Background The 2026 Andes virus (ANDV) outbreak linked to the cruise ship m/v Hondius triggered prolonged diagnostic follow-up of contacts in several countries. ANDV severity, long incubation, and person-to-person spread complicate case ascertainment. The cross-continent introduction of a rare transmissible virus with limited laboratory readiness is a paramount scenario in pandemic preparedness. To build response capacity under the increased regulatory demands of the European in-vitro diagnostic medical devices regulation (IVDR), we conducted an ad-hoc validation of an investigational RT-PCR product for ANDV detection. Methods Members of the EU Reference Laboratory for Vectorborne Viral Pathogens, the HERA network DURABLE, and the German network PAKOP did a rapid survey of hantavirus diagnostic capacity, a sequence-informed review of molecular in-house assays, and a decentralised validation of a strain-specific research-use-only (RUO) RT-PCR product. We tested biobanked patient samples, differential diagnostic materials, and virus cultures; analytical sensitivity was assessed by probit analysis of quantified in-vitro-transcribed RNA. Findings Networks enabled rapid exchange of methods and reference materials, revealing heterogeneous capacity. The RUO assay was evaluated on 302 clinical specimens and 120 cultured materials covering seven Orthohantavirus species and 67 non-hantavirus pathogens. No specificity issues arose; 37 of 38 ANDV samples were positive. Among non-Andes hantaviruses, only a Sin Nombre virus isolate was positive. The 95% limit of detection was 10·65 copies per reaction. Patient-derived material showed concordance with an established in-house RT-PCR. Interpretation Coordinated expert networks can deliver a swift laboratory response and generate ad-hoc validation data for diagnostic products entering regulatory channels. Network-based validation will probably become essential in future European outbreaks under the IVDR.

Abstract Sudan virus (SUDV) is a member of the family Filoviridae , which comprises highly pathogenic viruses associated with unusually high case fatality rates. The development of medical countermeasures against filoviruses, including antivirals, vaccines, and therapeutic antibodies, requires preclinical evaluation in suitable animal models. C57BL/6J IFNAR -/- mice, which lack the type I interferon (IFN-α/β) receptor, have been reported to be susceptible to filovirus infections, although their impaired innate immune response may represent a potential limitation of the model. Here, we show that IFNAR -/- mice constitute a suitable model for SUDV infection. Following infection, animals developed a clear clinical disease characterized by significant weight loss and pronounced changes in behaviour and appearance. Mice reached the predefined clinical endpoint 3–5 days post infection. Post mortem analysis of terminal samples revealed high viral loads and viral genome copies in all tested organs as well as in serum, indicating widespread systemic dissemination. Importantly, infection was associated with a marked increase in several key chemokines and cytokines linked to systemic inflammation, consistent with the development of a cytokine storm–like response. Together, these findings demonstrate that SUDV infection in IFNAR -/- mice induces systemic viral dissemination and a pronounced inflammatory response, supporting the suitability of this model for investigating filovirus pathogenesis and infection-associated immune dysregulation.

Abstract Crimean-Congo hemorrhagic fever virus (CCHFV) is the causative agent of a severe hemorrhagic fever in humans, associated with case fatality rates up to 40%. Due to the lack of approved vaccines or specific antiviral treatments, CCHFV is classified as a biosafety level 4 (BSL4) pathogen in most countries and designated a priority pathogen by the World Health Organization. To facilitate the preclinical assessment of medical countermeasures, we have established a murine model using C57BL/6J IFNAR −/− mice, which lack the IFNα/β receptor, infected with the phylogenetically distinct CCHFV strains Afghanistan09-2990 (Afg09) and Kosovo Hoti (Hoti). Infection with both CCHFV strains in IFNAR −/− mice resulted in significant weight loss, with Afg09 infection leading to more severe clinical disease. Quantitative analysis of viral RNA revealed widespread viral dissemination across multiple organs. Detection of infectious virus varied by organ and strain. These results independently validate previously reported disease phenotypes while providing new insight into strain-specific differences in CCHFV pathogenesis in IFNAR −/− mice. Thereby, these CCHFV models represent valuable tools for the evaluation of antiviral therapeutics and vaccine candidates, enabling the investigation of cross-lineage protection against genetically diverse CCHFV isolates.

