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  • 1
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    Online Resource
    Serological methods for detection of the causative agent of tularemia and their evaluation
    Central Research Institute for Epidemiology ; 2019
    In:  Journal of microbiology, epidemiology and immunobiology Vol. 96, No. 4 ( 2019-09-01), p. 32-38
    Details
    In: Journal of microbiology, epidemiology and immunobiology, Central Research Institute for Epidemiology, Vol. 96, No. 4 ( 2019-09-01), p. 32-38
    Abstract: Aim. A comparative study of serological methods for the detection of the causative agent of tularemia and their evaluation. Materials and methods. We used experimental diagnostic kits and test systems for the production of serological methods: indirect hemagglutination reaction (RGA); the reaction immunofluorescence (RIF); enzyme immunoassay (ELISA) using traditional microplate; IFA after selective concentration of the pathogen of tularemia in magnoimmunosorbents (MIS); microgravimetric analysis (MGA) based on piezoresistors (SP) and surface plasmon resonance (SPR). The experiments were carried out with homologous strains of tularemia microbe (test strains) and with strains of heterologous microorganisms in model experiments on tap water contaminated with different concentrations of the pathogen. Results. The parameters of each diagnostic method are determined and evaluated according to the following indicators: sensitivity (when working with pure cultures (test strains), contaminated samples of large volumes), specificity, time of setting and taking into account the results, informativeness, determining the modes of setting and accounting. Conclusion. The above diagnostic methods have their advantages and disadvantages. Therefore, when choosing a method, the researcher should be guided by the goals pursued. So, for screening studies it is advisable to carry out the formulation of ELISA, RIF, RGA, in identifying the pathogen in large volumes and contaminated samples, the effective use of selective concentration on MIS followed by the formulation of ELISA, to identify small amounts of samples and take into account the reaction in real time, it is possible to use MGA and SPR.
    Type of Medium: Online Resource
    ISSN: 2686-7613 , 0372-9311
    URL: Issue
    URL: Article
    DOI: 10.36233/0372-9311-2019-4
    DOI: 10.36233/0372-9311-2019-4-32-38
    Language: Unknown
    Publisher: Central Research Institute for Epidemiology
    Publication Date: 2019
    SSG: 12
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  • 2
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    Assessment of the Effectiveness of Using Magnoimmunosorbents for the Selective Concentration of Anthrax Agent Spores
    Russian Research Anti-Plague Institute Microbe ; 2023
    In:  Problems of Particularly Dangerous Infections , No. 3 ( 2023-10-03), p. 93-98
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 3 ( 2023-10-03), p. 93-98
    Abstract: The aim of the study was to assess the effectiveness of the developed anthrax magnoimmunosorbents (MIS) for the selective concentration of Bacillus anthracis spores and to increase the sensitivity of anthrax agent detection techniques, including when testing soil samples. Materials and methods. We used 10 vaccine strains of B. anthracis and 30 strains of closely related bacilli of the genus Bacillus ( B. cereus – 15, B. thuringiensis – 10, B. megaterium – 5) with typical species properties. The work was performed on three experimental batches of magnoimmunosorbents. DNA extraction and PCR setting was carried out in compliance with the instructions for reagent panel for B. anthracis DNA detection “ApliSens Bacillus anthracis -FRT”. Results and discussion. It is shown that when using MIS, the sensitivity of the cultural method is increased by at least 7 times (taking into account the possibility of sorption of 1–10 or more spores on a sorbent particle). The sensitivity of the PCR method is improved by 10 times and amounts to 50 B. anthracis spores per 1 ml for the samples concentrated with the help of MIS. The sensitivity of the bacteriological method using MIS increases by a factor of 7.5 when testing the artificially contaminated with B. anthracis soil samples. Hence, application of the developed MIS makes it possible to significantly enhance the sensitivity of anthrax agent detection methods and can be considered as an effective means of sample preparation for the investigation of environmental objects (soil).
