Oropouche Virus: An Emerging Orthobunyavirus.

On 2 February 2024, the Pan American Health Organization/World Health Organization issued an epidemiological alert on rising Oropouche virus (OROV) infections in South America. By 3 August 2024, this alert level had escalated from medium to high. OROV has been a public health concern in Central and South America since its emergence in Brazil in the 1960s. However, the 2024 outbreak marks a turning point, with the sustained transmission in non-endemic regions of Brazil, local transmission in Cuba, two fatalities and several cases of vertical transmission. As of the end of August 2024, 9852 OROV cases have been confirmed. The 2024 OROV outbreak underscores critical gaps in our understanding of OROV pathogenesis and highlights the urgent need for antivirals and vaccines. This review aims to provide a concise overview of OROV, a neglected orthobunyavirus.

ddPCR for the Detection and Absolute Quantification of Oropouche Virus.

BACKGROUND: Oropouche virus (OROV) is a segmented RNA virus belonging to the genus Orthobunyavirus in the family Peribunyaviridae. Herein, an in-house droplet digital PCR (ddPCR) assay was used for the detection and quantification of OROV. METHODS: The ddPCR reaction was assessed as duplex assay using the human housekeeping gene RPP30. Limit of detection (LoD) analysis was performed in whole blood, serum, and urine. The assay was executed on a total of 28 clinical samples (whole blood n = 9, serum n = 11, and urine n = 8), of which 16 specimens were tested positive at the routine molecular diagnostics (endpoint and real-time PCRs). RESULTS: The LoD of the ddPCR performed using 10-fold serial dilution of OROV detected up to 1 cp/µL in all the biological matrices. Compared to the routine molecular diagnostics, the ddPCR assay showed 100% sensitivity for whole blood and serum and 75% for urine, highlighting higher positive rate of ddPCR. CONCLUSION: We have established a quantitative RNA detection method of OROV with high sensitivity and specificity based on ddPCR. This test is capable of quantitatively monitoring the viral load of OROV and can contribute, in addition to laboratory diagnosis, to shed light on the pathogenesis, filling in the knowledge gaps of this neglected disease and to the vector control programs.

Oropouche Virus (OROV) in Pregnancy: An Emerging Cause of Placental and Fetal Infection Associated with Stillbirth and Microcephaly following Vertical Transmission.

Oropouche virus (OROV) is an emerging arbovirus endemic in Latin America and the Caribbean that causes Oropouche fever, a febrile illness that clinically resembles some other arboviral infections. It is currently spreading through Brazil and surrounding countries, where, from 1 January to 1 August 2024, more than 8000 cases have been identified in Bolivia, Brazil, Columbia, and Peru and for the first time in Cuba. Travelers with Oropouche fever have been identified in the United States and Europe. A significant occurrence during this epidemic has been the report of pregnant women infected with OROV who have had miscarriages and stillborn fetuses with placental, umbilical blood and fetal somatic organ samples that were RT-PCR positive for OROV and negative for other arboviruses. In addition, there have been four cases of newborn infants having microcephaly, in which the cerebrospinal fluid tested positive for IgM antibodies to OROV and negative for other arboviruses. This communication examines the biology, epidemiology, and clinical features of OROV, summarizes the 2023-2024 Oropouche virus epidemic, and describes the reported cases of vertical transmission and congenital infection, fetal death, and microcephaly in pregnant women with Oropouche fever, addresses experimental animal infections and potential placental pathology findings of OROV, and reviews other bunyavirus agents that can cause vertical transmission. Recommendations are made for pregnant women travelling to the regions affected by the epidemic.

(Re-)Emergence of Oropouche Virus (OROV) Infections: Systematic Review and Meta-Analysis of Observational Studies.

