AMENDMENTS TO THE CONSTITUTION OF THE RUSSIAN FEDERATION AS THE BASIS OF IDENTITY POLICY IN THE RUSSIAN FEDERATION

The article is devoted to the consideration of amendments to the Constitution of Russia approved during the vote on July 1, 2020, which set the main parameters of identity policy in our country. Special attention is paid to the reaction of public figures, experts and representatives of the leadership of the republics within the Russian Federation. During the discussion of the amendments, there was a strong public demand to clarify and fix in the form of constitutional provisions the main content of cultural and civilizational parameters of the Russian identity. The adoption of the amendments demonstrated broad public agreement on the constitutionally defined parameters of Russian national identity. The discussion showed an ambiguous attitude of ethnic activists in the republics to the set parameters of identity policy. And the leaders of a number of republics expressed their dissenting opinion, which reflected a certain tension over the inclusion of the concept of “state-forming people”in the article on the state language

Antigenic Components of Chemical Bivalent Cholera Vaccine, Methods of their Isolation and Control

The paper presents a review of the data on the methods of isolation and control of Vibrio cholerae antigens – cholerogen-anatoxin and O-antigens of Inaba and Ogawa – components of the oral bivalent chemical cholera vaccine produced by the RusRAPI “Microbe”, the only prophylactic drug against cholera registered in the territory of the Russian Federation. Currently, the vaccine is produced using the method of segregated manufacturing of cholerogenanatoxin and O-antigens Inaba and Ogawa with step-by-step control of their main properties, which ensures the production of a high-quality finished product. Ultrafiltration is an effective method for concentrating a semi-finished product, which helps to reduce losses and increases the yield of the final product. It remains promising to develop a method for gentle steril ization of O-antigens to maximize the preservation of specific activity. To control the specific activity of the antigenic components and the finished vaccine preparation, a complex of in vivo and in vitro methods is applied. However, the multi-stage process and duration, the use of several types of laboratory animals, as well as modern WHO requirements determine the need for the introduction of alternative in vitro control methods. The use of cell cultures as a replacement for the biological method appears prospective, and demonstrates a positive correlation with animal tests. To assess the activity of antigens, the use of an immunochemical method – dot-immunoassay with gold nanoparticles – is put forward, which will make it possible to harmonize the control method at all stages of the production process, as well as to determine the serovar specificity of Vibrio cholerae O-antigens. The development of molecular-genetic, microbiological, immunochemical methods is relevant for a more complete and comprehensive control of the main immunogens of industrial strains of cholera vibrio. The introduction of promising methods for obtaining antigens and monitoring their properties will allow for a more complete characterization of the component composition of the finished dosage form of the chemical cholera vaccine

CHARACTERISTICS OF THE PROTECTIVE ANTIGEN COMPLEX OBTAINED FROM FRANCISELLA TULARENSIS SSP. NOVICIDA

F. tularensis ssp. novicida, considered earlier as a representative of a separate species, has been recently classed among F. tularensis variety, based on the results of comparative analysis of 16S-ribosomal RNA. Subspecies novicida can cause disease only in immunocompromised humans and is low virulent for rabbits. Despite this, high rate of homology of the nucleotide sequence of F. tularensis intraspecific taxon is established. Objective of the study is to obtain protective surface antigen complex from F. tularensis ssp. novicida Utah 112 (ATTC 15 482) cells and investigate its properties. Materials and methods. Protein, carbohydrate, and lipid content of the antigen preparation was measured using conventional colorimetric methods, SDS-PAGE was conducted according to U.Laemmli, and immunoblotting – to H.Towbin. For purification and molecular mass determination column chromatography was applied. Immune-chromatographic activity was analyzed by immune-enzyme assay. Immunogenicity of the produced preparation was tested on scrub white mice, with LD50 and ED50 calculated according to Karber’s method. Results and conclusions. Carried out has been comparative analysis of physical-chemical, antigenic and bio-chemical peculiarities of the protective antigen complex obtained from F. tularensis Utah 112 cells and equivalent antigen complex obtained from the vaccine strain – F. tularensis 15 NIIEG. Protectivity of the preparation has been tested through inoculation of the immunized white mice with virulent F. tularensis 503/840 strain. Demonstrated have been distinctive features of the new preparation, by structure and composition, as compared to similar antigen from the vaccine producer strain, as well as the slowdown of its immunochemical and protective activities

Enhancement of Manufacturing Technology for Finished Dosage Form of Bivalent Chemical Tableted Cholera Vaccine

Objective of the study is to experimentally substantiate the possibility to improve manufacturing efficiency by means of mass reduction of a vaccine tablet from 300 to 100 mg. Materials and methods. Inaba O-antigen lyophilizate serves as the specific immunogenic component of the vaccine. Results and conclusions. It is identified that it is expedient to produce tablets of 6 mm in diameter. Justified is the quantitative content of additive substances (lactose monohydrate, micro-crystal cellulose, and polyvinylpyrolidone). Moreover, the studies have specified target values for technological parameters of such processes as fluid bed granulation of the formula with overfeed of the binder, tablet compression and enteric-coating (Acryl-eze) application to finished dosage form. Using Inaba O-antigen lyophilizate manufactured has been model experimental series of the vaccine. Investigated have been its characteristics. Verified vaccine quality indicators testify to the compliance of the product with the requirements of manufacturer’s pharmacopoeial monograph. The studies exercised showed the possibility in principle to enhance manufacturing efficacy through the decrement of additives amounts, and thus the mass of a vaccine tablet from 300 up to 100 mg

Development of Methodological Approaches for Examination of Particularly Dangerous Infectious Diseases Agents by Means of Atomic Power Microscopy

