[The viral protein NS1: a major player in the pathogenesis of orthoflaviviruses]. / La protéine virale NS1 : un acteur majeur de la pathogenèse des orthoflavivirus.

Orthoflaviviruses are enveloped positive-sense RNA viruses comprising numerous human pathogens transmitted by hematophagous arthropods. This includes viruses such as dengue virus, Zika virus, and yellow fever virus. The viral nonstructural protein NS1 plays a central role in the pathogenesis and cycle of these viruses by acting in two different forms: associated with the plasma membrane (NS1m) or secreted outside the cell (NS1s). The versatility of NS1 is evident in its ability to modulate various aspects of the infectious process, from immune evasion to pathogenesis. As an intracellular protein, it disrupts many processes, interfering with signaling pathways and facilitating viral replication in concert with other viral proteins. As a secreted protein, NS1 actively participates in immune evasion, interfering with the host immune system, inhibiting the complement system, facilitating viral dissemination, and disrupting the integrity of endothelial barriers. This review primarily aims to address the role of NS1 in viral pathogenesis associated with orthoflaviviruses.

Virulence and transmission vary between Usutu virus lineages in Culex pipiens.

Usutu virus (USUV) is a zoonotic arbovirus infecting mainly wild birds. It is transmitted by ornithophilic mosquitoes, mainly of the genus Culex from birds to birds and to several vertebrate dead-end hosts. Several USUV lineages, differing in their virulence have emerged in the last decades and now co-circulate in Europe, impacting human populations. However, their relative transmission and effects on their mosquito vectors is still not known. We thus compared the vector competence and survival of Culex pipiens mosquitoes experimentally infected with two distinct USUV lineages, EU2 and EU3, that are known to differ in their virulence and replication in vertebrate hosts. Infection rate was variable among blood feeding assays but variations between EU2 and EU3 lineages were consistent suggesting that Culex pipiens was equally susceptible to infection by both lineages. However, EU3 viral load increased with viral titer in the blood meal while EU2 viral load was high at all titers which suggest a greater replication of EU2 than EU3 in mosquito. While their relative transmission efficiencies are similar, at least at low blood meal titer, positive correlation between transmission and blood meal titer was observed for EU3 only. Contrary to published results in vertebrates, EU3 induced a higher mortality to mosquitoes (i.e. virulence) than EU2 whatever the blood meal titer. Therefore, we found evidence of lineage-specific differences in vectorial capacity and virulence to both the vector and vertebrate host which lead to balanced propagation of both viral lineages. These results highlight the need to decipher the interactions between vectors, vertebrate hosts, and the diversity of arbovirus lineages to fully understand transmission dynamics.

Co-circulation of two Alphaviruses in Burkina Faso: Chikungunya and O'nyong nyong viruses.

BACKGROUND: Chikungunya virus (CHIKV) and O'nyong nyong virus (ONNV) are phylogenetically related alphaviruses in the Semliki Forest Virus (SFV) antigenic complex of the Togaviridae family. There are limited data on the circulation of these two viruses in Burkina Faso. The aim of our study was to assess their circulation in the country by determining seroprevalence to each of the viruses in blood donor samples and by retrospective molecular and serological testing of samples collected as part of national measles and rubella surveillance. METHODOLOGY/PRINCIPAL FINDINGS: All blood donor samples were analyzed on the Luminex platform using CHIKV and ONNV E2 antigens. Patient samples collected during national measles-rubella surveillance were screened by an initial ELISA for CHIKV IgM (CHIKjj Detect IgM ELISA) at the national laboratory. The positive samples were then analyzed by a second ELISA test for CHIKV IgM (CDC MAC-ELISA) at the reference laboratory. Finally, samples that had IgM positive results for both ELISA tests and had sufficient residual volume were tested by plaque reduction neutralization testing (PRNT) for CHIKV and ONNV. These same patient samples were also analyzed by rRT-PCR for CHIKV. Among the blood donor specimens, 55.49% of the samples were positive for alphaviruses including both CHIKV and ONNV positive samples. Among patient samples collected as part of national measles and rubella surveillance, 3.09% were IgM positive for CHIKV, including 2.5% confirmed by PRNT. PRNT failed to demonstrate any ONNV infections in these samples. No samples tested by RT-qPCR. had detectable CHIKV RNA. CONCLUSIONS/SIGNIFICANCE: Our results suggest that CHIKV and ONNV have been circulating in the population of Burkina Faso and may have been confused with malaria, dengue fever or other febrile diseases such as measles or rubella. Our study underscores the necessity to enhance arbovirus surveillance systems in Burkina Faso.

