ABSTRACTBorna disease virus 1 (BoDV-1) was just recently shown to cause predominantly fatal encephalitis in humans. Despite its rarity, bornavirus encephalitis (BVE) can be considered a model disease for encephalitic infections caused by neurotropic viruses and understanding its pathomechanism is of utmost relevance. Aim of this study was to compare the extent and distribution pattern of cerebral inflammation with the clinical course of disease, and individual therapeutic procedures. For this, autoptic brain material from seven patients with fatal BVE was included in this study. Tissue was stained immunohistochemically for pan-lymphocytic marker CD45, the nucleoprotein of BoDV-1, as well as glial marker GFAP and microglial marker Iba1. Sections were digitalized and counted for CD45-positive and BoDV-1-positive cells. For GFAP and Iba1, a semiquantitative score was determined. Furthermore, detailed information about the individual clinical course and therapy were retrieved and summarized in a standardized way. Analysis of the distribution of lymphocytes shows interindividual patterns. In contrast, when looking at the BoDV-1-positive glial cells and neurons, a massive viral involvement in the brain stem was noticeable. Three of the seven patients received early high-dose steroids, which led to a significantly lower lymphocytic infiltration of the central nervous tissue and a longer survival compared to the patients who were treated with steroids later in the course of disease. This study highlights the potential importance of early high-dose immunosuppressive therapy in BVE. Our findings hint at a promising treatment option which should be corroborated in future observational or prospective therapy studies.ABBREVIATIONS: BoDV-1: Borna disease virus 1; BVE: bornavirus encephalitis; Cb: cerebellum; CNS: central nervous system; FL: frontal lobe; GFAP: glial fibrillary acid protein; Hc: hippocampus; Iba1: ionized calcium-binding adapter molecule 1; Iba1act: general activation of microglial cells; Iba1nod: formation of microglial nodules; IL: insula; Me: mesencephalon; Mo: medulla oblongata; OL: occipital lobe; pASS: per average of 10 screenshots; patearly: patients treated with early high dose steroid shot; patlate: patients treated with late or none high dose steroid shot; Po: pons; So: stria olfactoria; Str: striatum.

OBJECTIVE: Human Borna disease virus (BoDV-1) encephalitis is an emerging disease in Germany. This study investigates the spectrum of human BoDV-1 infection, characterizes anti-BoDV-1-antibodies and kinetics, and compares laboratory test performances.
METHODS: Three hundred four encephalitis cases, 308 nation-wide neuropsychiatric conditions, 127 well-defined psychiatric cases from Borna disease-endemic areas, and 20 persons with contact to BoDV-1 encephalitis patients or animals were tested for BoDV-1 infections by serology and PCR.
RESULTS: BoDV-1 infections were only found in encephalitis patients with residence in, or recent travel to, virus-endemic areas. Antibodies were detected as early as 12 days after symptom onset. Serum antibody levels correlated with disease duration. Serology was ordered after 50% of the disease duration had elapsed, reflecting low awareness. BoDV-1-antibodies were of IgG1 subclass, and the epitope on BoDV-1 antigens was determined. Specificity of the indirect immunofluorescence antibody test (IFAT) and lineblot (LB) from serum and cerebrospinal fluid (CSF), as well as PCR testing from CSF, was 100%. Sensitivity, depending on first or all samples, reached 75-86% in serum and 92-94% in CSF for the IFAT, and 33-57% in serum and 18-24% in CSF for the LB. Sensitivity for PCR in CSF was 25-67%. Positive predictive values were 100% each, while negative predictive values were 99% (IFAT), 91-97% (LB), and 90% (PCR).
CONCLUSIONS: There is no hint that BoDV-1 causes other diseases than encephalitis in humans. Awareness has to be increased in virus-endemic areas. Tests are robust but lack sensitivity. Detection of IgG1 against specific peptides may facilitate diagnosis. Screening of healthy individuals is likely not beneficial.

In 2018, Borna Disease Virus 1 (BoDV-1) was confirmed as a human zoonotic pathogen causing rare but fatal encephalitis in Germany. While diagnostic procedures and the clinical picture have been described, epidemiology remains mysterious. Though endemic areas and a natural reservoir host have been identified with the shrew Crocidura leucodon shedding virus in secretions, transmission events, routes and risk factors are unclear. We performed the first comprehensive epidemiological study, combining a large case series with the first case-control study: We interviewed family members of 20 PCR-confirmed BoDV-1 encephalitis cases deceased in 1996-2021 with a standardized questionnaire covering medical history, housing environment, profession, animal contacts, outdoor activities, travel, and nutrition. Cases\' median age was 51 (range 11-79) years, 12/20 were female, and 18/20 lived in the federal state of Bavaria in Southeastern Germany. None had a known relevant pre-existing medical condition. None of the interviews yielded a transmission event such as direct shrew contact, but peridomestic shrew presence was confirmed in 13 cases supporting environmental transmission. Residency in rural areas endemic for animal BoDV-1 was the common denominator of all cases. A subsequent individually matched case-control study revealed residence close to nature in a stand-alone location or on the fringe of the settlement as a risk factor for disease in multivariable analysis with an adjusted OR of 10.8 (95% CI 1.3-89.0). Other variables including keeping cats were not associated with disease. Targeted prevention, future post-exposure-prophylaxis, and timely diagnosis remain challenging.

The newly described zoonotic variegated squirrel bornavirus 1 (VSBV-1) in German squirrel holdings has been associated with the death of three private owners and one zoo animal caretaker (confirmed cases). Epidemiological investigations were severely impeded by the general lack of data on holdings of the putative reservoir hosts, the family Sciuridae. To fill this lack of data for detailed epidemiological investigations of the captive squirrel population, a register of private and zoological squirrel holdings was established. The findings show a broad variety of kept species and their frequency distribution. By contacting the different stakeholders via Web-based social groups and societies, information passed in both directions so that disease awareness could be raised and participants could be recruited for further studies. Cross-sectional studies revealed a prevalence of VSBV-1-positive subpopulations of 0% (95% CI 0%-6.2%) among private squirrel collections and 1.9% (95% CI: 0%-9.9%) among zoos in Germany. The approach presented here can be transferred to other populations of non-traditional pets, which may be equally difficult to monitor, in the case of an emerging zoonotic infectious disease.