Ecology and geography of Cache Valley virus assessed using ecological niche modeling.

BACKGROUND: Cache Valley virus (CVV) is an understudied Orthobunyavirus with a high spillover transmission potential due to its wide geographical distribution and large number of associated hosts and vectors. Although CVV is known to be widely distributed throughout North America, no studies have explored its geography or employed computational methods to explore the mammal and mosquito species likely participating in the CVV sylvatic cycle. METHODS: We used a literature review and online databases to compile locality data for CVV and its potential vectors and hosts. We linked location data points with climatic data via ecological niche modeling to estimate the geographical range of CVV and hotspots of transmission risk. We used background similarity tests to identify likely CVV mosquito vectors and mammal hosts to detect ecological signals from CVV sylvatic transmission. RESULTS: CVV distribution maps revealed a widespread potential viral occurrence throughout North America. Ecological niche models identified areas with climate, vectors, and hosts suitable to maintain CVV transmission. Our background similarity tests identified Aedes vexans, Culiseta inornata, and Culex tarsalis as the most likely vectors and Odocoileus virginianus (white-tailed deer) as the most likely host sustaining sylvatic transmission. CONCLUSIONS: CVV has a continental-level, widespread transmission potential. Large areas of North America have suitable climate, vectors, and hosts for CVV emergence, establishment, and spread. We identified geographical hotspots that have no confirmed CVV reports to date and, in view of CVV misdiagnosis or underreporting, can guide future surveillance to specific localities and species.

Broad-spectrum activity against mosquito-borne flaviviruses achieved by a targeted protein degradation mechanism.

Viral genetic diversity presents significant challenges in developing antivirals with broad-spectrum activity and high barriers to resistance. Here we report development of proteolysis targeting chimeras (PROTACs) targeting the dengue virus envelope (E) protein through coupling of known E fusion inhibitors to ligands of the CRL4CRBN E3 ubiquitin ligase. The resulting small molecules block viral entry through inhibition of E-mediated membrane fusion and interfere with viral particle production by depleting intracellular E in infected Huh 7.5 cells. This activity is retained in the presence of point mutations previously shown to confer partial resistance to the parental inhibitors due to decreased inhibitor-binding. The E PROTACs also exhibit broadened spectrum of activity compared to the parental E inhibitors against a panel of mosquito-borne flaviviruses. These findings encourage further exploration of targeted protein degradation as a differentiated and potentially advantageous modality for development of broad-spectrum direct-acting antivirals.

Diversity and molecular characterization of insect-specific flaviviruses in mosquitoes (Diptera: Culicidae) collected in central and northern Argentina.

The genus Flavivirus comprises approximately 80 different viruses. Phylogenetic relationships among its members indicate a clear ecological separation between those viruses transmitted by mosquitoes, ticks, with no known vector, and insect-specific Flaviviruses. The diversity and phylogenetic relationships among insect-specific flaviviruses circulating in the central and northern regions of Argentina were studied by performing molecular detection and characterization of the NS5 protein gene in mosquitoes collected in Córdoba, Chaco and Tucumán provinces. Overall, 68 out of 1776 pools were positive. CxFV, KRV and CFAV circulate in the 3 studied provinces. Several mosquito species (Aedes aegypti, Culex bidens, Cx. dolosus, Cx. interfor, Cx. quinquefasciatus, Cx. saltanensis, Haemagogus spegazzini) were found infected. A wide circulation of CxFV was observed in the central-northern region of Argentina. CxFV strains detected in our study clustered with strains circulating in Santa Fe and Buenos Aires provinces (Argentina), and other countries such as Indonesia, Mexico, Uganda and Taiwan. The presence of these viruses in mosquitoes could play an important role from the public health perspective, because it has been shown that previous CxFV infection can increase or block the infection of the mosquito by other pathogenic flaviviruses.

Ecological Requirements for Abundance and Dispersion of Brazilian Yellow Fever Vectors in Tropical Areas.

