Evolutionary traits of Tick-borne encephalitis virus: Pervasive non-coding RNA structure conservation and molecular epidemiology

Abstract

Tick-borne encephalitis virus (TBEV) is the aetiological agent of tick-borne encephalitis, an infectious disease of the central nervous system that is often associated with severe sequelae in humans. While TBEV is typically classified into three subtypes, recent evidence suggests a more varied range of TBEV subtypes and lineages that differ substantially in the architecture of their 3' untranslated region (3'UTR). Building on comparative genomic approaches and thermodynamic modelling, we characterize the TBEV UTR structureome diversity and propose a unified picture of pervasive non-coding RNA structure conservation. Moreover, we provide an updated phylogeny of TBEV, building on more than 220 publicly available complete genomes, and investigate the molecular epidemiology and phylodynamics with Nextstrain, a web-based visualization framework for real-time pathogen evolution.

Keywords: Tick-borne encephalitis virus; conserved RNA structure; molecular epidemiology; untranslated region.

Figure 1.

Overview phylogeny of TBEV, depicting the topological arrangement of subtypes TBEV-FE, TBEV-Sib, and TBEV-Eur (all shown sensu stricto here), as well as novel lineages that do not cluster with these subtypes. Strain 178–79 and the Baikalean lineage share ancestral roots with TBEV-FE. The Himalayan lineage is located ancestral to the clades encompassing TBEV-FE and TBEV-Sib subtypes. The European TBEV strains form a separate clade, with the recently detected Western European and N5-17 lineages being clearly separated from the established TBEV-Eur subtype. The midpoint-rooted maximum likelihood tree is based on 256 complete TBEV genomes. Bootstrap values are shown for branches with a support lower than 100.

Figure 2.

Secondary structure prediction of the 5ʹ-terminal 159 nt of TBEV strain Neudoerfl (NC_001672.1), comprising the 5ʹUTR and the distal portion of the capsid protein coding region. The canonical 5ʹUTR organization of TBEV encompasses a Y-shaped SLA element (orange), as well as the hairpin loops CSA (blue), and CSB (green). The start codon is highlighted in red.

Figure 3.

Annotated 3ʹUTRs of representative TBEV strains. Strains were selected to cover the complete 3ʹUTR diversity found in publicly available genome data. 3ʹUTR schemes are plotted to scale, highlighting the variable length and architectural organization of individual strains. Coloured boxes represent evolutionarily conserved, structured RNAs. Truncated sequences from incomplete sequencing are marked by an asterisk. The phylogenetic tree on the right has been computed from full genome nucleotide multiple sequence alignments of 28 TBEV strains. Identifiers show strain name, and lineage association where available, i.e. TBEV-FE Clusters I/II/III (C-I/C-II/C-III), the two Baikalean subtypes TBEV-Bkl-1 (Bkl-1), and TBEV-Bkl-2 (Bkl-2), and TBEV-Sib lineages Baltic (Bal), Zausaev (Zau), Vasilchenko (Vas), Bosnia (Bos), Obskaya (Obs). Accession numbers and 3ʹUTR lengths are listed in Supplementary Table S1.

Figure 4.

Diverging 3ʹUTR architecture in four representative TBEV strains, Neudoerfl (A), TBEV-2871 (B), 886–84 (C), and Senzhang (D). Core and variable regions are underlined in red and grey, respectively. Structural homologs of evolutionarily conserved RNA elements are depicted in the same colour for all strains (matching the colours used in Fig. 3). Structural components that are not found in all homologs, such as closing stems of some CSL4 and CSL2 elements, or additional short stem loops that are only predicted in particular strains, are not considered conserved.

Figure 5.

Panel a: Visualization of the TBEV phylodynamics in Nextstrain and Panel b: Unrooted phylogenetic tree, exposing the divergence of TBEV subtypes and novel lineages. c Geographic spread of TBEV across Eurasia.