Tick-borne encephalitis virus (TBEV) is an emerging pathogen that initially causes flu-like symptoms and can progress to central nervous system (CNS) infections. Tick-borne encephalitis (TBE) is an endemic disease in southern coastal counties with regular human cases, while the causative agent, TBEV, is prevalent in ticks in most of the coastal regions of Norway. This study was aimed to understand TBEV infection status across Norway including both TBE endemic and non-endemic areas. For this, we analyzed a total of 1940 residual serum samples from 19 counties of Norway (as of 2016). The samples were initially screened by ELISA, followed by virus neutralization tests for TBEV confirmation. We found a similar TBEV seroprevalence of 1.7% in TBE endemic and 1.6% in non-endemic areas. Since TBE cases are only reported from endemic regions, our findings suggest a potential subclinical or asymptomatic infection and underdiagnosis in non-endemic areas. Notably, only 43% of the ELISA-positive samples were confirmed by virus neutralization tests indicating that not all ELISA positives are true TBEV infections. Additionally, 137 samples of patients presenting with symptoms of CNS infections from a non-endemic area were included. Of these samples, 11 ELISA-positive samples were analyzed for cross-reactivity among flaviviruses. Cross-reactivity was detected with Dengue virus, West Nile Virus, and non-specific reactions. This underscores the importance of using multiple diagnostic tests to confirm TBEV infections. None of the patients with CNS infection was found to be TBE positive, and in the whole cohort, we found a low TBEV seroprevalence of 0.7%.

Tick-borne encephalitis virus (TBEV) is a medically important orthoflavivirus endemic across Europe and Asia, transmitted primarily by Ixodes ticks but also through ingestion of unpasteurized milk from infected ruminants. While milk-borne human infections are well documented, experimental evidence for natural mother-to-offspring transmission in livestock remains scarce. Here, we experimentally assessed the potential for lactogenic transmission of TBEV in sheep. Eight ewes were infected subcutaneously with the European subtype TBEV strain Hochosterwitz and co-housed with 16 three-week-old lambs (two per ewe) for an 18-day observation period. Clinical parameters, hematology, and virological and serological profiles were monitored. All animals remained asymptomatic, with no hematological abnormalities. TBEV RNA was detected in ewes' serum and milk from day one post-infection, and in lamb serum from day five. Virus-specific antibodies emerged in ewes from day four and in lambs from day 13. These findings provide direct experimental evidence that TBEV can be transmitted via milk from ewes to their suckling lambs throughout the lactation period, underscoring a potential but underappreciated route for virus transmission in endemic areas.