Oropouche Virus (OROV; genus of Orthobunyavirus) is the causal agent of Oropouche Fever (OF). Due to the lack of specific signs and symptoms and the limited availability of diagnostic tests, the actual epidemiology of OROV infections and OF has been extensively disputed. In this systematic review with meta-analysis, a literature search was carried out in PubMed, Scopus, EMBASE, and MedRxiv in order to retrieve relevant articles on the documented occurrence of OROV infections. Pooled detection rates were then calculated for anti-OROV antibodies and virus detection (i.e., viral RNA detected by viral cultures and/or real-time polymerase chain reaction [RT-qPCR]). Where available, detection rates for other arboviruses (i.e., Dengue [DENV], Chikungunya [CHKV], and Zika Virus [ZIKV]) were calculated and compared to those for OROV. A total of 47 studies from South America and the Caribbean were retrieved. In individuals affected by febrile illness during OROV outbreaks, a documented prevalence of 0.45% (95% confidence interval [95%CI] 0.16 to 1.12) for virus isolation, 12.21% (95%CI 4.96 to 27.09) for seroprevalence (including both IgM and IgG class antibodies), and 12.45% (95%CI 3.28 to 37.39) for the detection of OROV-targeting IgM class antibodies were eventually documented. In the general population, seroprevalence was estimated to be 24.45% (95%CI 7.83 to 55.21) for IgG class antibodies. The OROV detection rate from the cerebrospinal fluids of suspected cases of viral encephalitis was estimated to be 2.40% (95%CI 1.17 to 5.03). The occurrence of OROV infections was consistently lower than that of DENV, CHKV, and ZIKV during outbreaks (Risk Ratio [RR] 24.82, 95%CI 21.12 to 29.16; RR 2.207, 95%CI 1.427 to 3.412; and RR 7.900, 95%CI 5.386 to 11.578, respectively) and in the general population (RR 23.614, 95%CI 20.584 to 27.129; RR 3.103, 95%CI 2.056 to 4.685; and RR 49.500, 95%CI 12.256 to 199.921, respectively). In conclusion, our study stresses the possibly high underestimation of OROV prevalence in the general population of South America, the potential global threat represented by this arbovirus infection, and the potential preventive role of a comprehensive "One Health approach".

Oropouche Fever, Cuba, May 2024.

Phylogenetic analyses showed that the virus responsible for a May 2024 Oropouche fever outbreak in Cuba was closely related to viruses from Brazil in 2023. Pools of Ceratopogonidae spp. biting midges and Culex quinquefasciatus mosquitoes were positive for Oropouche viral RNA. No cases were severe. Virus extension to new areas may increase case numbers and severity.

Perguntas e respostas sobre o vírus Oropouche/Preguntas y respuestas sobre el virus Oropouche/ Q&A – Oropouche fever

Washington D.C., 24 de julho de 2024 (OPAS) [Atualizado em 26 de julho de 2024] – Em julho deste ano, a Organização Pan-Americana da Saúde (OPAS) emitiu alerta epidemiológico sobre um aumento nos casos notificados do vírus Oropouche (OROV) em cinco países (Brasil, Bolívia, Peru, Cuba e Colômbia) na Região das Américas. Washington D.C., 24 de julio de 2024 (OPS) [Actualizado el 26 de julio de 2024] – En julio de este año, la Organización Panamericana de la Salud (OPS) emitió una alerta epidemiológica sobre un aumento de casos reportados del virus Oropouche (OROV) en cinco países (Brasil, Bolivia, Perú, Cuba y Colombia) de la Región de las Américas. Washington D.C., 24 July 2024 (PAHO) [Updated 26 July 2024] – In July this year, the Pan American Health Organization (PAHO) issued an epidemiological alert on an increase in reported cases of Oropouche virus (OROV) in five countries (Brazil, Bolivia, Peru, Cuba and Colombia) in the Region of the Americas.

Activation of neurotoxic A1-reactive astrocytes by SFTS virus infection accelerates fatal brain damage in IFNAR1-/- mice.

Severe fever with thrombocytopenia syndrome (SFTS) has a high mortality rate compared to other infectious diseases. SFTS is particularly associated with a high risk of mortality in immunocompromised individuals, while most patients who die of SFTS exhibit symptoms of severe encephalitis before death. However, the region of brain damage and mechanisms by which the SFTS virus (SFTSV) causes encephalitis remains unknown. Here, we revealed that SFTSV infects the brainstem and spinal cord, which are regions of the brain associated with respiratory function, and motor nerves in IFNAR1-/- mice. Further, we show that A1-reactive astrocytes are activated, causing nerve cell death, in infected mice. Primary astrocytes of SFTSV-infected IFNAR1-/- mice also induced neuronal cell death through the activation of A1-reactive astrocytes. Herein, we showed that SFTSV induces fatal neuroinflammation in the brain regions important for respiratory function and motor nerve, which may underlie mortality in SFTS patients. This study provides new insights for the treatment of SFTS, for which there is currently no therapeutic approach.

A reporter Oropouche virus expressing ZsGreen from the M segment enables pathogenesis studies in mice.

