Neurotropic enteroviruses (Picornaviridae: Enterovirus): predominant types, basis of neurovirulence.

Enteroviruses are one of the most common causative agents of infectious diseases of the central nervous system. They are characterized by genetic variability, the ability to infect a wide range of cells, including brain microglial cells and astrocytes, and persist in the central nervous system tissue, causing delayed and chronic diseases. The review presents data on the basis of neurovirulence of non-polio enteroviruses and the most common pathogens causing enteroviral neuroinfections.

Phylodynamic characteristics of reassortant DS-1-like G3P[8]-strains of rotavirus type A isolated in Nizhny Novgorod (Russia).

Since 2013, there has been an increase in reports of the spread of a double intergroup reassortant strain of rotavirus type A (RVA) with the genotype G3P[8] and other genes belonging to the second genogroup I2-R2-C2-M2-A2-N2-T2-E2-H2. In our study, we provide a molecular genetic characterization of rotaviruses with genotype G3P[8]-I2 isolated in Nizhny Novgorod. In our study, we used RT-PCR, Sanger sequencing, RNA-PAGE methods. Phylogenetic and phylodynamic analysis were performed using the Bayesian approach. According to our study, there was a significant increase in the proportion of G3P[8] from 15% during the period of 2020-2021 to 53% during the period of 2021-2022 in Nizhny Novgorod, Russia. Phylogenetic analysis based on the VP4 gene revealed that DS-1-like RVAs isolated in Nizhny Novgorod belong to different clusters of the P[8]-3.1 lineage, with a level of variation ranging from 1.1% to 1.3%. Based on the VP6 gene, the equine-like RVAs identified by us carry genetic variants belonging to three distinct clusters of the lineage I2-V, with a variation level ranging from 2.0% to 4.5%. These data indicate the genotypic diversity of circulating DS-1-like G3 RVAs. Phylogenetic analysis of the VP7 gene allowed us to assign the isolates identified in our study to the G3-1 lineage. We estimated that the circulation of the most recent common ancestor of the spreading strains dates back to 2002. Additionally, we determined the typical level of mutations in the VP7 gene, which amounted to 2.14*10-3 substitutions/per site/per year.

The use of Dapagliflozin in Acute Decompensated Heart Failure: Results of the Randomized Study.

Aim      To determine the effect of sodium-glucose cotransporter-2 inhibitors (SGLT2i) on kidney function in acute decompensated heart failure (ADHF).Material and methods  A controlled randomized study on the dapagliflozin treatment in ADHF was performed. Patients were randomized to a main group (standard therapy supplemented with dapagliflozin) or a control group (standard therapy for ADHF). The primary endpoint was the development of acute kidney injury (AKI). 200 patients were included (mean age, 74±12 years; 51% men). 31% of patients had type 2 diabetes mellitus (DM2). Mean left ventricular ejection fraction (LV EF) was 47±14 %; in 44.5% of patients, LV EF was less than 45%. Median concentration of N-terminal pro-brain natriuretic peptide (NT-proBNP) was 5225 [3120; 9743] pg / ml, glomerular filtration rate (GFR) was 51 [38; 64] ml / min / 1.73 m2.Results In-hospital mortality was 6.5%. Analysis of the dynamics of body weight loss showed significant differences (4200 [2925; 6300] g vs. 3000 [1113; 4850] g; p=0.011) in favor of the dapagliflozin group. The requirement for increasing the daily dose of furosemide and adding an another class diuretic (thiazide or acetazolamide) did not differ between the groups. However, median furosemide dose during the stay in the hospital was lower in the dapagliflozin group (80 [67; 120] mg vs. 102 [43; 120] mg; p=0.016). At 48 hours after randomization, GFR significantly decreased in the dapagliflozin group (-5.5 [-11; 3] ml/min/ 1.73 m2) compared to the control group (-0.3 [-4; 5] ml / min/1.73 m2, р=0.012). Despite this, GFR did not differ between the groups at discharge (51 [41; 66] ml/min/1.73 m2 and 49 [38; 67] ml/min/1.73 m2, respectively; p = 0.84). In the dapagliflozin group, frequency of AKI episodes was not increased compared to the control group (13 and 9.4%, respectively; p = 0.45).Conclusion      The dapagliflozin treatment in ADHF is associated with more pronounced body weight loss and lower average doses of loop diuretics during the period of stay in the hospital, with no associated clinically significant impairment of renal function.

