Dynamics of Vector-Host Interactions in Avian Communities in Four Eastern Equine Encephalitis Virus Foci in the Northeastern US

Background: Eastern equine encephalitis (EEE) virus (Togaviridae, Alphavirus) is a highly pathogenic mosquito-borne zoonosis that is responsible for occasional outbreaks of severe disease in humans and equines, resulting in high mortality and neurological impairment in most survivors. In the past, human disease outbreaks in the northeastern U.S. have occurred intermittently with no apparent pattern; however, during the last decade we have witnessed recurring annual emergence where EEE virus activity had been historically rare, and expansion into northern New England where the virus had been previously unknown. In the northeastern U.S., EEE virus is maintained in an enzootic cycle involving the ornithophagic mosquito, Culiseta melanura, and wild passerine (perching) birds in freshwater hardwood swamps. However, the identity of key avian species that serve as principal virus reservoir and amplification hosts has not been established. The efficiency with which pathogen transmission occurs within an avian community is largely determined by the relative reservoir competence of each species and by ecological factors that influence contact rates between these avian hosts and mosquito vectors. Methodology and principle findings: Contacts between vector mosquitoes and potential avian hosts may be directly quantified by analyzing the blood meal contents of field-collected specimens. We used PCR-based molecular methods and direct sequencing of the mitochondrial cytochrome b gene for profiling of blood meals in Cs. melanura, in an effort to quantify its feeding behavior on specific vertebrate hosts, and to infer epidemiologic implications in four historic EEE virus foci in the northeastern U.S. Avian point count surveys were conducted to determine spatiotemporal host community composition. Of 1,127 blood meals successfully identified to species level, >99% of blood meals were from 65 avian hosts in 27 families and 11 orders, and only seven were from mammalian hosts representing three species. We developed an empirically informed mathematical model for EEE virus transmission using Cs. melanura abundance and preferred and non-preferred avian hosts. To our knowledge this is the first mathematical model for EEE virus, a pathogen with many potential hosts, in the northeastern U.S. We measured strong feeding preferences for a number of avian species based on the proportion of mosquito blood meals identified from these bird species in relation to their observed frequencies. These included: American Robin, Tufted Titmouse, Common Grackle, Wood Thrush, Chipping Sparrow, Black-capped Chickadee, Northern Cardinal, and Warbling Vireo. We found that these bird species, most notably Wood Thrush, play a dominant role in supporting EEE virus amplification. It is also noteworthy that the competence of some of the aforementioned avian species for EEE virus has not been established. Our findings indicate that heterogeneity induced by mosquito host preference, is a key mediator of the epizootic transmission of vector-borne pathogens. Conclusion and significance: Detailed knowledge of the vector-host interactions of mosquito populations in nature is essential for evaluating their vectorial capacity and for assessing the role of individual vertebrates as reservoir hosts involved in the maintenance and amplification of zoonotic agents of human diseases. Our study clarifies the host associations of Cs. melanura in four EEE virus foci in the northeastern U.S., identifies vector host preferences as the most important transmission parameter, and quantifies the contribution of preference-induced contact heterogeneity to enzootic transmission. Our study identifies Wood Thrush, American Robin and a few avian species that may serve as superspreaders of EEE virus. Our study elucidates spatiotemporal host species utilization by Cs. melanura in relation to avian host community. This research provides a basis to better understand the involvement of Cs. melanura and avian hosts in the transmission and ecology of EEE virus and the risk of human infection in virus foci

Host Feeding Patterns of Culex Mosquitoes and West Nile Virus Transmission, Northeastern United States

Culex salinarius is a bridge vector to humans, while Cx. pipiens and Cx. restuans are more efficient enzootic vectors

Population genomics of Culiseta melanura, the principal vector of Eastern equine encephalitis virus in the United States.

