In this study, tick-borne encephalitis (TBE) viruses from Siberia and far-eastern Asia were characterized in order to determine virus subtype distribution. TBE viruses were isolated from ticks (Ixodes persulcatus) collected in the far-eastern (Khabarovsk and Vladivostok) and Siberian (Irkutsk) regions of Russia in 1999. Phylogenetic analysis showed that isolates formed distinct clusters of far-eastern and Siberian subtypes. There was also a minor difference in antigenicity between the Irkutsk isolates and other TBE virus strains, as demonstrated by the reactivity of monoclonal antibodies. Amino acid alignments of the E gene showed that the Irkutsk isolates had a single amino acid change at position 234 (Q or H); this amino acid position is considered to be a 'signature' of Siberian subtype TBE viruses. Strains isolated in Irkutsk also exhibited equivalent or somewhat higher virulence in mice compared with far-eastern TBE virus isolates. All viruses isolated in this study (i.e. far-east Asian and Siberian isolates) have 3' non-coding regions (NCRs) of almost the same length, which contrasts with the various sizes of 3'NCRs of other TBE viruses strains reported previously. The data presented in this study show that the 3'NCR is uniform among TBE viruses isolated from Siberia and far-eastern Asia and that the 3'NCR is essential for TBE virus growth in tick and/or rodent host cells.