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.

Tick-borne phleboviruses represent emerging pathogens with zoonotic potential, yet their distribution across Asian Russia remains poorly characterized. This study investigated the prevalence, genetic diversity, and evolutionary dynamics of phleboviruses in 1,078 individual Ixodes persulcatus ticks collected from 143 locations across Asian Russia during summer 2023. Samples underwent PCR screening, high-throughput sequencing for genome reconstruction, phylogenetic analysis, and AlphaFold protein structural modeling. We detected 27 phleboviral isolates belonging to Phlebovirus mukawaense (MKWV), Mudanjiang phlebovirus, Onega tick phlebovirus, and Gomselga virus, with prevalence rates of 0.6%, 0.1%, 0.3%, and 1.5%, respectively. Phylogenetic analysis suggested Gomselga virus belongs within the MKWV species complex. Complete coding sequences of Russian MKWV isolates enabled high-confidence structural predictions for key proteins. The predicted tertiary structure of the MKWV nucleoprotein exhibits strong similarity to those of Rift Valley fever virus and Toscana virus, despite limited sequence identity. The MKWV nucleoprotein features a conserved globular core with a well-defined RNA-binding cleft. Segment-specific phylogenetic incongruence among Primorsky MKWV isolates indicated potential reassortment events. Additionally, coinfections involving MKWV and Alongshan virus, Borrelia miyamotoi, or Rickettsia spp. were identified. These findings significantly expand the known geographic range and genetic diversity of tick-borne phleboviruses in Russia. The integration of genomic and structural data provides a robust framework for functional annotation and highlights the evolutionary stability of essential viral domains. Further research should focus on virus isolation and investigating antigenic properties and replication capacity in mammalian cells.

High-throughput sequencing methods have made it possible to identify numerous novel tick-borne viruses that are potentially pathogenic to humans. Among these, Songling virus (Orthonairovirus songlingense, SGLV) has been associated with febrile illness in patients following tick bites in China, but its geographic distribution outside China remains largely unexplored. In this study, we aimed to detect SGLV circulation in ticks across Asian Russia, focusing on regions bordering China. A total of 3444 adult ticks representing six species were collected from 170 locations across 11 regions during the summer of 2024. SGLV RNA was detected in Haemaphysalis concinna ticks, with 11 positive specimens yielding an SGLV RNA prevalence rate of 2.2%. Positive ticks were found in four regions, with the highest positivity rate (5.8%) recorded in Amur Oblast, which directly borders China. The detection of SGLV in the Republic of Altai represents the westernmost record of this virus to date. Full-length nucleoprotein-coding sequences obtained for all Russian isolates revealed up to 1.2% nucleotide divergence. Phylogenetic analysis confirmed that all Russian SGLV isolates belong to Orthonairovirus songlingense, with the Altai SGLV isolate showing genetic similarity to a human-derived Chinese SGLV isolate. Co-infections with Rickettsia heilongjiangensis were detected in four SGLV-positive ticks, highlighting the potential for simultaneous pathogen transmission. These findings establish the first evidence of SGLV circulation in Russia across a wide geographic range and underscore the need for differential diagnosis of febrile illnesses following tick bites in this region.

This study reports the design, synthesis, and comprehensive biological evaluation of a new class of orthopoxvirus inhibitors utilizing natural bicyclic monoterpenoid scaffolds as strategic replacements for synthetic polycyclic cores. A focused library of amides based on camphor, apo-/isocamphane, and nopol-derived amines was synthesized and screened for activity against vaccinia (VACV), cowpox (CPXV), ectromelia (ECTV) and variola (VARV) viruses. Structure-activity relationship (SAR) analysis identified the importance of a para-substituted electron-withdrawing group on the benzamide moiety, with the apocamphanyl amine series yielding the most potent candidate, compound 13d. Compound 13d exhibited potent in vitro antiviral activity with IC50 values of 0.32 μM (VACV), 4.43 μM (CPXV), 3.28 μM (ECTV), and 1.6 μM (VARV) coupled with high selectivity indices (up to 1226). Further profiling revealed metabolic stability of 13d in mouse blood and favorable pharmacokinetics in mice following a single intragastric dose (100 mg kg-1), characterized by rapid absorption and a long elimination half-life (T 1/2 ∼ 6.7 h), supportive of once-daily dosing. Computational studies involving pharmacophore analysis, molecular docking, and MD simulations elucidated the binding mode to the p37 phospholipase domain and rationalized the observed SAR, highlighting the critical role of hydrophobic interactions and stereochemistry. These findings validate the bicyclic monoterpenoid platform as a promising strategy for discovering potent, metabolically stable anti-orthopoxvirus agents and establish 13d as a compelling lead candidate for further preclinical development.