The atomic power microscopy (APM) is used to study the cell surface structure of particularly dangerous infectious diseases agents and to carry out the morphometric analysis. APM shows similar results with scanning electron microscopy. However, its application makes it possible to avoid time-consuming and labour-intensive procedures of samples preparing for testing by fixation, dehydration and sputtering of conducting layer. Methodological approach has been elaborated for preparing and analysis of samples of agents of particularly dangerous infectious diseases by means of APM. This approach includes a selection of optimal substrate, mode of disinfection and scanning of samples

Deployment of Ultrafiltration for Concentrating and Purification of Antigens

Represented is domestic and foreign literature review dedicated to usage of ultrafiltration for concentrating and purification of antigens. Discussed are the issues of deployment of various ultrafiltration techniques. It is determined that filtering in the tangential mode by means of modules with flat-frame filtering elements is among the prospective ones. Demonstrated is the impact of such technological specifications as concentration rate, pressure, temperature, and membrane nominal cut-off on molecular mass on the quality of target products, the time elapsed, and preparation losses decrease (increase). Literature data analysis proves to be useful for the selection of the proper procedure for concentrating and purification of protective antigens of bacterial and viral origins. In addition, it allows for taking into account the parameters under discussion when developing specific manufacturing technologies for diagnostic and preventive medical immunobiological preparation production

Experimental Substantiation of the Possibility to Use Finite Cell Line CHO-K1 for Determination of Specific Activity of Components of Chemical Cholera Vaccine

Objective was to experimentally substantiate the possibility to use the finite cell line CHO-K1 for measuring specific activity of cholera toxin and component of the vaccine choleragen-anatoxin in the process of chemical cholera vaccine manufacturing. Materials and methods. The studies involved the finite cell line CHO-K. The registration of results of bio-indication method was performed visually with the help of inverted microscope and photometrically - in colorimetric test for the assessment of metabolic activity of the cells at the wave length of 595 nm. Results and discussion. The proposed method allows for determining the toxin-production activity of Vibrio cholerae 569B strain during submerged cultivation in bioreactor and specific activity of choleragen-anatoxin by anatoxin binding measuring using cell cultures. The results correlate with the data obtained using intradermal Craig’s technique, GM1-ELISA and radial passive immune hemolysis (RPIH). Introduction of cell culture method into practice will provide for significant decrease in the volumes of usage of animals at the stages of manufacturing of chemical bivalent cholera vaccine

Investigating the stability of the Properties of Vibrio cholerae strains – Producers of Active Components of the Chemical Cholera Vaccine

One of the key requirements to producer strains used in the manufacturing of immunobiological preparations is their stability, which consists in maintaining the main cultural, morphological, physiological, and productive properties in a series of generations. This paper describes a comprehensive methodological approach to testing strain stability using in vitro techniques.The purpose of this study was to conduct an integrated analysis of the stability in the strains that produce active components of the chemical cholera vaccine when preparing seed material and at the stage of cultivation.Materials and methods. Toxigenic strains of Vibrio сholerae 569B of the classical biovar, serovar Inaba and V. сholerae M-41 of the classical biovar, serovar Ogawa were used in the work. Cell morphology was monitored through light and transmission electron microscopy. Atomic force microscopy was applied to measure the main parameters of the bacterial cell. The strains were tested for the presence of ctxA gene in the chromosome using the “GenChol” test system with electrophoretic registration of results. Whole genome sequencing of the strains was performed on the Ion Torrent PGM platform using the Ion 318 Chip Kit and the Ion PGM Hi-Q View Chef 400 Kit. To determine the specific activity of cholera toxin and O-antigen, a DOT immunoassay with a conjugate based on staphylococcal protein A and colloidal gold nanoparticles was applied.Results and discussion. The stability of the main properties of industrial V. сholerae strains – producers of the active components of the chemical cholera vaccine has been confirmed using microbiological, immunochemical, molecular-genetic methods and microscopic analysis at all stages of cultivation, and the prospects for using the integrated methodological approach experimentally substantiated. Tailoring of these methods will make it possible to control the stability of producer strains, optimize cultivation conditions and, as a result, increase the yield of the necessary antigenic component of the vaccine

Mathematical Model of Kinetics of O-Antigen Accumulation in the Process of Periodic Submerged Cultivation of <I>Vibrio cholerae</I> M-41 Ogava with Limitation on Carbon Substrate

Developed is the system of differential equations that characterize kinetics of biomass growth, glucose utilization and O-antigen accumulation in the process of submerged cultivation of V. cholerae strain M-41 Ogawa. The parameters of the mathematic model are identified. Using the developed software in Mathcad 15.0 determined are kinetic constants and coefficients. The mathematical model is demonstrated to describe adequately O-antigen biosynthesis process. The received data can be used in large-scale technology of V. cholerae strain M-41 submerged cultivation

Mathematical Model of Kinetics of Antigens Accumulation in the Process of Periodical Submerged Cultivation of <I>Vibrio cholerae </I>569В Inaba with Limitation as Regards Carbonic Substrate

Presented is mathematical model of kinetics of the process of O-antigen and cholera toxin synthesis during periodical submerged cultivation of V. cholerae 569В Inaba with limitation as regards carbonic substrate. The proposed model is based upon analysis of experimental data on V. cholerae 569В Inaba biomass and antigens accumulation, rate of growth and antigens release, and glucose utilization. Using Mathcad 15.0 software calculated are coefficients of differential equations entering into the mathematical model. Comparison of predicted and experimental data demonstrates that relative error of determination of concentrations of the synthesized substances, glucose and cholera vibrio is between 5 and 20 %. The proposed model permits to determine maximum output of final products and specify the parameters of cultivation process performance at different initial conditions