Usutu Virus Research Articles

Usutu virus-induced meningoencephalitis in immunocompetent mice is characterized by the recruitment of mononuclear cells and a proinflammatory T helper 1 response.

Usutu virus (USUV) is an arbovirus and has emerged as a potential cause of encephalitis in humans and other vertebrates. The increasing detection of USUV in mosquitoes and birds across Africa and Central Europe, along with the lack of specific treatments or vaccines for many encephalitic orthoflaviviruses, underscores the need for focused research. In this study, we developed a USUV infection model in immunocompetent C57BL/6 mice (8-12 weeks old) to characterize disease development and associated inflammatory mechanisms. Mice were intracranially infected with 104 PFU of USUV, leading to neurological symptoms such as hunched posture, paralysis, conjunctivitis, and eventual death by day 6 post-infection. Meningeal cell infiltration and microglia activation were most prevalent in mouse brains; however, neuronal loss was not observed at the peak of the disease, which coincided with increased viral load and leukocyte infiltration. The immune response in the brain was marked by the systematic recruitment and activation of macrophages, neutrophils, and T lymphocytes. A noticeable shift was seen in CD4+ T cells toward T helper 1 (Th1) polarization, which corroborates a massive increase in the expression of Th1-associated cytokines and chemokines at the peak of infection, indicative of an augmented proinflammatory state. Additionally, a rise in regulatory T cells was observed, peaking on day 6 post-infection. These findings highlight the dynamic nature of the host response to USUV infection, enhance our understanding of the disease pathogenesis, and address the scarcity of immunocompetent experimental models for the investigation of neglected emerging flaviviruses.IMPORTANCEMosquito-borne viruses, including USUV, are maintained in nature through complex cycles involving arthropod vectors and vertebrate hosts. A comprehensive understanding of USUV biology and host-pathogen interactions is crucial for developing effective treatments, which necessitates reliable experimental models (G. J. Sips, J. Wilschut, and J. M. Smit, Rev Med Virol 22:69-87, 2012, https://doi.org/10.1002/rmv.712; T. C. Pierson and M. S. Diamond, Nat Microbiol 5:796-812, 2020, https://doi.org/10.1038/s41564-020-0714-0). The establishment of a USUV infection model in immunocompetent adult mice brings new perspectives on the inflammatory component of viral encephalitis, which is difficult to study in mice lacking antiviral interferon responses. Moreover, USUV is an emerging viral disease lacking therapeutic and preventive measures. The interplay of USUV pathogenesis and the host's immune response indicates that lymphocytes and monocytes participate in USUV infection in this model and could be explored in search of treatments targeting immunopathogenic processes triggered by infection.

Detection of Bagaza Virus in Europe: A Scoping Review

The Bagaza virus (BAGV) belongs to the genus Orthoflavivirus (Ntaya serocomplex) and emerged in Europe, Spain, in 2010. The natural transmission cycle of this virus is perpetuated by Culex spp. mosquitoes and viraemic birds. The ability of BAGV to cause infection in several game birds from the family Phasianidae has been well-studied. BAGV is antigenically similar to other orthoflaviviruses from the Japanese encephalitis serocomplex, such as the West Nile and Usutu viruses, a circumstance which can lead to cross-reactivity in less specific serological techniques (e.g., ELISA). Severe implications in animal health has already been described, but some aspects of the dynamics of transmission and the limits of zoonotic potential of BAGV still need to be clarified. Further investigation focused on epidemiological surveillance in high-risk areas would be beneficial for prevention and control of new outbreaks. The present study is a systematic review of the BAGV reports in Europe.

