesek JD, Shriver WG. 2010. Epidemiology of West Nile virus: a silent epiornithic in northern Delaware in 2007 without associated human cases. J Am Mosq Control Assoc 26: 274-286. Gong H, DeGaetano AT, Harrington LC. 2010. Climate-based models for West Nile Culex mosquito vectors in the northeastern USA. Int J Biometeorology DOI 10.1007/s00484-010-0354-9. Hardstone MC, Huang X, Harrington LC, Scott JG. 2010. Differences in development, glycogen, and lipid content associated with cytochrome P450-mediated permethrin resistance in Culex pipiens quinquefasciatus (Diptera: Culicidae). J Med Entomol 47:188-198. Harvey WR, Okech BA, Linser PJ, Becnel JJ, Ahearn GA, Sterling KM. 2010. H(+) V-ATPase-energized transporters in brush border membrane vesicles from whole larvae of Aedes aegypti. J Insect Physiol. 56:1377-89. Hoel DF, Kline DL, Hogsette JA, Bernier UR, El-Hossary SS, Hanafi HA, Watany N, Fawaz EY, Furman BD, Obenauer PJ, Szumlas DE. 2010. Efficacy of commercial mosquito traps in capturing phlebotomine sand flies (Diptera: Psychodidae) in Egypt. J Med Entomol 47:1179-1184. Hoel D, Pridgeon JW, Bernier UR, Chauhan K, Meepagala K, Cantrell C. 2010. Departments of Defense and Agriculture team up to develop new insecticides for mosquito control. Wingbeats 21: 29-34. Katritzky AR, Wang Z, Slavov S, Dobchev DA., Hall CD, Tsikolia M, Bernier UR, Elejalde NM, Clark GG, Linthicum KJ. 2010. Novel carboxamides as potential mosquito repellents. J Med Entomol 47:924-938. Kaufman MG, Pelz-Stelinski K, Yee D, Juliano S, Ostrom M, Walker ED. 2010. Stable isotope analysis reveals detrital resource base sources of the tree hole mosquito, Aedes triseriatus. Ecol Entomol 35: 586593. Kesavaraju B, Afify A, Gaugler R. 2010. Growth and survival of the invasive Aedes albopictus larvae on Diospyrus virginiana (American persimmon) leaves. J Med Entomol (in press). Kesavaraju B, Afify A, Alto B, Gaugler R. 2010. Malathion influences competition between Aedes albopictus and Aedes japonicus. J Med Entomol (in press). Kimura M, Darbro JM, Harrington LC. 2010. Avian malaria parasites share congeneric mosquito vectors. J Parasitol 96:144-151. Larson RT, Lorch JM, Pridgeon JW, Becnel JJ, Clark GG, Lan Q. 2010. The biological activity of alpha-mangostin, a larvicidal botanic mosquito sterol carrier protein-2 inhibitor. J Med Entomol. 47:249-57. Logan JG, Cook JI, Mordue (Luntz) AJ, Kline DL. 2010. Understanding and exploiting olfaction for the surveillance and control of Culicoides biting midges. Chapter 10, pp. 217-246, In: Takken, W. & Knols, B.G.J. (eds), Ecology and control of vector-borne diseases, Vol. 2. Olfaction in vector-host interactions, Wagengen Academic Publishers. Lounibos LP, O'Meara GF, Juliano SA, Nishimura N, Escher RL, Reiskind MH, Cutwa M, Greene K. 2010. Differential survivorship of invasive mosquito species in South Florida cemeteries: do site-specific microclimates explain patterns of coexistence and exclusion? Ann Entomol Soc Amer 103:757-770. Molaei G, Cummings RF, Su T, Armstrong PM, Williams GA, Cheng ML, Webb JP, Andreadis TG. 2010. Vector-host interactions governing epidemiology of West Nile virus in southern California. Am J Trop Med Hyg 83:1269-1282. Müller GC, Junnila A, Qualls W, Kline DL, Allan S, Schlein Y, Xue R. 2010. Control of Culex quinquefasciatus in a storm drain system in Florida with attractive toxic sugar baits. Med Vet Entomol 24:346-351. Nelder M, Kesavaraju B, Farajollahi A, Healy S, Unlu I, Crepeau T, Fonseca D, Gaugler R. 2010. Suppressing Aedes albopictus, an emerging vector of dengue and chikungunya viruses, by a novel combination of a monomolecular film and insect-growth regulator. Amer J Trop Med & Hyg 82:831-7. Obenauer PJ, Allan SA, Kaufman PE. 2010. Aedes albopictus (Diptera: Culicidae) oviposition responses to organic infusions from common flora of suburban Florida. J Vector Ecol 35:1-6. Obenauer PJ, Kaufman PE, Kline DL, Allan SA. 2010. Detection of and monitoring for Aedes albopictus (Diptera: Culicidae) in suburban and sylvatic habitats in North Central Florida using four sampling techniques. Environ Entomol 39:1608-1616. Pelz-Stelinski K, Walker ED, Kaufman MG. 2010. Senescent leaf exudates increase mosquito survival and microbial activity. Ecol Entomol 35: 329-340. Price D, Gunther D, Gaugler R. 2010 . First collection records of Phlebotomine sand flies from New Jersey. J Med Entomol (in press). Pridgeon JW, Becnel JJ, Bernier UR, Clark GG, Linthicum KJ. 2010. Structure-activity relationships of 33 carboxamides as toxicants against female Aedes aegypti (Diptera: Culicidae). J Med Entomol. 47:172-8. Reiskind MH, Westbrook CJ, Lounibos LP. 2010. Exposure to chikungunya virus and adult longevity in Aedes aegypti (L.) and Aedes albopictus (Skuse). J Vector Ecol 35:61-68. Reiskind MH, Zarrabi AA. 2011. The Importance of an Invasive Tree Fruit as a Resource for Mosquito Larvae. J Vector Ecol (In Press). Reiskind MH, Zarrabi AA, Lounibos LP. 2010. Invasive leaf resources alleviate density dependence in the invasive mosquito, Aedes albopictus. Biol Invasions 12:2319-2328. Ruiz MO, Chaves LF, Hamer GL, Sun T, Brown WM, Walker ED, Haramis L, Goldberg TL, Kitron UD. 2010. Local impact of temperature and precipitation on West Nile virus infection in Culex species mosquitoes in northeast Illinois, U.S.A. Parasites & Vectors 3:19. PMID: 20302617. Tabanca N, Bernier UR, Tsikolia M, Becnel JJ, Sampson B, Werle C, Demrici B, Can Baser HC, Blythe EK, Pounders CT, Wedge DE. 2010. Eupatorium capillifolium essential oil: Chemical composition, antifungal activity, and insecticidal activity. NAt Prod Comm 5:1409-1415. Unlu, I, Farajollahi A, Healy S, Crepeau T, Bartlett-Healy K, Williges E, Strickman D, Gaugler R, Fonseca D. 2011. Area-wide management of the Asian tiger mosquito: establishment of study sites using geospatial tools, socio-economic factors and BG-Sentinel traps. Pest Mgt Sci (in press). Vossbrinck CR, Baker MD, Andreadis TG. 2010. Phylogenetic position of Octosporea muscaedomesticae (Microsporidia) and its relationship to Octosporea bayeri based on small subunit rDNA analysis. J Invertebr Pathol 105:366-370. Vrzal EM, Allan SA, Hahn DA. 2010. Effect of nutritional status and presence of amino acids in a sugar mixture on longevity of Culex quinquefasciatus. J. Insect Physiol 56:1659-1664. Walker E, Kaufman M, Merritt R. 2010. An acute trophic cascade among microorganisms in the tree hole ecosystem following removal of omnivorous mosquito larvae. Comm Ecol 11:171-178. Westbrook CJ, Reiskind MH, Pesko KN, Greene KE, Lounibos LP. 2010. Larval environmental temperature and the susceptibility of Aedes albopictus Skuse (Diptera: Culicidae) to chikungunya Virus. Vector-Borne Zoonotic Dis 10:241-247. Wiwatanaratanabutr I, Allan SA, Linthicum K. Kittayapong P. 2010. Strain-specific differences in mating, oviposition, and host-seeking behavior of Wolbachia-infected and uninfected Aedes albopictus. J Am Mosq Control Assoc. 26:265-273. Xue,R-D, Qualls WA, Kline DL, Zhao T-Y. 2010. Evaluation of Lurex", octenol and CO2 sachet as baits in mosquito pro traps against mosquitoes. J Am Mosq Control Assoc 26:344-345. Zhao L, Becnel JJ, Clark GG, Linthicum KJ, Chen J, Jin X. 2010. Identification and expression profile of multiple genes in response to magnesium exposure in Culex quinquefasciatus larvae. J. Med. Entomol 47: 1053-1061. Zhao L., Becnel JJ, Clark GG, Linthicum KJ. 2010. Expression of AeaHsp26 and AeaHsp83 in Aedes aegypti (Diptera: Culicidae) larvae and pupae in response to heat shock stress. J Med Entomol 47:367-75.

Impact Statements

Work carried out in 2010 as part of the Northeast Regional Research Project, NE-1043, provided valuable theoretical and practical knowledge to help manage mosquito-borne disease.
Research on new chemical insecticides is moving novel technologies from the bench to operational use.
Statewide mosquito and arbovirus surveillance programs provide an early warning system for intervention activities by local mosquito control agencies.
The feasibility of initiating large field cage experiments on the sterile insect technique was demonstrated.
A risk model was developed and made available via website to end-users (local mosquito control agencies) for predicting eastern equine encephalitis and West Nile virus.
The groundwork was prepared for analysis of proteins to target for disrupting mosquito reproduction. Novel molecular methods are being deployed to identify gene function in proteins with potential for mosquito control.
Research led to an enhanced understanding of mosquito biology and host-mosquito interactions across an array of ecosystems. Unraveling these ecological relationships is essential to developing economical and effective integrated pest management programs for mosquitoes.
Genetic engineering research is resulting in new bacterial toxins with enhanced virulence against mosquito larvae with potential for commercial development as new, low risk biological insecticides.
Provided for and encouraged environmentally sound, scientifically based, and professional control by local mosquito control agencies.

Date of Annual Report: 03/20/2012

Report Information

Annual Meeting Dates: 03/13/2012 - 03/13/2012
Period the Report Covers: 10/01/2010 - 09/01/2011

Participants

Anderson, John (john.f.anderson@ct.gov) Connecticut, Agricultural Experiment Station;
Andreadis, Theodore (theodore.andreadis@ct.gov) Connecticut, Agricultural Experiment Station;
Armstrong, Philip (philip.armstrong@ct,gov) Connecticut, Agricultural Experiment Station;
Fallon, Ann (fallo002@umn.edu) Minnesota, University of Minnesota;
Federici, Brian (brian.federici@ucr.edu) California, University of California-Riverside;
Harrington, Laura (lch27@cornell.edu) New York, Cornell University;
Daniel, Kline (dan.kline@ars.usda.gov) USDA, ARS, Gainesville, Florida;
Leisnham, Paul (leisnham@umd.edu) Maryland, University of Maryland;
Marcombe, Sebastien (sebastien.marcombe@rutgers.edu) New Jersey, Rutgers University;
Molaei, Goudarz (goudarz.molaei@ct.gov) Connecticut, Agricultural Experiment Station;
Robinson, Mark (mrobinson@nifa.usda.gov) NIFA, USDA;
Shepard, John (john.shepard@ct.gov) Connecticut, Agricultural Experiment Station;
Stafford, Kirby (kirby.stafford@ct.gov) Connecticut, Agricultural Experiment Station;
Thomas, Michael (michael.c.thomas@ct.gov) Connecticut, Agricultural Experiment Station;
Vossbrinck, Charles (charles.vossbrinck@ct.gov)Connecticut, Agricultural Experiment Station;

Participants Attachment:

Attachment

Brief Summary of Minutes

The annual meeting of the Multi-State Project NE1043, Biology, Ecology and Management of Emerging Disease Vectors was held on March 13, 2012 at The Connecticut Agriculture Experiment Station, New Haven, CT.

The following were in attendance: Federici (CA), Andreadis (CT), Anderson (CT), Armstrong (CT), Molaei (CT), Shepard (CT), Thomas (CT), Vossbrinck (CT); Leisnham (MD), Fallon (MN), Marcombe (NJ), Harrington (NY), Kline (ARS-Gainesville), Stafford (Administrative Advisor-CT), and Robinson (USDA-NIFA).

Dr. Theodore Andreadis (Chair) reviewed the agenda, local arrangements and format for the meeting. He circulated the Annual Report and noted that reports were submitted from 9 participating agencies (CT, MD, MN, NJ, NY, OK, WI, USDA-Gainesville). No reports were received or participation at the annual meeting from DL, KY, MA, NE, or TX. He indicated that there are currently 26 official members of the project representing 13 Experiment Stations/Universities and 2 USDA laboratories, the Medical and Veterinary Entomology Laboratory (Gainesville, FL) and the Grain Marketing and Production Research Center (Manhattan, KS). Notification of the meeting was sent to individuals within the CDC and US Military (Armed Forces Pest Management Board and Navy Marine Corps Public Health Center Detachment) but no representatives were able to attend. He announced that his 2yr-term as chair would terminate with this meeting and that elections for a new slate of officers (chair, vice-chair, and secretary) would be held at the final business meeting.