The use of Dapagliflozin in Acute Decompensated Heart Failure: Results of the Randomized Study.

Aim      To determine the effect of sodium-glucose cotransporter-2 inhibitors (SGLT2i) on kidney function in acute decompensated heart failure (ADHF).Material and methods  A controlled randomized study on the dapagliflozin treatment in ADHF was performed. Patients were randomized to a main group (standard therapy supplemented with dapagliflozin) or a control group (standard therapy for ADHF). The primary endpoint was the development of acute kidney injury (AKI). 200 patients were included (mean age, 74±12 years; 51% men). 31% of patients had type 2 diabetes mellitus (DM2). Mean left ventricular ejection fraction (LV EF) was 47±14 %; in 44.5% of patients, LV EF was less than 45%. Median concentration of N-terminal pro-brain natriuretic peptide (NT-proBNP) was 5225 [3120; 9743] pg / ml, glomerular filtration rate (GFR) was 51 [38; 64] ml / min / 1.73 m2.Results In-hospital mortality was 6.5%. Analysis of the dynamics of body weight loss showed significant differences (4200 [2925; 6300] g vs. 3000 [1113; 4850] g; p=0.011) in favor of the dapagliflozin group. The requirement for increasing the daily dose of furosemide and adding an another class diuretic (thiazide or acetazolamide) did not differ between the groups. However, median furosemide dose during the stay in the hospital was lower in the dapagliflozin group (80 [67; 120] mg vs. 102 [43; 120] mg; p=0.016). At 48 hours after randomization, GFR significantly decreased in the dapagliflozin group (-5.5 [-11; 3] ml/min/ 1.73 m2) compared to the control group (-0.3 [-4; 5] ml / min/1.73 m2, р=0.012). Despite this, GFR did not differ between the groups at discharge (51 [41; 66] ml/min/1.73 m2 and 49 [38; 67] ml/min/1.73 m2, respectively; p = 0.84). In the dapagliflozin group, frequency of AKI episodes was not increased compared to the control group (13 and 9.4%, respectively; p = 0.45).Conclusion      The dapagliflozin treatment in ADHF is associated with more pronounced body weight loss and lower average doses of loop diuretics during the period of stay in the hospital, with no associated clinically significant impairment of renal function.

Detection and full-genotype determination of rare and reassortant rotavirus A strains in Nizhny Novgorod in the European part of Russia.

Reassortant DS-1-like rotavirus A strains have been shown to circulate widely in many countries around the world. In Russia, the prevalence of such strains remains unclear due to the preferred use of the traditional binary classification system. In this work, we obtained partial sequence data from all 11 genome segments and determined the full-genotype constellations of rare and reassortant rotaviruses circulating in Nizhny Novgorod in 2016-2019. DS-1-like G3P[8] and G8P[8] strains were found, reflecting the global trend. Most likely, these strains were introduced into the territory of Russia from other countries but subsequently underwent further evolutionary changes locally. G3P[8], G9P[8], and G12P[8] Wa-like strains of subgenotypic lineages that are unusual for the territory of Russia were also identified. Reassortant G2P[8], G4P[4], and G9P[4] strains with one Wa-like gene (VP4 or VP7) on a DS-1-like backbone were found, and these apparently had a local origin. Feline-like G3P[9] and G6P[9] strains were found to be phylogenetically close to BA222 isolated from a cat in Italy but carried some traces of reassortment with human strains from Russia and other countries. Thus, full-genotype determination of rotavirus A strains in Nizhny Novgorod has clarified some questions related to their origin and evolution.

NGS Technology in Monitoring the Genetic Diversity of Cytomegalovirus Strains.

