Human seasonal coronaviruses (hCoVs) are a group of viruses that affect the upper respiratory tract. While seasonal patterns and the annual variability of predominant hCoV species are well-documented, their genetic and species diversity in St. Petersburg and across Russia remains largely unexplored. In this study, we developed a two-pool, long-amplicon (900–1100 bp) PCR primer panel for the whole-genome sequencing of four seasonal hCoV species. The panel was validated using nasopharyngeal swab samples collected within the Global Influenza Hospital Surveillance Network (GIHSN) project. Over a period of six epidemiological seasons from 2017 to 2023, we retrospectively analyzed 14,704 nasopharyngeal swabs collected from patients hospitalized in St. Petersburg clinics. Of these samples, 5010 (34.07%) tested positive for respiratory viruses, with 424 (2.88% of all samples) identified as seasonal human coronaviruses. The assessment of species diversity showed that predominant hCoV species alternate between seasons. Whole-genome sequences for 85 seasonal human coronaviruses (hCoVs) with >70% genome coverage were obtained, including 23 hCoV-OC43, 6 hCoV-HKU1, 39 hCoV-229E, and 17 hCoV-NL63. These represent the first near-complete genomes of seasonal hCoVs from the Russian Federation, addressing a significant gap in the genomic epidemiology of these viruses. A detailed phylogenetic analysis of the sequenced genomes was conducted, highlighting the emergence of hCoV-229E subclades 7b.1 and 7b.2, which carry numerous substitutions in the Spike protein. Additionally, we sequenced a historical hCoV-229E isolate collected in the USSR in 1979, the oldest sequenced 229E virus from Eurasia, and demonstrated that it belongs to Genotype 2. The newly developed PCR-based sequencing protocol for seasonal hCoVs is straightforward and well-suited for genomic surveillance, providing a valuable tool to enhance our understanding of the genetic diversity of human seasonal coronaviruses.

Human adenoviruses (HAdVs) are globally distributed pathogens capable of causing a wide range of clinical manifestations, particularly acute respiratory infections. However, their genomic diversity remains insufficiently characterized, with substantial geographic gaps in available sequence data, including for Russia, where only a few complete genomes have been deposited prior to this work. In this study, we analyzed more than 1200 PCR-positive respiratory specimens collected from hospitalized patients within routine surveillance projects and the Global Influenza Hospital Surveillance Network (GIHSN) across plenty of Russian regions during 2023–2024. Virus isolation followed by next-generation sequencing yielded 128 complete HAdV genomes representing species B, C, and D. The dataset included 27 B3, 9 B7, 44 B55, 12 C1, 16 C2, 4 C5, 7 C89, 5 C108, and one D109 genome, as well as three unassigned recombinant viruses with p89h5f5, p5h6f6 and p5h57f6 genomic structures (p, penton base; h, hexon; f, fiber). Phylogenetic analyses of whole genomes and capsid genes revealed extensive variability in immunogenic regions, particularly in species C, and identified clusters within B3 viruses. Notably, HAdV-D109 was identified in Russia, marking only the second reported detection of this genotype worldwide. Together, these findings substantially expand the currently available genomic landscape of HAdVs, highlighting the circulation of diverse and recombinant strains in Russia.

Human parainfluenza viruses 3 (hPIV3) are important pathogens, responsible for acute respiratory tract diseases, especially in young children. Information on hPIV3 circulation and their diversity pattern in Russia is limited. The aim of this study was to perform a molecular and genetic characterization of hPIV3 circulating in Saint Petersburg, Russia. From October 2017 to September 2023, 14,704 swabs were screened using real-time reverse transcription-PCR. A phylogenetic analysis of the complete hemagglutinin–neuraminidase (HN) gene was performed. Out of 1334 positive hPIV cases, hPIV3 was the most common subtype. Phylogenetic analysis of the studied and previously published HN sequences revealed four distinct genetic clusters, A, B, C, and D, with Cluster D being first delineated in this study. In addition, two newly subdivided genetic lineages, C5a and C5b, were documented. Phylogenetic analysis revealed that the analyzed Russian strains grouped into Cluster C and D; further subclusters C5a, C5b, C3b, C3e, and C3a. While three strains were classified within cluster D, the majority of isolates fell within subcluster C3a, followed by C5b. Taken together, these findings demonstrate the co-circulation of hPIV3 strains during the study period. This is the first study that describes the genetic and molecular aspects of hPIV3 circulating in Russia. Moreover, our results provide an up-to-date hPIV3 phylogenetic analysis.