Dr. Mark Robinson (USDA-NIFA representative) circulated his report and reviewed the FY2012 NIFA budget and AFRI research priority areas for 2012. He announced that the RFAs would be out this spring and would notify the chair of funding opportunities that pertain to members of the project. He noted that members could receive electronic notification of funding opportunities by registering at http://www.nifa.usda.gov/funding/notification.html. There was concern expressed by members that the current research priorities established by AFRI do not reflect the research interests and objectives of the project.

Dr. Kirby Stafford (Administrative Advisor) presented his administrative report and indicated that the current project will terminate on September 30, 2014. He presented a timetable for preparing a new project. New proposals require two years for approval. A request to re-write should be submitted in March 2013 and the reviewed proposal in March 2014.

A presentation was given by Dr. Philip Armstrong, Center for Vector Biology & Zoonotic Diseases, CAES entitled: Molecular Evolution of West Nile Virus in a Northern Temperate Region: Connecticut 1999-2008.

Brief research updates were given by each cooperator with active discussion ensuing. A group photo was taken.

Several in-depth presentations were made providing stimulus for further discussion and collaboration. 1.) Dr. Brian Federici (CA): current research on recombinant bacteria, Bacillus thuringiensis isrealensis Bacillus sphearicus, and a new research iniative to examine the genomics of fungal Coelomoyces parasites of mosquitoes; 2.) Dr. John Anderson (CT): research on the efficacy of a novel chemical attractant lure for Ochlerotatus japonicus, and vector competence evaluating the comparative efficiency of West Nile virus genotypes NY99 and WN02 transmission by Culex tarsalis and Culex salinarius 3.) Dr. Theodore Andreadis (CT): research on a novel microsporidian parasite imported from Japan on the invasive mosquito Ochlerotatus j. japonicus.

Nominations for new officers were solicited at the final business meeting. It was unanimously decided that the current Vice-Chair, Dr. Edward Walker (MI) would assume the Chair position. Dr. Daniel Kline (USDA-Gainesville) was elected Vice-Chair. It was suggested that next years meeting could be held in conjunction with the Annual Meeting of the American Mosquito Control Association that will be held February 24-28, 2013 in Atlantic City, NJ. However, it was decided the date and location of next years annual meeting would be at the discretion of the newly elected Chair who is responsible for coordinating such.

