[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.

Generation and Evaluation of Bispecific Anti-TNF Antibodies Based on Single-Chain VHH Domains.

Systemic cytokine inhibition may be an effective therapeutic strategy for several autoimmune diseases. However, recent studies suggest that tissue or cell type-specific targeting of certain cytokines, including TNF, may have distinct advantages and show fewer side effects. Here we describe protocols for generating and testing bispecific cytokine inhibitors using variable domain of single-chain antibodies from Camelidae (VHH) with a focus on cell-specific TNF inhibitors.

[Bispecific Antibodies: Formats and Areas of Application].

Bispecific antibodies capable of simultaneously binding two targets have been studied for many years with a view to their implementation in clinical practice. Unique biological and pharmacological properties, as well as the diversity of their formats, make it possible to consider bispecific antibodies as promising agents for use in various procedures: from visualization of intracellular processes to targeted anticancer therapy. Bispecific antibodies help to determine more precisely the therapeutic target, thereby increasing the efficiency of therapy and reducing the probability of side effects. The present review describes the main formats of bispecific antibodies, methods for their generation, and possibilities for practical application.

[Analysis of new variants of West Nile fever virus]. / Analiz novykh variantov virusa likhoradki Zapadnogo Nila.

The complete nucleotide sequences for 6 strains of the West Nile fever virus were determined. For the first time the complete nucleotide sequences of the Indian isolate and Krsn190 strain, that is the most far phylogenetically from all isolates known at present time were established. The scheme for separation of virus variants into 4 groups and criteria for determination the group to which the isolate belongs are suggested.

[Genome analysis of hepatitis C virus strain 274933RU isolated in Russian Federation]. / Analiz genoma izoliata 274933RU virusa gepatita C, vydelennogo na territorii Rossiiskoi Federatsii.

Five overlapping cDNA fragments of hepatitis C virus (HCV) isolate 274933RU, obtained by RT-PCR, were amplified and cloned. Complete nucleotide RNA sequence has been determined. The genomic organization of 274933RU was, from 5' to 3' terminals, 5' UTR (341 nt), polyprotein ORF (9033 nt), 3' UTR (40 nt except for the poly(U-UC) and polypyrimidine stretch), and X-tail (98 nt). Phylogenetic analysis of the core and NS5 genes showed that the isolated strain belonged to HCV 1b subtype.

[Genetic analysis of West Nile fever virus, isolated in the south of the Russian plain (Volgograd and Astrakhan regions) in 1999]. / Geneticheskii analiz virusov likhoradki Zapadnogo Nikia, vydelennykh na iuge Russkoi ravniny (Volgogradskaia i Astrakhanskaia oblasti) v 1999 g.

Two strains of West Nile virus, Vlg 27889 and Ast 986, were isolated from the brain of a dead man and from the blood of a patient, respectively, during an outbreak of serous meningitis and meningoencephalitis in July-September, 1999, in the Volgograd and Astrakhan regions. Analysis of parts of genome of the strains cloned from cell culture by reverse transcription and polymerase chain reaction demonstrated their identity and appurtenance to group I West Nile viruses.

[Determination of hepatitis G virus markers in various risk groups]. / Opredelenie markerov virusa gepatita G v razlichnykh gruppakh riska.

Study of prevalence of hepatitis G virus (HGV) markers in different risk groups showed the presence of HGV RNA in 3.2% blood donors, 24.2% patients with hepatitis C (HCV), and 28% patients with hemophilia. HGV antibodies were detected in 11.3% donors, 16.0% patients with HCV, 13.4% patients with hemophilia, and 8.5% HIV-infected subjects. Anti-E1 HGV were more often detected in the absence of HGV RNA. Antibodies to HGV E2 protein were significantly more often detected in adult HCV patients but not in adolescent patients aged 8-15 years.

[Comparative analysis of fragments of HGV NS5b-region isolated in Russia, Kazakhstan and Kyrgyztan]. / Sravnitel'nyi analiz fragmentov NS5B-regiona izoliatov HGV, obnaruzhennykh v Rossii, Kazakhstane i Kyrgyzstane.

Nucleotide sequence of a 356 nt fragment of the NS5 region of the hepatitis G virus (HGV) genome was determined in 7 strains isolated in Russia, Kazakhstan, and Kyrghyzstan. Philogenetic analysis showed that all isolated strains genetically differed from the West African strain described previously (type 1) and are closely related to strains isolated in the USA and South-Eastern Asia (type 2). Hence, the strains from Russia, Kazakhstan, and Kyrghyzstan belong to type 2 variant of HGV genome.