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Introduction. Molecular epidemiological monitoring of enterovirus infection (EVI) in the territories of the Russian Federation showed that coxsackievirus A6 (CVA-6) had been one of the most prevalent types of enteroviruses that circulated among the country population during last years and had caused majority of EVI outbreaks. Objective — to evaluate coxsackievirus A6 circulation in the territories of the Far Eastern Federal district (FEFD) in 2014–2019 utilizing methods of molecular genetics. Materials and methods. RT-PCR was employed to detect enterovirus RNA in biological material collected in 9 territories of the FEFD . In order to establish enterovirus types, amplification of positive samples was carried out to detect nucleotide sequence fragments of the VP1 and VP2 genes. Molecular genetic analysis of the Far Eastern CVA-6 strains was based on detection of nucleotide sequences of VP1 and 3Dpol gene fragments. Phylogenetic trees were constructed by the means of Bayesian phylogenetic methods. Results. Total 1773 nucleotide sequences of 43 types of non-polio enteroviruses were obtained in 2014-2019. Majority of the sequences belonged to coxsackievirus A6 (524; 29.5%). In the years of the highest CVA-6 detection an increase in EVI incidence as well as EVI outbreaks were observed in the territories of FEFD. The most prevalent manifestations of EVI caused by CVA-6 in FEFD were herpangina and exanthemic forms. Phylogenetic analysis showed that Far Eastern strains of CVA-6 during in the analyzed period of time belonged to D3 subgenotype that is dominant in the world. The circulation of several recombinant forms of CVA-6 (RF-A, -H, -L, -N, -R) was also registered. Conclusion. The genetic diversity of CVA-6 strains circulating in the territories of the FEFD in 2014–2019 revealed in this study requires further investigation in order to obtain new knowledge about the CVA-6 molecular epidemiology and improve the enterovirus surveillance system.

Enteroviruses comprise highly diverse group of single-stranded positive RNA viruses belonging to Enterovirus genus, Picornaviridae family. They are the most prevalent viruses worldwide highlighted by high resistance to environmental cues. Enteroviruses normally cause seasonal self-limiting infections, but also known as causative infectious agents of encephalitis, myocarditis, poliomyelitis, acute heart failure and sepsis. Enterovirus genetic plasticity contributes to widespread epidemics and sporadic outbreaks (e. g., outbreaks of Enterovirus D68 and Enterovirus 71). Type B Coxsackieviruses of Enterovirus B species is one of commonly identified infectious agents associated predominantly with mild upper respiratory and gastrointestinal illnesses. Nevertheless, Coxsackieviruses B3 infection can result in severe myocarditis leading ultimately to heart failure. The pathogenesis of Coxsackievirus B3-induced myocarditis is well known being mediated by both direct damage due to viral proteases and indirectly via secondary host immune responses. Despite success in preventive vaccination of some enterovirus infections that allowed to control some of them direct antiviral agents for treatment of enteroviral infection particularly Coxsackieviruses B3 myocardial infection are still in demand. In addition, no ongoing clinical trials for therapy or prevention of Coxsackieviruses B3 infection are available. Current treatment strategies are mainly aimed to stabilize patient condition and relieve discomfort condition. It seems that relatively small market for anti-enteroviral drugs prevents pharma industry from developing new drugs. The Coxsackieviruses B3 lifecycle have been extensively studied and potential targets for drug design have been identified. The aim of our review was to describe current state in the field of antiviral drug design combating Coxsackieviruses B3 infection emphasizing direct-acting antivirals, albeit paying some attention to host factor-targeting inhibitors (including compounds from medicinal plant extracts) as well. The following categories of direct Coxsackieviruses B3 inhibitors are discussed in detail: capsid binders (pleconaril and its derivatives), viral 3C protease inhibitors (rupintrivir and its analogs), drugs targeting viral replication (both nucleoside analogs and non-nucleoside inhibitors). Results of drug repurposing screens for amiloride, benzerazide, dibucaine and fluoxetine are also discussed.