Accomplishments

Objective 1. Strengthen basic and applied research on the mosquito, pathogen, hosts, and environmental factors that influence disease emergence. To determine whether West Nile virus (WNV) overwinters locally or is reseeded annually, we examined the patterns of viral lineage persistence and replacement in CT over 10 consecutive transmission seasons by phylogenetic analysis. We compared the full protein coding sequence among WNV isolates to search for evidence of convergent and adaptive evolution. Viruses sampled from CT segregated into a number of well-supported subclades by year of isolation with few clades persisting e2 years. Similar viral strains were dispersed in different locations across the state and divergent strains appeared within a single location during a single transmission season, implying widespread movement and rapid colonization of virus. Numerous amino acid substitutions arose in the population but only one change, VàA at position 159 of the envelope protein, became permanently fixed. Several instances of parallel evolution were identified in independent lineages, including one amino acid change in the NS4A protein that appears to be positively selected. Results suggest that annual reemergence of WNV is driven by both reintroduction and local-overwintering of virus. Despite ongoing diversification of WNV, most amino acid variants occurred at low frequencies and were transient in the virus population (CT). Cx. tarsalis is a superior horizontal and vertical vector of WNV compared with Cx. salinarius. Cx. salinarius transmitted WNV genotype NY99 (CT 2741-99 strain) horizontally to suckling mice at significantly lower rates than Cx. tarsalis and Cx. salinarius transmitted WNV genotype NY99 to offspring at a lower vertical transmission infection rate than Cx. tarsalis. Cx. tarsalis transmitted WNV genotypes NY99 and WN02 (CT S0084-08 strain) with equal efficiency. Daily percent horizontal transmission of genotype NY99 by Cx. tarsalis -infected per os and by intra-thoracic infection was not significantly different from daily transmission of genotype WN02. Findings do not support the previously published hypothesis that genotype NY99 was replaced in the New World by WN02 because of a shorter extrinsic incubation of WN02 (CT). A survey of mosquito larvae infected with microsporidia was conducted from 2005-08 in the Tomsk, Kemerovo and Novosibirsk regions of western Siberia, Russia. Twenty-one morphologically and genetically unique species of microsporidia were isolated from 9 species of mosquitoes including 14 new species of Amblyospora, a new genus and species, Novothelohania ovalae and 6 species of Amblyospora, Parathelohania and Trichoctosporea from which gene sequences had not been previously obtained. Detailed ultrastructure of meiospores revealed unique cytological features associated with the length, arrangement and ratio of broad to narrow coils of the polar filament, comparative thickness of the exospore and endospore, and overall size of each species reaffirming their value in distinguishing taxonomic relationships. SSU rDNA sequences were unique when compared with GenBank entries. Phylogenetic trees yielded similar topologies with a high degree of congruence between parasite and host at the generic level. Species that parasitize Aedes/Ochlerotatus and Culex mosquitoes segregate into distinct monophyletic groupings mirroring their host phylogeny, while species from Anopheles mosquitoes group as a sister clade basal to the entire group of mosquito-parasitic microsporidia as their Anopheles hosts cluster as a sister clade to the entire group of culicine mosquitoes. This provides strong evidence for host-parasite coevolution by descent at the generic level and limited host lineage switching between unrelated taxa. Among parasites of Aedes/Ochlerotatus and Anopheles mosquitoes, we found several instances where a single mosquito species serves as a host for two or more related species of microsporidia, an observation consistent with host switching and independent parasite speciation. Among the microsporidian parasites of Culex mosquitoes, we found only one parasite per host indicating a higher degree of host specificity and less host switching among parasites of this genus. Findings suggest a degree of host-parasite co-speciation with host switching occurring occasionally when the normal host is unavailable in the aquatic ecosystem. Frequency of host switching seems to be occurring in proportion to host relatedness and does not cross generic boundaries in this system (CT). The spread of exotic mosquito species into new environments can introduce shifts in mosquito populations and potentially alter public health risks to mosquito-borne diseases. The successful establishment of exotic species may occur due to their competitive advantage over other cohabitating species. We hypothesized that the recently introduced exotic mosquito Ae. japonicus would be a more effective competitor than Ae. atropalpus and Ae. triseriatus, and an equal competitor to Cx. pipiens based on larval abundance data within tire habitats. Impacts of competition were measured using the larval developmental rate and survival of larvae, adult mortality, wing length, and sex ratio. We found that intraspecific competition acted strongest against Ae. japonicus versus the other three resident mosquito species by delaying larval development and increasing adult mortality. Interspecific competition was generally weak and significant main effects were only detected for species and density. Results show that larval competition between Ae. japonicus and the three resident species was weak when present, indicating that other ecological or behavioral factors may be influencing the invasion success for Ae. japonicus in North America (CT). Life-table experiments were conducted to test for differences in the adult survival and reproductive schedules of Ae. albopictus females from three populations from the northern and southern extremes of the species distribution in North America. There were consistent differences between northern and southern populations in incidence of photoperiodically-induced egg diapause. Under short daylength, diapause eggs constituted twice the proportion of total viable eggs from northern females than southern females. There were no consistent differences between northern and southern populations in resource allocation between reproduction and maintenance, reproduction over time, and reproductive investment among offspring, and no apparent trade-offs between diapause incidence with reproduction or longevity. Results suggest that the main response of North American Ae. albopictus to unfavorable winter climates is via the life history strategy of producing diapausing eggs, rather than quantitative variation in reproduction, and that there are no detectable costs to adult survival (MD). A study was conducted to characterize mosquito communities among different habitats in historically ditched tidal salt marshes and adjacent wooded areas in a wetland management area on the Maryland Delmarva Peninsula. Wooded habitats had more total mosquitoes, were more frequently occupied by mosquitoes, and had higher densities of mosquitoes. The majority of larvae at the control site were Ae. sollicitans in marsh pannes while Cx. salinarius, An. bradleyi, Ae. cantator, and Ae. sollicitans were collected in high numbers from ditches at the experimental site. We found a difference in the proportion of marsh pannes occupied by Ae. sollicitans but not total mosquitoes sampled 45 days after spring tide events than on other occasions (MD). Carbon-based secondary or structural plant compounds usually increase when they are exposed to elevated atmospheric CO2, and may differentially affect container mosquito species that feed on litter that fall into water-filled containers. Competition between Ae. albopictus and Ae. triseriatus was tested in laboratory microcosms provisioned with litter grown in either elevated (1,000 ppm) or ambient CO2 (385 ppm). In additional microcosms, single-species cohorts of either Ae. albopictus or Ae. triseriatus were provided with increasing amounts of one of the two litter types or concentrations of commercial tannic acid to test potential toxicity effects of leaf-derived tannins. We found no differences in mosquito competition between litter types, with Ae. albopictus consistently superior to Ae. triseriatus. Single-species microcosms provisioned with elevated leaf litter had slower male development time than microcosms with ambient (lower-tannin) litter. Ae. albopictus and Ae. triseriatus also experienced similarly higher mortality and slower development with increasing commercial tannic acid concentrations. These results suggest that possible litter grown in high CO2 conditions may suppress mosquito production by increasing the toxicity of leaf litter, but that it would not affect population-level competition between Ae. albopictus and Ae. triseriatus (MD). We conducted yard surveys for mosquito larval habitats paired with knowledge, attitude, and practice questionnaires administered to residents among six neighborhoods in Washington DC that vary in socioeconomic status. Household income was related to overall knowledge and attitudes of residents towards mosquitoes, but that knowledge and attitudes were not related with source reduction as measured through questionnaire responses or actual numbers of water-holding containers in their yard. Numbers of water-holding containers was strongly positively related to exposure to Ae. albopictus, Cx. pipiens and pupae but not related to whether or not households practiced source reduction. Households that reported practicing source reduction had lower Cx. pipiens infestation and abundances but no difference in Ae. albopictus or pupal infestation or abundance. Results suggest large households are a greater source of vector mosquitoes and should be targeted in education campaigns, and that self-perceived source reduction of residents may not reduce urban mosquito habitat (MD). The risk presented to humans by circulation of WNV in the Culex bird enzootic cycle established in metropolitan areas of the upper Midwest was continued. Culex Flavivirus was positively associated with WNV infection in a case-control study of mosquito pools collected from an endemic focus of WNV transmission in Chicago. This study provides direct evidence that insect-specific flaviviruses may influence human health risk by enhancing WNV infection in mosquitoes that are epidemic bridge vectors to human hosts (MI). Analysis of molecular genetic diversity in 9 marker regions of 5 genes within the bacteriophage WO region of the Wolbachia pipentis genome revealed high diversity of wPip in the population of Cx. pipiens sampled in metropolitan Chicago, IL. Wolbachia endosymbionts may modulate propensity to infection with this virus and may affect population structure of their mosquito hosts. Multiple regression showed that markers associated with gene Gp2d were significantly associated with ancestry of individuals to form molestus or form pipiens, as determined by prior microsatellite allele frequency analysis. Data suggest that certain wPip molecular genetic types are associated with genetic substructuring in the Cx. pipiens complex of Chicago, and that the association extends to host preference. The influence of host diversity on multi-host pathogen persistence and transmission is confounded by the large number of species and complex biological interactions that comprise the transmission system. Although diversity and composition of host communities have been hypothesized to affect intensity of transmission, the effect of host community structure on the transmission of vector-borne pathogens remains largely unknown. Results suggest that host community measures, such as species diversity, may not be useful indicators of transmission risk at fine spatial scales in vector-borne disease systems (MI). Culex mosquitoes carry an intracellular symbiont, W. pipientis, a bacterium which can potentially be manipulated to control vector populations by spreading a reproductive distortion called cytoplasmic incompatibility. We obtained a Cx. pipiens colony from Ohio State University and, for comparative purposes, have developed a sister colony cured of Wolbachia infection. By comparing protein bands from dissected testes, we identified proteins produced in response to Wolbachia infection. Efforts are underway to clone two Wolbachia DNA binding proteins that are implicated in the reproductive distortion known as cytoplasmic incompatibility. Parallel investigations with Wolbachia-infected cell lines are used to identify Wolbachia-inducible protein expression in undifferentiated cells that do not engage in gamete production. We are investigating methods to modulate expression of these proteins by testing effects of specific components, including the DNA synthesis inhibitor mimosine and the mitochondrial inhibitor, paraquat, in the culture medium. We are finalizing methodology for quantifying Wolbachia infections using fluorescence activated cell sorting (MN). Ae. japonicus, which was first collected during summer 2010, was again collected during summer 2011 in eastern NE. Specimens were collected using CDC-light traps baited with carbon dioxide. Mosquitoes were collected in four sites within Lincoln, NE. The importance of Ae. japonicus to vector disease ecology and epidemiology in eastern NE is not known. (NE). Using a panel of seven microsatellite loci, we confirmed the existence of two abundant genetic forms in specimens originally collected in 19992000 that matches the disjunctive distribution of mitochondrial haplotypes. To examine the distribution of the two genetic types across PA we created a fine scale genetic map of Ae. japonicus using specimens collected from 54 Pennsylvania counties in 20022003. We also made direct comparisons between collections in 19992000 and new collections made in 20042005 obtained from the same areas in the northeastern US. We observed that the strong association between mtDNA haplotype and microsatellite signature seen in 19992000 had weakened significantly by 2002 across PA, a trend continued to some extent in 20042005 in PA, NJ, and NY, indicating that once easily distinguishable separate introductions are merging. The two expanding genetic forms create a complex correlation between spatial and genetic distances. The existence of multiple introductions would be obscured without sampling early and across time with highly polymorphic molecular markers. Results provide a high resolution analysis of the spatial and temporal dynamics of a newly introduced disease vector and argue that successive introductions may be a common pattern for invasive mosquitoes (NJ) We documented the potential importance of eastern red cedar, Juniperus virginiana leaves and fruit as a larval resource for the invasive Ae. albopictus in OK. We examined how different leaf species commonly found in FL interact to affect competition within and between Ae. albopictus and Aedes aegypti. We have also continued investigations into the effects of temperature and food on growth in larval mosquitoes (OK). Laboratory and field tests of potential biological control agents were conducted for control of Cx. pipiens. The key goals of this project are to test locally adapted fathead minnows and copepods for control of Cx. pipiens/restuans. Fish are used in aboveground breeding sites while copepods are used in the underground catch basin system. Fish were not released in 2011 because of extended dry conditions which limited the extent of targeted wet sites and the occurrence of larvae in those sites. Belowground sites did begin to generate toward the end of the season and these were used for trials of copepod releases. We tested the efficacy of copepods as predators of Culex or Anopheles larvae. Acanthocyclops trajani and Macrocyclops albidus were tested against An. stephensi and Cx. pipiens. M. albidus females killed 50% of Culex larvae within 24 hr while A. trajani killed only 35% of larvae. Results for An. stephensi were similar. We tested whether releases of M. albidus could affect larval Culex in catch basins (WI). We released M. albidus, into storm drain catch basins in Madison, WI at the beginning of August. We monitored these sites along with untreated control sites until the end of September. The catch basins were sampled every week using a mosquito dipper, and immature mosquitoes were collected, counted, and reared to adulthood for identification of the species. All adult mosquitoes collected were identified as Cx. pipiens. Copepods were collected during the sampling and identified as M. albidus or A. vernalis or inconsequential species. Catch basins were infested with mosquito larvae. M. albidus failed to establish in all but one location. This single location, however, maintained and multiplied the number of M. albidus with many being collected every week. M. albidus colonized an adjacent catch basin after approximately 1 month and their numbers continued to increase throughout the survey. Citywide water main flushing after our introductions took place may have resulted in the flushing of M. albidus from the test sites. Heavy rainfall events may have also flushed these copepods from their new habitats on more than one occasion. The A. vernalis colony was created from the catch basin samples and these copepods were frequently found in our sites. They appear better adapted to the catch basin environment than M. albidus (WI). Objective 2. Use knowledge of mosquito, pathogen, vertebrate reservoir, and environment interactions to enhance ability to predict conditions leading to disease. Seasonal epizootics of vector-borne pathogens infecting multiple species are ecologically complex and difficult to forecast. Pathogen transmission potential within the host community is determined by the relative abilities of host species to maintain and transmit the pathogen and by ecological factors influencing contact rates between hosts and vectors. Increasing evidence of strong feeding preferences by a number of vectors suggests that the host community experienced by the pathogen may be very different from the local host community. We developed an empirically informed transmission model for WNV in 4 sites using Cx. pipiens and preferred and non-preferred avian hosts. We measured strong feeding preferences for American robins (Turdus migratorius) by Cx. pipiens, quantified as the proportion of Cx. pipiens blood meals from robins in relation to their abundance (feeding index). The model accurately predicted WNV prevalence in Cx. pipiens at 3 of 4 sites. Sensitivity analysis revealed feeding preference was the most influential parameter on intensity and timing of peak WNV infection in Cx. pipiens and a threshold feeding index for transmission was identified. Findings indicate host preference induced contact heterogeneity is a key mediator of vector-borne pathogen epizootics in multi-species host communities, and should be incorporated into multi-host transmission models (CT). We studied the impacts of landscape heterogeneity and climatic variability on the richness and diversity patterns of mosquitoes and on the abundance and WNV infection rate of Cx. pipiens. Heterogeneity in the landscape was the best predictor of both mosquito species richness and diversity, with the most heterogeneous landscapes harboring the largest number of species. In general there were no changes in species richness over the years that could be associated with weather patterns and climatic variability. Although MIR was independent of mosquito diversity, it was associated with overall mosquito abundance, which had a convex association with species richness (i.e. abundance increases to a point after which it decreases as function of species richness). Results highlight the importance of considering dominant vector species as part of a community of vectors, whose biodiversity patterns can directly or indirectly impact the risk of infectious disease transmission (MI). We examined vector competence of Cx. pipiens and Cx. restuans mosquitoes for WNV collected over 2 years from 16 local sites across 2 states to measure spatial and temporal variation in vector competence. We found extreme variation in vector competence with 0-52% of mosquitoes transmitting WNV at a single site between different sampling periods, and similar variation across populations. We also found that vector competence within a smaller geographic range (counties within a state) tended to vary somewhat synchronously, supporting environmental or population genetic drivers of variability in vector competence. In support of genetic influences, we found significant genetic variance between mosquitoes that became infected with WNV after feeding on infected blood, and those that did not. These results highlight the spatio-temporal variability in vector competence in mosquito populations and indicate that temporal variability may be influenced by local processes, such as population genetics and environmental factors. It is important that consideration of vector competence in risk estimates take this variability into account when interpreting any single measurement (NJ). Chikungunya (CHIK) fever is a mosquito-borne viral infection that has started to invade temperate countries showing a surprising potential for geographic expansion. We developed a model for disease introduction based on virus introduction by one individual. Our study combines a climate-based mosquito population dynamics stochastic model with an epidemiological model to identify temporal windows that have epidemic risk. We ran this model with temperature data from different US locations (combined with strain-specific Ae. albopictus development and CHIK virus transmission rates) to study the geographic sensitivity of epidemic potential. We found that in locations with marked seasonal variation in temperature there also was a season of epidemic risk matching the period of the year in which mosquito populations survive and grow. I n these locations controlling mosquito population sizes might be an efficient strategy. But, in other locations where the temperature is enough for mosquito development all year around the epidemic risk is high and (practically) constant. In these locations, mosquito population control alone might not be an efficient strategy control and other strategies should be implemented to complement it. Results suggest that in the event of an introduction and establishment of CHIK in the US endemic and epidemic regions would emerge initially, primarily defined by environmental factors controlling annual mosquito population cycles. These regions should be identified to plan different intervention measures. In addition, reducing vector:human ratios can lower the probability and magnitude of outbreaks for regions with strongly marked seasonal temperature patterns (NY). We continued extending our work on the landscape ecology of mosquito assemblages, transmission of dog heartworm, larval ecology, and mosquito oviposition behavior. We found highly significant segregation of mosquito species assemblages at scales of less than 100m, with different groups of species found in pasture, deciduous forest, and pasture invaded with eastern red cedar J. virginiana. We found significantly different rates of dog heartworm in vectors in urban compared to rural sites, and variable rates of dog heartworm and other parasites in coyotes across Oklahoma (OK). Objective 3. Develop strategies to control mosquito vectors. Catch basins are a major source of Cx. pipiens, Cx. restuans, and Ae. japonicus in northeastern U.S. VectoBac® CG (ai. Bacillus thuringiensis israelensis (Bti),VectoLex® CG (ai, Bacillus sphaericus (Bs), and VectoBac® 12AS (ai, Bti), each applied at maximum label rates (VectoBac CG, 1.8g per catch basin, VectoLex CG, 1.8g per catch basin, and VectoBac 12 AS, 0.193 ml per catch basin) significantly reduced numbers of larvae for one week. The dosages on the labels for treatment of mosquito larvae in catch basins, where mosquito breeding is continuous, are not adequate for providing long-term control in northeastern US without the need for frequent retreatment. When applied at 3 times the maximum label rate, VectoLex CG, VectoBac 12AS, and VectoBac CG significantly reduced numbers of larvae for 5, 4, and 2 weeks, respectively. A single application of VectoMaxTM WSP (ai, Bti + Bs) per catch basin significantly reduced numbers of 3rd and 4th instar larvae and healthy pupae in catch basins but numbers of 3rd and 4th instars in treated catch basins at 21 days post-treatment had increased to 40% of the numbers in untreated catch basins. A second treatment of 1 pouch per catch basin reduced the numbers of 3rd and 4th instars and healthy pupae to near zero. VectoMax applied as 1 pouch per catch basin on July 1 and again on August 18 significantly reduced numbers of healthy pupae throughout the summer until the end of September. A second application of VectoMax to catch basins is likely needed during summer, when rainfall averages 13.7 inches during June through September, to keep numbers of Culex and Ae. japonicus significantly reduced to lower risk of human exposure to WNV. The application of one NatularTM XRT tablet, each weighing approximately 40.5 g (ai, 6.25% spinosad), to individual catch basins in significantly reduced total numbers of larvae for 5 weeks (CT). Funded by the Area-wide management of the Asian tiger mosquito (USDA-ARS) we have tested, optimized, and shared with the mosquito control community multiple methodologies to control Ae. albopictus. We have made multiple presentations at AMCA, ESA, NJMCA, ASTMH (NJ, ARS-Gainesville, FL). A broad based approach to new toxicant discovery includes screening of conventional chemical libraries and structure activity relationship analysis, evaluation of registered compounds for mosquitocidal activity and bioassays of compounds derived from natural products. In addition, we are exploring gene silencing using RNAi technology as a method to knock down critical proteins in mosquitoes with possible applications for control (ARS-Gainesville, Fl). Studies on mosquito pathogenic viruses and microsporidia are focused on pathogen/host interactions. A cell culture system for the mosquito iridescent IIV-3 has been developed for conducting functional genomic investigations. Studies investigating the immune response in mosquitoes challenged with a baculovirus (CuniNPV) have found that a reaper ortholog (mx) is induced but apoptosis is blocked in permissive hosts by an unknown mechanism. Complete genomic sequences for three mosquito microsporidia are nearly completed. Sequencing of the genomes of Edhazardia aedis and Vavria culicis are completed and annotation is in the final stages. Anncallia algerae sequencing is underway. The annotation of all three species is expected to be completed in late 2012 (ARS-Gainesville, FL). Objective 4. Enhance surveillance technologies for mosquitoes and mosquito-borne pathogens. Mosquitoes transmit a number of distinct viruses including important human pathogens such as WNV, dengue virus, and chickungunya virus. Many of these viruses have intensified in their endemic ranges and expanded to new territories, necessitating effective surveillance and control programs to respond to these threats. One strategy to monitor virus activity involves collecting large numbers of mosquitoes from endemic sites and testing them for viral infection. We describe how to handle, process, and screen field-collected mosquitoes for infectious virus by Vero cell culture assay. Mosquitoes are sorted by trap location and species, and grouped into pools containing d50 individuals. Pooled specimens are homogenized in buffered saline using a mixer-mill and the aqueous phase is inoculated onto confluent Vero cell cultures. Cell cultures are monitored for cytopathic effect from days 3-7 post-inoculation and any viruses grown in cell culture are identified by the appropriate diagnostic assays. By utilizing this approach, we have isolated 9 different viruses from mosquitoes collected in CT, and among these, 5 are known to cause human disease. Three of these viruses (WNV Potosi virus, and La Crosse virus) represent new records for North America or the New England region since 1999. The ability to detect a wide diversity of viruses is critical to monitoring both established and newly emerging viruses in the mosquito population (CT). In rural areas, dry ice availability can be a problem. Octenol has a stable shelf life, does not need to be refrigerated and is commercially available. Last year, we compared a widely-available octenol gel product (Nosquito octenol lure 24%) versus dry ice in CDC light traps. The findings demonstrated that overall mosquito and Culex spp. capture was not statistically significant using octenol and Culex spp. and total mosquito captures were significantly higher using dry ice compared to the octenol lure. We also continued studies assessing the use of octenol as a chemoattractant. Our objective was to assess if different concentrations and forms of the compound influence Culex spp. collections. Treatments include single vs. multiple octenol gels, octenol gel vs. liquid octenol, and low vs. high diffusion of liquid octenol (NE). We used two mitochondrial loci (nicotinamide adenine dinucleotide dehydrogenase subunit 4 and cytochrome oxidase II) and a nuclear locus (28S-D2 spacer) for a total of 1337 bp to evaluate the relationships among the four subspecies of Ae. japonicus which was recently introduced into the US and has been expanding rapidly. We included in our analysis a morphologically very closely related species, Ae. koreicus, as well as 3 more distantly related species: Ae. togoi, Ae. hatorii, and Ae. vexans. We found that the 4 subspecies in the Ae. japonicus complex are genetically distinct but form a monophyletic group that includes Ae. koreicus, suggesting the need for a taxonomic reconsideration of the group. We found the 2 southern subspecies are more closely related to each other than to any of the remaining subspecies or to Ae. koreicus and may indicate an ancient north9south split of the lineage. Considering the overlap between Ae. j. japonicus and Ae. koreicus, but the stronger association between Ae. koreicus and humans, we are surprised it also has not expanded from its original range. As a proactive reaction to this possibility, we designed and tested a DNA-based rapid assay to differentiate Ae. koreicus from some of the species with which it may be confused in the US. Belgian and Italian researchers have used our rapid assay to conclude that Ae. koreicus has expanded to Belgium and Italy. These Aedes are putative vectors of several important viral encephalitides (NJ). We developed new methodology to identify the bloodmeals of Ae. albopictus using blocking primers to avoid nuclear copies and developed a new high throughput qPCR methodology to identify large numbers of container Aedes eggs (NJ). We examined the importance of physical size characteristics of larval habitats in oviposition choice in Ae. albopictus. This information can be applied to the generation of species specific gravid trapping (OK). We continued our phonological survey of the flowering plants of North Central FL. Naturally occurring, landscape and nursery plants were all included in this survey. Olfactometer studies using both male and female Ae. albopictus were conducted utilizing intact flowers of many different flowering plants. Positive responses were obtained from many of the plants, but the two which resulted in the best responses were the butterfly bush (Buddleja davidi) and golden rod (Solidago spp.). Additional olfactometer studies were conducted with these two plant types utilizing intact flowers, solvent extracts of whole flowers and individual compounds/blends that were isolated and identified by gas chromatography/mass spectrometry (ARS-Gainesville, FL). Objective 5. Develop strategies for sustainable mosquito control by including training at all levels. Andrea Egizi, graduate student, obtained an Ecology and Evolution Graduate Program Small Grants Award, Understanding the successful establishment of a temperate mosquito on a tropical island and received a Buell Award, Department of Ecology, Evolution and Natural Resources Opening the black box: Microbial metagenomics of larval mosquito feeding. George Condon, graduate student, started in September 2011 and submitted an NSF Graduate Research Fellowship entitled "High resolution spatial and temporal population genetics of invasive species". Rafael Valentin, undergraduate student will start as a graduate student in the fall of 2012. He is a co-author on a paper using qPCR for high throughput container Aedes egg identification. Rebekah Heiry, undergraduate student has been working with Dr. Fonseca since June 2011. She is now a student technician. Talks by Dr. Fonseca outside of the scientific meeting circuit aimed at enhancing understanding of vector-borne diseases and mosquito control. May, 2011: Cervia, Ravenna, Italy, Meeting entitled "Emerging vector borne disease: the role of Aedes mosquitoes". Talk entitled "The Asian tiger mosquito in the United States: past, present, and future". May 17 and May 25, 2011: two 3-hour presentations entitled "Enlightened mosquito control" to the Rutgers Environmental Stewards, at the Atlantic County Utility Authority, NJ and the Duke Farms, Hillsborough, NJ. August, 2011: presented two talks in Ghana, Africa during the workshop on Genetics and Diseases Control (NJ). Dr. Sebastien Marcombe, a postdoc funded by the Area-wide management of the Asian tiger mosquito, Ae. albopictus started May 2011 and has uncovered extensive patterns of insecticide resistance and reduced susceptibility in populations of Ae. albopictus (NJ). Dr. Jiawu Xu funded in part by AW-ATM and in part by a new NIH-R21, has optimized 8 microsatellite loci for the Asian tiger mosquito. The NextGen sequencing project of Ae. albopictus to develop a high resolution SNP array is progressing, now funded by an NIH R21 (NJ). One masters student was finished in OK in 2011, Ms. Kelsey Paras. Ms. Paras is continuing her education at the Ohio State University, pursuing a DVM. Dr. Valerie OBrien continued her post-doctoral position in mosquito ecology during 2011 (OK). We co-instructed an all-day short course with Chris Lesser (Manatee County, FL, Mosquito Abatement District) on The Asian Tiger Mosquito: Biology & Control, at the Florida Mosquito Control Associations Dodd Short Courses (USDA-ARS, Gainesville, FL).