BACKGROUND:Eastern Equine Encephalitis (EEE) (Togaviridae, Alphavirus) is a highly pathogenic mosquito-borne arbovirus that circulates in an enzootic cycle involving Culiseta melanura mosquitoes and wild Passeriformes birds in freshwater swamp habitats. Recently, the northeastern United States has experienced an intensification of virus activity with increased human involvement and northward expansion into new regions. In addition to its principal role in enzootic transmission of EEE virus among avian hosts, recent studies on the blood-feeding behavior of Cs. melanura throughout its geographic range suggest that this mosquito may also be involved in epizootic / epidemic transmission to equines and humans in certain locales. Variations in blood feeding behavior may be a function of host availability, environmental factors, and/or underlying genetic differences among regional populations. Despite the importance of Cs. melanura in transmission and maintenance of EEE virus, the genetics of this species remains largely unexplored. METHODOLOGY AND PRINCIPLE FINDINGS:To investigate the occurrence of genetic variation in Cs. melanura, the genome of this mosquito vector was sequenced resulting in a draft genome assembly of 1.28 gigabases with a contig N50 of 93.36 kilobases. Populations of Cs. melanura from 10 EEE virus foci in the eastern North America were genotyped with double-digest RAD-seq. Following alignment of reads to the reference genome, variant calling, and filtering, 40,384 SNPs were retained for downstream analyses. Subsequent analyses revealed genetic differentiation between northern and southern populations of this mosquito species. Moreover, limited fine-scale population structure was detected throughout northeastern North America, suggesting local differentiation of populations but also a history of ancestral polymorphism or contemporary gene flow. Additionally, a genetically distinct cluster was identified predominantly at two northern sites. CONCLUSION AND SIGNIFICANCE:This study elucidates the first evidence of fine-scale population structure in Cs. melanura throughout its eastern range and detects evidence of gene flow between populations in northeastern North America. This investigation provides the groundwork for examining the consequences of genetic variations in the populations of this mosquito species that could influence vector-host interactions and the risk of human and equine infection with EEE virus

SNP dataset

This is the primary SNP dataset for Soghigian et al. and contains more than 40,000 SNPs used in the analysis of eastern populations of <i>Culiseta mealnura</i>. These SNPs were called from a draft genome developed for the project and described in Soghigian et al. using the Stacks parameter r set to 0.75, requiring loci be present in at least 75% of samples

Insights into the recent emergence and expansion of eastern equine encephalitis virus in a new focus in the Northern New England USA

Abstract Background Eastern equine encephalomyelitis virus (EEEV) causes a highly pathogenic zoonosis that circulates in an enzootic cycle involving the ornithophagic mosquito, Culiseta melanura, and wild passerine birds in freshwater hardwood swamps in the northeastern U.S. Epidemic/epizootic transmission to humans/equines typically occurs towards the end of the transmission season and is generally assumed to be mediated by locally abundant and contiguous mammalophagic “bridge vector” mosquitoes. Methods Engorged mosquitoes were collected using CDC light, resting box, and gravid traps during epidemic transmission of EEEV in 2012 in Addison and Rutland counties, Vermont. Mosquitoes were identified to species and blood meal analysis performed by sequencing mitochondrial cytochrome b gene polymerase chain reaction products. Infection status with EEEV in mosquitoes was determined using cell culture and RT-PCR assays, and all viral isolates were sequenced and compared to other EEEV strains by phylogenetic analysis. Results The host choices of 574 engorged mosquitoes were as follows: Cs. melanura (n = 331, 94.3 % avian-derived, 5.7 % mammalian-derived); Anopheles quadrimaculatus (n = 164, 3.0 % avian, 97.0 % mammalian); An. punctipennis (n = 56, 7.2 % avian, 92.8 % mammalian), Aedes vexans (n = 9, 22.2 % avian, 77.8 % mammalian); Culex pipiens s.l. n = 6, 100 % avian); Coquillettidia perturbans (n = 4, 25.0 % avian, 75.0 % mammalian); and Cs. morsitans (n = 4, 100 % avian). A seasonal shift in blood feeding by Cs. melanura from Green Heron towards other avian species was observed. EEEV was successfully isolated from blood-fed Cs. melanura and analyzed by phylogenetic analysis. Vermont strains from 2012 clustered with viral strains previously isolated in Virginia yet were genetically distinct from an earlier EEEV isolate from Vermont during 2011. Conclusions Culiseta melanura acquired blood meals primarily from birds and focused feeding activity on several competent species capable of supporting EEEV transmission. Culiseta melanura also occasionally obtained blood meals from mammalian hosts including humans. This mosquito species serves as the primary vector of EEEV among wild bird species, but also is capable of occasionally contributing to epidemic/epizootic transmission of EEEV to humans/equines. Other mosquito species including Cq. perturbans that feed more opportunistically on both avian and mammalian hosts may be important in epidemic/epizootic transmission under certain conditions. Phylogenetic analyses suggest that EEEV was independently introduced into Vermont on at least two separate occasions