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.

Skin tropism during Usutu virus and West Nile virus infection: an amplifying and immunological role.

Usutu virus (USUV) and West Nile virus (WNV) are closely related emerging arboviruses belonging to the Flavivirus genus and posing global public health concerns. Although human infection by these viruses is mainly asymptomatic, both have been associated with neurological disorders such as encephalitis and meningoencephalitis. Since USUV and WNV are transmitted through the bite of an infected mosquito, the skin represents the initial site of virus inoculation and provides the first line of host defense. Although some data on the early stages of WNV skin infection are available, very little is known about USUV. Herein, USUV-skin resident cell interactions were characterized. Using primary human keratinocytes and fibroblasts, an early replication of USUV during the first 24 hours was shown in both skin cells. In human skin explants, a high viral tropism for keratinocytes was observed. USUV infection of these models induced type I and III interferon responses associated with upregulated expression of various interferon-stimulated genes as well as pro-inflammatory cytokine and chemokine genes. Among the four USUV lineages studied, the Europe 2 strain replicated more efficiently in skin cells and induced a higher innate immune response. In vivo, USUV and WNV disseminated quickly from the inoculation site to distal cutaneous tissues. In addition, viral replication and persistence in skin cells were associated with an antiviral response. Taken together, these results provide a better understanding of the pathophysiology of the early steps of USUV infection and suggest that the skin constitutes a major amplifying organ for USUV and WNV infection.IMPORTANCEUsutu virus (USUV) and West Nile virus (WNV) are closely related emerging Flaviviruses transmitted through the bite of an infected mosquito. Since they are directly inoculated within the upper skin layers, the interactions between the virus and skin cells are critical in the pathophysiology of USUV and WNV infection. Here, during the early steps of infection, we showed that USUV can efficiently infect two human resident skin cell types at the inoculation site: the epidermal keratinocytes and the dermal fibroblasts, leading to the induction of an antiviral innate immune response. Moreover, following cutaneous inoculation, we demonstrated that both viruses can rapidly spread, replicate, and persist in all distal cutaneous tissues in mice, a phenomenon associated with a generalized skin inflammatory response. These results highlight the key amplifying and immunological role of the skin during USUV and WNV infection.

Monitoring SARS-CoV-2 Seroprevalence in Domestics and Exotic Animals in Southern France.

Since late 2019, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has emerged as a significant global threat to public health. Responsible for the COVID-19 pandemic, this new coronavirus has prompted extensive scientific research to comprehend its transmission dynamics, especially among humans. However, as our understanding deepens, it becomes increasingly clear that SARS-CoV-2's impact goes beyond human populations. Recent investigations have illuminated the transmission of the virus between humans and various animal species, raising important questions about zoonotic spillover events and their potential implications for both human and animal health. Our study set out to investigate the prevalence of SARS-CoV-2 in domestic animals (dogs and cats) and zoo animals in the south of France in 2021 and 2022, covering pre-Omicron and Omicron waves. We identified evidence of SARS-CoV-2 antibodies not only in domestic dogs and cats but also in several mammals in zoos. This study shows the importance of implementing surveillance measures, including serological studies, to identify and monitor cases of SARS-CoV-2 infection in animals.

Rift Valley fever in West Africa: A zoonotic disease with multiple socio-economic consequences.

Rift Valley fever virus (RVFV) is an arbovirus that causes Rift Valley fever (RVF), a zoonotic disease that mainly affects domestic and wildlife ruminants and humans. The first epidemic in North-Western and West Africa occurred in Senegal and Mauritania in 1987, two countries where RVF is now endemic. Slaughterhouse workers, farmers, herders and veterinarians are at high risk of exposure to RVF. Beyond the health threat, RVF is considered to cause major socio-economic problems, specifically in developing countries where livestock farming and trade are important economic activities. Indeed, the mortality rate linked to RVF infection can reach 95-100% in newborns and young animals. In West Africa, livestock production is a key factor for food production and for national economics. Epizootics caused by RVF can therefore have serious socio-economic consequences by impacting multisectoral economics, the psycho-social health of pastoral communities, and food security. Improving prevention strategies against RVF, including vaccination, enhancing knowledge of RVF and correcting any inappropriate behaviors by populations of endemics areas, as well as better monitoring of RVF ecological factors are effective ways to better foresee and control outbreaks of RVF and its socio-economical side-effects in countries at high risk of occurrence of the disease.