In the Americas, wild yellow fever (WYF) is an infectious disease that is highly lethal for some non-human primate species and non-vaccinated people. Specifically, in the Brazilian Atlantic Forest, Haemagogus leucocelaenus and Haemagogus janthinomys mosquitoes act as the major vectors. Despite transmission risk being related to vector densities, little is known about how landscape structure affects vector abundance and movement. To fill these gaps, we used vector abundance data and a model-selection approach to assess how landscape structure affects vector abundance, aiming to identify connecting elements for virus dispersion in the state of São Paulo, Brazil. Our findings show that Hg. leucocelaenus and Hg. janthinomys abundances, in highly degraded and fragmented landscapes, are mainly affected by increases in forest cover at scales of 2.0 and 2.5 km, respectively. Fragmented landscapes provide ecological corridors for vector dispersion, which, along with high vector abundance, promotes the creation of risk areas for WYF virus spread, especially along the border with Minas Gerais state, the upper edges of the Serra do Mar, in the Serra da Cantareira, and in areas of the metropolitan regions of São Paulo and Campinas.

Antiviral Activities of Heparan Sulfate Mimetic RAFT Polymers Against Mosquito-borne Viruses.

Mosquito-borne viruses are a major worldwide health problem associated with high morbidity and mortality rates and significant impacts on national healthcare budgets. The development of antiviral drugs for both the treatment and prophylaxis of these diseases is thus of considerable importance. To address the need for therapeutics with antiviral activity, a library of heparan sulfate mimetic polymers was screened against dengue virus (DENV), Yellow fever virus (YFV), Zika virus (ZIKV), and Ross River virus (RRV). The polymers were prepared by RAFT polymerization of various acidic monomers with a target MW of 20 kDa (average Mn ∼ 27 kDa by GPC). Among the polymers, poly(SS), a homopolymer of sodium styrenesulfonate, was identified as a broad spectrum antiviral with activity against all the tested viruses and particularly potent inhibition of YFV (IC50 = 310 pM). Our results further uncovered that poly(SS) exhibited a robust inhibition of ZIKV infection in both mosquito and human cell lines, which points out the potential functions of poly(SS) in preventing mosquito-borne viruses associated diseases by blocking viral transmission in their mosquito vectors and mitigating viral infection in patients.

The effect of temperature on the boundary conditions of West Nile virus circulation in Europe.

West Nile virus (WNV) is a vector-borne flavivirus that causes an increasing number of human and equine West Nile fever cases in Europe. While the virus has been present in the Mediterranean basin and the Balkans since the 1960s, recent years have witnessed its northward expansion, with the first human cases reported in Germany in 2018 and the Netherlands in 2020. WNV transmission and amplification within mosquitoes are temperature-dependent. This study applies a mathematical modelling approach to assess the conditions under which WNV circulation occurs based on the proportion of mosquito bites on WNV-competent birds (dilution), vector-host ratios, mosquito season length and the observed daily temperature data. We modelled five distinct European regions where previous WNV circulation has been observed within the Netherlands, Germany, Spain, Italy, and Greece. We observed that the number of days in which the basic reproduction number (R0) is above one, increased over the last 40 years in all five regions. In the Netherlands, the number of days in which the R0 is above one, is 70% lower than in Spain. The temperature in Greece, Spain and Italy allowed for circulation under low vector-host ratios, and at a high dilution. On the other hand in the Netherlands and Germany, given the observed daily temperature, the thresholds for circulation requires a lower dilution and higher vector-host ratios. For the Netherlands, a short window of introductions between late May and mid-June would result in detectable outbreaks. Our findings revealed that the temperate maritime climate of the Netherlands allows WNV circulation primarily during warmer summers, and only under high vector-host ratios. This research contributes valuable insights into the dynamic relationship between temperature, vector properties, and WNV transmission, offering guidance for proactive strategies in addressing this emerging health threat in Europe.

Vegetation structure drives mosquito community composition in UK's largest managed lowland wetland.

PURPOSE: The rising burden of mosquito-borne diseases in Europe extends beyond urban areas, encompassing rural and semi-urban regions near managed and natural wetlands evidenced by recent outbreaks of Usutu and West Nile viruses. While wetland management policies focus on biodiversity and ecosystem services, few studies explore the impact on mosquito vectors. METHODS: Our research addresses this gap, examining juvenile mosquito and aquatic predator communities in 67 ditch sites within a South England coastal marsh subjected to different wetland management tiers. Using joint distribution models, we analyse how mosquito communities respond to abiotic and biotic factors influenced by wetland management. RESULTS: Of the 12 mosquito species identified, Culiseta annulata (Usutu virus vector) and Culex pipiens (Usutu and West Nile virus vector) constitute 47% of 6825 larval mosquitoes. Abundant predators include Coleoptera (water beetles) adults, Corixidae (water boatmen) and Zygoptera (Damselfy) larvae. Models reveal that tier 3 management sites (higher winter water levels, lower agricultural intensity) associated with shade and less floating vegetation are preferred by specific mosquito species. All mosquito species except Anopheles maculipennis s.l., are negatively impacted by potential predators. Culiseta annulata shows positive associations with shaded and turbid water, contrary to preferences of Corixidae predators. CONCLUSIONS: Tier 3 areas managed for biodiversity, characterised by higher seasonal water levels and reduced livestock grazing intensity, provide favourable habitats for key mosquito species that are known vectors of arboviruses, such as Usutu and West Nile. Our findings emphasise the impact of biodiversity-focused wetland management, altering mosquito breeding site vegetation to enhance vector suitability. Further exploration of these trade-offs is crucial for comprehending the broader implications of wetland management.

