Characterization of the adaptive immune response of donors receiving live anthrax vaccine

Live anthrax vaccine containing spores from attenuated strains STI-1 of Bacillus anthracis is used in Russia and former CIS (Commonwealth of Independent States) to prevent anthrax. In this paper we studied the duration of circulation of antibodies specific to spore antigens, the protective antigen (PA), the lethal factor (LF) and their domains (D) in donors’ blood at different times after their immunization with live anthrax vaccine. The relationship between the toxin neutralization activity level and the level of antibodies to PA, LF and their domains was tested. The effect of age, gender and number of vaccinations on the level of adaptive post-vaccination immune response has been studied. It was shown that antibodies against PA-D1 circulate in the blood of donors for 1 year or more after immunization with live anthrax vaccine. Antibodies against all domains of LF and PA-D4 were detected in 11 months after vaccination. Antibodies against the spores were detected in 8 months after vaccination. A moderate positive correlation was found between the titers of antibodies to PA, LF, or their domains, and the TNA of the samples of blood serum from the donors.

Evaluation of the AV7909 Anthrax Vaccine Toxicity in Sprague Dawley Rats Following Three Intramuscular Administrations

AV7909 is a next-generation anthrax vaccine under development for post-exposure prophylaxis following suspected or confirmed Bacillus anthracis exposure, when administered in conjunction with the recommended antibacterial regimen. AV7909 consists of the FDA-approved BioThrax® vaccine (anthrax vaccine adsorbed) and an immunostimulatory Toll-like receptor 9 agonist oligodeoxynucleotide adjuvant, CPG 7909 . The purpose of this study was to evaluate the potential systemic and local toxicity of AV7909 when administered via repeat intramuscular injection to the right thigh muscle (biceps femoris) to male and female Sprague Dawley rats. The vaccine was administered on Days 1, 15, and 29 and the animals were assessed for treatment-related effects followed by a 2-week recovery period to evaluate the persistence or reversibility of any toxic effects. The AV7909 vaccine produced no apparent systemic toxicity based on evaluation of clinical observations, body weights, body temperature, clinical pathology, and anatomic pathology. Necrosis and inflammation were observed at the injection sites as well as in regional lymph nodes and adjacent tissues and were consistent with immune stimulation. Antibodies against B. anthracis protective antigen (PA) were detected in rats treated with the AV7909 vaccine, confirming relevance of this animal model for the assessment of systemic toxicity of AV7909. In contrast, sera of rats that received saline or soluble CPG 7909 alone were negative for anti-PA antibodies. Overall, 3 intramuscular immunizations of Sprague Dawley rats with AV7909 were well tolerated, did not induce mortality or any systemic adverse effects, and did not result in any delayed toxicity.

A putative exosporium lipoprotein GBAA0190 of Bacillus anthracis as a potential anthrax vaccine candidate

Background Bacillus ancthracis causes cutaneous, pulmonary, or gastrointestinal forms of anthrax. B. anthracis is a pathogenic bacterium that is potentially to be used in bioterrorism because it can be produced in the form of spores. Currently, protective antigen (PA)-based vaccines are being used for the prevention of anthrax, but it is necessary to develop more safe and effective vaccines due to their prolonged immunization schedules and adverse reactions. Methods We selected the lipoprotein GBAA0190, a potent inducer of host immune response, present in anthrax spores as a novel potential vaccine candidate. Then, we evaluated its immune-stimulating activity in the bone marrow-derived macrophages (BMDMs) using enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Protective efficacy of GBAA0190 was evaluated in the guinea pig (GP) model. Results The recombinant GBAA0190 (r0190) protein induced the expression of various inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), and macrophage inflammatory protein-1α (MIP-1α) in the BMDMs. These immune responses were mediated through toll-like receptor 1/2 via activation of mitogen-activated protein (MAP) kinase and Nuclear factor-κB (NF-κB) pathways. We demonstrated that not only immunization of r0190 alone, but also combined immunization with r0190 and recombinant PA showed significant protective efficacy against B. anthracis spore challenges in the GP model. Conclusions Our results suggest that r0190 may be a potential target for anthrax vaccine.

World-famous scientist Leon Tsenkovsky (1822–1887): to the 200th anniversary of the father of the anthrax vaccine in the Russian Empire

The aim of the proposed article is to cover the life and scientific achievements of Leon Tsenkovsky (1822–1887), a prominent microbiologist, botanist, bacteriologist, traveler, theorist and scientist-practitioner, a talented teacher, creator of his own vaccine against anthrax in Russia, one of the most prominent researchers of primitive organisms at that time. The authors of the article came to the conclusion that the scientist occupies a forward position among the Polish scholars, who were working during the statelessness of Poland and brought glory to their homeland. The narration of the material is presented in the context of the main stages of the scientist's life – „Warsaw”, „Petersburg”, „Yaroslavl”, „Foreign”, „Odessa”, „Kharkiv”. Particular attention is paid to the Kharkiv period of the scientist's life, when he proved himself as a talented scientist-practitioner, creating the so-called „Tsenkovsky vaccine”, which was not inferior in effectiveness to Pasteur's invention. It is demonstrated that in addition to scientific activities, he was engaged in public and teaching activities at the Imperial University and Kharkiv Veterinary Institute. The memory of Leon Tsenkovsky in Kharkiv is immortalized on the pages of a calendar dedicated to prominent Polish scientists associated with the city, as well as in a commemorative plaque that will be unveiled at the former Kharkiv Veterinary Institute in autumn 2021 and dedicated to the prominent Poles, creators of veterinary medicine in the territory of East of Ukraine.