BACKGROUND:Recognition of the 2026 Bundibugyo virus (BDBV) disease outbreak was delayed by adverse political circumstances and logistical constraints. The deployed test was intended for Ebola virus rather than BDBV, a distinct viral species. A virus isolate representing the outbreak strain is unavailable, and regulatory demands on diagnostics have increased. We aimed to provide reference virus material and validate diagnostic tests to aid in regulatory clearance. METHODS:We generated an inactivated, patient-derived BDBV genome reference standard (GRS; 2·4 × 106 copies per mL) from a throat swab of a patient admitted to Charité-Universitätsmedizin Berlin, Berlin, Germany. We distributed the GRS internationally; tested its stability over 90 h at -80°C, 4°C, 20°C, and 37°C; and analytically validated four pilot real-time RT-PCR diagnostic tests. We assessed the sensitivity and run-to-run consistency of the diagnostic tests at the coordinating laboratory and specificity in a network of nine laboratories, using cultured pathogen materials (52-62 samples per test, representing 30 pathogens) and clinical leftover samples (120-172 samples per assay, representing 22 differential diagnostic aetiologies). Detection outcomes from all four tests were pooled in a probit model with assay class as a covariate to estimate the ratio of 95% limits of detection between BDBV-specific and broad-range assays. FINDINGS:Relative to storage at -80°C, GRS viral load after 90 h was reduced 1·15-fold (95% CI 0·96-1·37, p=0·12) at 4°C and at 20°C, and 2·67-fold (2·24-3·19, p<0·0001) at 37°C. 95% limits of detection were 8·4 copies per μL (95% CI 3·2-87·2) for the Altona Diagnostics RealStar Filovirus Screen RT-PCR Kit 1.0, 2·4 copies per μL (1·0-20·6) for the Altona Diagnostics RealStar Bundibugyo RT-PCR Kit 1.0, 11·9 copies per μL (4·7-139·4) for the Roche LightMix Modular Ebola Virus Test Kit, and 2·5 copies per μL (1·2-14·4) for the Roche LightMix Modular Bundibugyo Virus Test Kit. In the pooled probit model, the 95% limits of detection of BDBV-specific assays were 0·24 times (95% CI 0·13-0·47) that of broad-range assays-ie, approximately four times lower. Decentralised evaluation of specificity in nine laboratories was completed within 15 days, and no cross-reactivity occurred in cultured or clinical panels. INTERPRETATION:Decentralised evaluation of diagnostic test kits can facilitate emergency-use decisions, national authorisation, or regulatory review in ongoing outbreaks. This approach defines a course of action for laboratory networks to accelerate the development and assessment of diagnostic medical countermeasures. It bridges the interval between first outbreak recognition and availability of fully certified commercial diagnostic tests, especially for rare, high-consequence pathogens for which virus isolates or clinical materials are scarce. FUNDING:None.

Numbers of human encephalitis cases caused by infection with Borna disease virus 1 (BoDV1) increase continuously in endemic areas. The reservoir host of BoDV1 is the bicolored white-toothed shrew, albeit few naturally infected individuals of other shrew species have been detected. To establish a reliable experimental reservoir model, 15 greater white-toothed shrews were infected with a shrew-derived BoDV1 isolate by different inoculation routes (intracerebral, intranasal, oral, subcutaneous, and intraperitoneal) and monitored up to 41 days. Except for the oral route, all other animals (12/15) were successfully infected, and the majority of them displayed temporarily reduced feed intake and loss of body weight but no inflammatory lesions. Infectious virus was isolated from 11/12 infected animals. Viral RNA was demonstrated by qRT-PCR in the central nervous system (CNS) and the majority of organs. Immunohistochemistry demonstrated BoDV1 antigen in neurons and astrocytes in the CNS and peripheral nerves. High viral loads in the CNS and the spinal cord points towards spread from periphery to the CNS to enhance viral replication and subsequent centrifugal spread to organs capable of secretion and excretions. In general, successful experimental BoDV1 infection of shrews proves their usefulness as animal model, enabling further studies on maintenance, transmission, pathogenesis, and risk assessment for human spillover infections.