Enhancing Eastern Equine Encephalitis Virus Surveillance in New Jersey: Optimized Collection of Culiseta melanura.

Eastern equine encephalitis virus (EEEV) causes the most clinically severe neuroinvasive arboviral disease in the United States. The virus is endemic in eastern and Gulf Coast states and the Great Lakes region, causing cases annually. To detect EEEV circulation in its enzootic cycle before the virus infects humans and other mammals, mosquito control agencies in New Jersey have conducted mosquito surveillance using a series of permanent wooden resting box sites since 1975. We conducted 2 field studies, 1 evaluating resting traps and 1 evaluating efficacy of CO2 lures, to optimize collection of Culiseta melanura, the primary enzootic vector of EEEV. Resulting mosquito samples were subjected to molecular analysis to determine EEEV infection rates. Corrugated plastic boxes trapped more bloodfed Cs. melanura than other resting trap types (resting boxes, Centers for Disease Control and Prevention [CDC] resting traps, or fiber pots) and were similar to resting boxes in total number of female Cs. melanura caught. Further, non-baited CDC light traps were more successful in trapping host-seeking Cs. melanura than those baited with dry ice, a CO2 lure. The EEEV RNA was identified in Cs. melanura, Aedes vexans, Anopheles quadrimaculatus, and Uranotaenia sapphirina. Our findings indicate that corrugated plastic boxes and non-CO2 baited traps could improve detection of Cs. melanura. Mosquito control agencies are encouraged to periodically assess their surveillance strategy for EEEV.

Host influence on the eukaryotic virome of sympatric mosquitoes and abundance of diverse viruses with a broad host range.

Mosquitoes harbor a large diversity of eukaryotic viruses. Those viromes probably influence mosquito physiology and the transmission of human pathogens. Nevertheless, their ecology remains largely unstudied. Here, we address two key questions in virome ecology. First, we assessed the influence of mosquito species on virome taxonomic diversity and relative abundance. Contrary to most previous studies, the potential effect of the habitat was explicitly included. Thousands of individuals of Culex poicilipes and Culex tritaeniorhynchus, two vectors of viral diseases, were concomitantly sampled in three habitats over two years. A total of 95 viral taxa from 25 families were identified with meta-transcriptomics, with 75% of taxa shared by both mosquitoes. Viromes significantly differed by mosquito species but not by habitat. Differences were largely due to changes in relative abundance of shared taxa. Then, we studied the diversity of viruses with a broad host range. We searched for viral taxa shared by the two Culex species and Aedes vexans, another disease vector, present in one of the habitats. Twenty-six out of the 163 viral taxa were found in the three mosquitoes. These taxa encompassed 14 families. A database analysis supported broad host ranges for many of those viruses, as well as a widespread geographical distribution. Thus, the viromes of mosquitoes from the same genera mainly differed in the relative abundance of shared taxa, whereas differences in viral diversity dominated between mosquito genera. Whether this new model of virome diversity and structure applies to other mosquito communities remains to be determined.

Circulation of West Nile Virus and Usutu Virus in Europe: Overview and Challenges.

West Nile Virus (WNV) and Usutu Virus (USUV) are both neurotropic mosquito-borne viruses belonging to the Flaviviridae family. These closely related viruses mainly follow an enzootic cycle involving mosquitoes as vectors and birds as amplifying hosts, but humans and other mammals can also be infected through mosquito bites. WNV was first identified in Uganda in 1937 and has since spread globally, notably in Europe, causing periodic outbreaks associated with severe cases of neuroinvasive diseases such as meningitis and encephalitis. USUV was initially isolated in 1959 in Swaziland and has also spread to Europe, primarily affecting birds and having a limited impact on human health. There has been a recent expansion of these viruses' geographic range in Europe, facilitated by factors such as climate change, leading to increased human exposure. While sharing similar biological traits, ecology, and epidemiology, there are significant distinctions in their pathogenicity and their impact on both human and animal health. While WNV has been more extensively studied and is a significant public health concern in many regions, USUV has recently been gaining attention due to its emergence in Europe and the diversity of its circulating lineages. Understanding the pathophysiology, ecology, and transmission dynamics of these viruses is important to the implementation of effective surveillance and control measures. This perspective provides a brief overview of the current situation of these two viruses in Europe and outlines the significant challenges that need to be addressed in the coming years.