Studies of HIV-1 molecular epidemiology describe significant differences in HIV infection spread across geographical areas. We examined 80 HIV-1 samples from the Jewish Autonomous Region of Russia in 2024. HIV-1 genome sequences for 12 samples revealed a novel HIV-1 called CRF173_63A6. HIV-1 CRF173_63A6 was found to have arisen through recombination between a specific Russian A6 subtype and the recombinant virus CRF63_02A6, which is responsible for the PWID-associated HIV outbreak in the Siberian region of Russia. Phylogenetic analysis of pol sequences previously deposited in Genbank showed that the CRF173_63A6 samples we described are grouped into a common phylogenetic cluster that includes 54 HIV-1 samples isolated in the JAR and other areas of the Russian Far East, indicating a wide distribution of this virus genovariant. This study once again proves the significant contribution of the key PWID group not only to the development of local Russian HIV epidemics, but also to the change in the characteristics of the circulating virus population.

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.

Haseki tick virus (HSTV) is a recently discovered virus detected in human serum following tick bites, yet its protein repertoire remains uncharacterized. In this study, we applied an integrative approach based first on membrane topology prediction, followed by AI-based structural prediction and experimental validation to annotate the structural part of the HSTV polyprotein. For the first time, we recombinantly expressed one of the putative HSTV structural protein (SP1) and determined its overall architecture using small-angle X-ray scattering (SAXS). Structural comparisons of the AI-predicted HSTV SP1 models revealed only a vague resemblance to the pestiviral Erns and Npro. The strong agreement between experimental SAXS data and the AI-predicted HSTV SP1 model supported the conclusion that HSTV SP1 adopts a distinct spatial architecture in solution, one that is not captured by existing pestiviral structures but is reliably represented by modern AI-based prediction. Our findings indicate that HSTV SP1 adopts a fold not previously observed among characterized members of the Flaviviridae family. This work establishes a methodological pipeline for characterizing highly divergent viral proteins and provides the first insights into HSTV SP1, a virus with emerging zoonotic potential. These results lay the foundation for future functional and structural studies, diagnostic development, and evolutionary analyses of atypical Flaviviridae family members.

A novel flavi-like virus with a segmented genome—Alongshan virus (ALSV)—has been isolated from Ixodes ticks in Russia. In this study, 4458 ixodid ticks collected in 22 regions of Russia were tested for genetic markers of ALSV by RT PCR. The highest rates of ALSV infection in ticks were detected in the Republic of Khakassia (3.3%) and in Kemerovo Oblast (2.4%), while low infection rates were more typical in the European part of Russia (0.4–0.7%). Complete four-segment genomes of 20 ALSV isolates derived from 22 PCR-positive Ixodes persulcatus ticks were sequenced using a high-throughput approach. The nucleotide sequences for Asian ALSV isolates have a 94.5–96.5% identity to ALSV isolates previously found in China, with this range for the European isolates being 89–93%. This data, together with phylogenetic analysis, indicates the existence of Asian and European subtypes of ALSV, and these may be associated with I. persulcatus and I ricinus ticks. The obtained results express the spread of ALSV in Russia and also may be useful for the diagnosis, prophylactics, and treatment of this infection.

In this study, we investigated the features of co-infection with SARS-CoV-2 and the enterovirus vaccine strain LEV8 of coxsackievirus A7 or enterovirus A71 for Vero E6 cells and Syrian hamsters. The investigation of co-infection with SARS-CoV-2 and LEV-8 or EV-A71 in the cell model showed that a competitive inhibitory effect for these viruses was especially significant against SARS-CoV-2. Pre-infection with enteroviruses in the animals caused more than a 100-fold decrease in the levels of SARS-CoV-2 virus replication in the respiratory tract and more rapid clearance of infectious SARS-CoV-2 from the lower respiratory tract. Co-infection with SARS-CoV-2 and LEV-8 or EV-A71 also reduced the severity of clinical manifestations of the SARS-CoV-2 infection in the animals. Additionally, the histological data illustrated that co-infection with strain LEV8 of coxsackievirus A7 decreased the level of pathological changes induced by SARS-CoV-2 in the lungs. Research into the chemokine/cytokine profile demonstrated that the studied enteroviruses efficiently triggered this part of the antiviral immune response, which is associated with the significant inhibition of SARS-CoV-2 infection. These results demonstrate that there is significant viral interference between the studied strain LEV-8 of coxsackievirus A7 or enterovirus A71 and SARS-CoV-2 in vitro and in vivo.

