[The application of micro-cultural enzyme-linked immunosorbent assay of modified immunofluorescence technique for diagnostic of adenovirus infection.]

The particular conditions of professional activities of drafty military personnel determine wide-spread of of respiratory viruses in crew of Armed Forces. The frequent mixing of military staff conditions of carrying of infection agents , including adenoviruses. It is known, that up to 60% of acute respiratory viral disease in newly formed staff have adenovirus etiology. In these cases, the most frequently are isolated serotonins 4 and 7 of adenovirus. The diagnostic possibilities of monoclonal immunologic tests of indirect enzyme-linked immunosorbent assay and micro-cultural enzyme-linked immunosorbent assay for diagnostic of adenovirus infection are investigated. The analysis was applied to 40 clinical samples from patients with diagnosis of acute respiratory viral disease residing for treatment in military medical organizations during April-June 2014. The culture of cells A-549 infected with materials from patients was used for analysis. The evaluation of reproduction of adenovirus in infected culture of cells using both techniques was implemented by application of monoclonal antibodies to hexon of adenovirus on stage of detection. The availability of adenovirus was proved applying polymerase chain reaction in 20 samples; isolation of cell culture in 19 samples; indirect enzyme-linked immunosorbent assay - in 14 samples; micro-cultural enzyme-linked immunosorbent assay - in 14 samples. For detection of serotypes of adenovirus isolation of DNA and sequencing of 10 of analyzed samples positive for adenovirus according results of polymerase chain reaction were implemented. The results of phylogenetic analysis on site of gene string demonstrated belonging pofadenovirus out of all samples to serotype 4 (subgroup E). The sensitivity of indirect enzyme-linked immunosorbent assay and micro-cultural enzyme-linked immunosorbent assay in detection of adenovirus in cell cultures infected with materials from patients, in comparison with polymerase chain reaction, made up to 85% and 87% correspondingly. The specificity of both techniques reached 100%.

[THE APPLICATION OF DOT-TECHNIQUE FOR DETECTING ANTIGENS OF ADENOVIRUS IN CLINICAL SAMPLES].

The article substantiates possibility of application of point enzyme-linked immunosorbent assay (dot-technique) for detecting viral antigens in samples from patients. To diagnose adenovirus infection conjugate of virus-specific monoclonal antibodies and peroxidase of horse-radish were used The chromatographic rectification of conjugate from free peroxidase permits diminishing background coloring of nitrocellulose membrane and therefore to increase sensitivity. The application of direct conjugates on the basis of virus-specific monoclonal antibodies increases specifcity of dot-technique and significantly shortens time period of analysis. As in case of application of direct conjugates on the basis of polyclonal serum, samples from patients require preliminary processing with detergent for preventing non-specific reactions. The dot-technique demonstrates good coincidence with data of polymerase chain reaction and after clinical trials it can be used in diagnostic of human viral infections.

[New monoclonal kits for the diagnosis of the adenoviral infection].

Diagnostic properties of new monoclonal antibodies (MAbs) to hexon adenovirus antigen (AB) monoclonal ELISA kit for early diagnosis of adenoviral infection were tested. Developed ELISA kit and FITC-conjugate of new monocional antibodies for immunofluorescent analysis were used for detection of different types of adenoviruses in clinical materials. The availability of their use in clinical and epidemiological practice was validated.

The investigation of the dynamics of changes in neutralizing antibody titers against type 5 adenovirus in the context of vaccination against a new coronavirus infection.

This research focuses on analyzing the dynamics of neutralizing antibody (nAbs) titers against type 5 adenovirus (Ad5) in the adult population of Russia following vaccination against the novel coronavirus infection with recombinant adenovirus type-5 COVID-19 vaccine (CanSino Biologics, China). The impact of the Ad5 vector on nAb titers was investigated using 302 blood serum samples from individuals who received a single dose of the Ad5-nCoV vector vaccine. The research revealed that 33.8% of adults in Russia had pre-existing anti-Ad5 nAbs before the pandemic. Notably, 40% of vaccinated individuals did not exhibit an increase in nAbs titers upon receiving the Ad5-based vaccine. However, in the group with no or low titers of anti-Ad5 nAbs (1:10-1:40), a significant 8-16-fold increase in nAb titers to Ad5 was observed.

[The application of synthetic peptides to characterize the site-directed antiviral humoral immune response in patients with respiratory syncytial viral infection].