Publications

Anderson JF, Ferrandino FJ, Dingman DW, Main AJ, Andreadis TG, Becnel JJ. 2011. Control of catch basin mosquitoes in Connecticut with Bacillus thuringiensis israelensis, Bacillus sphearicus, and Spinosad. J Am Mosq Control Assoc 27:45-55. Armstrong PM, Andreadis TG, Finan S, Shepard JJ, Thomas MC, Anderson JF. 2011. Detection of infectious virus from field-collected mosquitoes by Vero cell culture assay. J. Visualized Exper. 52. http://www.jove.com/index/Details.stp?ID=2889,doi: 10.3791/2889. Armstrong PM, Vossbrinck CR, Andreadis TG, Anderson JF, Pesko KN, Newman RM, Lennon NJ, Birren BW, Ebel GD, Henn MR. 2011. Molecular Evolution of West Nile virus in a northern temperate region: Connecticut, USA 1999-2008. Virol 417:203-210. Andreadis TG. The contribution of Culex pipiens complex mosquitoes to transmission and persistence of West Nile virus in North America. J. Amer Mosq Control Assoc (in press). Andreadis TG, Simakova AV, Vossbrinck CR, Shepard JJ, Yurchenko YA. 2012. Ultrastructural characterization and comparative phylogenetic analysis of new Microsporidia from Siberian mosquitoes: evidence for coevolution and host switching. J Invertebr Pathol 109:59-75. Bartlett-Healy K, Hamilton G, Healy S, Crepeau T, Unlu I, Farajollahi A, Fonseca DM, Gaugler R, Clark GG, Strickman D. 2011. Source reduction behavior as an independent measurement of the impact of a public health education campaign in an integrated vector management program for the Asian tiger mosquito. Int J Environ Res Public Health 8:1358-1367. Becnel JJ, Pridgeon JW. 2011. Susceptibility of mosquito and lepidopteran cell lines to the mosquito iridescent virus (IIV-3) from Aedes taeniorhynchus. J Invertebr Pathol 108:40-45. Cameron EC, Wilkerson RC, Mogi M, Miyagi I, Toma T, Kim H-C, Fonseca DM. 2010. Molecular phylogenetics of Aedes japonicus, a disease vector that recently invaded Western Europe, North America, and the Hawaiian Islands. J Med Entomol 47:527-35. Carroll JF, Tabanca N, Kramer MH, Elejalde NM, Wedge DE, Bernier UR, Coy, MR, Becnel JJ, Demirci B, Can Baser KH, Zhang J, Zhang S. 2011. Essential oils of Cupressus funebris, Juniperus communis, and J. chinensis (Cupressaceae) as repellents against ticks (Acari: Ixodidae) and mosquitoes and toxicants against mosquitoes. J Vector Ecol. 36:258-268. Chaves L, Hamer GL, Walker ED, Brown WM, Ruiz MO, Kitron UD. 2011. Climatic variability and landscape heterogeneity impact urban mosquito diversity and vector abundance and infection. Ecosphere 2:70. Chuang, T-W, Knepper RG, Stanuszek WW, Walker ED, Wilson ML. 2011. Temporal and spatial patterns of West Nile virus transmission in Saginaw County, Michigan, 2003-2006. J Med Entomol 48:1047-1056. Farajollahi A, Fonseca DM, Kramer LD, Kilpatrick AM. 2011. "Bird biting" mosquitoes and human disease: A review of the role of Culex pipiens complex mosquitoes in epidemiology. Infect Genetics Evol 11:1577-1585. Fonseca DM, Widdel A, Spichiger S-E, Hutchinson M, Kramer LD. 2010. Fine-scale spatial and temporal population genetics of a new US mosquito reveal multiple introductions Molec Ecol 19:1559-1572. Gerenday A, Fallon AM. 2011. Increased levels of the cell cycle inhibitor protein, dacapo, accompany 20-hydroxyecdysone-induced G1 arrest in a mosquito cell line. Archs Insect Biochem Physiol 78:61-73. Hamer GL, Chaves LF, Anderson TK, Kitron UD, Brawn, JD, Ruiz MO, Loss SR, Walker ED, Goldberg TL. 2011. Spatial variation in mosquito host selection and host community competence within an urban focus of West Nile virus transmission. PLoS One 6: e23767. Hamer GL, Kelly PH, Focks DA, Goldberg TJ, Walker ED. 2011. Quantification of Culex mosquito production in urban catch basins using a novel emergence trap. J Am Mosq Control Assoc 27: 142-147. Hardstone MC, Andreadis TG. 2012. Weak larval competition between the invasive mosquito, Aedes japonicus japonicus (Diptera: Culicidae) and three resident container-inhabiting mosquitoes under standard laboratory conditions. J. Med. Entomol. (in press). Hellestad VJ, Witthuhn BA, Fallon AM. 2011. The insect repellent DEET (N,N-diethyl-3-methylbenzamide) increases synthesis of glutathione S-transferase in cultured mosquito cells. Cell Biol Toxicol 27:149-157. Huang S, Molaei G, Andreadis TG. 2011. Reexamination of Culex pipiens hybridization zone in the eastern United States by ribosomal DNA-based single nucleotide polymorphism markers. Am J Trop Med Hyg 85:434-441. Helinski MEH, Harrington LC. Considerations for male fitness in successful genetic vector control programs. In: Takken and Koenraadt (eds). The Ecology of Disease Vectors (in press). Kiang RK, Soebiyanto RP, Grieco JP, Achee NL, Harrington LC, Reisen WK, Anyamba1 A, Linthicum KJ, Pinzon JC, Zollner G, Colacicco-Mayhugh M. Chapter 6: Vectorborne infectious diseases and influenza (Harrington section: Remote sensing for dengue surveillance and control) In: ISPRS - Remote Sensing Applications. (in press). Kilpatrick AM, Fonseca DM, Ebel G, Reddy M, Kramer LD. 2010. Spatial and temporal variation in vector competence of Culex mosquitoes for West Nile virus. Am J Trop Med Hyg 83:607-613. Ledemsa N, Harrington LC. 2011. Topical Review: Vectors of Dog Heartworm in the United States: Vector status and factors effecting transmission efficiency. Topics in Companion Animal Medicine 26:178-185. Leisnham PT. 2011. Invasion of the Asian tiger mosquito, Aedes albopictus. In: Francis R, ed. A Handbook of Global Freshwater Invasive Species. London: Earthscan Pub. p 137148. Leisnham PT. 2011. Vulnerable populations and regions. In: Nriagu JO, ed. Encyclopedia of Environmental Health, Vol. 5. Burlington, VT: Elsevier. p 705714. Leisnham PT, Sandoval-Mohapatra S. 2011. Mosquitoes associated with ditch-plugged and control tidal salt marshes on the Delmarva Peninsula. Int J Environ Res Pub Health 8:3099-3113. Leisnham PT, Towler L, Juliano SA. 2011. Evolution of increased photoperiodic diapause aides invasion success of the mosquito Aedes albopictus (Diptera: Culicidae). Ann Entomol Soc Am 104: 1309-1318. Liu B, Becnel JJ, Zhang Y, Zhou L. 2011. Induction of reaper ortholog mx in mosquito midgut cells following baculovirus infection. Cell Death and Differentiation 18:1337-1345. Molaei, G, Huang S, Andreadis TG. 2012. Vector-host interactions of Culex pipiens complex mosquitoes in northeastern and southern USA. J Amer Mosq Control Assoc. (in press). Newman C, Cerutti F, Anderson T, Hamer G, Walker E, Kitron U, Ruiz M, Brawn J, Goldberg T. 2011. Culex flavivirus and West Nile virus mosquito co-infection and positive ecological association in Chicago, USA. Vector Borne Zoototic Dis 11:1099-1105. Reichard MV, Tiernan KE, Paras KL, Interisano M, Reiskind MW, Panciera RJ, Pozio E. 2011. Detection of Trichinella murrelli in coyotes (Canis latrans) from Oklahoma and North Texas. Veterinary Parasitol 182:368-371. Reiskind MH, Zarrabi AA. 2011. The importance of an invasive tree fruit as a resource for mosquito larvae. J Vector Ecol 36: 97-203. Reiskind MH, Zarrabi AA. 2012. Water surface area and depth determine oviposition choice in Aedes albopictus (Diptera: Culicidae). J Med Entomol 49: 71-76. Reiskind MH, Zarrabi AA, Lounibos LP. 2012 Effects of combination of leaf resources on competition in container mosquito larvae. Bull Entomol Res FirstView: 1-1. Simakova AV, Lukiantsev VV, Vossbrinck CR, Andreadis TG. 2011. Identification of mosquito-parasitic microsporidia, Amblyospora rugosa and Trichoctosporea pygopellita (Microsporidia: Amblyosporidae), from Acanthocyclops venustus and Acanthocyclops reductus (Copepoda: Cyclopidae), based on small subunit rDNA analysis. Parazitologiia 45:140-146. Simpson JE, Hurtado PJ, MedlockJ, Molaei G, Andreadis TG, Galvani AP, Diuk-Wasser MA. 2012. Vector host-feeding preferences drive transmission of multi-host pathogens: West Nile virus as a model system. Proc R Soc B 279:925-933 Sirot LK, Hardstone MC, Helinksi MEH, Marinotti O, Kimura M, Deewatthanawong P, Wolfner MF and LC Harrington. 2011. Towards an ejaculatome of the dengue vector mosquito: protein identification and potential functions. PLoS Neglected Trop Dis. 15;5(3):e989. Sobolev VS; Khan SI, Tabanca N, Wedge DE, Manly SP, Cutler SJ, Coy MR, Becnel JJ, Neff SA, Gloer JB. 2011. Biological activity of peanut (Arachis hypogaea) phytoalexins and selected natural and synthetic stilbenoids. J Agric Food Chem. 59:1673-1682. Tabanca N, Demirci B, Gürbüz, I, Demirci F; Becnel JJ, Wedge DE, Baser HKC. 2011. Essential oil composition of five collections of Achillea biebersteinii from Central Turkey and their antifungal and insecticidal activity. Natural Product Communic 6:701-706. Unlu I, Farajollahi A, Healy SP, Crepeau T, Bartlett-Healy K, Williges E, Strickman D, Clark GG, Gaugler R, Fonseca DM. 2011. Area-wide management of Aedes albopictus: choice of study sites based on geospatial characteristics, socioeconomic factors, and mosquito populations. Pest Manage Sci 67: 965-974. Valles SM, Becnel JJ, Pereira RM. 2011. Kneallhazia carolinensae sp. nov., a microsporidian pathogen of the thief ant, Solenopsis carolinensis. J Invertebr Pathol 108:59-62. Xu J, Fonseca DM. 2011. One-way sequencing of multiple amplicons from tandem repetitive mitochondrial DNA control region. Mitochondrial DNA 22:155-158. Zhao L, Chen J, Becnel JJ; Kline DL; Clark GG, Linthicum KJ. 2011. Identification and transcription profiling of trypsin in Aedes taeniorhynchus (Diptera: Culicidae): developmental regulation, blood feeding, and permethrin exposure. J Med Entomol 48:546-553.

