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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.

Relevance. The epidemiological situation in the world is characterized by an increase in the incidence of orthopoxvirus infections in humans and animals. In this regard, it is necessary to develop new safe vaccines against these infections.Aim. Conducting preclinical studies on the specific activity of the live vaccine against smallpox and other orthopoxvirus infections VACΔ6 based on the vaccinia virus (VACV) with six deleted virulence genes are presented.Matherials and methods. The studies were performed in accordance with the requirements of the Guidelines for conducting preclinical studies of drugs (immunobiological preparations), the State Pharmacopoeia XIII and the European Pharmacopoeia 7.0.Results and discussion. The vaccine strain VACΔ6 VACV showed significantly reduced neurovirulence in the model of intracerebral administration to suckling mice, and reduced inflammatorynecrotic activity in the model of intradermal administration to rabbits compared to the classical first­generation smallpox live vaccine approved for use in Russia. Preclinical studies of three series of the finished dosage form of the VAC∆6 vaccine showed its authenticity, thermal stability, non­pyrogenicity and safety. Double intradermal vaccination of rabbits at a dose of 106 PFU/animal, a 100% protective effect was provided against the intranasal infection of rabbits with VACV strain HB­-92 at a dose of 1995 LD50 and a double intradermal vaccination of mice at a dose of 106 PFU/animal, full protection was provided against the intranasal infection of mice with ectromelia virus strain K1 at a dose of 56 LD50.Conclusion. Based on the conducted complex of studies, it can be concluded that the created vaccine of the fourth generation VACΔ6 is safer compared to the live smallpox vaccine of the first generation and is not inferior to it in immunogenic and protective properties.

Due to cessation of mass smallpox vaccination in 1980, the collective immunity of humans against orthopoxvirus infections has virtually been lost. Therefore, the risk of spreading zoonotic human orthopoxvirus infections caused by monkeypox and cowpox viruses has increased in the world. First-generation smallpox vaccines based on Vaccinia virus (VAC) are reactogenic and therefore not suitable for mass vaccination under current conditions. This necessitates the development of modern safe live vaccines based on VAC using genetic engineering. We created the VACΔ6 strain by transient dominant selection. In the VACΔ6 genome, five virulence genes were intentionally deleted, and one gene was inactivated by inserting a synthetic DNA fragment. The virus was passaged 71 times in CV-1 cells to obtain the VACΔ6 strain from the VAC LIVP clonal variant. Such a long passage history might have led to additional off-target mutations in VACΔ6 compared to the original LIVP variant. To prevent this, we performed a genome-wide sequencing of VAC LIVP, VACΔ6, and five intermediate viral strains to assess possible off-target mutations. A comparative analysis of complete viral genomes showed that, in addition to target mutations, only two nucleotide substitutions occurred spontaneously when obtaining VACΔ4 from the VACΔ3 strain; the mutations persisting in the VACΔ5 and VACΔ6 genomes. Both nucleotide substitutions are located in intergenic regions (positions 1431 and 189738 relative to LIVP), which indicates an extremely rare occurrence of off-target mutations when using transient dominant selection to obtain recombinant VAC variants with multiple insertions/deletions. To assess the genome stability of the resulting attenuated vaccine strain, 15 consecutive cycles of cultivation of the industrial VACΔ6 strain were performed in 4647 cells certified for vaccine production in accordance with the “Guidelines for Clinical Trials of Medicinal Products”. PCR and sequencing analysis of six DNA fragments corresponding to the regions of disrupted genes in VACΔ6 showed that all viral DNA sequences remained unchanged after 15 passages in 4647 cells.

Pathogenic hantaviruses, belonging to the family Hantaviridae, genus Orthohantavirus, are widely spread in many regions of the world and cause hemorrhagic fever with renal syndrome (HFRS) in Europe and Asia. In the European Russia, the most active HFRS foci are located in the optimum habitat area of natural reservoir of the Puumala virus (PUUV), bank voles (Myodes glareolus), – in the Middle Volga and Cis-Urals. The largest number of cases of HFRS was registered in the Volga Federal District. In 2019, an outbreak of HFRS was registered among the residents of Saratov and the Saratov Region, the number of cases was 2702. Objective of the study was genetic identification of hantaviruses from HFRS patients and rodent carriers and phylogenetic analysis of full-size genomes from natural hosts during HFRS outbreak in the Saratov Region, 2019. Materials and methods. Blood samples of 8 HFRS patients from Saratov and 3 lung samples of bank voles captured in the suburb of Saratov were analyzed using reverse transcription polymerase chain reaction, followed by sequencing and phylogenetic analysis. Results and discussion. A total of 6 viral RNA isolates from HFRS patients were genetically typed, full-length RNA-isolate genomes were obtained for 3 natural carriers. Our data indicate that PUUV virus was associated with HFRS outbreak in Saratov. Genetic analysis revealed that the virus belonged to RUS lineage of PUUV, related most closely to strains from Udmurtia and Tatarstan and differed from strains circulating in the territory of Bashkortostan and Samara Region.

The eradication of smallpox has become one of the greatest successes of modern health science. This great achievement was made possible thanks to the widespread vaccination of the population. The last case of human infection with smallpox virus occurred in 1977. In 1980, at the 33rd session of the World Health Assembly, routine vaccination against that infection was recommended to be discontinued due to severe post-vaccination complications. However, humanity remains vulnerable to other orthopoxvirus infections closely related to smallpox virus. Recently, the cases of human infection with ortopoxviruses such as monkeypox virus, cowpox virus, vaccinia virus have become more frequent. Also, cases of infection of people with previously unknown orthopoxvirus species are recorded. Zoonotic orthopoxviruses pathogenic for humans, circulating in nature, require a detailed study and monitoring of the emergence of new strains. Their occurrence against the background of the cessation of planned vaccination of the population against smallpox virus can lead to the emergence of new highly pathogenic viruses. This review contains information on cases of human infection with orthopoxviruses around the world for the period 2008–2018. It also describes epidemiological anamnesis and the relations between cases of human infection in different countries due to the spread of viruses over a wide area, the movement of people between countries, population contacts with domestic and wild animals. Also, this paper provides information on the infection of people with previously unknown strains of orthopoxviruses.