Dose and strain dependent lethality of Usutu virus in an Ifnar−/− mouse model

Usutu virus (USUV) is a mosquito-borne zoonotic flavivirus with a geographic range that has expanded over recent years. Maintained in a transmission cycle between mosquito vectors and avian reservoirs the virus can cause large seasonal outbreaks in bird populations, but spillover into mammalian hosts has also been reported. While usually mild or asymptomatic in humans, neurological disorders are increasingly observed, which has boosted interest and the need for better understanding of the pathogenesis of various USUV lineages. In this study we inoculated interferon α/β receptor knockout (Ifnar−/−) mice with decreasing doses of USUV, monitoring symptoms and survival to determine a less lethal dose, and we directly compared isolates from three different viral lineages. We found that a Dutch isolate of USUV Africa-3 lineage is lethal at a dose of 20 pfu per mouse, which is considerably lower than what was anticipated based upon the literature. A Europe-2 strain showed an even higher virulence in this mouse model, compared to strains from Africa-3 and Europe-3 lineages—though this was not reflected in in vitro studies. These results enhance our understanding of the pathogenicity of different USUV strains and provide guidance for the use of low doses for inoculation in an Ifnar−/− animal model.

West Nile and Usutu viruses: current spreading and future threats in a warming northern Europe

Climate change heavily threatens planetary and human health. Arboviral infections are best studied using the One Health concept, due to their reliance on complex interactions between environmental factors, arthropod vectors and vertebrate hosts. This review focuses on two arboviruses, namely West Nile Virus (WNV) and Usutu Virus (USUV), both causing emerging public health issues in northern Europe. They are both maintained in an enzootic cycle involving birds and Culex spp mosquitoes. WNV has demonstrated its sensitivity to the consequences of climate change and there is already evidence that global warming contributes to its expansion in Europe. Future WNV indigenous transmission in northern Europe is therefore plausible. Usutu is a lesser known arbovirosis, sharing similar vectors and hosts as WNV. USUV has a similar geographic expansion to WNV, exhibiting some level of co-circulation. It is therefore crucial to monitor these viruses in the hitherto relatively spared regions of northern Europe.

Relevance of the synergy of surveillance and populational networks in understanding the Usutu virus outbreak within common blackbirds (Turdus merula) in Metropolitan France, 2018

Relevance of the synergy of surveillance and populational networks in understanding the Usutu virus outbreak within common blackbirds (Turdus merula) in Metropolitan France, 2018

Generation of Cas9 Knock-In Culex quinquefasciatus Mosquito Cells

Background/Objectives: Culex species mosquitoes are globally distributed and transmit several pathogens that impact animal and public health, including West Nile virus, Usutu virus, and Plasmodium relictum. Despite their relevance, Culex species are less widely studied than Aedes and Anopheles mosquitoes. To expand the genetic tools used to study Culex mosquitoes, we previously developed an optimized plasmid for transient Cas9 and single-guide RNA (sgRNA) expression in Culex quinquefasciatus cells to generate gene knockouts. Here, we established a monoclonal cell line that consistently expresses Cas9 and can be used for screens to determine gene function or antiviral activity. Methods: We used this system to perform the successful gene editing of seven genes and subsequent testing for potential antiviral effects, using a simple single-guide RNA (sgRNA) transfection and subsequent virus infection. Results: We were able to show antiviral effects for the Cx. quinquefasciatus genes dicer-2, argonaute-2b, vago, piwi5, piwi6a, and cullin4a. In comparison to the RNAi-mediated gene silencing of dicer-2, argonaute-2b, and piwi5, our Cas9/sgRNA approach showed an enhanced ability to detect antiviral effects. Conclusions: We propose that this cell line offers a new tool for studying gene function in Cx. quinquefasciatus mosquitoes that avoids the use of RNAi. This short study also serves as a proof-of-concept for future gene knock-ins in these cells. Our cell line expands the molecular resources available for vector competence research and will support the design of future research strategies to reduce the transmission of mosquito-borne diseases.

Molecular Xenomonitoring (MX) allows real-time surveillance of West Nile and Usutu virus in mosquito populations.