Identification of Bacteria and Viruses Associated with Patients with Acute Febrile Illness in Khon Kaen Province, Thailand.

The majority of cases of undifferentiated acute febrile illness (AFI) in the tropics have an undefined etiology. In Thailand, AFI accounts for two-thirds of illnesses reported to the Ministry of Public Health. To characterize the bacterial and viral causes of these AFIs, we conducted molecular pathogen screening and serological analyses in patients who sought treatment in Chum Phae Hospital, Khon Kaen province, during the period from 2015 to 2016. Through integrated approaches, we successfully identified the etiology in 25.5% of cases, with dengue virus infection being the most common cause, noted in 17% of the study population, followed by scrub typhus in 3.8% and rickettsioses in 6.8%. Further investigations targeting viruses in patients revealed the presence of Guadeloupe mosquito virus (GMV) in four patients without other pathogen co-infections. The characterization of four complete genome sequences of GMV amplified from AFI patients showed a 93-97% nucleotide sequence identity with GMV previously reported in mosquitoes. Nucleotide substitutions resulted in amino acid differences between GMV amplified from AFI patients and mosquitoes, observed in 37 positions. However, these changes had undergone purifying selection pressure and potentially had a minimal impact on protein function. Our study suggests that the GMV strains identified in the AFI patients are relatively similar to those previously reported in mosquitoes, highlighting their potential role associated with febrile illness.

Vector mosquito distribution and richness are predicted by socio-economic, and ecological variables.

Mosquitoes of vectorial importance represent a ubiquitous and constant threat of potentially devastating arboviral outbreaks. Our ability to predict such outcomes is still restricted. To answer this, we have used an extensive data collection of 23 vector and 233 non-vector mosquito species distributed throughout the Mexican territory and linked them to social and environmental factors. Our aim was to predict vector and non-vector mosquitoes' distribution and species richness based on socioeconomic and environmental data. We found that lack of health services, human population variation, ecological degradation, and urban-rural categorization contributed significantly to explain the distribution of vector mosquitoes. mosquitoes. This phenomenon is probably attributed to the degradation of natural ecosystems as it creates favorable conditions for the proliferation of vector mosquitoes. The richness of vector mosquitoes was similarly explained by most of these variables as well as altitude. As for non-vector mosquitoes, social marginalization, ecological degradation, anthropogenic impact, and altitude explain species richness and distribution. These findings illustrate the complex interaction of environmental and socioeconomic factors behind the distribution of mosquitoes, and the potential for arboviral disease outbreaks. Areas with human populations at highest risk for mosquito-borne diseases should be primary targets for vector control.

Metagenomic analysis of individual mosquito viromes reveals the geographical patterns and drivers of viral diversity.

Mosquito transmitted viruses are responsible for an increasing burden of human disease. Despite this, little is known about the diversity and ecology of viruses within individual mosquito hosts. Here, using a meta-transcriptomic approach, we determined the viromes of 2,438 individual mosquitoes (81 species), spanning ~4,000 km along latitudes and longitudes in China. From these data we identified 393 viral species associated with mosquitoes, including 7 (putative) species of arthropod-borne viruses (that is, arboviruses). We identified potential mosquito species and geographic hotspots of viral diversity and arbovirus occurrence, and demonstrated that the composition of individual mosquito viromes was strongly associated with host phylogeny. Our data revealed a large number of viruses shared among mosquito species or genera, enhancing our understanding of the host specificity of insect-associated viruses. We also detected multiple virus species that were widespread throughout the country, perhaps reflecting long-distance mosquito dispersal. Together, these results greatly expand the known mosquito virome, linked viral diversity at the scale of individual insects to that at a country-wide scale, and offered unique insights into the biogeography and diversity of viruses in insect vectors.

Impact of symbiotic insect-specific viruses on mosquito vector competence for arboviruses.

