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Anthrax is a particularly dangerous zooanthroponosis caused by the Gram-positive spore-forming bacterium Bacillus anthracis. This disease mainly affects hoofed herbivores, including those used in agriculture, but can occur in other animals and in humans. That is why the majority of studies of this infection are focused on anthrax in humans and farm animals, as the most important issues from a practical point of view. At the same time, the issues of anthrax epidemiology in natural ecosystems are described in the literature in insufficient detail and often very fragmentary. This paper provides a review of the literature describing the main pathways, patterns and risks of the spread of various clinical forms of anthrax infection in nature, primarily among wild animals. Among other things, we cover some non-notable aspects of intestinal infection with anthrax which, for instance, explain the difference in sensitivity to infection in herbivorous and carnivorous mammals and even different sex and age groups within the same species.

The review provides the data on the epidemic condition of tularemia foci in countries neighboring Russia and ongoing preventive measures in these territories. The highest tularemia incidence in the stated countries and in the world as a whole is observed in Finland. In 2019, 42 cases of human infection with tularemia pathogen were recorded in the territory of the Russian Federation, 85 % of which occur in three Federal Districts: North-Western, Central, and Siberian. Epizootic manifestations of infection of varying degrees of intensity were detected in 51 entities of the Federation. Against this background, sporadic cases of tularemia are registered in 17 regions of the country. The most severe epidemic complications continue in Karelia – 9 patients. Seventeen Francisella tularensis cultures were isolated from environmental objects in the Tver, Yaroslavl, Vologda Regions, in the Altai Territory and the Altai Republic. The paper gives an opinion on the regions in which epidemic complications of tularemia are most likely to occur in 2020.

Here, we review the data on mechanisms inhibiting cytotoxic effect of anthrax toxin on the immune system cells. Various disease forms, immunopathogenesis and contemporary methods for anthrax treatment are discussed. In addition, an anthrax toxin was outlined, whereas structural and functional organization of the protective antigen, lethal and edema factors was detailed. A mechanism for association of a protective antigen and lethal factor, protective antigen and edema factor leading to formation of a lethal toxin and edema toxin, respectively, was described. Participation of protective antigen domains in the process of interaction with surface receptors of imunocompetent cells as well as features of binding a protective antigen with lethal factor and edema factor are discussed. A mechanism of endosomal toxin complex internalization and subsequent transfer of effector molecules to the cytosol are described. Effects of the lethal factor and the edema factor on components of eukaryotic cells as well as cytotoxicity mechanisms are analyzed. The approaches to block anthrax toxin action at various stages of toxicoemia have been analyzed based on previously uncovered sequential signs of cytotoxic activity for Bacillus anthracis toxins. Currently available chimeric and humanized monoclonal antibodies are capable of neutralizing B. anthracis toxins at diverse assembly stages, particularly considering the drugs inhibiting: inter-receptor interaction between protective antigen with eukaryotic cells; furin-like enzymes activating prepore assembly; protective antigen oligomerization; binding of the lethal factor or edema factor to the protective antigen; translocation of the lethal factor or the edema factor into cell cytosol; transport of protective antigen with lethal factor or edema factor from endosomes; enzymatic activity of lethal factor or edematous factor. The anti-toxin agents approved for anthrax prevention and treatment in Russia and worldwide are discussed. The limitations of anti-toxin agents and perspectives for their improvement are also described including inhibition of lethal factor activity, interference with integration of toxin components, blockade of interactions between toxic complexes and immune cell receptors.

Objective of the study – assessment of epizootic and epidemic situation on tularemia in 2018 and forecasting the risk of infection in the territory of the Russian Federation in 2019. Analysis of epidemiological situation was carried out on the basis of the data from monitoring activities performed by the Rospotrebnadzor institutions and the data contained in the reports of the Irkutsk Research Anti-Plague Institute of Siberia and Far East, Plague Control Center, Federal Center of Hygiene and Epidemiology, as well as federal statistical survey forms No 5 “Information on preventive vaccination” and No 2 “Information on infectious and parasitic diseases” over the period of January-December 2018 in eight Federal Districts including 85 constituent entities. Given are the retrospective data on tularemia epidemic situation in the territory of the Russian Federation over the past decade. 1944 human tularemia cases were registered in Russia between 2009 and 2018, 1005 out of which occurred during epidemic outbreak in 2013 in Khanty-Manssiysk Autonomous Region. High sporadic and small cluster incidence was mainly observed in the territories of the North-estern and Siberian Federal Districts over the recent years. In 2018, 71 cases of human infection with tularemia agent were reported. Epizootic manifestations of varying degree of intensity were detected in 52 entities of Russia. Against that background, sporadic cases of human infection were registered in 19 regions of the country. For three years epidemic complications expressed to the maximum were observed in the Omsk Region – 18 cases of tularemia infection, and Karelia – 14 cases, respectively. 15 Francisella tularensis cultures were isolated from ambient environment objects in Pskov, Leningrad Regions, Altai Territory, Republics of Altai and Tuva. Conclusions have been drawn in relation to the regions where epidemic complications associated with tularemia are most likely to emerge in 2019.