Modern molecular genetic methods, massive parallel sequencing in particular, allow for genotyping of various pathogens with the aim of their epidemiological marking and improvement of molecular epidemiological surveillance of actual infections, including cytomegalovirus infection. The aim of the study is to evaluate the next-generation sequencing (NGS) technology for genotyping clinical isolates of cytomegalovirus (CMV). Materials and Methods: The object of the study were samples of biological substrates (leukocyte mass, saliva, urine) taken from patients who underwent liver and kidney transplantation. Detection of CMV DNA was carried out by a real-time PCR using commercial diagnostic AmpliSense CMV-FL test systems (Central Research Institute for Epidemiology, Moscow, Russia). DNA extraction was performed using DNA-sorb AM and DNA-sorb V kits (Central Research Institute for Epidemiology) in accordance with manufacturer's manual. The quality of the prepared DNA library for sequencing was assessed by means of the QIAxcel Advanced System capillary gel electrophoresis system (QIAGEN, Germany). Alignment and assembly of nucleotide sequences were carried out using CLC Genomics Workbench 5.5 software (CLC bio, USA). The sequencing results were analyzed using BLAST of NCBI server. Results: CMV DNA samples were selected for genotyping. The two variable genes, UL55(gB) and UL73(gN), were used for CMV genotype determination, which was performed using NGS technology MiSeq sequencer (Illumina, USA). Based on the exploratory studies and analysis of literature sources, primers for genotyping on the UL55(gB) and UL73(gN) genes have been selected and the optimal conditions for the PCR reaction have been defined. The results of sequencing the UL55(gB) and UL73(gN) gene fragments of CMV clinical isolates from recipients of solid organs made it possible to determine the virus genotypes, among which gB2, gN4c, and gN4b were dominant. In some cases, association of two and three CMV genotypes has been revealed. Conclusion: The application of the NGS technology for genotyping cytomegalovirus strains can become one of the main methods of CMV infection molecular epidemiology, as it allows for obtaining reliable results with a significant reduction in research time.

[Variability of genes encoding nonstructural proteins of rotavirus А (Reoviridae: Rotavirus: Rotavirus A) genotype G9P[8] during the period of dominance in the territory of Nizhny Novgorod (central part of Russia) (2011-2020)].

INTRODUCTION: In Russia, rotavirus A is the main cause of severe viral gastroenteritis in young children. The molecular features that allow a rotavirus of a particular genotype to gain an evolutionary advantage remain unclear, therefore, the study of the genetic diversity of rotaviruses based on genes encoding nonstructural proteins (NSPs) responsible for the reproduction of the virus in the cell is an urgent task. OBJECTIVE: To study the genetic diversity of rotaviruses of genotype G9P[8], which dominated Nizhny Novgorod in 20112020, based on genes encoding nonstructural proteins. MATERIALS AND METHODS: Rotavirus-positive samples were subjected to PCR-genotyping and sequencing of NSP1 NSP5 genes. Phylogenetic analysis was carried out in the MEGA X program. RESULTS: In the period 20112020, G9P[8] rotaviruses with four variants of the NSP2 gene were co-circulating in Nizhny Novgorod. New alleles were noted in 2012 (N1-a-III), 2016 (N1-a-IV) and in 2019 (N1-a-II). The appearance of new variants of other genes occurred in 2014 (E1-3, NSP4), 2018 (T1-a3-III, NSP3) and in 2019 (A1-b-II, NSP1). NSP2 gene had the most variable amino acid sequence (16 substitutions), 2 to 7 substitutions were observed in NSP1, NSP3 and NSP4, NSP5 was conservative. DISCUSSION: The results obtained are consistent with the literature data and indicate the participation of NSP genes in maintaining the heterogeneity of the rotavirus population. CONCLUSION: Until 2018, the genetic diversity of rotaviruses in Nizhny Novgorod was determined by the circulation of strains carrying several alleles of the NSP2 gene and conservative genes NSP1, NSP3NSP5. By the end of the study period, new variants of the genotype G9P[8] were formed in the population, carrying previously unknown combinations of alleles of nonstructural genes.

On the nature of picobirnaviruses.

