The avidity index (AI) of IgG to the RBD of SARS-CoV-2 was determined for 71 patients with a mild (outpatient) course of COVID-19, including 39 primarily and 36 secondarily reinfected, and 92 patients with a severe (hospital) course of COVID-19, including 82 primarily and 10 secondarily infected. The AI was shown to correlate with the severity of repeated disease. In the group of outpatients with a mild course, the reinfected patients had significantly higher median AIs than those with primary infections (82.3% vs. 37.1%, p < 0.0001). At the same time, in patients with a severe course of COVID-19, reinfected patients still had low-avidity antibodies (median AI of 28.4% vs. 25% in the primarily infected, difference not significant, p > 0.05). This suggests that the presence of low-avidity IgG to RBD during reinfection is a negative prognostic factor, in which a patient’s risk of developing COVID-19 in a severe form is significantly increased. Thus, patients with IgG of low avidity (AI ≤ 40%) had an 89 ± 20.5% chance of a severe course of recurrent COVID-19, whereas the detection of high-avidity antibodies (AI ≥ 50%) gave a probability of 94 ± 7.9% for a mild course of recurrent disease (p < 0.05).

Objectives. To evaluate the effect of cellular genes FLT4, Nup98, and Nup205 on the reproduction of the influenza A virus in A549 human lung cancer cell line.Methods. The work was carried out using the equipment of the center for collective use of the I.I. Mechnikov Research Institute of Vaccines and Sera (Russia). The virus-containing fluid was collected within three days from the moment of transfection and infection and the intensity of viral reproduction was assessed by viral titration and hemagglutination reaction. The viral RNA concentration was determined by real-time reverse-transcription polymerase chain reaction (RT-PCR). To calculate statistically significant differences between groups, the nonparametric Mann–Whitney test was used.Results. In cells treated with small interfering RNAs (siRNAs) targeted at FLT4, Nup98, and Nup205 genes, a significant decrease in their expression and indicators of viral reproduction (virus titer, hemagglutinating activity, viral RNA concentration) was observed at a multiplicity of infection (MOI) = 0.1. Additionally, it was found that a decrease in the expression of target genes using siRNA does not lead to a significant decrease in cell survival. The viral titer in cells treated with siRNA FLT4.2, Nup98.1, and Nup205 on the first day was lower by an average of 1.0 lg, and on the second and third days, by 2.2–2.3 lg, compared to cells treated with nonspecific siRNA. During real-time RT-PCR, a significant decrease in the concentration of viral RNA was observed with siRNA Nup98.1 (up to 190 times) and Nup205 (up to 30 times) on the first day, 26 and 29 times on the second day, and 6 and 30 times on the third day, respectively. For FLT4.2 siRNA, the number of viral RNA copies decreased by 23, 18, and 16 times on the first, second, and third days. Similar results were obtained when determining the hemagglutinating activity of the virus. The hemagglutinating activity on the third day most strongly decreased in cells treated with siRNA Nup205 and FLT4.2 (16 times). In cells treated with siRNA FLT4.1, Nup98.1, and Nup98.2, hemagglutinating activity decreased by 8 times.Conclusions. In the present study, three cellular genes (FLT4, Nup98, and Nup205) were identified—the decrease in the expression of which effectively suppresses viral reproduction— and the original siRNA sequences were obtained. The results obtained are important for creating therapeutic and prophylactic medication, whose action is based on the RNA interference mechanism.

Background: Influenza is one of the most common infectious diseases, which affect the lower respiratory tract, and can lead to serious complications including death. It is known that currently available therapeutic agents and vaccines do not provide 100% protection against influenza viruses. The development of drugs based on the RNA interference mechanism in the context of this problem is a promising area. This paper aims to assess the effect of FLT4, Nup98, and Nup205 cellular gene knockdown on the reproduction of influenza A virus in human lung cell culture. Materials and methods: Influenza virus strain A/WSN/1933 (St. Jude's Children's Research Hospital, USA) was used in this work as well as A549 cell culture (human lung adenocarcinoma, ATCC® CCL-185, USA) and MDCK cell culture (dog kidney cells, Institut Pasteur, France). Small interfering RNAs (siRNAs) (Syntol, Russia) were synthesized for targeting of the FLT4, Nup98, and Nup205 genes. Lipofectamin 2000 (Invitrogen, USA) was used for transfection. After 4 hours, the transfected cells were infected with the influenza virus at MOI = 0.1. Virus-containing fluid was collected within three days from the moment of transfection and the intensity of viral reproduction was assessed by CPE titration and hemagglutination reactions. Viral RNA concentration was determined by RT-PCR. Mann-Whitney U test was used for statistical analysis. Results: In cells treated with siRNA for FLT4, Nup98, and Nup205 genes, there was a significant decrease in the expression of target genes and indicators of viral reproduction (virus titer, hemagglutinating activity, viral RNA concentration) at MOI = 0.1, although the cell survival rate did not decrease significantly. On the first day, the viral titer in cells treated with declared siRNA was lower, on average, by 1 Lg, and on the second and third days, by 2.2-2.3 Lg, compared to cells treated with nonspecific siRNA. During RT-PCR, a significant decrease in the concentration of viral RNA with Nup98.1 and Nup205 siRNA was detected: up to 190 times and 30 times on the first day; 26 and 29 times on the second day; 6 and 30 times on the third day, respectively. For FLT4.2 siRNA, the number of viral RNA copies has decreased 23, 18, and 16 times on the first, second, and third days. Similar results were obtained while determining the hemagglutinating activity of the virus. The hemagglutinating activity decreased mostly (by 16 times) in cells treated with Nup205 and FLT4.2 siRNAs on the third day. In cells treated with FLT4.1, Nup98.1, and Nup98.2 siRNAs, the hemagglutinating activity decreased by 8 times. Conclutions: We identified a number of genes such as FLT4, Nup98, and Nup205, the decrease in the expression of which can effectively suppress viral reproduction. The original siRNA sequences were also obtained. These results are important for the creation of therapeutic and prophylactic agents, whose action is based on the RNA interference mechanism.