Crimean-Congo hemorrhagic fever virus (CCHFV) is the causative agent of a severe hemorrhagic fever in humans, associated with case fatality rates ranging from 10 to 40%. Due to the lack of approved vaccines or specific antiviral treatments, CCHFV is classified as a biosafety level 4 (BSL4) pathogen in most countries and designated a priority pathogen by the World Health Organization (WHO). To facilitate the preclinical assessment of medical countermeasures, we have established two murine models using C57BL/6J IFNAR−/− mice, which lack the IFNα/β receptor, infected with the phylogenetically distinct CCHFV strains Afghanistan09-2990 (Afg09) and Kosovo Hoti (Hoti). Infection with both CCHFV strains in IFNAR−/− mice resulted in significant weight loss, with Afg09 infection leading to more severe clinical disease. Quantitative analysis of viral RNA revealed widespread viral dissemination across multiple organs in both models. Detection of infectious virus varied by organ and strain. These results confirm and extend previous findings, providing a deeper understanding of CCHFV strain-specific pathogenesis in IFNAR−/− mice. Thereby, these mouse models represent valuable tools for the evaluation of antiviral therapeutics and vaccine candidates, enabling the investigation of cross-lineage protection against genetically diverse CCHFV isolates. ### Competing Interest Statement The authors have declared no competing interest.

Objectives: Infectious diseases and high-consequence infectious diseases (HCID), are often present in febrile travelers. Multiplex polymerase chain reaction (PCR) may help to confirm or rule out HCIDs and thus prevent a delay in treatment or undue isolation. Methods: The BioFire® FilmArray® Global Fever Panel–RUO was evaluated vs conventional methods in diagnostics in febrile returning travelers. Results: Eighty-two patients and three simulated patients with HCIDs were analyzed. A total of 10 of the 19 possible pathogens were detected by multiplex PCR. In 30 samples, at least one pathogen was detected by the multiplex PCR, as compared to 35 with conventional diagnostics. The positive percentage agreement was 85.71% (69.74-95.19) overall: Crimean-Congo hemorrhagic fever virus 1/1, dengue virus 4/4, Ebola virus 1/1, Leptospira 1/2 (50%, 1.26-98.74), Marburg virus 1/1, Plasmodium spp. 22/23 (95.65%, 78.05-99.89), Plasmodium falciparum 19/20 (95%, 75.13-99.89), Plasmodium vivax/ovale 2/2, Salmonella enterica serovar typhi 0/2, and Salmonella enterica serovar Paratyphi 0/1. The overall negative percentage agreement was 96.0% (86.29-99.51). Conclusion: The multiplex PCR detected pathogens from blood with varying levels of specificity and sensitivity in less than 1 hour. For HCID, it could shorten the time to diagnosis. However, Salmonella enterica spp. or Leptospira spp. were detected infrequently.

[This corrects the article DOI: 10.1016/j.heliyon.2023.e19613.].