The picobirnaviruses (Picobirnaviridae, Picobirnavirus, PBVs) are currently thought to be animal viruses, as they are usually found in animal stool samples. However, no animal model or cell culture for their propagation has yet been found. In 2018, a hypothetical assumption about PBVs belonging to prokaryotic viruses was put forward and experimentally substantiated. This hypothesis is based on the presence of Shine-Dalgarno sequences in the genome of all PBVs before three reading frames (ORF) at the ribosomal binding site, with which the prokaryotic genome is saturated, while in the eukaryotic genome such regions occur with low frequency. The genome saturation with the Shine-Dalgarno sequences, as well as the preservation of this saturation in the progeny, according to scientists, allows us to attribute PBVs to prokaryotic viruses. On the other hand, there is a possibility that PBVs belong to viruses of eukaryotic hosts - fungi or invertebrates, since PBV-like sequences similar to the genome of fungal viruses from the families of mitoviruses and partitiviruses have been identified. In this regard, the idea arose that, in terms of reproduction mode, PBVs resemble fungal viruses. The divergence of views on the true PBV host(s) has sparked discussions among scientists and required further research to elucidate their nature. The review highlights the results of the search for a PBV host. The reasons for the occurrence of atypical sequences among the PBV genome sequences that use an alternative mitochondrial code of lower eukaryotes (fungi and invertebrates) for the translation of viral RNA-dependent RNA polymerase (RdRp) instead of the standard genetic code are analyzed. The purpose of the review was to collect arguments in support of the hypothesis about the phage nature of PBVs and to find the most realistic explanation of the reasons for identifying non-standard genomic sequences for PBVs. Based on the hypothesis about the genealogical relationship of PBVs with RNA viruses from other families with similar segmented genomes, such as Reoviridae, Cystoviridae, Totiviridae and Partitiviridae, virologists support the assumption of a decisive role in the origin of atypical PBV-like reassortment strains between PBVs and viruses of the listed families. The collected arguments given in this review indicate a high probability of a phage nature of PBVs. The data presented in the review show that the belonging of PBV-like progeny to prokaryotic or eukaryotic viruses is determined not only by its genome saturation level with a prokaryotic motif, standard or mitochondrial genetic code. The primary structure of the gene encoding the viral capsid protein responsible for the presence or absence of specific proteolytic properties of the virus that determine its ability for independent horizontal transmission into new cells may also be a decisive factor.

An increase in prevalence of recombinant GII.3[P12] norovirus in sporadic acute diarrhea in children in Nizhny Novgorod, Russia, 2018-2021.

Noroviruses are important etiological agents causing acute intestinal infection in humans. In the last decades, the most common norovirus genotype was GII.4 despite a significant genetic diversity among strains, while the active circulation of noroviruses with other genotypes was observed periodically. This study shows an increase in the detection rate of recombinant GII.3[P12] norovirus in Nizhny Novgorod, Russia, from 6.8% in 2018-2019 to 34.9% in 2020-2021. We performed a phylogenetic analysis based on the nucleotide sequences of noroviruses possessing this genotype obtained in this work, as well as presented in the GenBank database. It has been shown that the circulation of GII.3[P12] noroviruses in the study area was the result of several independent introductions, either directly from the Western Pacific region, or through the Asian part of Russia. The polyphyletic origin, the geographical expansion, and the growth of the epidemic significance of the recombinant GII.3[P12] noroviruses were noted.

[Detection SARS-CoV-2 (Coronaviridae: Coronavirinae: Betacoronavirus: Sarbecovirus) in children with acute intestinal infection in Nizhny Novgorod during 2020-2021].

INTRODUCTION: The novel coronavirus infection COVID-19 is a major public health problem worldwide. Several publications show the presence of gastrointestinal (GI) symptoms (nausea, vomiting, and diarrhea) in addition to respiratory disorders.The aim of this study was the monitoring of RNA of COVID-19 pathogen, coronavirus SARS-CoV-2 (Coronaviridae: Coronavirinae: Betacoronavirus; Sarbecovirus) in children hospitalized with acute intestinal infection (AII), with following molecular-genetic characterization of detected strains. MATERIAL AND METHODS: Fecal samples of children with AII hospitalized in infectious hospital of Nizhny Novgorod (Russia) in the period from 01.07.2020 to 31.10.2021 were used as material for the study. Viral RNA detection was performed by real-time polymerase chain reaction (RT-PCR). The nucleotide sequence of S-protein gene fragment was determined by Sanger sequencing. RESULTS AND DISCUSSION: SARS-CoV-2 genetic material was detected in 45 out of 2476 fecal samples. The maximum number of samples containing RNA of the virus occurred in November 2020 (detection rate of 12.2%). In 20.0% of cases, SARS-CoV-2 RNA was detected in combination with rota-, noro-, and adenoviruses. 28 nucleotide sequences of S-protein gene fragment complementary DNA (cDNA) were determined. Phylogenetic analysis showed that the studied SARS-CoV-2 strains belonged to two variants. Analysis of the S-protein amino acid sequence of the strains studied showed the absence of the N501Y mutation in the 2020 samples, which is a marker for variants with a high epidemic potential, called variants of concern (VOC) according to the World Health Organization (WHO) definition (lines Alpha B.1.1.7, Beta B.1.351, Gamma P.1). Delta line variant B.1.617.2 was identified in two samples isolated in September 2021. CONCLUSION: The detection of SARS-CoV-2 RNA in the fecal samples of children with AII, suggesting that the fecal-oral mechanism of pathogen transmission may exist, determines the necessity to optimize its monitoring and to develop an algorithm of actions with patients with signs of AII under the conditions of a novel coronavirus infection pandemic.