Mosquitoes are vectors for arboviruses, such as dengue, Zika, and Chikungunya. Symbiotic interactions can affect the intrinsic ability of mosquitoes to acquire and transmit arboviruses, referred to as vector competence. Insect-specific viruses (ISVs) are commonly found in symbiotic associations with mosquitoes in the wild and can affect many aspects of mosquito biology. Here, we review current knowledge on the effects of symbiotic ISV-mosquito interactions on vector competence. We discuss potential mechanisms underlying these interactions and their implications for shaping new biological control strategies. Finally, we highlight the need for field data analyzing the circulation of ISVs in mosquitoes associated with mechanistic studies in the laboratory.

Multiplex PCR method for MinION sequencing of Bagaza virus isolated from wild caught mosquitoes in South Africa.

Bagaza virus (BAGV) is a mosquito-borne orthoflavivirus known to occur in regions of southern Europe, Africa, India and the Middle East. The virus has been associated with neurological disease and fatalities in various wild bird species. Association with human disease is not confirmed although limited serological evidence has suggested human infection. Surveillance programs for screening mosquitoes for evidence of arbovirus infection play an important role in providing information regarding the circulation and spread of viruses in specific regions. BAGV was detected in a mosquito pool during surveillance of mosquitoes collected in central South Africa between November 2019 and March 2023. Homogenized mosquito pools were screened for flaviviral RNA using conventional RT-PCR and virus isolation was attempted on positive samples. BAGV was detected and subsequently isolated using cell culture. A multiplex tiling PCR method for targeted enrichment using a PCR based or amplicon sequencing approach of the complete genome of BAGV was developed and optimized. Primers were designed using alignment of complete genome sequence data retrieved from GenBank to identify suitable primer sites that would generate overlapping fragments spanning the complete genome. Six forward primers and eight reverse primers were identified that target the complete genome and amplified nine overlapping fragments, that ranged in length from 1954 to 2039 with an overlap ranging from 71 to 711 base pairs. The design strategy included multiple forward and reverse primer pairs for the 5' and 3' ends. Phylogenetic analysis with other isolates was performed and BAGV isolate VBD 74/23/3 was shown to share high similarity with previous BAGV isolates from all regions, with genetic distance ranging from 0.026 to 0.083. VBD 74/23/3 was most closely related to previous isolates from southern Africa, ZRU96/16/2 isolated from a post-mortem sample from a pheasant in 2016 and MP-314-NA-2018 isolated from mosquitoes in northwestern Namibia with genetic distance 0.0085 and 0.016 respectively. Currently there is limited complete genome sequence data available for many of the arboviruses circulating in Africa. The multiplex tiling method provided a simple and cost-effective method for obtaining complete genome sequence. This method can be readily applied to other viruses using sequence data from publicly available databases and would have important application facilitating genomic surveillance of arboviruses in low resource countries.

Screening and identification of persistent viruses in cell lines derived from medically important arthropods.

Arthropod-derived cell lines serve as crucial tools for studying arthropod-borne viruses (arboviruses). However, it has recently come to light that certain cell lines harbor persistent infections of arthropod-specific viruses, which do not cause any apparent cytopathic effects. Moreover, some of these persistent viral infections either inhibit or promote the growth of arboviruses. Therefore, it is of utmost importance to identify the presence of such persistent viruses and understand their impact on arboviral infections. In this study, we conducted a comprehensive virome analysis of several arthropod-derived cell lines, including mosquito-derived NIID-CTR, Ar-3, MSQ43, NIAS-AeAl-2, CCL-126 cells, and tick-derived IDE8 cells, along with flesh fly-derived NIH-Sape-4 cells. The aim was to determine if these cells were infected with persistent viruses. The results revealed the presence of 15 persistent viruses in NIID-CTR, Ar-3, MSQ43, NIAS-AeAl-2, and IDE8 cells. Among these, 11 were already known arthropod-specific viruses, while the remaining 4 were novel viruses belonging to Orthophasmavirus, Rhabdoviridae, Totiviridae, and Bunyavirales. In contrast, CCL-126 and NIH-Sape-4 cells appeared to be free of viral infections. This study provides valuable insights into the diversity and latency of arthropod-specific viruses within arthropod-derived cell lines. Further investigations are required to explore persistent viral infections in other arthropod-derived cell cultures and their effects on arbovirus replication. Understanding these factors will enhance the accuracy and reliability of experimental data obtained using these cell lines.