Human rhinovirus picornain 3C is a high-value commercial cysteine protease, which is widely used to remove affinity tags and fusion proteins during the purification of the target proteins. A variant of rhinovirus A28 picornain 3C produced in this study is not annotated in the NCBI databases, shares 79% sequence identity in the PDB, and was not previously used in the protein engineering. A protocol was developed for the isolation and purification of the protein to use it in structural studies. The initial crystallization conditions were found. The determination and analysis of the structure of rhinovirus A28 picornain 3C will provide new possibilities for performing basic research on the evolution of proteolytic enzymes and for the design of the optimal variant of this protease.

Introduction. Since early May 2022, more than 90,000 cases of monkeypox virus infection have been reported in more than 70 countries around the World. This is the largest outbreak of monkeypox ever recorded outside of Africa. The aim of the study is to confirm the first case of monkeypox in Russia, to isolate and sequence a new strain of monkeypox virus (MPXV), and to assess its sensitivity to the 7-[N-(4-trifluoromethylbenzoyl)-hydrazinocarbonyl]-tricyclo-[3.2.2.0^2,4]non-8-en-6-carboxylic acid (NIOCH-14) antipox drug. Materials and methods. The biological materials obtained from the affected area of the skin (contents of vesicles), a nasopharyngeal smear, sputum and venous blood from a patient with suspected monkeypox were used. The disease was confirmed by PCR followed by determination of the nucleotide sequence of viral DNA by sequencing. Isolation of the new MPXV strain from clinical samples was carried out in Vero E6 cells. The antiviral effectiveness of NIOCH-14 against the new MPXV strain was assessed using an adapted spectrophotometric method. Results. A diagnostic study of the biological samples of a patient who returned from a tourist trip to European countries with complaints of skin rashes all over the body revealed MPXV DNA. A new strain of MPXV was isolated from vesicles in Vero E6 cells, and the genomic sequence MPXV-pustule S45 was assembled using high-throughput parallel sequencing (NGS). Discussion. The effectiveness of the finished dosage form of NIOCH-14 against the new strain of MPXV based on the results of determining the 50% virus inhibitory concentration (IC50) was 0.02 μg/mL, and the selectivity index (SI) was 15,000. Conclusion. In this study, the pathogen of monkeypox was detected and identified using real-time PCR, NGS and electron microscopy, and the first imported case of this disease in Russia was confirmed. It has been proven that the drug NIOCH-14 exhibits high antiviral activity in vitro against the new MPXV strain.

Scientific interest in orthopoxvirus infections and search for new highly effective compounds possessing antiviral activity against orthopoxviruses have significantly increased as a result of worldwide mpox outbreak in 2022. The present work deals with the synthesis of new 2-arylimidazoles exhibiting in vitro activity not only against the vaccinia virus, cowpox virus and ectromelia (mousepox) virus but also against the variola virus. Among the imidazole derivatives under consideration (1-hydroxyimidazoles, 1-methoxyimidazoles, 1-benzyloxyimidazoles, and imidazole N-oxides), the most promising antiviral activity is demonstrated by 1-hydroxyimidazoles, which may exist as two prototropic tautomers. Both of these tautomers may be manifested in different crystal structures of these compounds, according to single-crystal X-ray diffraction analysis, while predominantly one of them (N-hydroxy-tautomeric form) is present in DMSO-d 6 solutions and in the gaseous state, as shown by NMR spectroscopy and quantum-chemical calculations. The leader compound 1-hydroxy-2-(4-nitrophenyl)imidazole 4a demonstrated the highest selectivity indices against the vaccinia virus (SI = 1072) and the variola virus (SI = 373).

Introduction. Obtaining stabilized recombinant HIV-1 Env trimers that have a close to the native conformation is one of the directions in the field of development of vaccines against HIV-1. The aim of the study was to obtain and characterize the stabilized trimer Env of HIV-1 SOSIP.664 based on the circulating genetic variant of the recombinant form CRF63_02A6. Materials and methods. Bioinformatics resources were used to design the trimer Env gene based on the HIV-1 recombinant genetic variant CRF63_02A6. The designed gene was synthesized and cloned as part of an integration plasmid vector. A stable producer of trimer Env was obtained by transfection of the CHO-K1 cell line using the developed plasmid vector. Purification of the protein complex was performed using affinity chromatography and gel filtration. Antigenic properties of trimer Env were studied using immunochemical analysis using broadly neutralizing HIV-1 monoclonal antibodies (bnAbs). Results. A new variant of the stabilized trimer Env of the surface glycoprotein of the recombinant form CRF63_02A6 HIV-1 SOSIP.664 was designed, including additional stabilizing modifications. Based on the CHO-K1 cell line, a stable producer was obtained and a purification protocol for the designed trimer Env was developed. It was found that the trimer Env CRF63_02A6 SOSIP.664 is effectively recognized by bnAbs 2G12, VRC01 and PGT126. Conclusion. The obtained results indicate the prospects for further study of the structural features of the trimer Env CRF63_02A6 SOSIP.664, as well as its immunogenicity and the possibility of using it as a vaccine antigen.