[Construction of norovirus (Caliciviridae: Norovirus) virus-like particles containing VP1 of the Echovirus 30 (Picornaviridae: Enterovirus: Enterovirus B)].

INTRODUCTION: Enterovirus (nonpolio) infection is widespread all over the world, registered as sporadic cases and large-scale outbreaks and can cause severe lesions such as serous meningitis. Epidemiological studies have shown that enterovirus (Picornaviridae; Enterovirus) variant Echovirus 30 (E30) is the most frequently detected variant in patients with enterovirus meningitis in the Russian Federation. However, no vaccines to prevent the disease caused by this pathogen have been developed so far. One of the promising modern trends in terms of creating vaccine preparations is the use of virus-like particles (VLPs), including chimeric ones containing the biological structures of viruses belonging to different species.The aim of this work was to obtain norovirus (Caliciviridae; Norovirus) VLPs displaying enterovirus Echovirus E30 full-length VP1 on the surface. MATERIAL AND METHODS: The nucleotide sequences of VP1 protein of norovirus genotype GII.4 and VP1 E30 of genotype h circulating in Russia were used. The SN-VP1E30 protein was constructed, in which the shell (S) and the hinge regions of the norovirus VP1 are fused into one molecule with the full-length VP1 of the E30 virus. The protein was expressed in E. coli, purified using affinity chromatography, and characterized by polyacrylamide gel electrophoresis (PAGE) and immunoblotting. VLPs were visualized by electron microscopy. RESULTS: The S N-VP1E30 protein expressed in E. coli as insoluble form, so the conditions for SN-VP1E30 solublisation were defined. Sucrose has been shown to significantly increase the efficiency of renaturation. Electrophoretic mobility comparison of denatured and non-denatured SN-VP1E30 demonstrated that most monomers form high molecular weight compounds. Electron microscopy showed that renatured SN-VP1E30 spontaneously forms empty virus-like particles about 50 nm in diameter. CONCLUSION: Chimeric protein SN-VP1E30 self-assemble into VLPs displaying the VP1 protein of E30 variant that is highly prevalent in Russia. Further immunological research is necessary to characterize VLPs potential for development of the vaccine for enteroviral meningitis prevention.

[Molecular monitoring of the rotavirus (Reoviridae: Sedoreovirinae: Rotavirus: Rotavirus A) strains circulating in Nizhny Novgorod (2012-2020): detection of the strains with the new genetic features].

INTRODUCTION: The pentavalent rotavirus vaccine has been registered in Russia, however, the vaccination coverage remains low, and an annual increase in the incidence of rotavirus infection is unavoidable. In this regard, molecular monitoring of rotaviruses in order to search for new variants possessing epidemic potential is an urgent task. MATERIAL AND METHODS: PCR genotyping and VP4 and VP7 genes sequencing were used to characterize rotaviruses circulating in Nizhny Novgorod in 2012-2020. The phylogenetic analysis of the strains was carried out using the BEAST software package. RESULTS: The spectrum included 17 genotypes with predominance of G9P[8] (37,4%). Detected in this study genotypes G1P[4], G1P[9], G2P[8], G4P[4], G4P[6], G8P[8], and G9P[4] were not previously identified in Nizhny Novgorod. The circulation of DS-1-like strains possessing genotypes G1P[8], G3P[8], G8P[8], or G9P[8] and a short RNA pattern had been shown. Rotaviruses of the common genotypes were genetically heterogeneous and belonged to different phylogenetic lineages and/or sublineages (P[4]-IV-a; P[4]-IV-b; P[8]-3.1; P[8]-3.3; P[8]-3.4 and P[8]-3.6; G1-I; G1-II; G2-IVa-1; G2-IVa-3; G3-1; G3-3; G4-I-c; G9-III; G9-VI). DISCUSSION: These results extend the available data on the genotypic structure of rotavirus populations in Russia and show the genetic diversity of viral strains. G3P[8] DS-1-like viruses were representatives of the G3-1 lineage, new for the territory of Russia, and had the largest number of amino acid substitutions in the VP7 antigenic epitopes. CONCLUSION: The emergence and spread of strains with new genetic features may allow rotavirus to overcome the immunological pressure formed by natural and vaccine-induced immunity, and maintain or increase the incidence of rotavirus infection.