INTRODUCTION. Developing novel medicines based on non-pathogenic enterovirus strains exhibiting oncotropic and oncolytic properties represents an up-to-date and safe approach to complex cancer treatment and postoperative metastasis prevention. Safety pharmacology studies are a necessary step in the preclinical development of medicinal products.AIM. The study aimed to investigate the single and repeated-dose general toxicity, local tolerance, safety pharmacology, and pyrogenicity of medicinal products based on non-pathogenic LEV4, LEV7, LEV8, LEV14, and Russo enterovirus strains as part of preclinical safety studies.MATERIALS AND METHODS. The study used medicinal products of highly purified group A, B, and C enteroviruses at a titre of 2×107–5×108 CPD50/mL (CPD50 is a cytopathogenic dose of the virus causing 50% cell lysis) and normal saline as a diluent. The viruses were propagated in Vero cells. The safety study used 220 male and female BALB/c mice, 440 male and female Wistar rats, and 18 male Soviet chinchilla rabbits. The study animals received an intravenous dose of 1×105 or 1×106 CPD50/animal once (single-dose toxicity) or weekly for 90 days (repeated-dose toxicity). Clinical examination, laboratory testing, and necropsy were performed on days 45 and 91 of the experiment. Statistical data processing was performed using Prism 8.0 software (GraphPad Software, Inc., USA).RESULTS. Upon single administration of each of the five enterovirus medicinal products to mice and rats, the authors observed complete survival, upward trends in body weight gain, and no gross or histopathological changes in the brain, spleen, liver, kidneys, lungs, or at the injection site. Upon repeated administration at the study doses, the medicinal products caused no functional changes in the organs and systems. All the studied parameters were within the normal physiological ranges for male and female rats. Histopathological examination revealed no pathological changes or specific cytolytic and/or cytopathic effects. No local irritation was observed. None of the investigational medicinal products showed pyrogenicity.CONCLUSIONS. The obtained preclinical results demonstrate the safety of antineoplastic agents based on live non-pathogenic LEV4, LEV7, LEV8, LEV14, and Russo enteroviruses.

Here we present the data on foreign research publications describing healthcare-associated enteroviral (nonpolio) infections (HAI) sought in the Worldwide Database for Nosocomial Outbreaks (Institut für Hygiene und Umweltmedizin, Universitȁtmedizincomplex “Charite”, Germany) as well as PubMed search engine (The United States National Library), covering 1936–2017 timeframe. The publications retrieved contained the data on 28 nosocomial outbreaks caused by Enterovirus A (EV-A71), В (Echoviruses 11, 17, 18, 30, 31, 33, Coxsackie viruses А9, В2, В5) and D (EV-D68). It was discovered that the majority of the nosocomial enteroviral (non-polio) outbreaks occurred in obstetric hospitals and neonatal units so that children were mainly maternally infected. In addition, a case associated with intrauterine infection was described. It was shown that outbreaks might be started by an infected child at the incubation period. Single publications reported nosocomial outbreaks in geriatric hospitals. Generally, nosocomial enteroviral (non-polio) outbreaks were characterized by polymorphic clinical picture caused by any certain pathogen serotype and within a single site of the infection. Few lethal outcomes were recorded. Enterovirus B species dominated among identified etiological agents. Violated hospital hygiene and infection control contributing to spread of infection were among those found in neonatal units: putting used diapers out on baby bed prior disposal, sharing bathtub, toys and household objects as well as poor hand hygiene in medical workers. One of the measures recommended to improve diagnostics of enteroviral (non-polio) infections was virology screening of children with suspected sepsis in case of unidentified etiology. It was established that etiological decoding of nosocomial outbreaks was impossible without applying pathogen-specific diagnostic tools, mainly nested RT-PCR and direct sequencing of followed by subsequent phylogenetic analysis.