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2023-3-93-98
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2023
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  • 3
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    Online Resource
    Assessment of the Application of Erythrocytal Diagnosticum (Lyophilizate) in Detecting Tularemia Agent in Natural Foci
    Russian Research Anti-Plague Institute Microbe ; 2022
    In:  Problems of Particularly Dangerous Infections , No. 4 ( 2022-01-26), p. 79-83
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 4 ( 2022-01-26), p. 79-83
    Abstract: Tularemia is a zoonotic disease with a wide geographical dissemination, and its causative agent Francisella tularensis can be used as a bioterrorism agent. The aim of the study was to evaluate the use of a set of reagents “Erythrocytic immunoglobulin dry tularemia diagnosticum” (“DET-Ig”) with the help of control test strains and field material from natural tularemia foci. Materials and methods. Using the introduced erythrocyte diagnosticum, we studied the decontaminated cultures of test strains ( F. tularensis Miura, F. tularensis 55, F. tularensis Schu, F. tularensis 15 NIIEG, Brucella abortus 544, B. melitensis 16-M, B. suis 1330, and Yersinia enterocolitica 64, Y. enterocolitica 178, Y. enterocolitica 383) and environmental samples suspected of containing F. tularensis . Results and discussion. It has been proven that the developed diagnosticum is specific, sensitive, and easy to use for routine diagnostics of tularemia. In the course of laboratory tests of the experimental series of the DET-Ig reagent kit, the possibility of qualitative determination of the tularemia agent in bacterial cultures, biological material and environmental samples in the reaction of indirect hemagglutination was demonstrated. Comparison of the results of use of erythrocyte diagnosticum in liquid and lyophilized forms showed the advantages of drugs after lyophilization: the possibility of transportation and long-term storage at any temperature conditions in various climatic conditions; the setting of the reaction is possible without the use of special diluents. The guaranteed storage term is set for two years (observation period). The results obtained indicate the prospects of introducing the developed drug into healthcare practice.
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2021-4-79-83
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2022
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  • 4
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    Online Resource
    Development of a protective lyophilisation medium and conditions to stabilise the erythrocyte diagnostic preparation of tularaemia immunoglobulin
    SCEEMP ; 2022
    In:  Biological Products. Prevention, Diagnosis, Treatment Vol. 22, No. 2 ( 2022-06-02), p. 196-207
    Details
    In: Biological Products. Prevention, Diagnosis, Treatment, SCEEMP, Vol. 22, No. 2 ( 2022-06-02), p. 196-207
    Abstract: Liquid erythrocyte diagnostic preparations have a practical disadvantage; i.e., long-distance transportation involving possible non-compliance with cold-chain requirements may result in a complete loss of biological activity. A lyophilisation technology is necessary to ensure that the preparations retain their original properties for a long time. The aim of the work was to develop a protective medium and conditions for lyophilisation to stabilise the erythrocyte diagnostic preparation of tularaemia immunoglobulin. Materials and methods : Gelatin, thiourea, trehalose, sucrose, dextran, and Tween 80 were used as excipients for protective media. The authors used nine strains of homologous and heterologous microorganisms of different genera and species to control the lyophilised diagnostic preparation sensitivity and specificity. Evaluation of the main stability-related quality attributes (appearance of the dried preparation, loss on drying, solubility, appearance after reconstitution, appearance after settling, sensitivity, specificity) considered the temperatures specific to the climatic zones where the in vitro diagnostics is intended to be marketed and used. Results : The authors developed protective stabilising media with different compositions, used them in freeze-drying of the preparation and carried out control testing. The most promising was the lyophilisation medium containing a smaller amount of ingredients —6% of dextran, 0.06% of Tween 80 and up to 0.01% of sodium azide—as it was the simplest one to prepare and ensured complete preservation of the quality attributes. The authors carried out practical evaluation of lyophilisation procedures, and the 12–14-hour procedure proved to be the most cost-effective. Conclusions : The results of long-term, or real time, and accelerated stability testing of the lyophilised diagnostic preparation demonstrated the possibility of two-year storage at a labelled temperature of 2–8 °C, as well as at elevated and low temperatures of 30±2 °С and –18 °С, respectively. The tests showed no negative effects of the temperatures on the controlled quality attributes.