Spatial scale influences the distribution of viral diversity in the eukaryotic virome of the mosquito Culex pipiens.

Our knowledge of the diversity of eukaryotic viruses has recently undergone a massive expansion. This diversity could influence host physiology through yet unknown phenomena of potential interest to the fields of health and food production. However, the assembly processes of this diversity remain elusive in the eukaryotic viromes of terrestrial animals. This situation hinders hypothesis-driven tests of virome influence on host physiology. Here, we compare taxonomic diversity between different spatial scales in the eukaryotic virome of the mosquito Culex pipiens. This mosquito is a vector of human pathogens worldwide. The experimental design involved sampling in five countries in Africa and Europe around the Mediterranean Sea and large mosquito numbers to ensure a thorough exploration of virus diversity. A group of viruses was found in all countries. This core group represented a relatively large and diverse fraction of the virome. However, certain core viruses were not shared by all host individuals in a given country, and their infection rates fluctuated between countries and years. Moreover, the distribution of coinfections in individual mosquitoes suggested random co-occurrence of those core viruses. Our results also suggested differences in viromes depending on geography, with viromes tending to cluster depending on the continent. Thus, our results unveil that the overlap in taxonomic diversity can decrease with spatial scale in the eukaryotic virome of C. pipiens. Furthermore, our results show that integrating contrasted spatial scales allows us to identify assembly patterns in the mosquito virome. Such patterns can guide future studies of virome influence on mosquito physiology.

Rift Valley fever virus modulates apoptosis and immune response during infection of human astrocytes.

Rift Valley fever (RVF) is an arboviral disease of zoonotic origin that causes recurrent epidemics in Africa, the Arabic Peninsula, and islands of the South West of the Indian Ocean. RVF occurs mainly in livestock but also affects humans with severe clinical manifestations, including neurological disorders. However, human neuropathogenesis of Rift Valley fever virus (RVFV) is still poorly characterized. To study the interactions between RVFV and the central nervous system (CNS), we focused on RVFV infection of astrocytes, the major glial cells of the CNS that have several supporting roles including immune response regulation. We confirmed the permissiveness of astrocytes to RVFV infection and highlighted a strain-dependent infectivity. We showed that RVFV infection of astrocytes induced cell apoptosis and observed that the RVFV Non-Structural protein NSs, a known virulence factor, potentially delayed apoptosis by sequestrating activated-caspase 3 in the nucleus. Our study also showed that RVFV-infected astrocytes upregulated expression of genes associated with inflammatory and type I interferon responses at the mRNA level, but not at the protein level. This inhibition of immune response is potentially due to a NSs-dependent mechanism of mRNA nuclear export inhibition. Together, these results highlighted the direct impact of RVFV infection on the human CNS through the induction of apoptosis and a possible inhibition of early-onset immune responses that are crucial for the host survival.

One Health Approach to Arbovirus Control in Africa: Interests, Challenges, and Difficulties.

The "One Health" concept considers that human and animal health, and ecosystems are closely related and aims to make a link between ecology and human and veterinary medicine. Due to the explosion in population growth along with the geographic and climatic conditions (equatorial and/or tropical climate), Africa is becoming a major hotspot for various socio-health issues associated with infectious diseases, including arboviruses. The incontestable advantages of a One Health approach in Africa lie in the fight against pathogens, such as arboviruses, and in the preservation of environmental, animal, and human health to ensure that the increasing high needs of this population are met as well as their protection against potential epidemics. The One Health strategy gives us a glimpse of the difficulties and challenges that the African continent faces. The importance of this approach in Africa is to establish guidelines and strategies for effective solutions and changes in behavior and harmful activities. Overall, the establishment of high-quality global health policies in the framework of the global health standards program would provide healthy and sustainable human-animal-environmental interactions for the welfare of all.