The aim of this study was to investigate the outbreak of aseptic meningitis in theNovosibirskregion in 2008–2009. We studied genetic diversity and molecular-epidemiological characteristics of human enteroviruses that caused aseptic meningitis. Materials and metods. In the present study we investigated samples of cerebrospinal fluid from 199 patients with a diagnosis “aseptic meningitis”, based on the clinical characteristics of the disease (headache, stiff neck, fever, nausea, vomiting), and confirmed by laboratory tests of spinal fluid (lymphocyte cell count > 10 cells/ml).All samples were tested by PCR for RNA of Enterovirus and Flavivirus and DNA Myc. tuberculosis, Borrelia spp., Neisseria spp. In the samples there were not found RNA Flavivirus, DNA Myc. tuberculosis, Borrelia spp., Neisseria spp., but in 73 samples (37%) was identified RNA enterovirus (EV). Determination of nucleotide sequences of 5’UTR and VP1-region of EV revealed that they belong to the following genotypes: the highest percentage was presented by genotype ECHO 30 (62%); another genotypes were Cox A2 (8%), Cox A4 (5%), Cox A14 (3%), Cox A16 (5%), Cox B5 (8%), ECHO 6 (3%), ECHO 9 (3% ) and ECHO 25 (3%). In 2008 most of the EV that caused the symptoms of aseptic meningitis belonged to genotype ECHO 30 (76%). In 2009 the clinical specimens containing genotype ECHO 30 were not found, but the largest percentage of EV belonged to genotypes Cox A2 (33%) and Cox A4 (22%). Thus, in 2008 we recorded outbreak of aseptic meningitis, the major etiological factor was enterovirus ECHO 30. And the rise of the incidence of aseptic meningitis in 2009 is related to the circulation of new genotypes of EV. The investigated strains were deposited in an international database GenBank under accession numbers KP258231-KP258235, HM559584.

A study of the antiviral activity of antioxidants against viral infections is believed to be essential for creating complex antiviral agents. Dihydroquercetin is considered as the most active antioxidant extracted from larix gmelinii. In this work, we present results of experiments of the antiviral properties of dihydroquercetin against a member of the family picornaviridae – Coxsackievirus B4 in vitro. We have estimated that dihydroquercetin reduces viral titers at 100 µg/ml concentration as compared with control of virus. We have shown using the plaque assay that cpe of virus is reduced in the presence of dihydroquercetin at 100 µg/ml. Study of the phase of viral life cycle, in which dihydroquercetin acted, demonstrated that the highest efficacy of the antiviral therapy was reached at early stages of virus reproduction (1-3 hours post infection). These results show that dihydroquercetin has antiviral property against Coxsackievirus B4. This drug and other antioxidants can be tested as inhibitors of viral replication.

In this article, two different methods for the detection of enteroviruses in the fresh water were performed and compared. All collected enteroviruses were identified using molecular typing. The possible reasons of water contamination were discussed in conclusions.

Abstract. The analysis of epidemic process and definition of the etiology of the seasonal increase of serosal meningitis incidence in the Novgorod oblast in September-November, 2008 has been carried out. It was shown that in the specified period 73 persons had enteroviral meningitis. The proportion of children under 17 years old in the group of patients was 89.9%. The etiological agents of disease (enteroviruses ECHO 6 и ECHO 30) were detected by the virological and molecular-biological methods. It was established that two genetic variants of ECHO 6 and one variant of ECHO 30 co-circulated in the Novgorod oblast. The last virus variant was closely related to the strain isolated from enteroviral meningitis patients in Nizhniy Novgorod in 2007. The obtained data support the idea about necessity of systematic epidemiological and virological surveillance on enteroviral infection to get new information about circulation of non-poliomyelitis enteroviruses in the population and to define peculiarities of epidemic process development.

In material from 1086 patients who underwent heart valve disease surgery, traditional hystologic assessment, argyrophilic nucleolar organizer analysis, with calculation of mean Ag granule number per endotheliocyte and fibroblast nucleus; immuno-hystochemical assay with monoclone antibodies to Enteroviruses, vimentin, desmin, SMA, SMA, Ki-67-, bcl-2-antigens; print-smear cytology; and valve PCR were performed. Enterovirus replication was observed in endotheliocytes, fibroblasts, and smooth muscle cells in all cases of active rheumatism. Endothelial dysfunction was associated with Enterovirus infection.

Enterovirus infections are a group of acute infectious diseases caused by enteroviruses (including Coxsackie A and B viruses, ECHO viruses), which clinically present symptoms of damage to the central nervous system, cardiovascular system, gastrointestinal tract, muscular system, mucous membranes and skin, fever. This article presents a clinical case of patient L. , 12 years old, who admitted to an otorhinolaryngologist with clinical manifestations of herpangina. The diagnosis was confirmed by PCR. The patient was prescribed, adequate rehydration, diet with the exclusion of salty, spicy and fried foods, restriction of physical activity, exclusion of thermal procedures, Benzydamine Spray (Oralsept) 0.255 mg/dose, 6 doses 3 times/day, topically, on demand and inosine pranobex (Groprinosin) in a daily dose of 50 mg/kg of body weight: 1 tablet 500 mg 4 times a day for 7 days (at the rate of 1 tablet of 500 mg per 10 kg of body weight; for a patient weighing 41 kg - 4 tablets per day). On the 10th day from the onset of the disease, the docter noted a complete regression of clinical symptoms and the patient was discharged with recovery.