    Type of Medium: Online Resource
    ISSN: 2619-1156 , 2221-996X
    URL: Article
    DOI: 10.30895/2221-996X-2022-22-2-196-207
    Language: Unknown
    Publisher: SCEEMP
    Publication Date: 2022
    detail.hit.zdb_id: 3070755-9
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  • 5
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    Using CAST-test to investigate human specific hypersensitivity to the anthrax pathogen
    SPb RAACI ; 2020
    In:  Medical Immunology (Russia) Vol. 22, No. 5 ( 2020-12-01), p. 1017-1024
    Details
    In: Medical Immunology (Russia), SPb RAACI, Vol. 22, No. 5 ( 2020-12-01), p. 1017-1024
    Abstract: We present the results of applying functional cytometric test of antigen-stimulated activation basophils to assess specific immunological reactivity in the people with anthrax, and immunized with anthrax vaccine. As a criterion for antigen-specific basophil activation, we measured expression of the CD63 membrane receptor, which reflects the process of anaphylactic basophil degranulation. To determine spontaneous and antigen-induced activation of basophils (CCR3+CD63+), a FlowCAST reagent kit (Buhlmann laboratories AG, Switzerland) was used. Anthraxin, an experimental anthrax allergen (a hydrolysate the Bacillus anthracis STI-1 strain), manufactured by the Stavropol Anti-Plague Institute, was used as a specific antigen. As based on clinical and experimental data, a threshold value of 〉 10% of anthraxin-activated (CCR3+CD63+) basophils was accepted for the in vitro immunodiagnostic CAST test, as a laboratory criterion for the subjects exhibiting specific immune response, i.e., IgE-mediated sensitization. It was shown that, in anthrax patients within one week after onset of the disease (3-7 days), a positive CAST result was obtained in 92.3% cases; the levels of specific basophil activation with anthraxin averaged 37.9% (12.01 ÷ 78.9%). Immunological examination of individuals three weeks (21 days) after vaccination against anthrax revealed CAST-positivity in all the vaccinated persons. Intensity of anthraxin-induced basophil activation the vaccinated subjects was ranged from 10.87 to 30.03%, averaging 17.86%. The overall values of spontaneous and specific activation ranged within 12.39 ÷ 41.46%. The study opens prospectives for implementation of basophil antigenic activation test in the Flow CAST format in diagnostics of anthrax and to identify specific immune rearrangements after vaccination in humans, as an index of actual vaccination rates. Usage of CAST test with anthraxin makes it possible to identify anthrax patients at the early stages (2-4 days after onset of the disease) including, among patients with an increased CCR3+CD63+ background values, evaluation of immunological efficiency in the cohorts at risk for vaccination. At the same time, it was found that a significant decrease in diagnostic sensitivity of CAST test could be observed in the patients immune to anthrax pathogen who received intensive antibacterial and pathogenetic therapy at the early stages of infection, including glucocorticosteroids (anti-inflammatory drugs) and desensitizing agents that inhibit the degree of hypersensitivity development and its expression.
    Type of Medium: Online Resource
    ISSN: 2313-741X , 1563-0625
    URL: Article
    DOI: 10.15789/1563-0625-UCT-2058
    Language: Unknown
    Publisher: SPb RAACI
    Publication Date: 2020
    detail.hit.zdb_id: 3041761-2
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  • 6
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    Risk Management in the Manufacturing and Use of the Medical Product for 〈i〉in vitro〈/i〉 Diagnostics: Reagent Kit “Lyophilized Erythrocyte Antigenic Tularemia Diagnosticum”
    Russian Research Anti-Plague Institute Microbe ; 2022
    In:  Problems of Particularly Dangerous Infections , No. 2 ( 2022-07-13), p. 115-121
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 2 ( 2022-07-13), p. 115-121
    Abstract: The aim of the work was to implement the risk management strategies in the manufacturing and use of medical products for in vitro diagnostics by the example of the experimental series of the reagent kit “Lyophilized erythrocyte antigenic tularemia diagnosticum”. Materials and methods. We tested experimental series of the reagent panel “Lyophilized erythrocyte antigenic tularemia diagnosticum”. To carry out the identification, assessment and analysis of risks regarding the considered medical product, failure mode and effects analysis (FMEA) was proposed and adapted under production conditions. Identification of risks associated with manufacturing and control of medical products for in vitro diagnostics was carried out using technological regulations, standard operational procedures and manufacturing notes. Results and discussion. The main outcome of the study is the development of a corrective actions system aimed at reducing the risks and ensuring consistent monitoring. The proposed schemes for carrying out the risk management process can be used as standard ones in the design and development of medical products for in vitro diagnostics, taking into account the specifics of each individual manufacturing. The reporting documents developed within the framework of the system are applicable during the inspection of good manufacturing practice and in terms of completing the registration profile of a diagnostic product with subsequent registration in the healthcare system of the Russian Federation.