Impact Statements

Work carried out in 2011 as part of the Northeast Regional Research Project, NE-1043, provided valuable theoretical and practical knowledge to help manage mosquito-borne disease.
New insights were obtained on the manner in which mosquito-borne viruses overwinter, persist and circulate in mosquito populations in the northeastern US.
Research resulted in a new understanding of the host selection patterns and genetic structure of Culex pipiens populations the primary vector mosquito species for West Nile virus.
New biological control agents were discovered and characterized using molecular methods, and existing biologic agents were evaluated in a variety of mosquito breeding habitats.
Empirically informed transmission models were developed for predicting West Nile virus foci on the basis of mosquito feeding, variation in local bird species availability, landscape heterogeneity, and local temperature and precipitation.
A climate-based mosquito stochastic and epidemiological model was developed to evaluate the epidemic potential of Chikungunya virus in the event of an introduction into the US.
The ecology, behavior, and genetic structure of two invasive mosquitoes, Aedes albopictus and Aedes japonicus were studied. Understanding these ecological relationships is essential to developing economical and effective integrated pest management programs for their control.
New methods for mosquito trapping in the field and virus isolation and identification in the laboratory are is being evaluated to enhance mosquito and arbovirus surveillance programs.
Members are actively participating in graduate student and postdoctoral training programs in vector biology and mosquito and reporting research findings within the scientific community and with the general public.

Date of Annual Report: 12/22/2014

Report Information

Annual Meeting Dates: 02/27/2013 - 02/27/2013
Period the Report Covers: 10/01/2010 - 09/01/2013

Participants

Andreadis, Theodore (theodore.andreadis@ct.gov) Connecticut, Agricultural Experiment Station (Absent);
Armstrong, Philip (philip.armstrong@ct,gov) Connecticut, Agricultural Experiment Station (Present);
Becnel, Jimmy (email), USDA, Gainesville, F;
Dobson, Stephen (email) Kentucky, University of Kentucky (Present);
Fallon, Ann (fallo002@umn.edu) Minnesota, University of Minnesota (Absent);
Federici, Brian (brian.federici@ucr.edu) and Margaret Wirth, California, University of California-Riverside (Present);
Fonseca, Dina (email), New Jersey, Rutgers University (Present);
Gaugler, Randy (email) New Jersey, Rutgers University (Present);
Harrington, Laura (lch27@cornell.edu) New York, Cornell University (Absent);
Daniel, Kline (dan.kline@ars.usda.gov) USDA, ARS, Gainesville, Florida (Present);
Leisnham, Paul (leisnham@umd.edu) Maryland, University of Maryland (Present);
Stafford, Kirby (kirby.stafford@ct.gov) Connecticut, Agricultural Experiment Station (Absent);
Thomas, Michael (michael.c.thomas@ct.gov) Connecticut, Agricultural Experiment Station (Absent);
Vossbrinck, Charles (charles.vossbrinck@ct.gov)Connecticut, Agricultural Experiment Station (Absent);
Walker, Ned (walker@msu.edu) Michigan, Michigan State University (walker@msu.edu) Michigan AgBioResearch (Present);
Strickman, Daniel (daniel.strickman@ars.usda.gov) USDA, ARS, National Program Leader, Program 104: Veterinary, Medical, and Urban Entomology, Beltsville, Maryland. (Present);
Cortinas, Roberto (rcortinas@unl.edu) University of Nebraska Lincoln

Brief Summary of Minutes

The annual meeting of the Multi-State Project NE1043, Biology, Ecology and Management of Emerging Disease Vectors was held on February 27, 2013, at the Trump Taj Mahal hotel in Atlantic City, New Jersey. The meeting was held in conjunction with the annual conference of the American Mosquito Control Association.

The following were in attendance: Walker (MI), Armstrong (CT), Dobson (KY), Federici (CA), Wirth (CA), Leisnham (MD), Gaugler (NJ), Fonseca (NJ), Kline (ARS-Gainesville),.

Dr. Ned Walker (Chair) reviewed the agenda, local arrangements and format for the meeting. He circulated annual reports and noted that reports were submitted from 11 participating agencies (CT, MD, MN, NJ, NY, OK, WI, MI, NE, KY, USDA-Gainesville). The agenda included an overview, a session of individual project reports before, during, and after a buffet lunch, and a general session on status of the project and project renewal opportunities.

Dr. Kirby Stafford (Administrative Advisor) was unable to be present, however, he provided in writing a notification of the potential for renewal of the project and that the date for renewal was fast approaching. He further provided the following guidance.

“Ideally, a request to rewrite would be submitted at this meeting, but it won’t be accepted without a mid-term review and there is little time between February 27 and March 11, especially with me gone for three days the first week in March. A request can be submitted at the summer meeting and still keep on track, but in this case it might help to start work on a proposal ahead of time. The group needs to seriously think about whether to try and renew as a multi-state research project or as a coordinating committee. From what I can tell, the group is really functioning as a coordinating committee (CC) as your research activities are not jointly planned and multistate. However, while a CC will cover travel and getting together as a group, I believe it won’t cover research expenses. I have to double check that. We haven’t used CCs much in the northeast region. [They are as follows:] Multistate Research Coordinating Committees (CC) and Education/Extension and Research Activity (ERA): The membership of a CC or an ERA is made up of an AA, CSREES representative, scientists, and as applicable, extension specialists and/or extension agents. A CC or ERA provides opportunity for scientists, specialists, and others to work cooperatively to solve problems that concern more than one state, share research data, and coordinate research and other types of activities. This is presently one of the most common mechanisms for functionally integrated activities such as the regional IPM programs.

[By contrast,] The membership of a Multistate Research Project is called the technical committee, and is made up of SAES scientists, an AA, CSREES representative, other public and private sector scientists, and as applicable, extension specialists and/or extension agents. This type of activity involves cooperative, jointly planned research employing multidisciplinary approaches in which a SAES, working with other SAESs, the Agricultural Research Service (ARS), or a college or university, cooperates to solve problems that concern more than one state and usually more than one region. In addition, the following must be demonstrated in the project proposal:

1. The objectives are clearly focused.

2. Each participant listed has direct involvement in the accomplishment of the stated objectives.

3. The project is multistate and multidisciplinary.

4. The project proposal has been peer-reviewed.

5. The proposed project is oriented toward accomplishment of specific outcomes and impacts and based on priorities developed from stakeholder input.

6. The project is responsive to CSREES goals.

This guidance statement was read aloud by Dr. Walker, followed by discussion. There was no consensus regarding continuation of the NE

Brief research updates were given by each cooperator with active discussion ensuing. A group photo was taken.

Minutes Attachment:

Attachment

Accomplishments

Objective 1. Strengthen basic and applied research on the mosquito, pathogen, hosts, and environmental factors that influence disease emergence. To determine whether West Nile virus (WNV) overwinters locally or is reseeded annually, we examined the patterns of viral lineage persistence and replacement in CT over 10 consecutive transmission seasons by phylogenetic analysis. We compared the full protein coding sequence among WNV isolates to search for evidence of convergent and adaptive evolution. Viruses sampled from CT segregated into a number of well-supported subclades by year of isolation with few clades persisting e2 years. Similar viral strains were dispersed in different locations across the state and divergent strains appeared within a single location during a single transmission season, implying widespread movement and rapid colonization of virus. Numerous amino acid substitutions arose in the population but only one change, VàA at position 159 of the envelope protein, became permanently fixed. Several instances of parallel evolution were identified in independent lineages, including one amino acid change in the NS4A protein that appears to be positively selected. Results suggest that annual reemergence of WNV is driven by both reintroduction and local-overwintering of virus. Despite ongoing diversification of WNV, most amino acid variants occurred at low frequencies and were transient in the virus population (CT). Cx. tarsalis is a superior horizontal and vertical vector of WNV compared with Cx. salinarius. Cx. salinarius transmitted WNV genotype NY99 (CT 2741-99 strain) horizontally to suckling mice at significantly lower rates than Cx. tarsalis and Cx. salinarius transmitted WNV genotype NY99 to offspring at a lower vertical transmission infection rate than Cx. tarsalis. Cx. tarsalis transmitted WNV genotypes NY99 and WN02 (CT S0084-08 strain) with equal efficiency. Daily percent horizontal transmission of genotype NY99 by Cx. tarsalis -infected per os and by intra-thoracic infection was not significantly different from daily transmission of genotype WN02. Findings do not support the previously published hypothesis that genotype NY99 was replaced in the New World by WN02 because of a shorter extrinsic incubation of WN02 (CT). A survey of mosquito larvae infected with microsporidia was conducted from 2005-08 in the Tomsk, Kemerovo and Novosibirsk regions of western Siberia, Russia. Twenty-one morphologically and genetically unique species of microsporidia were isolated from 9 species of mosquitoes including 14 new species of Amblyospora, a new genus and species, Novothelohania ovalae and 6 species of Amblyospora, Parathelohania and Trichoctosporea from which gene sequences had not been previously obtained. Detailed ultrastructure of meiospores revealed unique cytological features associated with the length, arrangement and ratio of broad to narrow coils of the polar filament, comparative thickness of the exospore and endospore, and overall size of each species reaffirming their value in distinguishing taxonomic relationships. SSU rDNA sequences were unique when compared with GenBank entries. Phylogenetic trees yielded similar topologies with a high degree of congruence between parasite and host at the generic level. Species that parasitize Aedes/Ochlerotatus and Culex mosquitoes segregate into distinct monophyletic groupings mirroring their host phylogeny, while species from Anopheles mosquitoes group as a sister clade basal to the entire group of mosquito-parasitic microsporidia as their Anopheles hosts cluster as a sister clade to the entire group of culicine mosquitoes. This provides strong evidence for host-parasite coevolution by descent at the generic level and limited host lineage switching between unrelated taxa. Among parasites of Aedes/Ochlerotatus and Anopheles mosquitoes, we found several instances where a single mosquito species serves as a host for two or more related species of microsporidia, an observation consistent with host switching and independent parasite speciation. Among the microsporidian parasites of Culex mosquitoes, we found only one parasite per host indicating a higher degree of host specificity and less host switching among parasites of this genus. Findings suggest a degree of host-parasite co-speciation with host switching occurring occasionally when the normal host is unavailable in the aquatic ecosystem. Frequency of host switching seems to be occurring in proportion to host relatedness and does not cross generic boundaries in this system (CT). The spread of exotic mosquito species into new environments can introduce shifts in mosquito populations and potentially alter public health risks to mosquito-borne diseases. The successful establishment of exotic species may occur due to their competitive advantage over other cohabitating species. We hypothesized that the recently introduced exotic mosquito Ae. japonicus would be a more effective competitor than Ae. atropalpus and Ae. triseriatus, and an equal competitor to Cx. pipiens based on larval abundance data within tire habitats. Impacts of competition were measured using the larval developmental rate and survival of larvae, adult mortality, wing length, and sex ratio. We found that intraspecific competition acted strongest against Ae. japonicus versus the other three resident mosquito species by delaying larval development and increasing adult mortality. Interspecific competition was generally weak and significant main effects were only detected for species and density. Results show that larval competition between Ae. japonicus and the three resident species was weak when present, indicating that other ecological or behavioral factors may be influencing the invasion success for Ae. japonicus in North America (CT). Life-table experiments were conducted to test for differences in the adult survival and reproductive schedules of Ae. albopictus females from three populations from the northern and southern extremes of the species distribution in North America. There were consistent differences between northern and southern populations in incidence of photoperiodically-induced egg diapause. Under short daylength, diapause eggs constituted twice the proportion of total viable eggs from northern females than southern females. There were no consistent differences between northern and southern populations in resource allocation between reproduction and maintenance, reproduction over time, and reproductive investment among offspring, and no apparent trade-offs between diapause incidence with reproduction or longevity. Results suggest that the main response of North American Ae. albopictus to unfavorable winter climates is via the life history strategy of producing diapausing eggs, rather than quantitative variation in reproduction, and that there are no detectable costs to adult survival (MD). A study was conducted to characterize mosquito communities among different habitats in historically ditched tidal salt marshes and adjacent wooded areas in a wetland management area on the Maryland Delmarva Peninsula. Wooded habitats had more total mosquitoes, were more frequently occupied by mosquitoes, and had higher densities of mosquitoes. The majority of larvae at the control site were Ae. sollicitans in marsh pannes while Cx. salinarius, An. bradleyi, Ae. cantator, and Ae. sollicitans were collected in high numbers from ditches at the experimental site. We found a difference in the proportion of marsh pannes occupied by Ae. sollicitans but not total mosquitoes sampled 45 days after spring tide events than on other occasions (MD). Carbon-based secondary or structural plant compounds usually increase when they are exposed to elevated atmospheric CO2, and may differentially affect container mosquito species that feed on litter that fall into water-filled containers. Competition between Ae. albopictus and Ae. triseriatus was tested in laboratory microcosms provisioned with litter grown in either elevated (1,000 ppm) or ambient CO2 (385 ppm). In additional microcosms, single-species cohorts of either Ae. albopictus or Ae. triseriatus were provided with increasing amounts of one of the two litter types or concentrations of commercial tannic acid to test potential toxicity effects of leaf-derived tannins. We found no differences in mosquito competition between litter types, with Ae. albopictus consistently superior to Ae. triseriatus. Single-species microcosms provisioned with elevated leaf litter had slower male development time than microcosms with ambient (lower-tannin) litter. Ae. albopictus and Ae. triseriatus also experienced similarly higher mortality and slower development with increasing commercial tannic acid concentrations. These results suggest that possible litter grown in high CO2 conditions may suppress mosquito production by increasing the toxicity of leaf litter, but that it would not affect population-level competition between Ae. albopictus and Ae. triseriatus (MD). We conducted yard surveys for mosquito larval habitats paired with knowledge, attitude, and practice questionnaires administered to residents among six neighborhoods in Washington DC that vary in socioeconomic status. Household income was related to overall knowledge and attitudes of residents towards mosquitoes, but that knowledge and attitudes were not related with source reduction as measured through questionnaire responses or actual numbers of water-holding containers in their yard. Numbers of water-holding containers was strongly positively related to exposure to Ae. albopictus, Cx. pipiens and pupae but not related to whether or not households practiced source reduction. Households that reported practicing source reduction had lower Cx. pipiens infestation and abundances but no difference in Ae. albopictus or pupal infestation or abundance. Results suggest large households are a greater source of vector mosquitoes and should be targeted in education campaigns, and that self-perceived source reduction of residents may not reduce urban mosquito habitat (MD). The risk presented to humans by circulation of WNV in the Culex bird enzootic cycle established in metropolitan areas of the upper Midwest was continued. Culex Flavivirus was positively associated with WNV infection in a case-control study of mosquito pools collected from an endemic focus of WNV transmission in Chicago. This study provides direct evidence that insect-specific flaviviruses may influence human health risk by enhancing WNV infection in mosquitoes that are epidemic bridge vectors to human hosts (MI). Analysis of molecular genetic diversity in 9 marker regions of 5 genes within the bacteriophage WO region of the Wolbachia pipentis genome revealed high diversity of wPip in the population of Cx. pipiens sampled in metropolitan Chicago, IL. Wolbachia endosymbionts may modulate propensity to infection with this virus and may affect population structure of their mosquito hosts. Multiple regression showed that markers associated with gene Gp2d were significantly associated with ancestry of individuals to form molestus or form pipiens, as determined by prior microsatellite allele frequency analysis. Data suggest that certain wPip molecular genetic types are associated with genetic substructuring in the Cx. pipiens complex of Chicago, and that the association extends to host preference. The influence of host diversity on multi-host pathogen persistence and transmission is confounded by the large number of species and complex biological interactions that comprise the transmission system. Although diversity and composition of host communities have been hypothesized to affect intensity of transmission, the effect of host community structure on the transmission of vector-borne pathogens remains largely unknown. Results suggest that host community measures, such as species diversity, may not be useful indicators of transmission risk at fine spatial scales in vector-borne disease systems (MI). Culex mosquitoes carry an intracellular symbiont, W. pipientis, a bacterium which can potentially be manipulated to control vector populations by spreading a reproductive distortion called cytoplasmic incompatibility. We obtained a Cx. pipiens colony from Ohio State University and, for comparative purposes, have developed a sister colony cured of Wolbachia infection. By comparing protein bands from dissected testes, we identified proteins produced in response to Wolbachia infection. Efforts are underway to clone two Wolbachia DNA binding proteins that are implicated in the reproductive distortion known as cytoplasmic incompatibility. Parallel investigations with Wolbachia-infected cell lines are used to identify Wolbachia-inducible protein expression in undifferentiated cells that do not engage in gamete production. We are investigating methods to modulate expression of these proteins by testing effects of specific components, including the DNA synthesis inhibitor mimosine and the mitochondrial inhibitor, paraquat, in the culture medium. We are finalizing methodology for quantifying Wolbachia infections using fluorescence activated cell sorting (MN). Ae. japonicus, which was first collected during summer 2010, was again collected during summer 2011 in eastern NE. Specimens were collected using CDC-light traps baited with carbon dioxide. Mosquitoes were collected in four sites within Lincoln, NE. The importance of Ae. japonicus to vector disease ecology and epidemiology in eastern NE is not known. (NE). Using a panel of seven microsatellite loci, we confirmed the existence of two abundant genetic forms in specimens originally collected in 19992000 that matches the disjunctive distribution of mitochondrial haplotypes. To examine the distribution of the two genetic types across PA we created a fine scale genetic map of Ae. japonicus using specimens collected from 54 Pennsylvania counties in 20022003. We also made direct comparisons between collections in 19992000 and new collections made in 20042005 obtained from the same areas in the northeastern US. We observed that the strong association between mtDNA haplotype and microsatellite signature seen in 19992000 had weakened significantly by 2002 across PA, a trend continued to some extent in 20042005 in PA, NJ, and NY, indicating that once easily distinguishable separate introductions are merging. The two expanding genetic forms create a complex correlation between spatial and genetic distances. The existence of multiple introductions would be obscured without sampling early and across time with highly polymorphic molecular markers. Results provide a high resolution analysis of the spatial and temporal dynamics of a newly introduced disease vector and argue that successive introductions may be a common pattern for invasive mosquitoes (NJ) We documented the potential importance of eastern red cedar, Juniperus virginiana leaves and fruit as a larval resource for the invasive Ae. albopictus in OK. We examined how different leaf species commonly found in FL interact to affect competition within and between Ae. albopictus and Aedes aegypti. We have also continued investigations into the effects of temperature and food on growth in larval mosquitoes (OK). Laboratory and field tests of potential biological control agents were conducted for control of Cx. pipiens. The key goals of this project are to test locally adapted fathead minnows and copepods for control of Cx. pipiens/restuans. Fish are used in aboveground breeding sites while copepods are used in the underground catch basin system. Fish were not released in 2011 because of extended dry conditions which limited the extent of targeted wet sites and the occurrence of larvae in those sites. Belowground sites did begin to generate toward the end of the season and these were used for trials of copepod releases. We tested the efficacy of copepods as predators of Culex or Anopheles larvae. Acanthocyclops trajani and Macrocyclops albidus were tested against An. stephensi and Cx. pipiens. M. albidus females killed 50% of Culex larvae within 24 hr while A. trajani killed only 35% of larvae. Results for An. stephensi were similar. We tested whether releases of M. albidus could affect larval Culex in catch basins (WI). We released M. albidus, into storm drain catch basins in Madison, WI at the beginning of August. We monitored these sites along with untreated control sites until the end of September. The catch basins were sampled every week using a mosquito dipper, and immature mosquitoes were collected, counted, and reared to adulthood for identification of the species. All adult mosquitoes collected were identified as Cx. pipiens. Copepods were collected during the sampling and identified as M. albidus or A. vernalis or inconsequential species. Catch basins were infested with mosquito larvae. M. albidus failed to establish in all but one location. This single location, however, maintained and multiplied the number of M. albidus with many being collected every week. M. albidus colonized an adjacent catch basin after approximately 1 month and their numbers continued to increase throughout the survey. Citywide water main flushing after our introductions took place may have resulted in the flushing of M. albidus from the test sites. Heavy rainfall events may have also flushed these copepods from their