Enhancement of Risk for Lyme Disease by Landscape Connectivity, New York, New York, USA

Most tickborne disease studies in the United States are conducted in low-intensity residential development and forested areas, leaving much unknown about urban infection risks. To understand Lyme disease risk in New York, New York, USA, we conducted tick surveys in 24 parks throughout all 5 boroughs and assessed how park connectivity and landscape composition contribute to Ixodes scapularis tick nymphal densities and Borrelia burgdorferi infection. We used circuit theory models to determine how parks differentially maintain landscape connectivity for white-tailed deer, the reproductive host for I. scapularis ticks. We found forested parks with vegetated buffers and increased connectivity had higher nymph densities, and the degree of park connectivity strongly determined B. burgdorferi nymphal infection prevalence. Our study challenges the perspective that tickborne disease risk is restricted to suburban and natural settings and emphasizes the need to understand how green space design affects vector and host communities in areas of emerging urban tickborne disease

Vector-Host Interactions of Culiseta melanura in a Focus of Eastern Equine Encephalitis Virus Activity in Southeastern Virginia.

Eastern equine encephalitis virus (EEEV) causes a highly pathogenic mosquito-borne zoonosis that is responsible for sporadic outbreaks of severe illness in humans and equines in the eastern USA. Culiseta (Cs.) melanura is the primary vector of EEEV in most geographic regions but its feeding patterns on specific avian and mammalian hosts are largely unknown in the mid-Atlantic region. The objectives of our study were to: 1) identify avian hosts of Cs. melanura and evaluate their potential role in enzootic amplification of EEEV, 2) assess spatial and temporal patterns of virus activity during a season of intense virus transmission, and 3) investigate the potential role of Cs. melanura in epidemic/epizootic transmission of EEEV to humans and equines. Accordingly, we collected mosquitoes at 55 sites in Suffolk, Virginia in 2013, and identified the source of blood meals in engorged mosquitoes by nucleotide sequencing PCR products of the mitochondrial cytochrome b gene. We also examined field-collected mosquitoes for evidence of infection with EEEV using Vector Test, cell culture, and PCR. Analysis of 188 engorged Cs. melanura sampled from April through October 2013 indicated that 95.2%, 4.3%, and 0.5% obtained blood meals from avian, mammalian, and reptilian hosts, respectively. American Robin was the most frequently identified host for Cs. melanura (42.6% of blood meals) followed by Northern Cardinal (16.0%), European Starling (11.2%), Carolina Wren (4.3%), and Common Grackle (4.3%). EEEV was detected in 106 mosquito pools of Cs. melanura, and the number of virus positive pools peaked in late July with 22 positive pools and a Maximum Likelihood Estimation (MLE) infection rate of 4.46 per 1,000 mosquitoes. Our findings highlight the importance of Cs. melanura as a regional EEEV vector based on frequent feeding on virus-competent bird species. A small proportion of blood meals acquired from mammalian hosts suggests the possibility that this species may occasionally contribute to epidemic/epizootic transmission of EEEV