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2022-2-115-121
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2022
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  • 7
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    Qualitative Indicators of Experimental Brucellosis Antigen Preparations Designed for Cellular Tests in vitro
    Russian Research Anti-Plague Institute Microbe ; 2020
    In:  Problems of Particularly Dangerous Infections , No. 3 ( 2020-10-22), p. 83-88
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 3 ( 2020-10-22), p. 83-88
    Abstract: In order to develop the most diagnostically informative methods for carrying out antigen-stimulated cellular tests in vitro a careful selection of stimulating agent (antigen) is required, possessing an adequate activating potential and providing specificity of the reaction. Objective of the study was to identify the qualitative indicators of experimental batches of brucellosis antigen preparations designed for cellular tests in vitro. Materials and methods . Initially we produced antigen complexes of brucellosis microbe on the basis of the vaccine strains of three epidemically significant Brucella species (B. abortus, B. melitensis, B. suis). Quantitative determination of WsAg and PPBC proteins of experimental preparation series was performed applying capillary electrophoresis. Qualitative composition was assessed through ion exchange liquid chromatography with refractometric detection. Results and discussion . We have specified physical-chemical features, investigated chromatographic profiles and composition of protein fractions, as well as tried the produced experimental batches of brucellosis antigen preparations. After analyzing the defined protein and polysaccharide composition of the obtained WsAg samples, one can conclude that WsAg preparation cannot be used for cellular tests as the probability of non-specific lymphocyte reaction manifestation in vitro was experimentally proven. By contrast, complex brucellosis antigen preparation PPBC has an expressed specific activity and specificity under in vitro conditions and the prospects to be used when developing methodological approaches for laboratory diagnosis of brucellosis and assessment of de facto immunity rate in risk contingents after vaccination. The obtained parameters will allow for proper quality provision when manufacturing the developed experimental PPBC preparation designed for cellular tests in vitro, taking into account modern validation and standardization regulations. 
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2020-3-83-88
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2020
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  • 8
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    Experimental Peroxidase Conjugate for Detection of Specific Antibodies to Anthrax Agent in Enzyme Immunoassay
    Russian Research Anti-Plague Institute Microbe ; 2022
    In:  Problems of Particularly Dangerous Infections , No. 2 ( 2022-07-12), p. 94-100
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 2 ( 2022-07-12), p. 94-100
    Abstract: Anthrax poses a pressing issue for veterinary medicine and public health in many countries, including the Russian Federation, which necessitates the improvement and development of new, sensitive and specific diagnostic tools. The aim of the work was to create an experimental peroxidase conjugate for the detection of specific antibodies to the anthrax pathogen and to optimize the conditions for performing enzyme immunoassay (ELISA). Materials and methods. The peroxidase conjugate was constructed using horseradish peroxidase and Staphylococcus aureus protein A (Sigma-Aldrich, USA). Bacterial antigens isolated from strains of Bacillus anthracis 55ΔTPA-1Spo, B. anthracis Sterne 34 F2 were used as sensitizing agents. The developed experimental batches of the conjugate were tested in ELISA for the ability to bind antibodies in the blood sera of anthrax patients and vaccinated individuals. The sensitivity, specificity, and accuracy of the method were calculated using the built-in functions of the ROCR software package. Results and discussion. The peroxidase conjugate to detect specific antibodies to the anthrax pathogen in the study of clinical material has been developed; conditions for the ELISA performance have been optimized. To interpret the results of the study, a threshold value of the positivity coefficient was used, below which the result was considered negative, and at an equal or higher value, positive. The test demonstrated significant differences in the “positivity coefficient” indicator for the “Healthy”/“Sick” and “Healthy”/“Vaccinated” groups, while the differences between the “Sick”/“Vaccinated” groups were statistically insignificant. The maximum accuracy of the method was observed at blood serum dilutions of 1:250 and 1:500. 100 % intra-run, run-to-run and series-to-series reproducibility has been established for all positive samples. The sensitivity and specificity of the experimental peroxidase conjugates were 100 and 95.8 %, respectively, and the accuracy was 97.6 %.