West Nile Virus (WNV) and Usutu virus (USUV) circulate through complex cryptic transmission cycles involving mosquitoes as vectors, birds as amplifying hosts and several mammal species as dead-end hosts. Both viruses can be transmitted to humans through mosquito bites, which can lead to neuroinvasive and potentially fatal disease. Notably, WNV can also be transmitted through blood donations and organ transplants. The high proportion of asymptomatic infections caused by these viruses and their cryptic enzootic circulation make their early detection in the environment challenging. Viral surveillance in France still heavily relies on human and animal surveillance, i.e. late indicators of viral circulation. Entomological surveillance is a method of choice for identifying virus circulation ahead of the first human and animal cases and to reveal their genetic identity, but performing molecular screening of vectors is expensive, and time-consuming. Here we show substantial WNV and USUV co-circulation in Atlantic seaboard of France between July and August 2023 using a non-invasive MX (Molecular Xenomonitoring) method that use trapped mosquito excreta. MX offers significant advantages over traditional entomological surveillance: it is cost-effective and efficient, enabling viral RNA screening from a community of trapped mosquitoes via their excreta, which can be transported at room temperature. Additionally, MX extends the longevity of trapped mosquitoes, enhancing virus detection and simplifying logistics, and is easy to implement without requiring specialized skills. At the crossroads between entomological and environmental surveillance, MX can detect the circulation of zoonotic pathogens in the environment before cases are observed in humans and horses, enabling the timely alerts to health policy makers, allowing them to take suitable control measures.

Evaluation of Protocols for DNA Extraction from Individual Culex pipiens to Assess Pyrethroid Resistance Using Genotyping Real-Time Polymerase Chain Reaction.

Culex pipiens is a major vector of pathogens, including West Nile and Usutu viruses, that poses a significant public health risk. Monitoring pyrethroid resistance in mosquito populations is essential for effective vector control. This study aims to evaluate four DNA extraction protocols-QIAsymphony, DNAzol® Direct reagent, PrepMan® Ultra Sample Preparation Reagent (USPR), and Chelex® 100-to identify an optimal method to extract DNA from individual Culex pipiens, as part of a high-throughput surveillance of pyrethroid resistance using Real-Time Genotyping PCR. The target is the L1014F mutation in the voltage-sensitive sodium channel (VSSC) gene, which confers knockdown (kdr) resistance to pyrethroids. Mosquitoes were collected from wintering and summer habitats in Lazio and Tuscany, Italy, and DNA was extracted using the four methods. The quality, quantity, extraction time, and cost of the DNA were compared among the various methods. The PrepMan® USPR protocol was the most efficient, providing high-quality DNA with a 260/280 purity ratio within the optimal range at the lowest cost and in a short time. This method also demonstrated the highest amplification success rate (77%) in subsequent real-time PCR assays, making it the preferred protocol for large-scale genotyping studies.

Exposure to West Nile Virus in Wild Lagomorphs in Spanish Mediterranean Ecosystems.

West Nile virus (WNV) is the most widely distributed mosquito-borne flavivirus. Over the past decade, its spread across Europe has raised significant concerns for both public and animal health. Although WNV exposure has been evidenced in various wild mammal species in Spain, no seroepidemiological studies have been conducted on this flavivirus in wild lagomorphs so far. This study aimed to assess WNV exposure in European wild rabbit (Oryctolagus cuniculus) and Iberian hare (Lepus granatensis) populations inhabiting Spanish Mediterranean ecosystems. Sera from 540 wild lagomorphs (399 European wild rabbit and 141 Iberian hares), from 106 hunting grounds distributed throughout Andalusia (southern Spain), were collected between the 2018/2019 and 2022/2023 hunting seasons. Antibodies against flavivirus were detected by blocking enzyme-linked immunosorbent assay (bELISA) in 5.0% (27/540; 95% CI: 3.2-6.8) of the wild lagomorphs. Exposure to WNV was confirmed in 4.8% (19/394; 95% CI: 2.7-6.9) of wild rabbits and 0.7% (1/141; 95% CI: 0.0-2.1) of Iberian hares by virus microneutralisation test. Anti-WNV antibodies were found in wild lagomorphs sampled from three (2.8%) hunting grounds located in western Andalusia during the seasons 2020-2021 and 2021-2022. Remarkably, this spatiotemporal distribution overlaps with the largest outbreak of WNV in Spain. Antibodies against Usutu virus and Bagaza virus were not detected in the wild lagomorph populations analysed. This study constitutes the first report of WNV exposure in wild rabbit in Spain and in Iberian hare worldwide. While these species seem not play a primary role in the epidemiology of the virus, they could serve as sentinel for monitoring WNV in Iberian Mediterranean ecosystems.