Human parainfluenza viruses (hPIVs) are major contributors to respiratory tract infections in young children worldwide. Despite their global significance, genomic surveillance of hPIV1 and hPIV2 had not previously been conducted in Russia. This study aimed to develop a robust amplicon-based sequencing protocol for these viruses. The designed primer sets were tested on clinical samples containing hPIV RNA to evaluate their performance and efficiency. Sequencing results demonstrated high-quality genome data and efficient amplification across various Ct values. As a result, 41 hPIV1 and 13 hPIV2 near-complete genome sequences were successfully obtained from clinical specimens collected in Saint Petersburg (Russia). Phylogenetic analysis of the HN gene sequences showed that Russian hPIV1 strains clustered into clades II and III, while hPIV2 strains were distributed between clusters G1a and G3. The whole-genome-based trees confirmed the same distribution of the strains. These findings highlight the potential of our primer panels and contribute to a better understanding of the molecular characteristics and phylogenetic diversity of circulating hPIV strains. Notably, this study presents the first evolutionary analysis of hPIVs in Russia.

Introduction. Human metapneumovirus (hMPV) holds significant epidemiological importance, being a dominant cause of lower respiratory tract infections in children under two years of age and individuals over 65. Multiple infections with hMPV throughout a person’s life are possible due to the antigenic and genetic variability of the virus. However, the genetic variability of hMPV circulating in Russia remains unexplored. Aim of the study. The aim of this study was to test a protocol for whole-genome sequencing of hMPV to assess the genetic diversity of metapneumoviruses circulating in certain regions of Russia. Materials and methods. Nasopharyngeal swabs were collected from patients of different ages with acute respiratory viral infections (ARVI) tested positive for hMPV using polymerase chain reaction (PCR). From some of the samples, viral isolates were obtained in cell culture. Whole-genome sequencing was performed on both swabs and isolates using the MiSeq Illumina platform, followed by phylogenetic analysis. Results. For the first time in Russia, whole-genome sequencing of 44 hMPV strains circulating from 2017 to 2024 was conducted. Their genetic group affiliation was described, with the A2b2 clade shown to dominate. It was confirmed that the greatest variability among genes encoding viral surface proteins was observed in the G gene, while changes in the F gene were minimal during the studied period. Conclusion. The study provides insights into the genetic diversity of hMPV strains circulating in various regions of the Russian Federation. Understanding the genetic variability of hMPV is crucial for comprehending viral evolution, transmission dynamics, and mechanisms of immune evasion, which influence the development of vaccines and antiviral drugs.

Introduction. Acute respiratory infections (ARI) are one of the main causes of morbidity and mortality from infectious diseases in the world. Respiratory infections can be caused by pathogens of various etiologies: viruses, bacteria, mycoplasmas, etc. Rapid and accurate identification of pathogens, such as bacteria, in biological samples is an important task, for which 16S rRNA gene sequencing using new generation platforms is used.The objective was a comparative analysis of the qualitative characteristics of the oropharyngeal microbiome of healthy volunteers and patients with ARI of unknown etiology based on the 16S rRNA gene sequencing.Methods and materials. Using V3–V4 region of 16S rRNA Illumina MiSeq sequences from oropharyngeal swabs, we analyzed the microbiome of hospitalized patients with ARI symptoms and healthy patients.Results. In this study, we conducted V3–V4 region of 16S rRNA sequencing analyses of the oropharyngeal samples from 116 hospitalized patients with ARI symptoms and 81 healthy patients. Patients with ARI exhibited higher abundance of opportunistic pathogens, particularly Staphylococcus, Ralstonia, Aeribacillus, Acinetobacter baumannii, Methylobacterium-Methylorubrum, Rhodococcus equi. In the control samples, normal commensal respiratory tract microbiota, such as Neisseria, Prevotella, Fusobacterium, Veilonella was dominated.Conclusions. The microbiota samples of hospitalized patients with ARI showed a predominance of opportunistic and potentially pathogenic microbiota, while normal representatives of the respiratory tract microbiota predominate in healthy volunteers. For a more detailed analysis, data on the species composition of the microbiota is required, which can be obtained by sequencing the complete sequence of the 16S rRNA gene.