Extensive spread of tick-borne diseases poses a significant problem for public health and the health of the population living in endemic areas.The aim of the study was to search, analyze genetic material and identify new viral agents of the Phenuiviridae family in taiga ticks collected in Asian regions of Russia using the method of high throughput sequencing.Materials and methods. The study involved 1460 taiga ticks collected in suburban areas of the Tomsk, Yekaterinburg and Primorsky Territory. The genetic material isolated from ticks was sequenced using Illumina technology followed by phylogenetic analysis.Results and discussion. Analysis of the sequencing results made it possible to detect extended nucleotide sequences of the L-gene fragment characteristic of the Phenuiviridae family viruses. We were able to identify 20 nucleotide sequences the length of 250 bp on average in homogenates of Ixodes persulcatus ticks. Eighteen isolates have been identified as members of the genus Uukuvirus and two isolates have been assigned to the genus Phlebovirus, Phenuiviridae family. Phylogenetic analysis has shown that all isolates of the genus Uukuvirus fall under the cluster of Tacheng tick virus 2 belonging to the species Tacheng uukuvirus. They form a separate phylogenetic group which is closely related to two Romanian variants of 2019. Tacheng tick virus 2 was detected in all three surveyed regions of the Asian part of Russia. Two Tomsk isolates of phlebovirus were classified as Sara tick phlebovirus and they clustered with two isolates of phleboviruses from Karelia. Thus, the genetic material of Tacheng tick virus 2 and Sara tick phlebovirus belonging to two genera of the family Phenuiviridae was found in I. persulcatus ticks collected in three geographically different regions of the Asian part of Russia.

— The new coronavirus pneumonia has rapidly spread around the world. The World Health Organization emphasized that the SARS-CoV-2 coronavirus spreads mainly between people who are in close contact with each other, as well as in the case of touching contaminated surfaces followed by touching the eyes, nose, or mouth without first cleaning the hands. Possible permanent sources of the spread of the virus can be gathering of patients in hospitals in the case of noncompliance with the requirements for organizing the functioning of a hospital. Meteorological conditions can be a key factor influencing the spread of the virus in the case of an accidental release of virus-containing aerosol from such a hospital. Simulations are carried out with modern methods for solving a system of differential equations of the atmospheric boundary layer, which are adapted to describe the distribution of harmful atmospheric impurities over a real complex terrain considering urban buildings of various heights, forests, reservoirs, changing meteorological conditions, and many other factors.

Background: Since most of the modern human population has no anti-smallpox immunity, it is extremely important to develop and implement effective drugs for the treatment of smallpox and other orthopoxvirus infections. The objective of this study is to determine the main characteristics of the chemical substance NIOCH-14 and its safety and bioavailability in the body of laboratory animals. Methods: The safety of NIOCH-14 upon single- or multiple-dose intragastric administration was assessed according to its effect on the main hematological and pathomorphological parameters of laboratory mice and rats. In order to evaluate the pharmacokinetic parameters of NIOCH-14 administered orally, a concentration of ST-246, the active metabolite of NIOCH-14, in mouse blood and organs was determined by tandem mass spectrometry and liquid chromatography. Results: The intragastric administration of NIOCH-14 at a dose of 5 g/kg body weight caused neither death nor signs of intoxication in mice. The intragastric administration of NIOCH-14 to mice and rats at doses of 50 and 150 µg/g body weight either as a single dose or once daily during 30 days did not cause animal death or critical changes in hematological parameters and the microstructure of internal organs. The tissue availability of NIOCH-14 administered orally to the mice at a dose of 50 µg/g body weight, which was calculated according to concentrations of its active metabolite ST-246 for the lungs, liver, kidney, brain, and spleen, was 100, 69.6, 63.3, 26.8 and 20.3%, respectively. The absolute bioavailability of the NIOCH-14 administered orally to mice at a dose of 50 µg/g body weight was 22.8%. Conclusion: Along with the previously determined efficacy against orthopoxviruses, including the smallpox virus, the substance NIOCH-14 was shown to be safe and bioavailable in laboratory animal experiments.