Oropouche fever caused by Oropouche virus (OROV) is a significant zoonosis in Central and South America. Despite its public health significance, we lack high-throughput diagnostics, therapeutics, and a comprehensive knowledge of OROV biology. Reporter viruses are valuable tools to rapidly study virus dynamics and develop neutralization and antiviral screening assays. OROV is a tri-segmented bunyavirus, which makes generating a reporter virus challenging, as introducing foreign elements into the viral genome typically affects fitness. We previously demonstrated that the non-structural gene NSm on the OROV medium (M) segment is non-essential for replication in vitro. Taking advantage of this, we have now generated a recombinant OROV expressing fluorescent protein ZsGreen in place of NSm. This reporter OROV is both stable and pathogenic in IFNAR-/- mice and provides a powerful tool for OROV pathogenesis studies and assay development.IMPORTANCEEmerging and reemerging infectious agents such as zoonotic bunyaviruses are of global health concern. Oropouche virus (OROV) causes recurring outbreaks of acute febrile illness in the Central and South American human populations. Biting midges are the primary transmission vectors, whereas sloths and non-human primates are their reservoir hosts. As global temperatures increase, we will likely see an expansion in arthropod-borne pathogens such as OROV. Therefore, developing reagents to study pathogen biology to aid in identifying druggable targets is essential. Here, we demonstrate the feasibility and use of a fluorescent OROV reporter in mice to study viral dynamics and pathogenesis. We show that this reporter OROV maintains characteristics such as growth and pathogenicity similar to the wild-type virus. Using this reporter virus, we can now develop methods to assist OROV studies and establish various high-throughput assays.

Rab27a GTPase and its effector Myosin Va are host factors required for efficient Oropouche virus cell egress.

Oropouche fever, a debilitating illness common in South America, is caused by Oropouche virus (OROV), an arbovirus. OROV belongs to the Peribunyaviridae family, a large group of RNA viruses. Little is known about the biology of Peribunyaviridae in host cells, especially assembly and egress processes. Our research reveals that the small GTPase Rab27a mediates intracellular transport of OROV induced compartments and viral release from infected cells. We show that Rab27a interacts with OROV glycoproteins and colocalizes with OROV during late phases of the infection cycle. Moreover, Rab27a activity is required for OROV trafficking to the cell periphery and efficient release of infectious particles. Consistently, depleting Rab27a's downstream effector, Myosin Va, or inhibiting actin polymerization also hinders OROV compartments targeting to the cell periphery and infectious viral particle egress. These data indicate that OROV hijacks Rab27a activity for intracellular transport and cell externalization. Understanding these crucial mechanisms of OROV's replication cycle may offer potential targets for therapeutic interventions and aid in controlling the spread of Oropouche fever.

Study of Akabane disease in an Iranian dairy herd: a re-emerging disease.

Akabane virus is a teratogenic pathogen transmitted by Culicoides spp. to ruminants. The virus induces anomalies in the central nervous system in the developing fetus, resulting in arthrogryposis-hydranencephaly (A-H) syndrome. During three outbreaks of the disease (2002, 2013, and 2020), 77 calves were born in Varamin, Iran, with A-H syndrome. The presenting neurologic signs were categorized into three main groups, as common, less common, and uncommon signs. The common signs were unawareness of the surroundings, blindness, deep depression, partial failure of suckling, and unintelligent behavior. The less commonly noted signs were hyperexcitability, regurgitation, head pressing, compulsive walking, and kicking, while the uncommon signs comprised protrusion of the tongue, making sounds resembling barking, carnivore-like milk drinking, and deafness. Arthrogryposis, dome-shaped skull, kyphosis, torticollis, lordosis, scoliosis, and spina bifida were the diagnosed skeletal defects. Upon necropsy, hydranencephaly, hydrocephaly, and microencephaly were seen in the calves presenting neurologic signs, while astrocytosis, astrogliosis, focal gliosis, perivascular, perineuronal, and submeningeal edema, perivascular cuffing, non-suppurative meningitis, non-suppurative encephalitis and lymphoplasmacytic infiltration, and perivascular and parenchymal hemorrhage were seen in samples obtained from the brains. RT-PCR detected Akabane virus in the brain tissues of the affected calves. This is the first clinical study of Akabane disease in calves in Iran.

N-glycosylation of viral glycoprotein is a novel determinant for the tropism and virulence of highly pathogenic tick-borne bunyaviruses.