new habitats on more than one occasion. The A. vernalis colony was created from the catch basin samples and these copepods were frequently found in our sites. They appear better adapted to the catch basin environment than M. albidus (WI). Objective 2. Use knowledge of mosquito, pathogen, vertebrate reservoir, and environment interactions to enhance ability to predict conditions leading to disease. Seasonal epizootics of vector-borne pathogens infecting multiple species are ecologically complex and difficult to forecast. Pathogen transmission potential within the host community is determined by the relative abilities of host species to maintain and transmit the pathogen and by ecological factors influencing contact rates between hosts and vectors. Increasing evidence of strong feeding preferences by a number of vectors suggests that the host community experienced by the pathogen may be very different from the local host community. We developed an empirically informed transmission model for WNV in 4 sites using Cx. pipiens and preferred and non-preferred avian hosts. We measured strong feeding preferences for American robins (Turdus migratorius) by Cx. pipiens, quantified as the proportion of Cx. pipiens blood meals from robins in relation to their abundance (feeding index). The model accurately predicted WNV prevalence in Cx. pipiens at 3 of 4 sites. Sensitivity analysis revealed feeding preference was the most influential parameter on intensity and timing of peak WNV infection in Cx. pipiens and a threshold feeding index for transmission was identified. Findings indicate host preference induced contact heterogeneity is a key mediator of vector-borne pathogen epizootics in multi-species host communities, and should be incorporated into multi-host transmission models (CT). We studied the impacts of landscape heterogeneity and climatic variability on the richness and diversity patterns of mosquitoes and on the abundance and WNV infection rate of Cx. pipiens. Heterogeneity in the landscape was the best predictor of both mosquito species richness and diversity, with the most heterogeneous landscapes harboring the largest number of species. In general there were no changes in species richness over the years that could be associated with weather patterns and climatic variability. Although MIR was independent of mosquito diversity, it was associated with overall mosquito abundance, which had a convex association with species richness (i.e. abundance increases to a point after which it decreases as function of species richness). Results highlight the importance of considering dominant vector species as part of a community of vectors, whose biodiversity patterns can directly or indirectly impact the risk of infectious disease transmission (MI). We examined vector competence of Cx. pipiens and Cx. restuans mosquitoes for WNV collected over 2 years from 16 local sites across 2 states to measure spatial and temporal variation in vector competence. We found extreme variation in vector competence with 0-52% of mosquitoes transmitting WNV at a single site between different sampling periods, and similar variation across populations. We also found that vector competence within a smaller geographic range (counties within a state) tended to vary somewhat synchronously, supporting environmental or population genetic drivers of variability in vector competence. In support of genetic influences, we found significant genetic variance between mosquitoes that became infected with WNV after feeding on infected blood, and those that did not. These results highlight the spatio-temporal variability in vector competence in mosquito populations and indicate that temporal variability may be influenced by local processes, such as population genetics and environmental factors. It is important that consideration of vector competence in risk estimates take this variability into account when interpreting any single measurement (NJ). Chikungunya (CHIK) fever is a mosquito-borne viral infection that has started to invade temperate countries showing a surprising potential for geographic expansion. We developed a model for disease introduction based on virus introduction by one individual. Our study combines a climate-based mosquito population dynamics stochastic model with an epidemiological model to identify temporal windows that have epidemic risk. We ran this model with temperature data from different US locations (combined with strain-specific Ae. albopictus development and CHIK virus transmission rates) to study the geographic sensitivity of epidemic potential. We found that in locations with marked seasonal variation in temperature there also was a season of epidemic risk matching the period of the year in which mosquito populations survive and grow. I n these locations controlling mosquito population sizes might be an efficient strategy. But, in other locations where the temperature is enough for mosquito development all year around the epidemic risk is high and (practically) constant. In these locations, mosquito population control alone might not be an efficient strategy control and other strategies should be implemented to complement it. Results suggest that in the event of an introduction and establishment of CHIK in the US endemic and epidemic regions would emerge initially, primarily defined by environmental factors controlling annual mosquito population cycles. These regions should be identified to plan different intervention measures. In addition, reducing vector:human ratios can lower the probability and magnitude of outbreaks for regions with strongly marked seasonal temperature patterns (NY). We continued extending our work on the landscape ecology of mosquito assemblages, transmission of dog heartworm, larval ecology, and mosquito oviposition behavior. We found highly significant segregation of mosquito species assemblages at scales of less than 100m, with different groups of species found in pasture, deciduous forest, and pasture invaded with eastern red cedar J. virginiana. We found significantly different rates of dog heartworm in vectors in urban compared to rural sites, and variable rates of dog heartworm and other parasites in coyotes across Oklahoma (OK). Objective 3. Develop strategies to control mosquito vectors. Catch basins are a major source of Cx. pipiens, Cx. restuans, and Ae. japonicus in northeastern U.S. VectoBac® CG (ai. Bacillus thuringiensis israelensis (Bti),VectoLex® CG (ai, Bacillus sphaericus (Bs), and VectoBac® 12AS (ai, Bti), each applied at maximum label rates (VectoBac CG, 1.8g per catch basin, VectoLex CG, 1.8g per catch basin, and VectoBac 12 AS, 0.193 ml per catch basin) significantly reduced numbers of larvae for one week. The dosages on the labels for treatment of mosquito larvae in catch basins, where mosquito breeding is continuous, are not adequate for providing long-term control in northeastern US without the need for frequent retreatment. When applied at 3 times the maximum label rate, VectoLex CG, VectoBac 12AS, and VectoBac CG significantly reduced numbers of larvae for 5, 4, and 2 weeks, respectively. A single application of VectoMaxTM WSP (ai, Bti + Bs) per catch basin significantly reduced numbers of 3rd and 4th instar larvae and healthy pupae in catch basins but numbers of 3rd and 4th instars in treated catch basins at 21 days post-treatment had increased to 40% of the numbers in untreated catch basins. A second treatment of 1 pouch per catch basin reduced the numbers of 3rd and 4th instars and healthy pupae to near zero. VectoMax applied as 1 pouch per catch basin on July 1 and again on August 18 significantly reduced numbers of healthy pupae throughout the summer until the end of September. A second application of VectoMax to catch basins is likely needed during summer, when rainfall averages 13.7 inches during June through September, to keep numbers of Culex and Ae. japonicus significantly reduced to lower risk of human exposure to WNV. The application of one NatularTM XRT tablet, each weighing approximately 40.5 g (ai, 6.25% spinosad), to individual catch basins in significantly reduced total numbers of larvae for 5 weeks (CT). Funded by the Area-wide management of the Asian tiger mosquito (USDA-ARS) we have tested, optimized, and shared with the mosquito control community multiple methodologies to control Ae. albopictus. We have made multiple presentations at AMCA, ESA, NJMCA, ASTMH (NJ, ARS-Gainesville, FL). A broad based approach to new toxicant discovery includes screening of conventional chemical libraries and structure activity relationship analysis, evaluation of registered compounds for mosquitocidal activity and bioassays of compounds derived from natural products. In addition, we are exploring gene silencing using RNAi technology as a method to knock down critical proteins in mosquitoes with possible applications for control (ARS-Gainesville, Fl). Studies on mosquito pathogenic viruses and microsporidia are focused on pathogen/host interactions. A cell culture system for the mosquito iridescent IIV-3 has been developed for conducting functional genomic investigations. Studies investigating the immune response in mosquitoes challenged with a baculovirus (CuniNPV) have found that a reaper ortholog (mx) is induced but apoptosis is blocked in permissive hosts by an unknown mechanism. Complete genomic sequences for three mosquito microsporidia are nearly completed. Sequencing of the genomes of Edhazardia aedis and Vavria culicis are completed and annotation is in the final stages. Anncallia algerae sequencing is underway. The annotation of all three species is expected to be completed in late 2012 (ARS-Gainesville, FL). Objective 4. Enhance surveillance technologies for mosquitoes and mosquito-borne pathogens. Mosquitoes transmit a number of distinct viruses including important human pathogens such as WNV, dengue virus, and chickungunya virus. Many of these viruses have intensified in their endemic ranges and expanded to new territories, necessitating effective surveillance and control programs to respond to these threats. One strategy to monitor virus activity involves collecting large numbers of mosquitoes from endemic sites and testing them for viral infection. We describe how to handle, process, and screen field-collected mosquitoes for infectious virus by Vero cell culture assay. Mosquitoes are sorted by trap location and species, and grouped into pools containing d50 individuals. Pooled specimens are homogenized in buffered saline using a mixer-mill and the aqueous phase is inoculated onto confluent Vero cell cultures. Cell cultures are monitored for cytopathic effect from days 3-7 post-inoculation and any viruses grown in cell culture are identified by the appropriate diagnostic assays. By utilizing this approach, we have isolated 9 different viruses from mosquitoes collected in CT, and among these, 5 are known to cause human disease. Three of these viruses (WNV Potosi virus, and La Crosse virus) represent new records for North America or the New England region since 1999. The ability to detect a wide diversity of viruses is critical to monitoring both established and newly emerging viruses in the mosquito population (CT). In rural areas, dry ice availability can be a problem. Octenol has a stable shelf life, does not need to be refrigerated and is commercially available. Last year, we compared a widely-available octenol gel product (Nosquito octenol lure 24%) versus dry ice in CDC light traps. The findings demonstrated that overall mosquito and Culex spp. capture was not statistically significant using octenol and Culex spp. and total mosquito captures were significantly higher using dry ice compared to the octenol lure. We also continued studies assessing the use of octenol as a chemoattractant. Our objective was to assess if different concentrations and forms of the compound influence Culex spp. collections. Treatments include single vs. multiple octenol gels, octenol gel vs. liquid octenol, and low vs. high diffusion of liquid octenol (NE). We used two mitochondrial loci (nicotinamide adenine dinucleotide dehydrogenase subunit 4 and cytochrome oxidase II) and a nuclear locus (28S-D2 spacer) for a total of 1337 bp to evaluate the relationships among the four subspecies of Ae. japonicus which was recently introduced into the US and has been expanding rapidly. We included in our analysis a morphologically very closely related species, Ae. koreicus, as well as 3 more distantly related species: Ae. togoi, Ae. hatorii, and Ae. vexans. We found that the 4 subspecies in the Ae. japonicus complex are genetically distinct but form a monophyletic group that includes Ae. koreicus, suggesting the need for a taxonomic reconsideration of the group. We found the 2 southern subspecies are more closely related to each other than to any of the remaining subspecies or to Ae. koreicus and may indicate an ancient north9south split of the lineage. Considering the overlap between Ae. j. japonicus and Ae. koreicus, but the stronger association between Ae. koreicus and humans, we are surprised it also has not expanded from its original range. As a proactive reaction to this possibility, we designed and tested a DNA-based rapid assay to differentiate Ae. koreicus from some of the species with which it may be confused in the US. Belgian and Italian researchers have used our rapid assay to conclude that Ae. koreicus has expanded to Belgium and Italy. These Aedes are putative vectors of several important viral encephalitides (NJ). We developed new methodology to identify the bloodmeals of Ae. albopictus using blocking primers to avoid nuclear copies and developed a new high throughput qPCR methodology to identify large numbers of container Aedes eggs (NJ). We examined the importance of physical size characteristics of larval habitats in oviposition choice in Ae. albopictus. This information can be applied to the generation of species specific gravid trapping (OK). We continued our phonological survey of the flowering plants of North Central FL. Naturally occurring, landscape and nursery plants were all included in this survey. Olfactometer studies using both male and female Ae. albopictus were conducted utilizing intact flowers of many different flowering plants. Positive responses were obtained from many of the plants, but the two which resulted in the best responses were the butterfly bush(Buddleja davidi) and golden rod (Solidago spp.). Additional olfactometer studies were conducted with these two plant types utilizing intact flowers, solvent extracts of whole flowers and individual compounds/blends that were isolated and identified by gas chromatography/mass spectrometry (ARS-Gainesville, FL). Objective 5. Develop strategies for sustainable mosquito control by including training at all levels. Andrea Egizi, graduate student, obtained an Ecology and Evolution Graduate Program Small Grants Award, Understanding the successful establishment of a temperate mosquito on a tropical island and received a Buell Award, Department of Ecology, Evolution and Natural Resources Opening the black box: Microbial metagenomics of larval mosquito feeding. George Condon, graduate student, started in September 2011 and submitted an NSF Graduate Research Fellowship entitled "High resolution spatial and temporal population genetics of invasive species". Rafael Valentin, undergraduate student will start as a graduate student in the fall of 2012. He is a co-author on a paper using qPCR for high throughput container Aedes egg identification. Rebekah Heiry, undergraduate student has been working with Dr. Fonseca since June 2011. She is now a student technician. Talks by Dr. Fonseca outside of the scientific meeting circuit aimed at enhancing understanding of vector-borne diseases and mosquito control. May, 2011: Cervia, Ravenna, Italy, Meeting entitled "Emerging vector borne disease: the role of Aedes mosquitoes". Talk entitled "The Asian tiger mosquito in the United States: past, present, and future". May 17 and May 25, 2011: two 3-hour presentations entitled "Enlightened mosquito control" to the Rutgers Environmental Stewards, at the Atlantic County Utility Authority, NJ and the Duke Farms, Hillsborough, NJ. August, 2011: presented two talks in Ghana, Africa during the workshop on Genetics and Diseases Control (NJ). Dr. Sebastien Marcombe, a postdoc funded by the Area-wide management of the Asian tiger mosquito, Ae. albopictus started May 2011 and has uncovered extensive patterns of insecticide resistance and reduced susceptibility in populations of Ae. albopictus (NJ). Dr. Jiawu Xu funded in part by AW-ATM and in part by a new NIH-R21, has optimized 8 microsatellite loci for the Asian tiger mosquito. The NextGen sequencing project of Ae. albopictus to develop a high resolution SNP array is progressing, now funded by an NIH R21 (NJ). One masters student was finished in OK in 2011, Ms. Kelsey Paras. Ms. Paras is continuing her education at the Ohio State University, pursuing a DVM. Dr. Valerie OBrien continued her post-doctoral position in mosquito ecology during 2011 (OK). We co-instructed an all-day short course with Chris Lesser (Manatee County, FL, Mosquito Abatement District) on The Asian Tiger Mosquito: Biology & Control,at the Florida Mosquito Control Associations Dodd Short Courses (USDA-ARS, Gainesville, FL).