[Phylodynamic characteristics of the Russian population of rotavirus А (Reoviridae: Sedoreovirinae: Rotavirus) based on the VP6 gene].

INTRODUCTION: Rotavirus A is one of the leading causes of acute gastroenteritis in children in the first years of life. Rotavirus infection is currently classified as a preventable infection. The most abundant rotavirion protein is VP6. MATERIAL AND METHODS: Phylogenetic analysis and calculation of phylodynamic characteristics were carried out for 262 nucleotide sequences of the VP6 gene of rotavirus species A, isolated in Russia, using the BEAST v.1.10.4 software package. The derivation and analysis of amino acid sequences was performed using the MEGAX program. RESULTS: This study provides phylodynamic characteristics of the rotaviruses in Russia based on the sequences coding VP6 protein. Bayesian analysis showed the circulation of rotaviruses of three sublineages of genotype I1 and three sublineages of genotype I2 in Russia. The level of accumulation of mutations was established, which turned out to be similar for genotypes I1 and I2 and amounted to 7.732E-4 and 1.008E-3 nucleotides/site/year, respectively. The effective population sizes based on nucleotide sequences of the VP6 I1 and I2 genotypes are relatively stable while after the 2000s there is a tendency of its decreasing. Comparative analysis of the amino acid sequences in the region of the intracellular neutralization sites A (231-260 aa) and B (265-292 aa) made it possible to reveal a mutation in position V252I in a proportion of Russian strains of genotype I1 some strains of genotypes I1 and I2 had mutation I281V. These substitutions were not associated with any sublineages to which the strains belong. The analysis of three T-cell epitopes revealed four amino acid differences (in aa positions 305, 315, 342, 348) that were associated with the first or second genogroup. CONCLUSION: Based on the phylodynamic characteristics and amino acid composition of antigenic determinants, it was concluded that the VP6 protein is highly stable and could potentially be a good model for development of a rotavirus vaccine.

Long-term monitoring of G1P[8] rotaviruses circulating without vaccine pressure in Nizhny Novgorod, Russia, 1984-2019.

The G1P[8] genotype is one of the most common among rotaviruses circulating in the last 40 years. Therefore, this genotype is a component of rotavirus vaccines licensed throughout the world. This paper presents the results of a 35-year (1984-2019) observation of the circulation of G1P[8] rotaviruses among children under 14 in one region (Nizhny Novgorod, Russia) without vaccine pressure. Several complementary approaches were used: RNA electropherotyping by polyacrylamide gel electrophoresis, PCR genotyping, and cDNA sequencing of rotavirus VP4 and VP7 genes. A total of 8375 rotavirus-positive samples were examined, and the proportion of genotype G1P[8] rotaviruses was 39.9% (4.3-98.9%). Two cycles of high circulation activity (1984-1993 and 1993-2007) and one cycle of low activity (2007-2019) were noted. Phylogenetic analysis revealed the presence of rotaviruses of two VP4 gene lineages (P[8]-1 and P[8]-3) and two VP7 gene lineages (sublineages IA, IB, ID, II-B, II-C, and II- E). The prolonged circulation of rotaviruses of only one sublineage (G1-II-E) and then a change of the prevailing sublineage within the G1-II lineage (from E to C) during the active circulation were shown. Since 2011, when the circulation intensity of G1P[8] rotaviruses was low, the appearance of strains of the G1-I lineage and their co-circulation with strains of the G1-II lineage were observed in the population.

Picobirnaviruses: prevalence, genetic diversity, detection methods.