Emerging mosquito-borne flaviviruses.

Flaviviruses comprise a genus of enveloped, positive-sense, single-stranded RNA viruses typically transmitted between susceptible and permissive hosts by arthropod vectors. Established flavivirus threats include dengue viruses (DENV), yellow fever virus (YFV), Zika virus (ZIKV), and West Nile virus (WNV), which continue to cause over 400 million infections annually and are significant global health and economic burdens. Additionally, numerous closely related but largely understudied viruses circulate in animals and can conceivably emerge in human populations. Previous flaviviruses that were recognized to have this potential include ZIKV and WNV, which only became extensively studied after causing major outbreaks in humans. More than 50 species exist within the flavivirus genus, which can be further classified as mosquito-borne, tick-borne, insect-specific, or with no known vector. Historically, many of these flaviviruses originated in Africa and have mainly affected tropical and subtropical regions due to the ecological niche of mosquitoes. However, climate change, as well as vector and host migration, has contributed to geographical expansion, thereby posing a potential risk to global populations. For the purposes of this minireview, we focus on the mosquito-borne subgroup and highlight viruses that cause significant pathology or lethality in at least one animal species and/or have demonstrated an ability to infect humans. We discuss current knowledge of these viruses, existing animal models to study their pathogenesis, and potential future directions. Emerging viruses discussed include Usutu virus (USUV), Wesselsbron virus (WSLV), Spondweni virus (SPOV), Ilheus virus (ILHV), Rocio virus (ROCV), Murray Valley encephalitis virus (MVEV), and Alfuy virus (ALFV).

How Usutu virus resists ISG20-mediated restriction

How Usutu virus resists ISG20-mediated restriction

Circulation of West Nile virus and Usutu virus in birds in Germany, 2021 and 2022

Background Usutu virus (USUV) and West Nile virus (WNV) are zoonotic arthropod-borne orthoflaviviruses. The enzootic transmission cycles of both include Culex mosquitoes as vectors and birds as amplifying hosts. For more than 10 years, these viruses have been monitored in birds in Germany by a multidisciplinary network. While USUV is present nationwide, WNV used to be restricted to the central-east. Methods In 2021 and 2022, over 2300 live bird blood samples and organs from over 3000 deceased birds were subjected to molecular and serological analysis regarding the presence of WNV and USUV. The samples were collected at sites all over Germany. Results Circulation of both viruses increased in 2022. For USUV, the nationwide presence of lineages Africa 3 and Europe 3 reported in previous years was confirmed. Lineage Europe 2, formerly restricted to the German east, was able to expand westward. Nonetheless, USUV neutralizing antibody (nAb) detection rates remained low (< 9%). Years 2021 and 2022 were characterized by stable enzootic circulation of WNV lineage 2, dominated by one previously identified subcluster (95% of generated sequences). In 2022, >20% of birds in the endemic region in eastern Germany carried nAb against WNV. Serological data also indicate expanding WNV circulation west and south of the known hotspots in Germany. Conclusions USUV circulates enzootically nationwide. Emergence of WNV at several new locations in Germany with a potential increase in human infections may be imminent. In this context, wild bird monitoring serves as a capable early warning system in a One Health setting.

First evidence of circulation of multiple arboviruses in Algeria.