Development of a Simple Method for the Specific Detection of Anthrax Antibody in Sera by Colored Slide Agglutination Test

This study was aimed to identify humoral immune response against anthrax vaccine in mice model by using colored slide agglutination test and detection of field infectivity of anthrax. The field isolates of B. anthracis (n=05) and F34 stern strain vaccine was isolated on agar plates in order to carry out the slide agglutination test. The field isolates of B. anthracis and vaccine bacteria grew on PLET agar medium produced roughly circular and creamy white colonies with ground glass appearance. The bacteria on sheep blood agar media produced rough, sticky, white-gray non hemolytic colonies. Colony polymerase chain reaction (PCR) protocol was adapted to detect fragments of pX01 (596bp) and pX02 (777bp) plasmid of virulent field isolates of B. anthracis. The fragment of pX01 plasmid was only detected in vaccine bacteria. Growth of a field isolate and a vaccine bacteria were colored with crystal violet and used in slide agglutination test to detect anthrax antibodies. The anti-anthrax antibody was prepared by immunizing female mice with 100ül anthrax vaccine through subcutaneous route. Tail bleed were collected on day 0, 30, 60, 120 and 180 of immunization. Cardiac bleed was collected on day 180 of immunization for extensive study. 25μl of diluted (1:10, 1:20, 1:50 and 1:100) antisera and 25μl colored antigen was mixed together onto a clean slide at room temperature and the results was red following 5min, 10mins, 15mins and 20mins of reaction. Unstained antigens and non-immunized sera from the mice were used as control. Results of slide agglutination test showed that the colored vaccine bacteria and field isolates clumped the mice anti-antisera (day 30, 60, and 120) at 1:20 dilution as seen in naked eye but the reaction was seen only at 1:10 dilution while colorless antigens were used. Under microscopic investigation of slide agglutination test, the reaction was read up to 1:100 dilutions with the sera collected at day 30, 60, 120 and 180 of immunization. The Anthrax Sterne strain vaccine induced anti-anthrax immunity in mice that was detected until day 180 of immunization. The clumping reaction was distinct while colored anthrax antigen was used in slide agglutination tests. The colored slide agglutination tests protocol developed in this study can be used to detect anti-anthrax immune response and anthrax bacteria in the field condition with minimum laboratory facilities. Res. Agric., Livest. Fish.7(3): 497-506, December 2020

Theoretical and Experimental Substantiation of Alternative Methods for Quality Control of Live Anthrax Vaccine

Preventive immunisation against anthrax is carried out in accordance with the national Immunisation Schedule for Epidemic Settings. The vaccination is performed using a live vaccine—a freeze-dried suspension of Bacillus anthracis STI-1 vaccine strain spores in a stabilizing media. Improvement of the quality control of immunobiological medicines is a pressing issue and an integral part of the quality management system.The aim of study was to streamline quality control of live anthrax vaccine in terms of the following test parameters: identification and specific activity (total spore concentration).Materials and methods: identification and specific activity (total spore concentration) tests were performed for samples of live anthrax vaccine, batch 266, produced by the 48 Central Scientific Research Institute. The identification test was performed using the B. anthracis immunochromatography test kit for express detection and identification of anthrax pathogen spores produced by the State Research Center for Applied Microbiology and Biotechnology (Obolensk). The specific activity (total spore concentration) was assessed by the visual method and calculated in the Goryaev chamber using the industry reference standard of bacterial suspension turbidity equivalent to 10 IU—OSO 42-28-85 (by the Scientific Centre for Expert Evaluation of Medicinal Products). The number of live spores in live anthrax vaccine was determined by the microbiological method (by inoculating media). The statistical processing of the results was performed using Excel and Statistica 10.0.Results: the authors provided theoretical and experimental substantiation to support the feasibility of using immunochromatography as an alternative identification test method for live anthrax vaccine. Test samples dilutions of 108 microbial cells per millilitre and 109 microbial cells per millilitre are used in the test. The authors developed a test procedure for determination of the total spore concentration (specific activity) in live anthrax vaccine using an industry reference standard of turbidity equivalent to 10 IU, and proposed a formula for calculation of the total spore concentration.Conclusions: the developed test procedures could be recommended for inclusion in the live anthrax vaccine specification files as alternative methods of quality control.