This article presents a general overview of the prevalence, genetic diversity and detection methods of picobirnaviruses (PBVs), which are small, non-enveloped icosahedral viruses with a segmented double-stranded RNA genome consisting of two segments taxonomically related to the genus Picobirnavirus of the family Picobirnaviridae. This review of scientific papers published in 1988-2019 provides data on the PBV distribution in the nature and a broad host range. PBV infection is characterized as opportunistic, the lack of understanding of the etiological role of PBVs in diarrhea is emphasized, since these viruses are detected both in symptomatic and asymptomatic cases. The concept of PBV infection as a chronic disease caused by a long-lasting persistence of the virus in the host is considered. Such factors as stress syndrome, physiological conditions, immune status and host age at the time of primary PBV infection influence the virus detection rate in humans and animals. The possible zoonotic nature of human PBV infection is noted due to the capacity for interspecies PBV transmission acquired during evolution as a result of the reassortment of the genome segments of different viruses infecting the same host. Data providing evidence that PBVs belong to eukaryotes and a challenging hypothesis stating that PBVs are bacterial viruses are presented. The need to intensify work on PBV detection because of their wide distribution, despite the complexity due to the lack of the cultivation system, is emphasized. Two strategies of RT-PCR as main PBV detection methods are considered. The genomes of individual representatives of the genus isolated from different hosts are characterized. Emphasis is placed on the feasibility of developing primers with broader specificity for expanding the range of identifiable representatives of the genus PBV due to a huge variety of their genotypes. The importance of effective monitoring of PBV prevalence for studying the zoonotic and anthroponotic potential using metagenomic analysis is highlighted, and so is the possibility of using PBV as a marker for environmental monitoring.

[Identification of rotavirus I- and E-genotypes by multiplex PCR method.]

INTRODUCTION: In recent years the presence of reassortant rotavirus strains is increasingly mentioned in the world due to the application of the full-genome based classification system. Information on the circulation of such strains in the territory of Russia is limited. The aim of this work was the development of the approach for determination of genotypes of segments encoding VP6 (I) and NSP4 (E) to reveal reassortant strains. MATERIAL AND METHODS: Rotavirus-positive samples were studied by means of nucleotide sequencing and multiplex PCR. Phylogenetic analysis was conducted using the Bayesian approach. RESULTS: Three alleles of the VP6 gene (I1-1, I2-IV, I2-VII) and seven alleles of the NSP4 gene (E1-I, E1-III, E2-VI, E2-VII, E2-X, E2-XII, E3) were detected on the base of nucleotide sequences of Nizhny Novgorod rotaviruses. Taking into account these results, the oligonucleotide primers specific to genotypes I1, I2, I3 and E1, E2, E3 were designed. Optimal conditions for multiplex PCR were chosen. The method was tested using the strains collected in Nizhny Novgorod in 2018. The diversity of I and E genotypes was determined and various combinations with G and P genotypes were identified. DISCUSSION: G9-P[8]-I1-E1 rotaviruses were predominant (32.7 %) and G2-P[4]-I2-E2 rotaviruses were in second place (29.1 %). Strains with genotypes G4-P[8]-I1-E2, G3-P[8]-I2-E2 and G2-P[4]-I2-E1 were detected sporadically. They had genes of two rotavirus genogroups, so can be considered to be reassortant. CONCLUSION: The proposed approach is a useful tool for the characterization of rotaviruses in the conditions of the beginning of vaccination against rotavirus infection in Russia.

Phylogenetic comparison of the VP7, VP4, VP6, and NSP4 genes of rotaviruses isolated from children in Nizhny Novgorod, Russia, 2015-2016, with cogent genes of the Rotarix and RotaTeq vaccine strains.

Group A rotaviruses (RVA) are one of the leading causes of gastroenteritis in young children worldwide. The introduction of universal mass vaccination around the world has contributed to a reduction in hospitalizations and outpatient visits associated with rotavirus infection. Continued surveillance of RVA strains is needed to determine long-term effects of vaccine introduction. In the present work, we carried out the analysis of the genotypic diversity of RVA strains isolated in Nizhny Novgorod (Russia) during the 2015-2016 epidemic season. Also we conducted a comparative analysis of the amino acid sequences of T-cell epitopes of wild-type and vaccine (RotaTeq and Rotarix) strains. In total, 1461 samples were examined. RVAs were detected in 30.4% of cases. Rotaviruses with genotype G9P[8] (40.5%) dominated in the 2015-16 epidemic season. Additionally, RVAs with the following genotypes were detected: G4P[8] (25.4%), G1P[8] (13%), G2P[4] (3.2%). Rotaviruses with genotypes G3P[9], G6P[9], and G1P[9] totaled 3%. The number of partially typed and untyped RVA samples was 66 (14.9%). The findings of a RVA of G6P[9] genotype in Russia were an original observation. Our analysis of VP6 and NSP4 T-cell epitopes showed highly conserved amino acid sequences. The found differences seem not to be caused by the immune pressure but were rather related to the genotypic affiliations of the proteins. Vaccination against rotavirus infection is not included in the national vaccination schedule in Russia. Monitoring of the genotypic and antigenic diversity of contemporary RVA will allow providing a comparative analysis of wild-type strains in areas with and without vaccine campaign.