Emerging Diseases/Viruses Prevention, Control, Surveillance, and One Health.

Emerging diseases have posed a constant threat and major challenge to human health throughout our history [...].

Differential effects of Usutu and West Nile viruses on neuroinflammation, immune cell recruitment and blood-brain barrier integrity.

Usutu (USUV) and West Nile (WNV) viruses are two closely related Flavivirus belonging to Japanese encephalitis virus serogroup. Evidence of increased circulation of these two arboviruses now exist in Europe. Neurological disorders are reported in humans mainly for WNV, despite the fact that the interaction and effects of viral infections on the neurovasculature are poorly described, notably for USUV. Using a human in vitro blood-brain barrier (BBB) and a mouse model, this study characterizes and compares the cerebral endothelial cell permissiveness, innate immunity and inflammatory responses and immune cell recruitment during infection by USUV and WNV. Both viruses are able to infect and cross the human BBB but with different consequences. We observed that WNV infects BBB cells resulting in significant endothelium impairment, potent neuroinflammation and immune cell recruitment, in agreement with previous studies. USUV, despite being able to infect BBB cells with higher replication rate than WNV, does not strongly affect endothelium integrity. Importantly, USUV also induces neuroinflammation, immune cell recruitment such as T lymphocytes, monocytes and dendritic cells (DCs) and was able to infect dendritic cells (DCs) more efficiently compared to WNV, with greater propensity for BBB recruitment. DCs may have differential roles for neuroinvasion of the two related viruses.

Usutu, West Nile, and Tick-Borne Encephalitis Viruses.

The beginning decades of the 21st century have been marked by multiple emergence and re-emergence phenomena of viral diseases [...].

Screening of Circulation of Usutu and West Nile Viruses: A One Health Approach in Humans, Domestic Animals and Mosquitoes in Burkina Faso, West Africa.

Usutu virus (USUV) and West Nile virus (WNV) are phylogenetically closely related arboviruses. These viruses mainly follow an enzootic cycle involving mosquitoes and birds, but they occasionally infect humans and other mammals, inducing neurotropic disorders. Since the discovery of USUV, only two human cases have been reported in Africa, including one in Burkina Faso in 2004. Since then, no studies have been conducted to measure the extent of the circulation of this virus in Burkina Faso, and no study regarding the circulation of WNV has been conducted. Our study aimed to determine the seroprevalence of USUV and WNV in blood donations and in animals (horses, dogs, chickens and pigeons) and to perform molecular screening in patients with febrile fever and in Culex quinquefasciatus and Aedes aegypti mosquitoes. The prevalence of USUV and WNV was studied by serological (ELISA and microneutralization tests) and molecular analyses (RT-qPCR) of mosquito, dog, domestic bird, horse, and human samples in Burkina Faso between 2019 and 2021. We detected a very active transmission of both viruses in Burkina Faso. WNV and USUV seroprevalence is particularly high in humans (19.16% and 14.17%, respectively) and horses (17.28% and 6.17%). Molecular screening did not detect WNV or USUV in the mosquito or human samples tested. Our study shows an active spread of USUV and WNV in Burkina Faso, especially for WNV. This study highlights the value of developing surveillance programs to better prevent, detect, and alert people to USUV and WNV circulation in both primary and incidental hosts.

Special Issue "Emerging Diseases/Viruses: Prevention, Control, Surveillance, and One Health".

Zoonotic diseases account for at least 60% of all infectious diseases and no less than two-thirds of new emerging ones, which underlines the importance of monitoring them as early as possible [...].

Serological Evidence of Zika Virus Circulation in Burkina Faso.