Introduction. Varicella-Zoster virus (VZV) is a highly contagious alpha-herpes virus. The diagnosis of chickenpox remains a difficult task especially in cases of breakthrough chickenpox, so the development of reliable laboratory tests is necessary. The simplest and most sensitive serological test for detecting antibodies in human and animal sera is the passive hemagglutination reaction (PHAR). The aim. To develop of a simple, highly sensitive and specific serological tests for the detection of antibodies to VZV in human and animal blood sera. Materials and methods. Human and animal cell cultures; various strains of VZV; human and animal immune sera; monoclonal antibody to VZV glycoprotein (GP) E. Formalin-treated erythrocytes of sheep, chickens and goats, sensibilised with GP of VZV from a virus-containing supernatant were used for PHAR. Results. Cell cultures with the maximum cytopathic effect at VZV infection were selected. A simple original method for obtaining virus-specific VZV GPs using lectins has been developed. Purified GPs were obtained by their elution from sheep erythrocytes after adsorption. The activity of VZV GP was confirmed in PHAR by an antibody diagnostic assay using formalin-treated sheep erythrocytes sensibilised using monoclonal antibodies to GP E of the “vOka” VZV strain (USA). Using GPs from different VZV strains, PHAR test and GP-based enzyme-linked immunosorbent assay (gpELISA) have been developed to detect antibodies in human and animal immune sera. These tests have high sensitivity, specificity and lack of cross-reactivity. Conclusion. A highly specific, sensitive and reproducible tests for the detection of antibodies to VZV have been developed.

The aim of this work was to study the capacity of human T cells immune for SARS-CoV-2, to produce IFNγ, a marker of T cell immunity, in response to stimulation by a peptide pool in whole blood. Eighty samples of whole blood were received from the volunteers with known medical history in 2021, and 258 volunteers were examined in September-October 2022. In this study, 2 detection techniques were used, i.e.: (1) solid-phase enzyme immunoassay to determine antibodies of class G to RBD SARS-CoV-2; (2) IGRA test to determine IFNγ produced by antigen-specific lymphocytes in response to their stimulation by viral antigen. The parameters of the IGRA test were optimized with a sample of 80 specimens from the volunteers. The threshold value of the IFNγ level was determined (4.85 pg/mL), at the diagnostic specificity of 100% (80.6-100), and diagnostic sensitivity of 92.19% (83-96.6%), 95% CI. The study was further continued with a sample of 258 volunteers. Of them, 28.7%, did not exceed the threshold level of IFNγ after stimulation, according to results of the IGRA test. Meanwhile, all volunteers exhibited class G antibodies to RBB SARS-CoV-2. There was no correlation between the levels of antibodies and the level of interferon response in the entire group. When comparing IgG antibody levels and the amplitude of IFNγ (if exceeding the baseline level) in the groups differing in the time of the last vaccination, the median values of the parameters were slightly higher for the subgroup which was revaccinated 1-2 months before the study, while a significant difference between these subgroups was revealed only when evaluating IFNγ, pg/mL (Mann–Whitney criterion, p = 0.0321). According to the results of the study, it can be assumed that all the patients in the sample who were vaccinated and had COVID-19 infection, showed a humoral immune response. However, about a third of them lacked cellular immunity to SARS-CoV-2. There was no correlation between the levels of antibodies and the level of interferon response (Spearman’s criterion). Revaccination within previous 1-2 months has been shown to promote the increased amplitude of interferon response.

Introduction. Epstein–Barr virus (EBV) is a widespread gamma-herpesvirus associated with a number of malignancies, including nasopharyngeal carcinoma and gastric cancer (GC/EBV+). The primary oncogenic protein of EBV is latent membrane protein 1 (LMP1). Genetic variability of the LMP1 protein affects its oncogenic activity and clinical manifestations. Objective. To investigate the optimal threshold value for EBV viral load, determine the correlation between high viral load and the detection of the LMP1 gene in the blood of patients with GC/EBV+, and to identify mutations in the C-terminal domains of the LMP1 gene that may affect protein function. Materials and methods. Total DNA was extracted from 227 blood samples of GC patients. The LMP1 gene was amplified using nested PCR. EBV DNA viral load in blood samples was analyzed by qPCR, followed by receiver operating characteristic (ROC) analysis and interquartile range (IQR) assessment. A selection criterion for positive DNA samples (EBV+) was established based on the mean viral load and ROC threshold. Results. ROC curve analysis and descriptive statistics identified 23 EBV-positive (EBV+) DNA samples, with the following results: AUC (area under the curve) = 0.83, optimal threshold 526.92 copies/reaction, sensitivity 0.69, specificity 0.87, and mean viral load: 513.5 copies/reaction. A statistically significant association was found between detection of the LMP1 gene in the blood and viral load level (χ2 test p = 0.018; Mann–Whitney U test p 0.0001). Additionally, synonymous and nonsynonymous mutations were identified in regions of LMP1 gene coding the C-terminal activating domains (CTAR1 and CTAR2). Conclusion. The findings highlight the heterogeneous molecular nature of EBV-associated gastric cancer, including genetic mutations in the LMP1 gene.