Algeria like other North African countries is experiencing recurrent episodes of West Nile Virus (WNV) emergences and new health threats associated with the introduction of Aedes albopictus in 2010 are to be feared. To improve the surveillance of mosquito-borne pathogens, we performed a study using innovative tools based on multiplex molecular methods. We combined two approaches: a high-throughput chip based on the BioMark Dynamic array system to detect arboviruses in mosquitoes, and a set of immunologic methods (ELISA, microsphere immunoassays (MIA) and virus microneutralization tests (MNT)) for serological surveys in animal hosts. We investigated two distinct regions: a first zone located in the coastal humid region and a second one in the Saharan desert region. We collected a total of 1,658 mosquitoes belonging to nine different species and found predominantly Culex pipienss. l. (56.5%) and Cx. perexiguus (27.5%). From 180 pools of 10 mosquitoes, we detected four arboviruses: Banna virus (BAV), chikungunya virus (CHIKV), Sindbis virus (SINV), and Usutu virus (USUV). Moreover, we examined 389 blood samples from equids and poultry and found that 52.4% were positive for flavivirus antibodies in ELISA, while 30.8% were positive for WNV and two chickens and two equids were positive for USUV by MNT and MIA respectively. To our knowledge, this is the first report of five arboviruses circulating in Algeria, with three reported for the first time (CHIKV, BAV, and USUV). Our study brings evidence that reinforcing surveillance using more discriminant tools may help in anticipating future emergences and propose adapted control measures.

Experimental arboviral infection of mosquito larvae: A novel approach for vector competence studies

Vector competence studies in mosquitoes present valuable opportunities to explore arboviral transmission and virus-vector interactions. However, oral infection studies in mosquitoes can be challenging. An alternative approach is to infect mosquitoes during their aquatic larval stage, resulting in the emergence of infected adults. To investigate the potential of this method, Culex pipiens biotype molestus larvae were infected with Usutu virus (USUV, Orthoflavivirus usutuense). For this purpose, larvae were exposed to USUV-infected mammalian and mosquito cell cultures for 24 h before being reared to adults. Subsequent analysis via RT-qPCR revealed that the Culex larvae successfully acquired USUV from the infected cells and exhibited high susceptibility to infection. Immediately after emergence, 32.10 % (26/81) of male and 41.03 % (16/39) of female mosquitoes tested positive for USUV RNA. Notably, females that were incubated for 15 days post-emergence demonstrated even higher infection rates, reaching 100.00 % (23/23). In addition, viral RNA and infectious particles were detected in some saliva samples, indicating the potential for transmission. This experimental infection of mosquito larvae thus offers the opportunity to produce infected adult mosquitoes for studies enhancing our understanding of virus-vector interactions, co-infections, and transmission routes. Such research contributes to better public health strategies addressing arboviral diseases.

Auto-Abs neutralizing type I IFNs in patients with severe Powassan, Usutu, or Ross River virus disease.

Arboviral diseases are a growing global health concern. Pre-existing autoantibodies (auto-Abs) neutralizing type I interferons (IFNs) can underlie encephalitis due to West Nile virus (WNV) (∼40% of patients) and tick-borne encephalitis (TBE, due to TBE virus [TBEV]) (∼10%). We report here that these auto-Abs can also underlie severe forms of rarer arboviral infections. Auto-Abs neutralizing high concentrations of IFN-α2, IFN-β, and/or IFN-ω are present in the single case of severe Powassan virus (POWV) encephalitis studied, two of three cases of severe Usutu virus (USUV) infection studied, and the most severe of 24 cases of Ross River virus (RRV) disease studied. These auto-Abs are not found in any of the 137 individuals with silent or mild infections with these three viruses. Thus, auto-Abs neutralizing type I IFNs underlie an increasing list of severe arboviral diseases due to Flaviviridae (WNV, TBEV, POWV, USUV) or Togaviridae (RRV) viruses transmitted to humans by mosquitos (WNV, USUV, RRV) or ticks (TBEV, POWV).

The Re-Emergence of Neuroinvasive Flaviviruses in Croatia During the 2022 Transmission Season.