Publications

1. Anderson JF, McKnight S, Ferrandino FJ. 2012. Aedes japonicus japonicus and associated woodland species attracted to Centers for Disease Control and Prevention miniature light traps baited with carbon dioxide and the TrapTech Mosquito Lure. J Amer Mosq Control Assoc 28:184-191. 2. Andreadis TG. 2012. The contribution of Culex pipiens complex mosquitoes to transmission and persistence of West Nile virus in North America. J Amer Mosq Control Assoc 28s:137-151. 3. Andreadis TG, Armstrong PM. 2012. 2012: a record year for West Nile virus activity in Connecticut. CT Weekly Agric Report 92:1-4. 4. Andreadis TG, Shepard JJ, Thomas MC. 2012. Field observations on the overwintering ecology of Culiseta melanura in the northeastern United States. J Amer Mosq Control Assoc 28:286-291. 5. Andreadis TG, Simakova AV, Vossbrinck CR, Shepard JJ, Yurchenko YA. 2012. Ultrastructural characterization and comparative phylogenetic analysis of new Microsporidia from Siberian mosquitoes: evidence for coevolution and host switching. J Invertebr Pathol 109:59-75. 6. Armstrong PM, Prince N, Andreadis TG. 2012. Development of a multi-target TaqMan assay to detect eastern equine encephalitis virus variants in mosquitoes. Vector-Borne Zoonotic Dis 12:872-876. 7. Hardstone MC, Andreadis TG. 2012. Weak larval competition between the invasive mosquito, Aedes japonicus japonicus (Diptera: Culicidae) and three resident container-inhabiting mosquitoes under standard laboratory conditions. J Med Entomol 49:277-285. 8. Molaei, G, Huang S, Andreadis TG. 2012. Vector-host interactions of Culex pipiens complex mosquitoes in northeastern and southern USA. J Amer Mosq Control Assoc 28:127-136. 9. Molaei G, Andreadis TG, Armstrong PM, Thomas MC, Deschamps T, Cuebas-Incle, E, Montgomery W, Osborne M, Smole S, Matton P, Andrews W, Best C, Cornine III F, Bidlack E, Texeira T. 2012. Vector-host interactions and epizootiology of eastern equine encephalitis virus in Massachusetts, USA. Vector-Borne Zoonotic Dis (in press). 10. Morningstar RJ, Hamer, GL, Goldberg TL, Huang S, Andreadis TG, Walker ED. 2012. Diversity of Wolbachia pipientis strain wPip in a genetically admixtured, above-ground Culex pipiens (Diptera: Culicidae) population: association with form molestus ancestry and host selection patterns. J Med Entomol 49:474-481. 11. Simpson JE, Hurtado PJ, Medlock J, Molaei G, Andreadis,TG, Galvani AP, Diuk-Wasser MA. 2012. Vector host-feeding preferences drive transmission of multi-host pathogens: West Nile virus as a model system. Proc R Soc B 279:925-933. 12. Hamer GL, Donovan DJ, Hood-Nowotny R, Kaufman MG, Goldberg TL, Walker ED. Evaluation of a stable isotope method to mark naturally-breeding larval mosquitoes for adult dispersal studies. J Med Entomol 2012; 49: 61-70. 13. Gardner A, Hamer G, Hines A, Newman C, Walker ED, Ruiz M. Temporal change in precipitation and temperature affects habitat for larval Culex (Diptera: Culicidae) in storm water catch basins in suburban Chicago, USA, 2010. J Med Entomol 2012; 49: 270-276. 14. Morningstar RJ, Hamer GL, Goldberg TL, Huang S, Andreadis TG, Walker ED. Diversity of Wolbachia pipientis strain wPip in a genetically admixtured, above-ground Culex pipiens (Diptera: Culicidae) population: association with form molestus ancestry and host selection patterns. J Med Entomol 2012; 49: 474-81. 15. Chuang T-W, Knepper RG, Stanuszek WW, Walker ED, Wilson ML. Cross-correlation map analyses show weather variation influences on mosquito abundance patterns in Saginaw County, Michigan, 1989 -2005. J Med Entomol 2012; 49: 851-58. 16. Kaufman MG, Stanuszek WW, Brouhard EA, Knepper RG, Walker ED. Establishment of Aedes japonicus japonicus and its colonization of container habitats in Michigan. J Med Entomol 2012; 49: 1307-17. 17. Versteirt V, De Clercq EM, Fonseca DM, Pecor J, Schaffner F, Coosemans M, Van Bortel W. 2012. Bionomics of the established exotic mosquito species Aedes koreicus in Belgium, Europe. Journal of Medical Entomology. 49(6): 1226-1232. 18. Mogi M, Armbruster P, Fonseca DM. 2012. Analyses of the northern distribution limit of Aedes albopictus (Diptera: Culicidae) with a simple thermal index. Journal of Medical Entomology. 49(6): 1233-1243. 19. Strickman D, Fonseca DM. 2012. Autogeny in Culex pipiens complex mosquitoes from the San Francisco Bay Area. American Journal of Tropical Medicine and Hygiene. 87(4):719-26. 20. Versteirt V, Pecor J, Fonseca DM, Coosemans M, Bortel WV. 2012. Confirmation of Aedes koreicus (Diptera: Culicidae) in Belgium and description of morphological differences between Korean and Belgian specimens validated by molecular identification. Zootaxa. 3191:21-32. 21. Ledemsa, N and Harrington, L.C. 2011. Topical Review: Vectors of Dog Heartworm in the United States: Vector status and factors effecting transmission efficiency. Topics in Companion Animal Medicine. Volume 26 (4): 178-185. 22. Ruiz-Moreno D, Vargas IS, Olson KE, Harrington LC. 2012. Modeling Dynamic Introduction of Chikungunya Virus in the United States. PLoS Negl. Trop. Dis. 6(11): e1918. doi:10.1371/journal.pntd.0001918 23. Brown HE, Harrington LC, Kaufman PE, McKay T, Bowman DD, Nelson CT, Wang D and R. Lund 2012. Key Factors Influencing Canine Heartworm, Dirofilaria immitis in the United States. Paras. Vect. 5(1):245. 24. Menda G, Uhr JH, Wyttenbach RA, Vermeylen FM, Smith DM, Harrington LC, and RR Hoy. 2012. Associative learning in the dengue vector mosquito, Aedes aegypti: Avoidance of a previously attractive odor or surface color that is paired with an aversive stimulus. J. Exper Biol doi:10.1242/jeb.074898 25. Helinksi MEH, Deewatthanawong P, Sirot LK, Wolfner MF and LC Harrington. 2012. Duration and dose-dependency of female sexual receptivity responses to seminal fluid proteins in Aedes albopictus and Ae. aegypti mosquitoes. J Insect Phys. 58(10):1307–1313 26. Helinski MEH and LC Harrington. 2012. The role of male harassment on female fitness for the dengue vector, Ae. aegypti. Behavioral Ecology and Sociobiology. DOI: 10.1007/s00265-012-1365-9 27. Beckmann, J. F. and Fallon, A. M. Decapitation improves detection of Wolbachia pipientis (Rickettsiales: Rickettsiaceae) in Culex pipiens Linnaeus (Diptera: Culicidae) mosquitoes by the polymerase chain reaction. J. Med. Entomol., 49, 1103-1108, 2012. NIHMS432423 28. Fallon, AM, Baldridge, GD, Higgins, LA, Witthuhn, BA. Wolbachia from the planthopper Laodelphax striatellus establishes a robust, persistent, streptomycin resistant infection in clonal mosquito cells. In vitro cell develop biol--Animal, 49, 66-73, 2013; chosen for feature article, In Vitro Reports Society for In Vitro Biology Newsletter. 29. Beckmann, JF, Markowski TW, Witthuhn BA, Fallon, AM. Detection of the Wolbachia encoded DNA binding protein, HU beta, in mosquito gonads. Insect Biochem Mol Biol, in press. NIHMS432148. 30. Coutinho, C.J.P., Alves, R., Sanscrainte, N.D., Viviani, A.S., Paulo Souza, P.A., Carvalho-Mello, I., Becnel, J.J. 2012. Occurrence and phylogenetic characterization of a baculovirus isolated from Culex quinquefasciatus in São Paulo State, Brazil Archives Virology 157: 1741-1745. 31. Coy, M. R. Sanscrainte, N. D., Chalaire, K. C., Inberg, A., Maayan, I., Glick, E., Paldi, N., Becnel, J. J. 2012. Gene silencing in adult Aedes aegypti mosquitoes through oral delivery of double-stranded RNA. J. Applied Entomol. 136: 741-748. DOI: 10.1111/j.1439-0418.2012.01713.x . 32. Cuomo CA, Desjardins CA, Bakowski MA, Goldberg J, Ma AT, Becnel JJ, Didier ES, Fan L, Heiman DI, Levin JZ, Young S, Zeng Q, Troemel ER. 2012. Microsporidian genome analysis reveals evolutionary strategies for obligate intracellular growth. Genome Research. 22: 2478-2488. doi/10.1101/gr.142802.112 33. Muttis, E., Miele, S.A.B., Belaich, M.N., Micieli, M.V., Becnel, J.J. Ghiringhelli P.D. and García, J.J. 2012. First record of a mosquito iridescent virus in Culex pipiens L. (Diptera: Culicidae) Archives Virology 157: 1569-1571. 34. Ozek, G., Ishmuratova,M., Tabanca, N., Radwan, M. M., Goger, F., Ozek, T., Wedge, D. E., Becnel, J. J., Cutler, S. J., Can Baser, K. H. 2012. One-step multiple component isolation from the oil of Crinitaria tatarica (Less.) Sojak by preparative capillary gas chromatography with characterization by spectroscopic and spectrometric techniques and evaluation of biological activity J. Sep. Sci., 35, 650-660. 35. Rosa, L. H., Tabanca, N., Techen, N., Wedge, D. E., Pan, Z., Bernier, U. R., Becnel, J. J., Elejalde, N. M., Walker, L. A., Moraes, R. M. 2012. Fungal Diversity Associated with Micropropagated Plants of Echinacea purpurea (L.) Moench American Journal of Plant Sciences, 3: 1105-1114. 36. Sims, K., Becnel, J. J. and Funderburk, J. 2012. The morphology and biology of the entomophilic Thripinema fuscum (Tylenchida: Allantonematidae), and the histopathological effects of parasitism on the host Frankliniella fusca (Thysanoptera: Thripidae), Journal of Natural History, 46:17-18, 1111-1128. 37. Sterling,K. M., Okech, B. A., Xiang, M. A., Linser, P. J., Price, D. A., VanEkeris, L., Becnel, J. J. and Harvey, W. H. 2012. High affinity 3H-Phe uptake by brush border membrane vesicles from whole larvae of Aedes aegypti (AaBMVw). Journal of Insect Physiology 58, 580-589. 38. Diaz-Mendoza M, Bideshi DK, Ortego F, Farinos GF, Federici BA. 2012. The 20-kDa chaperone-like protein of Bacillus thuringiensis spp. israelensis enhances yield, crystal size and solubility of Cry3A. Letts Appl Microbiol 54: 88-95. 39. Barboza-Corona JE. Park HW, Bideshi DK, Federici BA. 2012. The 60-kDa protein encoded by orf2 in the cry19A operon of Bacillus thuringiensis subsp. jegathesan functions like a C-terminal crystallization domain. Appl Environ Microbiol 78:2005-2012. 40. Diaz-Mendoza M, Bideshi DK, Federici BA. 2012. A 54-kDa protein encoded by pBtoxis is required for parasporal body structural integrity in Bacillus thuringiensis subsp. israelensis. J Bacteriol 194:1562-1571. 41. Wirth MC, W. E. Walton WE, Federici BA. 2012. Inheritance, stability, and dominance of resistance in in Culex quinquefasciatus Say (Diptera: Culicidae) selected with the three Cry toxins of Bacillus thuringiensis subsp. israelensis. J Med Ent 49:886-894. 42. Leisnham, P.T. and S.A. Juliano. 2012. Impacts of Climate, Land Use, and Biological Invasion on the Ecology of Immature Aedes Mosquitoes: Implications for La Crosse Emergence. EcoHealth 9: 217-228. 43. Andrews ES, Crain PR, Fu Y, Howe DK & Dobson SL (2012) Introduced Wolbachia Infection Affects Production of Reactive Oxygen Species and Brugia pahangi Survival within Aedes polynesiensis. PLoS Pathogens. In Press. 44. Calvitti M, Moretti R, Skidmore AR & Dobson SL (2012) Wolbachia strain wPip yields a pattern of cytoplasmic incompatibility enhancing a Wolbachia-based suppression strategy against the disease vector Aedes albopictus. Parasites & vectors. In Press. 45. Chambers EW, Bossin H, Ritchie SA, Russell RC & Dobson SL (2012) Landing response of Aedes polynesiensis mosquitoes to colored targets. Medical and Veterinary Entomology. In Press. 46. Mains JW, Brelsfoard CL, Crain PR, Huang Y & Dobson SL (2012) Population impacts of Wolbachia on Aedes albopictus. Ecological Applications. In Press.

Publications Attachment:

Attachment

Impact Statements

Work carried out in 2011 as part of the Northeast Regional Research Project, NE-1043, provided valuable theoretical and practical knowledge to help manage mosquito-borne disease.
New insights were obtained on the manner in which mosquito-borne viruses overwinter, persist and circulate in mosquito populations in the northeastern US.
Research resulted in a new understanding of the host selection patterns and genetic structure of Culex pipiens populations the primary vector mosquito species for West Nile virus.
New biological control agents were discovered and characterized using molecular methods, and existing biologic agents were evaluated in a variety of mosquito breeding habitats.
Empirically informed transmission models were developed for predicting West Nile virus foci on the basis of mosquito feeding, variation in local bird species availability, landscape heterogeneity, and local temperature and precipitation.
A climate-based mosquito stochastic and epidemiological model was developed to evaluate the epidemic potential of Chikungunya virus in the event of an introduction into the US.
The ecology, behavior, and genetic structure of two invasive mosquitoes, Aedes albopictus and Aedes japonicus were studied. Understanding these ecological relationships is essential to developing economical and effective integrated pest management programs for their control.
New methods for mosquito trapping in the field and virus isolation and identification in the laboratory are is being evaluated to enhance mosquito and arbovirus surveillance programs.
Members are actively participating in graduate student and postdoctoral training programs in vector biology and mosquito and reporting research findings within the scientific community and with the general public.