Marburg virus (MARV) is a causative agent of a severe hemorrhagic fever with high fatality rates endemic in central Africa. Current outbreaks of MARV in Equatorial Guinea and Tanzania underline the relevance of MARV as a public health emergency pathogen. In 2021, the first known human MARV case was confirmed in Guinea, West Africa. Since no infectious virus could be isolated from that fatal case in 2021, we generated recombinant (rec) MARV Guinea by reverse genetics in order to study and characterize this new MARV, which occurred in West Africa for the first time, in terms of its growth properties, detection by antibodies, and therapeutic potential compared to known MARV strains. Our results showed a solid viral replication of recMARV Guinea in human, bat, and monkey cell lines in comparison to other known MARV strains. We further demonstrated that replication of recMARV Guinea in cells can be inhibited by the nucleoside analogue remdesivir. Taken together, we could successfully reconstitute de novo the first West African MARV from Guinea showing similar replication kinetics in cells compared to other central African MARV strains. Our reverse genetics approach has proven successful in characterizing emerging viruses, especially when virus isolates are missing and viral genome sequences are incomplete.

OBJECTIVES:Scrub typhus is an emerging infectious disease in Asia caused by Orientia tsutsugamushi (Ot). From Nepal, only scant data on the genetic epidemiology of this agent is available, and determinants of immunoregulation are poorly understood. METHODS:Patients (n = 238) referred to the National Public Health Laboratory (Kathmandu, Nepal) from all over Nepal for suspected scrub typhus were enrolled upon positive immunoglobulin (Ig)M testing between July and October 2015. From Ot 16S and 47 kD polymerase chain reaction (PCR)-positive samples, the variable domain I of the 56 kD gene was sequenced and phylogenetically analyzed. T helper (Th) cell-associated cytokines (n = 13) and chemokines (n = 12) were quantified by multiplex bead arrays. RESULTS:In 93/238 (39.1%) IgM-positive samples, Ot DNA was detected by quantitative PCR. Phylogenetic analysis of 56 kD sequences revealed seven distinct clusters, six of them with high homologies to strains detected in other countries. The Th1-related cytokines interferon-γ and C-X-C motif chemokine ligand 10 were strongly upregulated and correlated with bacteremia, while levels of Th2-associated chemokines were reduced. Bacteremia also correlated with concentrations of interleukin (IL)-6 and IL-10 but not tumor necrosis factor-α. CONCLUSION:We identified a considerable genetic heterogeneity of human-pathogenic Ot strains circulating in Nepal. Acute Nepalese scrub typhus patients showed strong Th1 but impaired Th2 responses, especially on the chemokine level.

The urgent need for vaccines against Ebola virus (EBOV) was underscored by the large outbreak in West Africa (2014–2016). Since then, several promising vaccine candidates have been tested in pre-clinical and clinical studies. As a result, two vaccines were approved for human use in 2019/2020, of which one includes a heterologous adenovirus/Modified Vaccinia virus Ankara (MVA) prime-boost regimen. Here, we tested new vaccine candidates based on the recombinant MVA vector, encoding the EBOV nucleoprotein (MVA-EBOV-NP) or glycoprotein (MVA-EBOV-GP) for their efficacy after homologous prime-boost immunization in mice. Our aim was to investigate the role of each antigen in terms of efficacy and correlates of protection. Sera of mice vaccinated with MVA-EBOV-GP were virus-neutralizing and MVA-EBOV-NP immunization readily elicited interferon-γ-producing NP-specific CD8+ T cells. While mock-vaccinated mice succumbed to EBOV infection, all vaccinated mice survived and showed drastically decreased viral loads in sera and organs. In addition, MVA-EBOV-NP vaccinated mice became susceptible to lethal EBOV infection after depletion of CD8+ T cells prior to challenge. This study highlights the potential of MVA-based vaccines to elicit humoral immune responses as well as a strong and protective CD8+ T cell response and contributes to understanding the possible underlying mechanisms.

