Problems of Virology

The global rise in antimicrobial resistance (AMR) poses an urgent threat to public health, and novel alternatives to traditional antibiotics are needed. One of the most promising options is bacteriophages, viruses that infect and destroy bacteria. Once overshadowed by the discovery of antibiotics, phage therapy is now regaining attention, driven by advances in genomics, synthetic biology, and targeted medicine. This review examines the biology, diversity, and therapeutic use of bacteriophages in treating bacterial infections, especially those caused by multidrug-resistant pathogens. It also discusses how phages act through natural mechanisms, such as lytic enzymes (holins, endolysins, and muralysins), and highlights new genetic engineering techniques, such as CRISPR-Cas systems, phage recombineering, and synthetic genome reboots. In addition to clinical applications, we evaluate phages as biocontrol agents for food safety, environmental sanitation, and biofilm management. Additionally, the article explores key issues in phage therapy, including regulatory frameworks, formulation stability, dynamics of phage-host resistance, and the importance of rapid diagnosis. When properly integrated into modern health and biotechnology practices, bacteriophages offer significant potential and a sustainable solution to the global challenge of antimicrobial resistance.

Introduction. The spread of tick-borne encephalitis virus (Orthoflavivirus encephalitidis) continues. Existing vaccines have several limitations. Research new vaccine platforms will enable the development of more effective strategies of disease prevention.

Materials and methods. The constructs for producing vaccine preparations were obtained using the methods described earlier. Study of the effectiveness of candidate vaccines based on chimeric yellow fever virus, adenovirus serotype 25 and mRNA encoding a fragment of the tick-borne encephalitis viral polyprotein corresponding to the PrM and E regions, immunogenicity and protective efficacy was performed using BALB/c mice. The neutralizing activity assessment of animal sera and mice challenge were performed using the tick-borne encephalitis virus strain Sofjin-Chumakov.

Results. It was established that immunization of animals with mRNA encoding the PrM and E regions of the viral polyprotein of tick-borne encephalitis virus resulted in the induction of a higher level of neutralizing antibodies compared to the other vaccine platforms studied in the immunization schedule used. In addition, this platform provided 100% protection of animals from the lethal infection caused by the tick-borne encephalitis virus.

Conclusion. The study demonstrated that delivery of the nucleotide sequence encoding the PrM and E regions of the tick-borne encephalitis virus polyprotein within mRNA elicits a pronounced humoral immune response during animal immunization. These results demonstrate the fundamental feasibility of effectively expressing the same tick-borne encephalitis virus antigen in various vaccine platforms, each with its own advantages and limitations.

Objective. To evaluate how influenza infection affects the hypolipidemic effect of a novel combined drug – Prenophytol based on polyisoprenoids (sodium polyprenyl phosphate and beta-sitosterol) in mice with experimental dyslipidemia.

Materials and methods. In BALB/c mice with experimentally induced dyslipidemia, the impact of Prenophytol on lipid metabolism parameters was assessed. Currently the drug is at the stage of clinical trials (Roszdravnadzor's permission to conduct clinical trials No. 362 dated 08/14/2025). A subset of animals was infected with influenza A/California/07/2009 (H1N1) virus.

Results. Administration of the drug to dyslipidemia mice resulted in a significant reduction in total cholesterol, triglycerides, and LDL levels, along with a significant increase in HDL. However, in the presence of influenza infection, the hypolipidemic effect of Prenophytol was markedly attenuated: only cholesterol levels decreased significantly, whereas changes in triglycerides, LDL, and HDL did not reach statistical significance.

Conclusions. Influenza infection diminishes the therapeutic efficacy of the Prenophytol in the treatment of dyslipidemia. These findings indicate that concomitant viral infections must be taken into account when designing dyslipidemia treatment regimens and conducting clinical trials of this drug.

Introduction. Powassan virus (POWV) is a tick-borne orthoflavivirus capable of causing neurological diseases. In the Russian Far East, the area affected by POWV overlaps with that of another neurotropic orthoflavivirus, tick-borne encephalitis virus (TBEV). Currently, there are no differentiated test systems for specific detection of POWV in the presence of TBEV.

The aim of this work is to create differentiating test systems for the detection of POWV in a mixture with TBEV and other orthoflaviviruses.

Materials and methods. We constructed a genome alignment of eight POWV and five TBEV strains to select differentiating primers using MEGA X. To assess the specificity of the assay, we used POWV, several TBEV strains, and other orthoflaviviruses. PCR and qPCR were performed with the selected oligonucleotides, and the specificity of amplification was verified by electrophoresis in agarose gel and Sanger sequencing. cDNA was obtained from virus-containing material by isolating RNA and performing a reverse transcription reaction.

Results. The selected oligonucleotides for the qPCR-based differentiation system ensured specific detection of POWV without detecting signals from other orthoflaviviruses. The assay’s limit of detection was 10² copies/sample. Analysis of mixed samples containing POWV and TBEV demonstrated no distortion of the measurement results. Furthermore, differentiating oligonucleotides were selected and tested to amplify extended regions of the POWV and TBEV genomes for subsequent sequencing and phylogenetic analysis.

Conclusion. A laboratory technique based on PCR has been developed that allows to detect the POWV in samples containing other orthoflaviviruses.

Introduction. Rotaviruses exhibit a high degree of variability, which is ensured by the process of reassortment and antigenic drift. This leads to significant genetic diversity, including the emergence of strains with rare and unusual genotypes. This study, for the first time, provides a molecular genetic characterization of a rare RVA strain with the genotype G2P[9] based on all genome segments.

Aim of the study. To conduct a molecular genetic characterization of a rare strain of RVA genotype G2P[9] based on all genome segments, as well as to compare the strain with previously characterized representatives of subgroup BA222 from Nizhny Novgorod.

Materials and methods. Rotavirus-positive stool samples from children were analyzed using PCR-genotyping and PAGE. For the isolate under study, cDNA fragments of each of the 11 genes (VP1–VP4, VP6, VP7, NSP1–NSP5) were sequenced. Nucleotide and amino acid sequence analysis and phylogenetic tree construction were performed using MEGA X.

Results. In the period 2022–2023, a single strain of the unique genotype G2P[9] was identified in Nizhny Novgorod, characterized by a “broad” RNA electrophoretype (G2-P[9]-I2-R2-C2-M2-A3-N2-T3-E3-H3). The study showed that the origin of this strain is associated with reassortment processes between Nizhny Novgorod Wa- and BA222-like RVA, as well as isolates of the G3P[9] genotype from China. The studied strain, along with rotaviruses from China, differed from the Nizhny Novgorod BA222-like strains by the presence of 43 nucleotide substitutions in nine genes. These differences resulted in ten amino acid substitutions, eight of which were radical. All radical substitutions were located in the functionally active regions of the VP1, VP3, and VP4 proteins, which play a key role in the replicative cycle of rotaviruses.

Discussion. We characterized a strain with the rare G2P[9] genotype based on its complete genotype for the first time in Russia and globally. These results expand our understanding of the diversity of reassortant rotaviruses and complement our knowledge of the genotypic structure of the rotavirus population in Nizhny Novgorod.

Conclusion. The wide genetic diversity of rotaviruses, maintained by reassortment and antigenic drift, may facilitate rotaviruses' ability to overcome immunological pressure. Therefore, continuous molecular monitoring of circulating rotavirus variants is necessary to monitor the emergence of new variants and assess changes in rotavirus virulence following reassortment processes.