A test of the sensitivity of seven colon cancer cell lines to a panel of 12 nonpathogenic human enteroviruses revealed significant differences in the ability of tumor cells to become infected and replicate different viral strains. Among the factors that can affect the sensitivity of cells to viruses are differences in the state of the mechanisms of antiviral protection, associated with a reaction to type I interferons. Using the two colon cancer cell lines CaCo2 and LIM1215 as a model, significant differences were revealed in the ability of cells to defend themselves against virus infection after 16 hours of treatment with 1000 units/mL of interferon-alpha. To study the effect of the state of the interferon response system, represented by the Jak/STAT signaling pathway, on the sensitivity of cells to different strains of enteroviruses, HEK293T cell lines were used. These are capable of supporting replication of each of the tested enteroviruses, as well as maintaining the ability to protect against viral infection after the treatment with interferon. Using the CRISPR/Cas9 system, HEK293T sublines with knockouts of the IFNAR1 and STAT2 genes were obtained. The sensitivity of control and knockout cells to infection with five strains of enteroviruses and the vesicular stomatitis virus was analyzed. It was noted that knockout of the IFNAR1 and STAT2 genes resulted in an increased sensitivity to all tested viruses. In knockout cells, the levels of reproduction of the vaccine derived of poliovirus type 1, Echoviruses 7 and 30, and Coxsackie viruses B5 and A7 were also significantly increased in comparison with the control HEK293T cells. Thus, deficiencies in the Jak/STAT signaling pathway in tumor cells lead to an overall increase in the sensitivity to oncolytic viruses.

The article reviews literature and proprietary data on the role of pathogens in the etiology of infectious and non-infectious uveitis. Infectious uveitis is caused by active intraocular replication of the virus (herpesvirus, acute stage of enterovirus), or by long-term persistence of the viruses in eye tissues (Fuchs syndrome associated with rubella virus, late complications of enterovirus uveitis). Clinical picture, severity, outcomes of infectious uveitis depend on the pathogen, adequacy of the immune response and genetic characteristics of the patient. Infections trigger the development of non-infectious uveitis, including autoimmune. Their trigger mechanisms involve antigenic mimicry, bystander activation, epitope spreading, presence of superantigens, intestinal microbiota. An uncontrolled, excessive host immune response contributes to cell destruction even after removal of the infection.

A full analysis has been conducted of the sequences and secondary structures of viral type-I or related IRESs identified in all of the elements that correspond to the previously described minimal fragment of the enterovirus C IRES, which mimics the glycine tRNA anticodon hairpin in the IRES structure and is necessary for the specific binding of glycyl-tRNA synthetase. Experiments on human glycyl-tRNA synthetase binding with the mRNA fragments of several taxonomically distant viruses showed that the binding constants of these complexes are similar. These results indicate that the regulation of translation initiation via glycyl-tRNA synthetase must be a universal mechanism for these viruses and the corresponding parts of their mRNAs must have similar spatial structures. Furthermore, at least one additional mRNA hairpin with the glycyl anticodon loop has been found in all analyzed viral type-I IRESs. It seems plausible that this extra hairpin is associated with the second RNA-binding site of the glycyl-tRNA synthetase dimer and stabilizes its complex with the viral mRNA.

The currently available structural information is insufficient for a detailed analysis of interactions between human glycyl-tRNA synthetase (GARS) and enterovirus IRESs. At the same time, this information is required in order to understand how this IRES trans-acting factor (ITAF) functions during viral mRNA translation, which is in turn crucial for the development of direct-action antiviral agents. In this paper, a theoretical model of the complex between a cadicivirus A IRES fragment and the anticodon-binding domain of human GARS is constructed using molecular dynamics simulation based on all of the available structural and biochemical data. The proposed model enables the structural interpretation of the previously obtained biochemical data.