(Re-)emerging arboviruses, such as tick-borne encephalitis virus (TBEV), West Nile virus (WNV), and Usutu virus (USUV), are continuously increasing in incidence. We analyzed the epidemiological characteristics of flavivirus infections in humans, sentinel animals, and mosquitoes detected in the 2022 transmission season in Croatia. From April to November 2022, 110 hospitalized patients with neuroinvasive diseases (NID) were tested for the presence of arboviruses. RT-qPCR was used to detect TBEV, WNV, and USUV RNA. An ELISA and virus neutralization tests were used for the detection of flavivirus antibodies. TBEV infection was confirmed in 22 patients with NID. WNV NID was detected in six patients. TBE showed male predominance (81.8%; male-to-female ratio of 4.5:1). All but one WNV patients were males. TBE occurred from April to August, with the majority of patients (83.3%) being detected during the May-June-July period. WNV infections were recorded in August and September. In addition to human cases, asymptomatic WNV infections (IgM positive) were reported in 10 horses. For the first time in Croatia, WNV NID was observed in one horse that presented with neurological symptoms. Furthermore, USUV was confirmed in one dead blackbird that presented with neurological symptoms. A total of 1984 mosquitoes were collected in the City of Zagreb. Two Ae. albopictus pools tested positive for flavivirus RNA: one collected in July (USUV) and the other collected in August (WNV). A phylogenetic analysis of detected human and avian strains confirmed WNV lineage 2 and the USUV Europe 2 lineage. The presented results confirm the endemic presence of neuroinvasive flaviviruses in continental Croatia. The continuous monitoring of virus circulation in humans, sentinel animals, and mosquitoes is needed to reduce the disease burden.

Serological studies on the presence of antibodies against West Nile virus in wild birds of the order Passeriformes in Ukraine

West Nile fever is a very dangerous zoonotic viral disease of animals and humans. It is a naturally occurring focal disease, the natural cycle of which involves a natural reservoir of pathogens, such as wild birds, and vectors, such as mosquitoes, ticks, etc. Today, the problem of West Nile fever is becoming increasingly relevant from an epidemiological point of view. Natural foci of this disease pathogen have been present in Ukraine for a long time in the southern and eastern regions, but now, due to climate change, there are changes in the ecology of both natural carriers and vectors, which significantly changes the epidemiological risks to humans. Over the past few years, in particular, in 2024, an increase in human cases, including fatalities, has been recorded in Ukraine. At the same time, there is a lack of up-to-date information on the circulation of West Nile virus and other flaviviruses (Usutu virus, etc.) in the natural reservoir and among vectors in Ukraine. Our research aimed to conduct serological monitoring in Ukraine among wild forest birds, which are one of the main natural reservoirs of the West Nile virus. During 2023-2024, 268 blood samples and 9 egg yolks of wild Passeriformes (families buntings, finches, true sparrows, tits, bushtits, shrikes, wagtails, Hirundinidae) and Piciformes were collected in Kharkiv, Kyiv, Poltava, Odesa, and Khmelnytsky regions. Blood serum and egg yolks were tested in the ELISA ID.Vet - ID Screen West Nile. It was found that antibodies to WN virus were present in blood samples from Great Tits (seroprevalence from 20% to 100% depending on the region), Song Thrush (60–100%), Blackbird (93%-100%), Chaffinch (100%), Goldfinch (100%), House Sparrow (100%), Greenfinch (10% and 100%), hawfinch (100%), Jays (50%), Field Sparrow (20% and 25%), and Great Reed warbler (40%), reed warbler (33.3%), Сommon whitethroat (25%), Robin (100%), Yellowhammer (100%), Blackcap (100%), Spotted flycatcher (100%), Pied flycatcher (100%), Thrush nightingale(100%). No antibodies to the WN virus were detected in the barn Swallow, Sedge Warbler, Paddyfield Warbler, Collared Flycatcher, Reed Bunting, Spanish Sparrow, Common Woodpecker, Goldfinch, Red-backed Shrike. Seropositivity was found in wild birds from all regions studied. The highest percentage of seropositive birds was found in Poltava (86%, 58%) and Khmelnytsky (67%) regions, and the lowest in Kyiv (9%) and Odesa (17.1%) regions. We also found a difference in seropositivity in different years. Thus, in 2023 it was 27.4%, and in 2024 it was 50.5%