Background: Q fever is a zoonotic infectious disease characterized by fever, malaise, chills, significant weakness, and muscle pain. In some cases, the disease can become chronic and affect the inner membranes of the heart, such as the valves, leading to endocarditis and a high risk of death. Coxiella burnetii (C. burnetii) is the primary causative agent of Q fever in humans. This study aims to monitor the presence of C. burnetii in ticks collected from small mammals and cattle in the Republic of Guinea (RG). Methods: Rodents were trapped in the Kindia region of RG during 2019–2020, and ticks were collected from cattle in six regions of RG. Total DNA was extracted using a commercial kit (RIBO-prep, InterLabService, Russia) following the manufacturer’s instructions. Real-time PCR amplification was conducted using the kit (AmpliSens Coxiella burnetii-FL, InterLabService, Russia) to detect C. burnetii DNA. Results and Conclusions: Bacterial DNA was detected in 11 out of 750 (1.4%) small mammals and 695 out of 9620 (7.2%) tick samples. The high number of infected ticks (7.2%) suggests that they are the main transmitters of C. burnetii in RG. The DNA was detected in the liver and spleen of a Guinea multimammate mouse, Mastomys erythroleucus. These findings demonstrate that C. burnetii is zoonotic in RG, and measures should be taken to monitor the bacteria’s dynamics and tick prevalence in the rodent population.

The Zika virus (ZIKV) is a widespread mosquito-borne pathogen. Phylogenetically, two lineages of ZIKV are distinguished: African and Asian–American. The latter became the cause of the 2015–2016 pandemic, with severe consequences for newborns. In West African countries, the African lineage was found, but there is evidence of the emergence of the Asian–American lineage in Cape Verde and Angola. This highlights the need to not only monitor ZIKV but also sequence the isolates. In this article, we present a case report of Zika fever in a pregnant woman from Guinea identified in 2018. Viral RNA was detected through qRT-PCR in a serum sample. In addition, the seroconversion of anti-Zika IgM and IgG antibodies was detected in repeated blood samples. Subsequently, the virus was isolated from the C6/36 cell line. The detected ZIKV belonged to the African lineage, the Nigerian sublineage. The strains with the closest sequences were isolated from mosquitoes in Senegal in 2011 and 2015. In addition, we conducted the serological screening of 116 blood samples collected from patients presenting to the hospital of Faranah with fevers during the period 2018–2021. As a result, it was found that IgM-positive patients were identified each year and that the seroprevalence varied between 5.6% and 17.1%.

The spread of the disease caused by monkeypox virus (MPox) since 2022 has shown the urgency of developing countermeasures. The development of modern methods of clinical laboratory diagnostics of MPox contributes to this. Enzyme-linked immunosorbent assay (ELISA) is an accessible and sensitive platform for developing diagnostic tools. Detection of MPox antigens using ELISA kits based on monoclonal antibodies (MAbs) is promising due to the quick time of analysis and minimal requirements for sample preparation. We have developed and deposited two strains of Escherichia coli that produce recombinant proteins. Mice were immunized with the AgPOX protein, which contains unique antigenic sequences of MPox. The Trx + A29 protein for selecting MAb producers includes the original amino acid sequence A29L. The absence of antibody crossover to Trx protein and native preparations of variola virus and vaccinia virus tested by ELISA. As a result of hybridization of splenocytes from immunized mice, MAb producers were obtained. Fifteen MAb-producing hybridomas were selected based on ELISA results with three specific MPox antigens and three nonspecific ones. Three hybridomas were selected for deposit according to the productivity criteria. The possibility of detection by means of its MAbs of the native MPox antigen at various concentrations was tested and method sensitivity was determined. The MAbs a-A29L_MPoxV of three hybridomas detected the native antigen MPox at a concentration of 102 PFU/mL. It is likely that the method is even more sensitive when selecting analysis conditions. Based on labeled MAbs a-A29L_MPoxV, it is possible to develop a sensitive and specific indirect two-step ELISA kit for immunodiagnostics of MPox.

Despite the fact that the variola virus is considered eradicated, the search for new small molecules with activity against orthopoxviruses remains an important task, especially in the context of recent outbreaks of monkeypox. As a result of this work, a number of amides of benzoic acids containing an adamantane fragment were obtained. Most of the compounds demonstrated activity against vaccinia virus, with a selectivity index SI = 18,214 for the leader compound 18a. The obtained derivatives also demonstrated activity against murine pox (250 ≤ SI ≤ 6071) and cowpox (125 ≤ SI ≤ 3036). A correlation was obtained between the IC50 meanings and the binding energy to the assumed biological target, the p37 viral protein with R2 = 0.60.