The current Severe acute respiratory syndrome related coronavirus 2 (SARS-CoV-2) pandemic is a public health emergency of international concern. Sensitive and precise diagnostic tools are urgently needed. In this study, we developed a SARS-CoV-2 spike (S1) protein enzyme-linked immunosorbent assay (ELISA) to detect SARS-CoV-2-specific antibodies. The SARS-CoV-2 S1 ELISA was found to be specific [97.8% (95% CI, 96.7% - 98.5%)], reproducible and precise (intra-assay coefficient of variability (CV) 5.3%, inter-assay CV 7.9%). A standard curve and the interpolation of arbitrary ELISA units per milliliter served to reduce the variability between different tests and operators. Cross-reactivity to other human coronaviruses was addressed by using sera positive for MERS-CoV- and hCoV HKU1-specific antibodies. Monitoring antibody development in various samples of twenty-three and single samples of twenty-nine coronavirus disease 2019 (COVID-19) patients revealed seroconversion and neutralizing antibodies against authentic SARS-CoV-2 in all cases. The comparison of the SARS-CoV-2 (S1) ELISA with a commercially available assay showed a better sensitivity for the in-house ELISA. The results demonstrate a high reproducibility, specificity and sensitivity of the newly developed ELISA, which is suitable for the detection of SARS-CoV-2 S1 protein-specific antibody responses.

Despite the recent availability of vaccines against severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), there is an urgent need for specific anti-SARS-CoV-2 drugs. Monoclonal neutralizing antibodies are an important drug class in the global fight against the SARS-CoV-2 pandemic due to their ability to convey immediate protection and their potential to be used as both prophylactic and therapeutic drugs. Clinically used neutralizing antibodies against respiratory viruses are currently injected intravenously, which can lead to suboptimal pulmonary bioavailability and thus to a lower effectiveness. Here we describe DZIF-10c, a fully human monoclonal neutralizing antibody that binds the receptor-binding domain of the SARS-CoV-2 spike protein. DZIF-10c displays an exceptionally high neutralizing potency against SARS-CoV-2, retains full activity against the variant of concern (VOC) B.1.1.7 and still neutralizes the VOC B.1.351, although with reduced potency. Importantly, not only systemic but also intranasal application of DZIF-10c abolished the presence of infectious particles in the lungs of SARS-CoV-2 infected mice and mitigated lung pathology when administered prophylactically. Along with a favorable pharmacokinetic profile, these results highlight DZIF-10c as a novel human SARS-CoV-2 neutralizing antibody with high in vitro and in vivo antiviral potency. The successful intranasal application of DZIF-10c paves the way for clinical trials investigating topical delivery of anti-SARS-CoV-2 antibodies.

Einleitung Eine Infektion mit Borna Disease Virus 1 verursacht bei Tieren, aber auch beim Menschen die Borna’sche Krankheit. Die Feldspitzmaus ist der Reservoirwirt.