Zika virus (ZIKV) and dengue virus (DENV) are two closely related members of the Flaviviridae family, both transmitted by mosquitoes of the genus Aedes, and are among the arboviruses most at risk to human health. Burkina Faso has been facing an upsurge in DENV outbreaks since 2013. Unlike DENV, there is no serological evidence of ZIKV circulation in humans in Burkina Faso. The main objective of our study was to determine the seroprevalence of ZIKV and DENV in blood donors in Burkina Faso. A total of 501 donor samples collected in the two major cities of the country in 2020 were first tested by a competitive enzyme-linked immunosorbent assay to detect flavivirus antibodies. Positive sera were then tested using Luminex to detect ZIKV and DENV antibodies and virus-specific microneutralization tests against ZIKV were performed. The ZIKV seroprevalence was 22.75% in the donor samples and we found seropositivity for all DENV-serotypes ranging from 19.56% for DENV-1 to 48.86% for DENV-2. Molecular analyses performed on samples from febrile patients and Aedes aegypti mosquitoes between 2019 and 2021 were negative. Our study showed the important circulation of ZIKV and DENV detected by serology although molecular evidence of the circulation of ZIKV could not be demonstrated. It is essential to strengthen existing arbovirus surveillance in Burkina Faso and more broadly in West Africa by focusing on fevers of unknown origin and integrating vector surveillance to assess the extent of ZIKV circulation and identify the circulating strain. Further studies are needed to better understand the epidemiology of this virus in order to define appropriate prevention and response methods.

Usutu Virus Infects Human Placental Explants and Induces Congenital Defects in Mice.

Usutu virus (USUV) is a neurotropic mosquito-borne flavivirus that has dispersed quickly in Europe these past years. This arbovirus mainly follows an enzootic cycle involving mosquitoes and birds, but can also infect other mammals, causing notably sporadic cases in humans. Although it is mainly asymptomatic or responsible for mild clinical symptoms, USUV has been associated with neurological disorders, such as encephalitis and meningoencephalitis, highlighting the potential health threat of this virus. Among the different transmission routes described for other flaviviruses, the capacity for some of them to be transmitted vertically has been demonstrated, notably for Zika virus or West Nile virus, which are closely related to USUV. To evaluate the ability of USUV to replicate in the placenta and gain access to the fetus, we combined the use of several trophoblast model cell lines, ex vivo human placental explant cultures from first and third trimester of pregnancy, and in vivo USUV-infected pregnant mice. Our data demonstrate that human placental cells and tissues are permissive to USUV replication, and suggest that viral transmission can occur in mice during gestation. Hence, our observations suggest that USUV could be efficiently transmitted by the vertical route.

One Health surveillance of West Nile and Usutu viruses: a repeated cross-sectional study exploring seroprevalence and endemicity in Southern France, 2016 to 2020.

BackgroundWest Nile virus (WNV) and Usutu virus (USUV), two closely related flaviviruses, mainly follow an enzootic cycle involving mosquitoes and birds, but also infect humans and other mammals. Since 2010, their epidemiological situation may have shifted from irregular epidemics to endemicity in several European regions; this requires confirmation, as it could have implications for risk assessment and surveillance strategies.AimTo explore the seroprevalence in animals and humans and potential endemicity of WNV and USUV in Southern France, given a long history of WNV outbreaks and the only severe human USUV case in France in this region.MethodsWe evaluated the prevalence of WNV and USUV in a repeated cross-sectional study by serological and molecular analyses of human, dog, horse, bird and mosquito samples in the Camargue area, including the city of Montpellier, between 2016 and 2020.ResultsWe observed the active transmission of both viruses and higher USUV prevalence in humans, dogs, birds and mosquitoes, while WNV prevalence was higher in horses. In 500 human samples, 15 were positive for USUV and 6 for WNV. Genetic data showed that the same lineages, WNV lineage 1a and USUV lineage Africa 3, were found in mosquitoes in 2015, 2018 and 2020.ConclusionThese findings support existing literature suggesting endemisation in the study region and contribute to a better understanding of USUV and WNV circulation in Southern France. Our study underlines the importance of a One Health approach for the surveillance of these viruses.

Role of Dendritic Cells in Viral Brain Infections.

To gain access to the brain, a so-called immune-privileged organ due to its physical separation from the blood stream, pathogens and particularly viruses have been selected throughout evolution for their use of specific mechanisms. They can enter the central nervous system through direct infection of nerves or cerebral barriers or through cell-mediated transport. Indeed, peripheral lymphoid and myeloid immune cells can interact with the blood-brain and the blood-cerebrospinal fluid barriers and allow viral brain access using the "Trojan horse" mechanism. Among immune cells, at the frontier between innate and adaptive immune responses, dendritic cells (DCs) can be pathogen carriers, regulate or exacerbate antiviral responses and neuroinflammation, and therefore be involved in viral transmission and spread. In this review, we highlight an important contribution of DCs in the development and the consequences of viral brain infections.