The recent discovery of the Yezo virus (YEZV) in Japan and China has raised particular concern due to its potential to cause human diseases ranging from mild febrile illnesses to severe neurological disorders. We report, for the first time, the detection of five YEZV isolates in I. persulcatus ticks from three regions of Russia. The analysis was performed using 5318 ticks of two Ixodes genus collected in 2024 from 23 regions of Russia. The minimum infection rate of YEZV in Russia among I. persulcatus ticks was 0.12% (95% CI: 0.05–0.28). The westernmost and northernmost YEZV detection points have been recorded. YEZV isolates circulating in Russia are genetically diverse. Protein domains of Russian YEZV isolates’ genomes were characterized using HMMER, AlphaFold 3, and InterProScan. The YEZV nucleoprotein (N) of Russian isolates has a racket-shaped structure with “head” and “stalk” domains similar to those of Orthonairovirus haemorrhagiae. The Lys261–Arg261 substitution in the YEZV N Chita 2024-1 isolate occurs in the α11 structure in the region of interaction with viral RNA. Our results show that the distribution area of YEZV is much wider than previously known, provide new data on complete YEZV genomes, extend our structural insight into YEZV N, and suggest a potential target for antiviral drug development to treat YEZV infection.

The recent discovery of Orthonairovirus songlingense (SGLV) and Norwavirus beijiense (BJNV) in China has raised significant concern due to their potential to cause severe human disease. However, little is known about the structural features and function of their nucleoproteins, which play a key role in the viral life cycle. By combining small-angle X-ray scattering (SAXS) data and AlphaFold 3 simulations, we reconstructed the BJNV and SGLV nucleoprotein structures for the first time. The SGLV and BJNV nucleoproteins have structures that are broadly similar to those of Orthonairovirus haemorrhagiae (CCHFV) nucleoproteins despite low sequence similarity. Based on structural analysis, several residues located in the positively charged region of BJNV and SGLV nucleoproteins have been indicated to be important for viral RNA binding. A positively charged RNA-binding crevice runs along the interior of the SGLV and BJNV ribonucleoprotein complex (RNP), shielding the viral RNA. Despite the high structural similarity between SGLV and BJNV nucleoprotein monomers, their RNPs adopt distinct conformations. These findings provide important insights into the molecular mechanisms of viral genome packaging and replication in these emerging pathogens. Also, our work demonstrates that experimental SAXS data can validate and improve predicted AlphaFold 3 structures to reflect their solution structure and also provides the first low-resolution structures of the BJNV and SGLV nucleoproteins for the future development of POC tests, vaccines, and antiviral drugs.

A large number of novel viruses potentially pathogenic for humans are discovered currently. Studying many of them by classical methods of virology is difficult due to the absence of a live viral particles or a sufficient amount of its genetic material. In this case, the use of modern methods of bioinformatics, synthetic and structural biology can help. Haseki tick virus (HSTV) is a recently discovered tick-borne unclassified ssRNA(+) virus. HSTV positive patients experienced fever and elevated temperature. However, at the moment, there is no information on the tertiary structure and functions of its proteins. In this work, we used AlphaFold 3 for annotation of HSTV protein functions, based on the principle that the tertiary structure of a protein is inextricably linked with its molecular function. We were the first to obtain models of tertiary structures and describe the putative functions of HSTV nonstructural proteins (NS3 helicase, NS3 protease, NS5 RNA-dependent RNA-polymerase, and NS5 methyltransferase). These proteins play a key role in the replication of the viral genome and can be targets for the development of direct-acting antiviral drugs.

The continuous evolution of SARS-CoV-2 necessitates broadly neutralizing antibodies for robust therapeutics. In this study, we isolated 1824 single B cells from a COVID-19 convalescent donor and obtained 91 pairs of nucleotide sequences potentially encoding VH/VL domains. From this library, we generated and evaluated a panel of 38 recombinant monoclonal antibodies. Among this panel, we identified DEV_K57 as a broadly neutralizing human IgG1 antibody targeting the SARS-CoV-2 RBD. DEV_K57 exhibited in vitro neutralization breadth against diverse variants, demonstrating subnanomolar IC₅₀ values. This in vitro efficacy was translated in vivo. In a Syrian hamster model of SARS-CoV-2 Delta infection, DEV_K57 significantly reduced viral RNA and infectious titers in both the upper and lower respiratory tracts, conferring a favorable safety profile with minimal histopathological damage. To elucidate its mechanism, we performed comprehensive biophysical and structural analyses. Biolayer interferometry revealed a high picomolar binding affinity to the RBD Delta. Furthermore, size-exclusion chromatography coupled with small-angle X-ray scattering and computational modeling uncovered a unique, stable dimeric architecture of the DEV_K57 Fab-RBD complex in solution, with a binding interface distinct from previously characterized structures. These findings underscore the value of rapid antibody isolation for pandemic preparedness against future emerging biological threats.

Currently, a large number of novel tick-borne viruses potentially pathogenic to humans are discovered. Studying many of them by classical methods of virology is difficult due to the absence of live viral particles or a sufficient amount of their genetic material. In this case, the use of modern methods of bioinformatics and synthetic and structural biology can help. Haseki tick virus (HSTV) is a recently discovered tick-borne unclassified ssRNA(+) virus. HSTV-positive patients experienced fever and an elevated temperature. However, at the moment, there is no information on the tertiary structure and functions of its proteins. In this work, we used AlphaFold 3 and other bioinformatic tools for the annotation of HSTV nonstructural proteins, based on the principle that the tertiary structure of a protein is inextricably linked with its molecular function. We were the first to obtain models of tertiary structures and describe the putative functions of HSTV nonstructural proteins (NS3 helicase, NS3 protease, NS5 RNA-dependent RNA-polymerase, and NS5 methyltransferase), which play a key role in viral genome replication. Our results may help in further taxonomic identification of HSTV and the development of direct-acting antiviral drugs, POC tests, and vaccines.

The Yezo virus (YEZV) is a recently discovered tick-borne orthonairovirus with pathogenic potential, causing acute febrile illness in humans. Viral nucleoproteins (N) play a key role in genome packaging, replication, and modulation of host immune responses, making their structural characterization essential for understanding viral pathogenesis and developing targeted countermeasures. However, the absence of structural data for YEZV proteins significantly hinders these efforts. This study presents the first solution structure of the YEZV N domain 1 (D1). A highly purified, soluble, tag-free recombinant YEZV N D1 was produced from the native sequence of the clinical YEZV isolate. The native-state conformation was resolved through an integrated approach combining size-exclusion chromatography coupled with small-angle X-ray scattering (SEC-SAXS), AlphaFold 3 structure prediction, and all-atom molecular dynamics simulations. The YEZV N D1 structure adopts a stable, predominantly α-helical globular fold that remains monomeric under near-physiological conditions. SEC-SAXS data show excellent agreement with computational models, revealing moderate conformational flexibility. The characterized recombinant YEZV N D1 and its first solution structure reported here providing essential insights into understanding of YEZV molecular architecture. These findings lay a foundation for rational serological assay development and structure-guided therapeutic design against this and other emerging orthonairoviruses.