Background The Middle East respiratory syndrome coronavirus (MERS-CoV) causes a respiratory disease with a case fatality rate of up to 35%. Given its potential to cause a public health emergency and the absence of efficacious drugs or vaccines, MERS is one of the WHO priority diseases warranting urgent research and development of countermeasures. We aimed to assess safety and tolerability of an anti-MERS-CoV modified vaccinia virus Ankara (MVA)-based vaccine candidate that expresses the MERS-CoV spike glycoprotein, MVA-MERS-S, in healthy adults. Methods This open-label, phase 1 trial was done at the University Medical Center Hamburg-Eppendorf (Hamburg, Germany). Participants were healthy men and women aged 18-55 years with no clinically significant health problems as determined during medical history and physical examination, a body-mass index of 18.5-30.0 kg/m(2) and weight of more than 50 kg at screening, and a negative pregnancy test for women. A key exclusion criterion was a previous MVA vaccination. For the prime immunisation, participants received doses of 1 x 10(7) plaque-forming unit (PFU; low-dose group) or 1 x 10(8) PFU (high-dose group) MVA-MERS-S intramuscularly. A second identical dose was administered intramuscularly as a booster immunisation 28 days after first injection. As a control group for immunogenicity analyses, blood samples were drawn at identical study timepoints from six healthy adults, who did not receive any injections. The primary objectives of the study were safety and tolerability of the two dosage levels and reactogenicity after administration. Immunogenicity was assessed as a secondary endpoint by ELISA and neutralisation tests. T-cell immunity was evaluated by interferon-.-linked enzyme-linked immune absorbent spot assay. All participants who were vaccinated at least once were included in the safety analysis. Immunogenicity was analysed in the participants who completed 6 months of follow-up. This trial is registered with ClinicalTrials.gov, NCT03615911, and EudraCT, 2014-003195-23 Findings From Dec 17, 2017, to June 5, 2018, 26 participants (14 in the low-dose group and 12 in the high-dose group) were enrolled and received the first dose of the vaccine according to their group allocation. Of these, 23 participants (12 in the low-dose group and 11 in the high-dose group) received a second dose of MVA-MERS-S according to their group allocation after a 28-day interval and completed follow-up. Homologous prime-boost immunisation with MVA-MERS-S revealed a benign safety profile with only transient mild-to-moderate reactogenicity. Participants had no severe or serious adverse events. 67 vaccine-related adverse events were reported in ten (71%) of 14 participants in the low-dose group, and 111 were reported in ten (83%) of 12 participants in the high-dose group. Solicited local reactions were the most common adverse events: pain was observed in 17 (65%; seven in the low-dose group vs ten in the high-dose group) participants, swelling in ten (38%; two vs eight) participants, and induration in ten (38%; one vs nine) participants. Headaches (observed in seven participants in the low-dose group vs nine in the high-dose group) and fatigue or malaise (ten vs seven participants) were the most common solicited systemic adverse events. All adverse events resolved swiftly (within 1-3 days) and without sequelae. Following booster immunisation, nine (75%) of 12 participants in the low-dose group and 11 (100%) participants in the high-dose group showed seroconversion using a MERS-CoV S1 ELISA at any timepoint during the study. Binding antibody titres correlated with MERS-CoV-specific neutralising antibodies (Spearman's correlation r=0.86 [95% CI 0.6960-0.9427], p=0.0001). MERS-CoV spike-specific T-cell responses were detected in ten (83%) of 12 immunised participants in the low-dose group and ten (91%) of 11 immunised participants in the high-dose group. Interpretation Vaccination with MVA-MERS-S had a favourable safety profile without serious or severe adverse events. Homologous prime-boost immunisation induced humoral and cell-mediated responses against MERS-CoV. A doseeffect relationship was demonstrated for reactogenicity, but not for vaccine-induced immune responses. The data presented here support further clinical testing of MVA-MERS-S in larger cohorts to advance MERS vaccine development.

In December 2019, a novel coronavirus named SARS-CoV-2 first reported in Wuhan, China, emerged and rapidly spread to numerous other countries globally, causing the current pandemic. SARS-CoV-2 causes acute infection of the respiratory tract (COVID-19) that can result in severe disease and lethality. Currently, there is no approved antiviral drug for treating COVID-19 patients and there is an urgent need for specific antiviral therapies and vaccines. In order for SARS-CoV-2 to enter cells, its surface glycoprotein spike (S) must be cleaved at two different sites by host cell proteases, which therefore represent potential drug targets. In the present study we investigated which host cell proteases activate the SARS-CoV-2 S protein in Calu-3 human airway epithelial cells. We show that S can be cleaved by both the proprotein convertase furin at the S1/S2 site and the transmembrane serine protease 2 (TMPRSS2) at the S2’ site. We demonstrate that TMPRSS2 is essential for activation of SARS-CoV-2 S in Calu-3 cells through antisense-mediated knockdown of TMPRSS2 expression. Further, we show that SARS-CoV-2 replication can be efficiently inhibited by two synthetic inhibitors of TMPRSS2 and also by the broad range serine protease inhibitor aprotinin. Additionally, SARS-CoV-2 replication was also strongly inhibited by the synthetic furin inhibitor MI-1851. Combining various TMPRSS2 inhibitors with MI-1851 produced more potent antiviral activity against SARS-CoV-2 than an equimolar amount of any single serine protease inhibitor. In contrast, inhibition of endosomal cathepsins by E64d did not affect virus replication. Our data demonstrate that both TMPRSS2 and furin are essential for SARS-CoV-2 activation in human airway cells and are promising drug targets for the treatment of COVID-19 either by targeting one of these proteases alone or by a combination of furin and TMPRSS2 inhibitors. Therefore, this approach has a high therapeutic potential for treatment of COVID-19.