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2022-2-94-100
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2022
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  • 9
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    STUDYING DEVELOPMENT OF POST-VACCINAL CELLULAR IMMUNITY AGAINST BRUCELLOSIS BY MEANS OF LYMPHOCYTE 〈i〉IN VITRO〈/i〉 TESTS USING AN EXPERIMENTAL ANTIGENIC COMPLEX
    SPb RAACI ; 2019
    In:  Medical Immunology (Russia) Vol. 21, No. 3 ( 2019-07-13), p. 547-554
    Details
    In: Medical Immunology (Russia), SPb RAACI, Vol. 21, No. 3 ( 2019-07-13), p. 547-554
    Abstract: Regulatory framework and methodological approaches to evaluation of immunological effects of vaccination against brucellosis are not established, and the degree of immunological post-vaccinal rearrangement is not yet developed. Due to leading role of cellular immunity in formation of immune protection against brucellosis, evaluation the cellular response in response to antigenic stimulation may be considered the most informative and objective approach to analysis of immune changes in the body during vaccination. In order to develop the most diagnostically informative methods for design of antigen-stimulation cell tests in vitro , a careful selection of a stimulating agent (antigen) is required, which should have a sufficient activating potential, thus providing specificity of reaction under in vitro conditions. The aim of the present study is to study the in vitro specific activity of a protein-polysaccharide antigenic complex from the Brucella abortus 19 BA strain (BrAg), and an opportunity of its application in order to assess the formation of post-vaccinal cellular immunity against brucellosis. The study was performed with white laboratory mice (n = 50) immunized with the Brucella abortus 19 BA strain. The control group (n = 50) consisted of laboratory mice that received a sterile saline solution in a volume of 0.5 ml. Blood samples were taken from immunized and control animals before vaccination, and 7, 14, 21, and 30 days after immunization. By means of flow cytometry, the activation molecules CD25, CD69, MHC II and CD95, expressed on T lymphocytes (CD3 + CD69 + , CD3 + CD25 + , CD3 + CD95 + , CD3 + MHC + ) were determined. To observe the development of immunity, the intensity of expression of T lymphocyte activation markers was calculated using the stimulation quotient. BrAg was used for specific in vitro stimulation of T lymphocytes. The liquid brucellosis allergen (brucellin) was used as an antigen for comparison, when studying opportunity of BrAg usage for assessing the postvaccinal immunity development. The following results were obtained: BrAg has pronounced specific activity, it did not cause non-specific in vitro reactions (activation) of T lymphocytes, thus enabling its application as a test antigen when evaluating development of adaptive vaccine immunity against brucella. Experimental testing of brucellosis antigen for carrying out the in vitro antigen-stimulated cellular reactions, aiming for evaluation of post-vaccinal immunity development against brucellosis, showed that the usage of BrAg promotes increase in diagnostic sensitivity of cellular reactions under in vitro experimental conditions. The applied experimental antigen is a quite promising tool for development of laboratory algorithms for brucellosis diagnostics, and assessment of actual vaccination efficiency in cohorts previously vaccinated against brucellosis.
    Type of Medium: Online Resource
    ISSN: 2313-741X , 1563-0625
    URL: Issue
    URL: Article
    DOI: 10.15789/1563-0625-2019-3
    DOI: 10.15789/1563-0625-2019-3-547-554
    Language: Unknown
    Publisher: SPb RAACI
    Publication Date: 2019
    detail.hit.zdb_id: 3041761-2
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  • 10
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    Online Resource
    To the 70th Anniversary of the Stavropol Research Anti-Plague Institute: Formation, Establishing, Operation, and Prospects of Development
    Russian Research Anti-Plague Institute Microbe ; 2022
    In:  Problems of Particularly Dangerous Infections , No. 1 ( 2022-04-20), p. 156-160
    Details
    In: Problems of Particularly Dangerous Infections, Russian Research Anti-Plague Institute Microbe, , No. 1 ( 2022-04-20), p. 156-160
    Abstract: .
    Type of Medium: Online Resource
    ISSN: 2658-719X , 0370-1069
    URL: Article
    DOI: 10.21055/0370-1069-2022-1-156-160
    Language: Unknown
    Publisher: Russian Research Anti-Plague Institute Microbe
    Publication Date: 2022
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