An escalating pandemic of the novel SARS-CoV-2 virus is impacting global health, and effective antivirals are needed. Umifenovir (Arbidol) is an indole-derivative molecule, licensed in Russia and China for prophylaxis and treatment of influenza and other respiratory viral infections. It has been shown that umifenovir has broad spectrum activity against different viruses. We evaluated the sensitivity of different coronaviruses, including the novel SARS-CoV-2 virus, to umifenovir using in vitro assays. Using a plaque assay, we revealed an antiviral effect of umifenovir against seasonal HCoV-229E and HCoV-OC43 coronaviruses in Vero E6 cells, with estimated 50% effective concentrations (EC50) of 10.0 ± 0.5 µM and 9.0 ± 0.4 µM, respectively. Umifenovir at 90 µM significantly suppressed plaque formation in CMK-AH-1 cells infected with SARS-CoV. Umifenovir also inhibited the replication of SARS-CoV-2 virus, with EC50 values ranging from 15.37 ± 3.6 to 28.0 ± 1.0 µM. In addition, 21–36 µM of umifenovir significantly suppressed SARS-CoV-2 virus titers (≥2 log TCID50/mL) in the first 24 h after infection. Repurposing of antiviral drugs is very helpful in fighting COVID-19. A safe, pan-antiviral drug such as umifenovir could be extremely beneficial in combating the early stages of a viral pandemic.

Introduction. Immunodeficiency underlying the development of severe forms of new coronavirus infection may be the result of mixed infection with SARS-CoV-2 and other pathogens, including Epstein–Barr virus (EBV).The aim is to study the prevalence and epidemiological features of co-infection with SARS-CoV-2 and EBV. Material and methods. A cross-sectional randomized study was conducted in Moscow region from March to May 2020. Two groups were examined for EBV-markers: hospital patients (n = 95) treated for SARS-CoV-2 infection and blood donors (n = 92).Results. With equal EBV prevalence the detection of active infection markers in donors (10.9%) was noticeably lower than in SARS-CoV-2 patients (80%). Significant differences in this indicator were also found when patients from subgroups with interstitial pneumonia with the presence (96.6%) and absence (97.2%) of SARS-CoV-2 in the nasopharyngeal smear were compared with the subgroup of patients with mild COVID-19 (43.3%). The average IgG VCA and IgG EBNA positivity coefficients in donor group were higher than in patient group (p 0.05). Patients with active EBV infection markers were significantly more likely to have pneumonia, exceeding the reference values of ALT and the relative number of monocytes (odds ratio – 23.6; 3.5; 9.7, respectively).Discussion. The present study examined the incidence and analyzed epidemiological features of active EBV infection in patients with COVID-19.Conclusion. A significantly higher rate of detection of active EBV infection markers in hospital patients indicates a combined participation SARS-CoV-2 and EBV in the development of interstitial pneumonia. Low levels of specific IgG EBV serve as predictors of EBV reactivation. Exceeding the reference values of ALT and the relative number of monocytes in patients should serve as a reason for examination for active EBV infection markers.

Introduction. The ability of SARS-CoV-2 antibodies to neutralize the virus is the primary indicator of their specific activity. The test for virus neutralizing antibodies (NAbs) is much needed in different biomedical studies.The aim of the study is to find optimum conditions for microscopic and spectrophotometric detection of SARSCoV-2 NAbs by inhibition of cytopathic effect (CPE) in cell cultures.Materials and methods. Blood sera collected from COVID-19 convalescent patients and healthy individuals (n = 96) were tested using the ELISA method. The SARS-CoV-2 coronavirus, Dubrovka strain (GenBank accession no. MW514307.1) was grown in culture medium of Vero cell line CCL-81 (ATCC). Real-time RT-PCR, ELISA, and Sanger sequencing were used for detection of the virus. The results of the neutralization test (NT) were assessed through the microscopic examination for CPE and by the methyl thiazolyl tetrazolium (MTT) assay.Results. SARS-CoV-2 was isolated from a COVID-19 patient and adapted to grow in cell culture. At a low dose of infection (MOI = 0.00001), the virus caused a pronounced CPE with the cell viability less than 3%, thus making it possible to assess NT results by CPE inhibition. The NT and ELISA-based comparative study of sera showed positive correlation between virus NAb titers and Nab titers to S-protein RBD (Spearman’s r = 0.714; p 0.001). The results of NAbs microscopic and spectrophotometric detection (the MTT assay) also demonstrated positive correlation (Spearman’s r = 0.963; p 0.05).Conclusion. The SARS-CoV-2 virus adapted to Vero cell culture served to develop a NAb titer assessment system, which can be used both in microscopic studies and for an MTT assay in spectrophotometric studies. The MTT assay provides automated reading of NT results, optimizes the statistical analysis of the obtained data, and minimizes subjectivity in assessment of results. Being a vital dye, MTT penetrates only viable cells, thus contributing to the reliability of the obtained results compared to other dyes.