Severe fever with thrombocytopenia syndrome (SFTS) virus, a tick-borne bunyavirus, causes a severe/fatal disease termed SFTS; however, the viral virulence is not fully understood. The viral non-structural protein, NSs, is the sole known virulence factor. NSs disturbs host innate immune responses and an NSs-mutant SFTS virus causes no disease in an SFTS animal model. The present study reports a novel determinant of viral tropism as well as virulence in animal models, within the glycoprotein (GP) of SFTS virus and an SFTS-related tick-borne bunyavirus. Infection with mutant SFTS viruses lacking the N-linked glycosylation of GP resulted in negligible usage of calcium-dependent lectins in cells, less efficient infection, high susceptibility to a neutralizing antibody, low cytokine production in macrophage-like cells, and reduced virulence in Ifnar-/- mice, when compared with wildtype virus. Three SFTS virus-related bunyaviruses had N-glycosylation motifs at similar positions within their GP and a glycan-deficient mutant of Heartland virus showed in vitro and in vivo phenotypes like those of the SFTS virus. Thus, N-linked glycosylation of viral GP is a novel determinant for the tropism and virulence of SFTS virus and of a related virus. These findings will help us understand the process of severe/fatal diseases caused by tick-borne bunyaviruses.

Replication of Akabane virus and related orthobunyaviruses in a fetal-bovine-brain-derived cell line.

Akabane virus (AKAV), Aino virus, Peaton virus, Sathuperi virus, and Shamonda virus are arthropod-borne viruses belonging to the order Elliovirales, family Peribunyaviridae, genus Orthobunyavirus. These viruses cause or may cause congenital malformations in ruminants, including hydranencephaly, poliomyelitis, and arthrogryposis, although their pathogenicity may vary among field cases. AKAV may cause relatively severe congenital lesions such as hydranencephaly in calves. Furthermore, strains of AKAV genogroups I and II exhibit different disease courses. Genogroup I strains predominantly cause postnatal viral encephalomyelitis, while genogroup II strains are primarily detected in cases of congenital malformation. However, the biological properties of AKAV and other orthobunyaviruses are insufficiently investigated in hosts in the field and in vitro. Here, we used an immortalized bovine brain cell line (FBBC-1) to investigate viral replication efficiency, cytopathogenicity, and host innate immune responses. AKAV genogroup II and Shamonda virus replicated to higher titers in FBBC-1 cells compared with the other viruses, and only AKAV caused cytopathic effects. These results may be associated with the severe congenital lesions in the brain caused by AKAV genogroup II. AKAV genogroup II strains replicated to higher titers in FBBC-1 cells than AKAV genogroup I strains, suggesting that genogroup II strains replicated more efficiently in fetal brain cells, accounting for the detection of the latter strains mainly in fetal infection cases. Therefore, FBBC-1 cells may serve as a valuable tool for investigating the virulence and tropism of the orthobunyaviruses for bovine neonatal brain tissues in vitro.

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.

Alongshan Virus Infection in Rangifer tarandus Reindeer, Northeastern China.

We investigated Alongshan virus infection in reindeer in northeastern China. We found that 4.8% of the animals were viral RNA-positive, 33.3% tested positive for IgG, and 19.1% displayed neutralizing antibodies. These findings suggest reindeer could serve as sentinel animal species for the epidemiologic surveillance of Alongshan virus infection.

Bluetongue Virus Serotype 3 and Schmallenberg Virus in Culicoides Biting Midges, Western Germany, 2023.

In October 2023, bluetongue virus serotype 3 (BTV-3) emerged in Germany, where Schmallenberg virus is enzootic. We detected BTV-3 in 1 pool of Culicoides biting midges collected at the time ruminant infections were reported. Schmallenberg virus was found in many vector pools. Vector trapping and analysis could elucidate viral spread.

Characterization of a natural 'dead-end' variant of Schmallenberg virus.