Case of thrombosis of the portal vein and its branches in a patient seropositive for Borrelia antigens.

Borreliosis after sucking ticks is an acute problem in the world. People do not go to doctors after that often, which leads to the development of various complications. Thrombosis of veins of various localization can be one of them. Thrombosis of the portal vein represents a significant problem too with high morbidity and mortality. The risk factors for splanchnic vein thrombosis include infections, but its relationship with borreliosis has not been studied. А 34-year-old man with chronic helicobacter-associated gastritis and gallstones was hospitalized due to development during the last 11 days of epigastric pain and fever to 38.7 °C after a picnic at the forest without a registered tick bite. The blood leukocytes were increased to 11.2*109/l, lymphocytes 70%, C-reactive protein 34.6 mg/l, procalcitonin 0.195 ng/ml. The multispiral computed tomography of the abdominal cavity revealed thrombosis of portal, lienalis and superior mesenteric veins. D-dimer was 1.98 mcg/ml, antithrombin III 75%. JACK2V617F, oncological, rheumatic, thrombophilia markers, blood and urine cultures were negative. A high concentration of anti-Borrelia burgdorferi IgM 62.2 U/ml and its increasing to 190 U/ml in dynamics was revealed at the immunofluorescence assay. Anti-Borrelia IgM to OspA, p31 and OspC, p25 were detected at the immunoblotting assay. Anticoagulation, doxycycline, detoxification therapy reduced pain and normalized temperature and inflammation markers. Vein thrombosis was not detected at the control tomography after 2 weeks. Despite that the combination of thrombosis and borreliosis is rare, it is necessary to screen for Borrelia antigens in patients with splanchnic vein thrombosis and fever.

Predominance of new G9P[8] rotaviruses closely related to Turkish strains in Nizhny Novgorod (Russia).

Genotype G9P[8] rotaviruses are rare in the territory of Russia. They were found in Nizhny Novgorod only in 2011-2012 for the first time, when their proportion was 25.9%. During the next two seasons, G9P[8] strains were detected in only 1.8% of cases. Their proportion substantially increased again in 2014, and they became predominant in the city by 2016. Phylogenetic analysis on the basis of gene VP7 nucleotide sequences showed that this increase was accompanied by the emergence of new strains in the population. These isolates were related to Turkish strains, but not to Russian ones detected earlier.

Detection and molecular characterization of reassortant DS-1-like G1P [8] strains of rotavirus A.

Group A rotaviruses (RVA) are the main cause of viral gastroenteritis in children worldwide. In this study we provide the molecular characteristics of reassortant DS-1-like G1P[8] RVA strains detected in Russia for the first time. Previously, such reassortant strains were detected in Japan and Thailand. The G1P[8] RVAs with DS-1-like short electropherotype RNA-PAGE were isolated from children hospitalised with an acute gastroenteritis during the 2013-2014 period. The DS-1-like G1P[8] strains accounted for 2.6% of all RVA strains detected continuously throughout the season. A phylogenetic analysis was made on the basis of the established nucleotide sequences of genes VP7, VP8* (VP4), VP6 and NSP4. The Nizhny Novgorod strains belong to G1-I and G1-II alleles of VP7 gene and to P[8]-3 allele of VP4. According to their VP6 sequences, two Russian samples clustered with the reassortant strains isolated in Japan, Thailand and Australia and two other strains were phylogenetically close to the typical G2P[4] DS-1-like RVA. Nucleotide sequences of G1P[8] strains that belong to NSP4 gene form a separate cluster from G3P[8] DS-1-like rotaviruses detected in Thailand and Australia. The RVA alleles included in Rotarix and RotaTeq vaccine strains were clustered separately from the studied reassortant RVAs. On the grounds of phylogenetic analysis we assume a polyphyletic origin of reassortants between Wa- and DS-1-like strains. Mutation rates evaluated by Bayesian inference in clusters with reassortant RVA strains were 1.004Е-3 (VP7), 1.227E-3 (VP4), 3.909E-4 (VP6), and 4.014Е-4 (NSP4). Analysis of tMRCA showed relatively contemporary origin of alleles DS-1-like G1P[8] rotaviruses: VP7 - 1998 (G1-I) and 1981 (G1-II), VP4 - 1998, VP6 - 1994, NSP4 - 1979.