Although WHO declared an end to the pandemic, COVID-19 remains a significant public health concern worldwide. Modern vaccines often induce either only humoral or only cellular immunity. Furthermore, new emergent epidemiologically significant SARS-CoV-2 variants and their spread considerably reduce the effectiveness of preventive vaccination. Therefore, there is an urgent need to improve the existing vaccines against COVID-19. One of the promising approaches to the solution of the problem is creation of a "universal" vaccine that would have a cross-protective activity against different antigenic variants of the virus. In this respect, the development of live attenuated vaccine is of special interest, as it can activate not only humoral, but also cell-mediated components of immunity, providing long-term immune response and cross-protection against different variants of the virus. This review highlights the existing approaches to producing attenuated SARS-CoV-2 strains and gives an assessment of their prospects for clinical use. Some researchers use methods of genetic engineering and reverse genetics such as site-directed mutagenesis and codon deoptimization for virus attenuation. Others tend to use traditional approaches focusing on producing virus mutants through extended passaging in cell culture under selective conditions. The gained experience demonstrates great prospects for development of highly effective live-attenuated vaccine against COVID-19.

Introduction. Animal models for SARS-CoV-2 infection, reproducing the clinical features of COVID-19 in humans, are important tools for studying the pathogenesis of the disease, transmission of the pathogen and are indispensable for testing antiviral drugs and vaccines. The aim of the study was to assess the virulence and tissue tropism for golden Syrian hamsters of SARS-CoV-2 strains belonging to different epidemiologically significant variants: Wuhan-like, Delta, Omicron BA.1.1 and Omicron BA.5.2. Materials and methods. Hamsters were intranasally infected with different SARS-CoV-2 strains. Virulence and tissue tropism of SARS-CoV-2 strains were assessed by comparing the dynamics of weight, viral load in organs and histopathological changes in lungs in infected and uninfected animals. Results. The Wuhan-like Dubrovka strain had the greatest virulence for hamsters, which was manifested by the development of severe pneumonia and a delay in weight gain by 14.6%, high virus content in the lungs, nasal passages and brain — 6.2, 5,9 and 3.7 lg TCID50/ml of homogenate, respectively. Presumably, it was the infection of the Wuhan-like virus of the central nervous system that negatively affected the weight and general condition of the animals. When hamsters were infected with viruses belonging to the Delta and Omicron variants, the observed minor weight loss in animals was uninformative, so indicators such as lung histopathology, viral load in the lungs, nasal passages, heart and other organs played a decisive role in assessing the virus pathogenicity. A score assessment of lung histopathology was of particular value in assessing the severity of pneumonia, since it reduced subjectivity in evaluating the results of histological examination and provided a semi-quantitative assessment of the pathological process. Conclusion. Despite the revealed lower virulence for hamsters of viruses belonging to the Delta and Omicron variants compared to the ancestral Wuhan virus, this animal model for COVID-19 retains its value for conducting preclinical trials of antiviral drugs.

Introduction. Due to the high clinical significance of herpesvirus diseases, the searching of fast and effective methods for their diagnosis remains relevant.The aim of the study was to evaluate the diagnostic efficiency of the loop-mediated isothermal amplification of DNA with real-time fluorescent detection (RT-LAMP) with SYTO-82 dye on a model of herpes simplex virus (HSV) infection.Materials and methods. A total of 44 urogenital swabs containing type 1 and type 2 HSV DNA and 43 swabs without HSV DNA, including 33 samples containing the DNA of cytomegalovirus, Epstein-Barr virus and herpesvirus type 6, were studied. For RT-LAMP, Bst 2.0 WarmStart DNA polymerase, SYTO-82 dye, LAMP primers were used.Results. The high efficiency of HSV DNA detection in the RT-LAMP reaction with SYTO-82 dye was shown. RT-LAMP in optimal conditions allowed to reduce reaction time for 2-3 times compared to real-time PCR (to 35 minutes). Analytical sensitivity of HSV type 1 and 2 detection in RT-LAMP was 103 copies of DNA/ml. The diagnostic sensitivity and specificity of the RT-LAMP diagnosis of HSV infection were 96% and 100%, respectively.Discussion. RT-LAMP method has a high sensitivity and specificity comparable to RTPCR, while the risk of false positive results obtaining is minimal.Conclusion. Thus, the reaction of RT-LAMP with SYTO-82 dye allows quickly, with high sensitivity and specificity to detect HSV DNA in clinical material and can be considered as a promising point-of-care testing method.