The article deals with the study of characteristics of epitope-specific humoral immune response to respiratory syncytial viral infection depending on nature of disease and patients' age. The couple serums from 226 children and adults with respiratory syncytial viral infection were analyzed. To detect in enzyme immunoassay the epitope-specific IgG the synthetic peptides were applied imitating the structure of functionally depended epitopes of F-protein of respiratory syncytial virus with amino acid sequences 221-232 (F = SP12), 479-491 (F-SP13) and G-protein with amino acid sequences 152-164 (G-H13), 184-198 (G-T15). The respiratory syncytial viral infection neutralizing antibodies were detected using the microneutralization reaction. The rate of seroconversions of epitope-specific IgG consisted 21-25% in children 3-18 years old under primary respiratory syncytial viral infection and increased up to 42-50% in children 3-18 years old and adults under recurrent episodes of diseases. In these groups, the seroconversions of respiratory syncytial virus neutralizing antibodies were observed in 42%, 65% and 58% of cases correspondingly. Independently of age of patients with diagnosed respiratory syncytial viral infection, the absence of conversions of neutralizing antibodies was statistically significant associated with the absence of response from epitope-specific IgG. The presence among B-cell epitopes of immune dominance of surface glycoproteins of respiratory syncytial virus in patients with recurrent (but not primary) respiratory syncytial viral infection is detected The hierarchy of epitope-specific immune response in case of complicated course of disease (response activity to epitopes: SP12 = G-T15 > G-H13) differed from the case of uncomplicated course of disease.

Acute viral infections with combined involvement of the respiratory and gastrointestinal tracts in children. Therapy with interferon.

We evaluated the percent of acute respiratory viral infections with gastrointestinal syndrome in the structure of morbidity in babies aging 6 months and elder. Therapeutic efficiency and safety of anaferon (pediatric formuation) as a component of complex therapy of acute respiratory viral infections with involvement of the gastrointestinal tract were proven; more rapid disappearance of all symptoms and improvement of the immune status parameters were demonstrated.

GENETIC DIVERSITY OF ADENOVIRUSES CIRCULATING AMONG THE MILITARY IN THE NORTH-WEST REGION.

The contribution of adenovirus (AV) infections to the overall structure of acute viral respiratory infections among young people of draft age can reach as high as 64.6%. Wide dissemination, the incidence of AV-associated pneumonias and lethal outcomes in the case of some complicated infections illustrate the urgency of studying the antigenic diversity of AVs circulating among the military. 991 nasopharyngeal swabs from patients hospitalized in military health facilities with symptoms of acute respiratory infections from 2014 to 2017 were detected by real-time PCR. Sanger sequencing was performed using forward and reverse primers matching the fiber gene. AVs were detected in 326 samples. In 80 of those, AVs were present in combination with other respiratory viruses, as follows: 26 with respiratory syncytial viruses (RSV), 49 with rhinoviruses, 2 with bocaviruses, 1 with RSV and rhinovirus, 1 with parainfluenza virus, and 1 with metapneumovirus. 31 samples were sequenced. Thirty AVs belonged to group E (serotype 4), and 1 AV belonged to group B (serotype 7).

[Characteristics of monoclonal antibodies to RS virus in immunoenzyme and immunofluorescence techniques]. / Kharakteristika monoklonal'nykh antitel k RS-virusu v immunofermentnykh i immunofliuorestsentnykh reaktsiiakh.

According to the competitive ELISA test data, new preparations of monoclonal antibodies (MAb) to respiratory syncytial virus (RSV) differed in their blocking activity and they were directed to 3 different virus epitopes. MAb 9C5 and MAb 131-2A (CDC, Atlanta) competed against each other strongly and they were directed to epitope F1a of RSV F-protein. MAbs 8C5 and 10D8 showed a two-way blocking and were presumably topologically linked. MAb 8B10 did not compete against other MAbs. The fact that MAbs 8B10, 9C5, and 10D8 may be used in indirect ELISA test to detect RSV antigens was shown. The specific activity of MAbs 9C5 was observed, by applying low antibody concentrations to ng/mg. The highest specific activity using the homologous pair MAbs-con 9C5 and 8C5 was observed in direct ELISA employing MAbs for capture and peroxidase conjugate of MAbs for detection. MAbs 8B10 were successfully used for direct (FITC-conjugate of MAbs) and indirect immunofluorescence detection of RSV antigens in the infected cell cultures and clinical materials from patients.

[Isolation and characteristics of anti-adenovirus monoclonal antibodies in immunoenzyme and immunofluorescent reactions]. / Poluchenie i kharakteristika monoklonal'nykh antitel k adenobirusu v immunofermentnykh i immunofluorestsentnykh reaktsiiakh.

New monoclonal antibodies (MAbs) to adenovirus hexon, highly active in ELISA and immunofluorescent analysis, were prepared. According to competitive ELISA, new MAbs differed in their blocking activity and were directed to 2 different hexon epitopes. MAb 3H8 did not modify antigen binding of the rest MAbs labeled with peroxidase (PAb x Pox), and none of unlabeled MAbs suppressed the reaction of MAb x Pox 3H8. MAbs 1E8 7F1, 1E11, and 3B1 reacted with each other but differed by the spectrum and level of competitive inhibition, which indicated that they were directed to different epitopes of adenovirus hexon. Comparison of the specific activity of MAbs 7F1 and 1E8 in direct immunofluorescent detection of adenovirus antigens in infected cell cultures and clinical materials from patients showed a good coincidence (90-97%) of the results with the IMAGEN Adenovirus test (Dako) and with polyclonal FITC conjugates to adenovirus hexon.