Background Emerging viruses like severe acute respiratory syndrome coronavirus (SARS-CoV), Crimean-Congo haemorrhagic fever virus (CCHFV) and Nipah virus (NiV) have been identified to pose a potential threat to transfusion safety. In this study, the ability of the THERAFLEX UV-Platelets and THERAFLEX MB-Plasma pathogen inactivation systems to inactivate these viruses in platelet concentrates and plasma, respectively, was investigated. Materials and methods Blood products were spiked with SARS-CoV, CCHFV or NiV, and then treated with increasing doses of UVC light (THERAFLEX UV-Platelets) or with methylene blue (MB) plus increasing doses of visible light (MB/light; THERAFLEX MB-Plasma). Samples were taken before and after treatment with each illumination dose and tested for residual infectivity. Results Treatment with half to three-fourths of the full UVC dose (0 center dot 2 J/cm(2)) reduced the infectivity of SARS-CoV (>= 3 center dot 4 log), CCHFV (>= 2 center dot 2 log) and NiV (>= 4 center dot 3 log) to the limit of detection (LOD) in platelet concentrates, and treatment with MB and a fourth of the full light dose (120 J/cm(2)) decreased that of SARS-CoV (>= 3 center dot 1 log), CCHFV (>= 3 center dot 2 log) and NiV (>= 2 center dot 7 log) to the LOD in plasma. Conclusion Our study demonstrates that both THERAFLEX UV-Platelets (UVC) and THERAFLEX MB-Plasma (MB/light) effectively reduce the infectivity of SARS-CoV, CCHFV and NiV in platelet concentrates and plasma, respectively.

Genomic surveillance during ebolavirus outbreaks to elucidate transmission chains and develop diagnostic tests is delayed by the laborious development of variant-specific laboratory assays. We developed a new protocol combining 31 parallel PCR assays with Illumina/MinION-based sequencing, allowing generic ebolavirus genomic surveillance, validated using cell culture-derived Ebola, Reston, Sudan and Taï Forest virus at concentrations compatible with patient viral loads. Our approach enables pre-emptive genomic surveillance of ongoing and future ebolavirus outbreaks irrespective of variant divergence.

Mammalian Bornavirus (BoDV-1) typically causes a fatal neurologic disorder in horses and sheep, and was recently shown to cause fatal encephalitis in humans with and without transplant reception. It has been suggested that BoDV-1 enters the central nervous system (CNS) via the olfactory pathway. However, (I) susceptible cell types that replicate the virus for successful spread, and (II) the role of olfactory ensheathing cells (OECs), remained unclear. To address this, we studied the intranasal infection of adult rats with BoDV-1 in vivo and in vitro, using olfactory mucosal (OM) cell cultures and the cultures of purified OECs. Strikingly, in vitro and in vivo, viral antigen and mRNA were present from four days post infection (dpi) onwards in the olfactory receptor neurons (ORNs), but also in all other cell types of the OM, and constantly in the OECs. In contrast, in vivo, BoDV-1 genomic RNA was only detectable in adult and juvenile ORNs, nerve fibers, and in OECs from 7 dpi on. In vitro, the rate of infection of OECs was significantly higher than that of the OM cells, pointing to a crucial role of OECs for infection via the olfactory pathway. Thus, this study provides important insights into the transmission of neurotropic viral infections with a zoonotic potential.