Schmallenberg virus (SBV) belongs to the Simbu serogroup within the family Peribunyaviridae, genus Orthobunyavirus and is transmitted by Culicoides biting midges. Infection of naïve ruminants in a critical phase of gestation may lead to severe congenital malformations. Sequence analysis from viremic animals revealed a very high genome stability. In contrast, sequence variations are frequently described for SBV from malformed fetuses. In addition to S segment mutations, especially within the M segment encoding the major immunogen Gc, point mutations or genomic deletions are also observed. Analysis of the SBV_D281/12 isolate from a malformed fetus revealed multiple point mutations in all three genome segments. It also has a large genomic deletion in the antigenic domain encoded by the M segment compared to the original SBV reference strain 'BH80/11' isolated from viremic blood in 2011. Interestingly, SBV_D281/12 showed a marked replication deficiency in vitro in Culicoides sonorensis cells (KC cells), but not in standard baby hamster kidney cells (BHK-21). We therefore generated a set of chimeric viruses of rSBV_D281/12 and wild-type rSBV_BH80/11 by reverse genetics, which were characterized in both KC and BHK-21 cells. It could be shown that the S segment of SBV_D281/12 is responsible for the replication deficit and that it acts independently from the large deletion within Gc. In addition, a single point mutation at position 111 (S to N) of the nucleoprotein was identified as the critical mutation. Our results suggest that virus variants found in malformed fetuses and carrying characteristic genomic mutations may have a clear 'loss of fitness' for their insect hosts in vitro. It can also be concluded that such mutations lead to virus variants that are no longer part of the natural transmission cycle between mammalian and insect hosts. Interestingly, analysis of a series of SBV sequences confirmed the S111N mutation exclusively in samples of malformed fetuses and not in blood from viremic animals. The characterization of these changes will allow the definition of protein functions that are critical for only one group of hosts.

Identification and characterization of three monoclonal antibodies targeting the SFTSV glycoprotein and displaying a broad spectrum recognition of SFTSV-related viruses.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel tick-borne viral pathogen that causes severe fever with thrombocytopenia syndrome (SFTS). The disease was initially reported in central and eastern China, then later in Japan and South Korea, with a mortality rate of 13-30%. Currently, no vaccines or effective therapeutics are available for SFTS treatment. In this study, three monoclonal antibodies (mAbs) targeting the SFTSV envelope glycoprotein Gn were obtained using the hybridoma technique. Two mAbs recognized linear epitopes and did not neutralize SFTSV, while the mAb 40C10 can effectively neutralized SFTSV of different genotypes and also the SFTSV-related Guertu virus (GTV) and Heartland virus (HRTV) by targeting a spatial epitope of Gn. Additionally, the mAb 40C10 showed therapeutic effect in mice infected with different genotypes of SFTSV strains against death by preventing the development of lesions and by promoting virus clearance in tissues. The therapeutic effect could still be observed in mice infected with SFTSV which were administered with mAb 40C10 after infection even up to 4 days. These findings enhance our understanding of SFTSV immunogenicity and provide valuable information for designing detection methods and strategies targeting SFTSV antigens. The neutralizing mAb 40C10 possesses the potential to be further developed as a therapeutic monoclonal antibody against SFTSV and SFTSV-related viruses.

First case of Oropouche fever detected in the international border region of the Colombian Amazon: clinical characteristics and molecular diagnosis.

OBJECTIVES: We report the first case of Oropouche fever detected in the border region of Colombia. METHODS: Using a multiplex real-time polymerase chain reaction (PCR), genetic sequencing and clinical characteristics during the dengue epidemic in 2019, a total of 175 samples were analysed, from cases notified to the system epidemiological surveillance such as dengue. FINDINGS: The Oropouche virus (OROV) isolate from Leticia belongs to lineage 2 according to both M and S genome segments maximum likelihood (ML) analysis, shares a common ancestor with samples obtained in Esmeraldas, Ecuador and Turbaco, Colombia. The patient: a woman resident in the border neighbourhood of the municipality of Leticia had the following symptoms: fever, headache, retro-orbital pain and myalgias. MAIN CONCLUSION: This cross-border surveillance can be useful to give an alert about the entry or exit of arboviruses circulation in the region, which are often underreported in public health surveillance systems.

Pathogenesis and virulence of Heartland virus.

Heartland virus (HRTV), an emerging tick-borne pathogenic bunyavirus, has been a concern since 2012, with an increasing incidence, expanding geographical distribution, and high pathogenicity in the United States. Infection from HRTV results in fever, thrombocytopenia, and leucopenia in humans, and in some cases, symptoms can progress to severe outcomes, including haemorrhagic disease, multi-organ failure, and even death. Currently, no vaccines or antiviral drugs are available for treatment of the HRTV disease. Moreover, little is known about HRTV-host interactions, viral replication mechanisms, pathogenesis and virulence, further hampering the development of vaccines and antiviral interventions. Here, we aimed to provide a brief review of HRTV epidemiology, molecular biology, pathogenesis and virulence on the basis of published article data to better understand this virus and provide clues for further study.