Adjusting susceptibilities of C57BL/6 mice to orthoflaviviruses for evaluation of antiviral drugs by altering the levels of interferon alpha/beta receptor function

The purpose of this study was to optimize the infectivity of four different orthoflaviviruses in mice for evaluating antiviral drugs by using wild-type mice with intact interferon responses, type 1 interferon alpha/beta receptor knockout mice, or by injecting wild type C57BL/6 mice with varying doses of anti-type 1 interferon receptor antibody (MAR1-5A3) to optimize the infectivity and lethality. West Nile virus productively infected wild-type C57BL/6 mice to cause lethality, whereas Usutu virus required a complete absence of type 1 interferon receptor function. Deer tick virus (lineage 2 Powassan virus) and Japanese encephalitis viruses required a dampening of type 1 interferon responses by adjusting the doses of MAR1-5A3 antibody injections. Challenge dose-responsive mortality, weight loss, and viral titers of these two viruses were observed if the type 1 interferon responses were dampened with MAR1-5A3. Conversely, without MAR1-5A3 injections, these disease phenotypes were not viral challenge dose-responsive. From these different interferon-responsive models, the appropriate lethality was identified to determine that 7-deaza-2’-C-methyladenosine has high efficacy for West Nile and Usutu viruses, and low efficacy for deer tick and Japanese encephalitis viruses.

UK mosquitoes are competent to transmit Usutu virus at native temperatures

Usutu virus (USUV) is an emerging zoonotic virus transmitted primarily by Culex mosquitoes. Since its introduction into Europe from Africa during the late 20th century, it has caused mortality within populations of passerine birds and captive owls, and can on occasion lead to disease in humans. USUV was first detected in the UK in 2020 and has become endemic, having been detected in either birds and/or mosquitoes every subsequent year. Importantly, the vector competence of indigenous mosquitoes for the circulating UK (London) USUV strain at representative regional temperatures is still to be elucidated. This study assessed the vector competence of five field-collected mosquito species/biotypes, Culex pipiens biotype molestus, Culex pipiens biotype pipiens, Culex torrentium, Culiseta annulata and Aedes detritus for the London USUV strain, with infection rates (IR) and transmission rates (TR) evaluated between 7 and 28 days post-infection. Infection and transmission were observed in all species/biotypes aside from Ae. detritus and Cx. torrentium. For Cx. pipiens biotype molestus, transmission potential suggests these populations should be monitored further for their role in transmission to humans. Furthermore, both Cx. pipiens biotype pipiens and Cs. annulata were shown to be competent vectors at 19 °C indicating the potential for geographical spread of the virus to other UK regions.

Epidemiology and Ecology of Usutu Virus Infection and Its Global Risk Distribution.

Usutu virus (USUV) is an emerging mosquito-transmitted flavivirus with increasing incidence of human infection and geographic expansion, thus posing a potential threat to public health. In this study, we established a comprehensive spatiotemporal database encompassing USUV infections in vectors, animals, and humans worldwide by an extensive literature search. Based on this database, we characterized the geographic distribution and epidemiological features of USUV infections. By employing boosted regression tree (BRT) models, we projected the distributions of three main vectors (Culex pipiens, Aedes albopictus, and Culiseta longiareolata) and three main hosts (Turdus merula, Passer domesticus, and Ardea cinerea) to obtain the mosquito index and bird index. These indices were further incorporated as predictors into the USUV infection models. Through an ensemble learning model, we achieved a decent model performance, with an area under the curve (AUC) of 0.992. The mosquito index contributed significantly, with relative contributions estimated at 25.51%. Our estimations revealed a potential exposure area for USUV spanning 1.80 million km2 globally with approximately 1.04 billion people at risk. This can guide future surveillance efforts for USUV infections, especially for countries located within high-risk areas and those that have not yet conducted surveillance activities.