168 cases of human infection with Francisella tularensis were registered in the Russian Federation in 2017. 80 % of the cases are accounted for by three Federal Districts: North-Western, North-Caucasian, and Siberian ones. Epizootic manifestations of the infection of varying intensity were reported in 61 constituent entities of the Russian Federation. High morbidity rates in 2017 were observed in the Stavropol Territory – 49 tularemia cases, in the Republic of Karelia – 40, Omsk Region – 20, and Saint Petersburg – 12. To assess epizootic and epidemic situation in each constituent entity of the Federation, the numbers of small mammals and their species composition, infection rates of rodents, ticks, mosquitoes, horseflies, hunting animals, nests, regurgitates of birds of prey and excrements of carnivores, surface water bodies, agent culture numbers, isolated from clinical samples, animals, ticks and water, as well as the data on incidence and vaccination of the population in the region were analyzed. The conclusion has been drawn in regard to the territories where epidemic complications are most likely to occur in 2018.

Bacillus cereus sensu lato group, which includes the causative agent of anthrax Bacillus anthracis, has a high degree of genetic similarity. However, in recent years, the development of molecular genetics has made it possible to find some molecular markers for identifying Bacillus anthracis in order to differentiate it from the closely related bacilli, get the clustering of the collection of strains, and describe the phylogenetic relationships between the clusters. This article deals with up-to-date methods of genotyping of Bacillus anthracis; it also evaluates its discrimination ability, informational value, validity of the results, efficiency, simplicity, and ease of use. In addition, international online resources and the possibility of their use for comparing collections of the B. anthracis strains isolated from various parts of the world are discussed.

Rationale: Enterococci are the leading cause of a number of nosocomial and community-acquired human diseases. In the last decade, these pathogens are becoming resistant to antibacterials, including vancomycin. Multidrug-resistant enterococci have been also isolated from agricultural animals in many countries worldwide, which raises concern of scientists because of possible horizontal transfer of resistance genes. Aim : To assess antibacterial sensitivity of Enterococcus spp. isolates collected from the poultry in the Russian Federation from 2013 to 2016, and to identify vancomycin-resistance genes in their genomes. Materials and methods : Eighty-seven enterococci isolates belonging to E. faecalis (n = 47, 54%), E. faecium (n = 25, 28.7%) and other species (n = 15, 17.2%) were collected from clinical samples of 297 heads of poultry (liver, lungs, heart, spleen, contents of the nasal and sinus cavities) from 17 poultry farms of the Northwest, Central, Volga, Ural and Southern Federal districts of the Russian Federation. Sensitivity of enterococci to antibacterials was determined by disk-diffusion and broth microdilution methods. Vancomycin resistance genes van was detected by polymerase chain reaction with specific primers. Results : Most enterococci isolates were resistant to erythromycin (74/87, 85.1%), gentamicin (70/87, 80.5%), ceftriaxone (61/87, 70.1%), ciprofloxacin (56/87, 64.4%), tetracycline (57/87, 65.5%), and rifampicin (48/87, 55.2%), fewer ones to trimethoprim (38/87, 43.7%), ampicillin (28/87, 32.2%), linezolid (15/87, 17.2%) and chloramphenicol (5/87, 5.7%). The vanC type genes ( vanC1 and vanC2/3 ) were identified in 10 isolates. Vancomycin minimal inhibitory concentrations for these isolates were 2 to 8 mg/L. E. faecium with vanC1 gene was isolated from poultry probably for the first time ever. Conclusion : Commercial poultry in the Russian poultry farms is an important reservoir and source of antibiotic-resistant enterococci populations, including enterococci carrying vanC1 and vanC2/3 vancomycin resistance genes.

Objective of the study was to identify phenotypic properties and genetic peculiarities of Bacillus anthracis strains, isolated during the outbreak of anthrax in the territory of Yamal in 2016. Materials and methods . Investigated were the strains of anthrax agent, applying basic and subsequent identification tests and canSNP-, MLVA-genotyping methods and whole genome sequencing. Results and conclusions . The results showed the identity of the phenotypic properties, canSNPand MLVA25-genotypes, and profiles of whole genome-sequencing, regardless of the source of the strains isolation. Confirmed was a common source of human infection. Defined were phylogenetic interrelations of the tested strains and their position in global B. anthracis population. For the first time ever explored was variability of the gene pattern, associated with pathogenicity, and demonstrated – the efficiency of the proposed algorithm for genetic typing.

123 cases of human infection with Francisella tularensis were registered in the territory of the Russian Federation in 2016. 90 % of the total number was reported by three Federal Districts: North-Western, Siberian, and Central ones. Epizootic manifestations of varying intensity were detected in 56 regions. A considerable number of tularemia cases were observed in the Republic of Karelia, Nenets Autonomous District, Omsk and Ryazan Regions, and in Saint-Petersburg. Conspicuous are some recent peculiarities of tularemia manifestations: non-specificity of clinical symptoms, especially at early stages of the disease, their increasing variability against the background of mixed infections with tularemia and other natural-focal infections agents. The most expressed symptoms are fever and lymphatic adenitis; the patients are diagnosed with tularemia during initial days of the sought treatment in 40-45 % of the cases. Based on the data on morbidity rates in humans, epizootiological condition of natural tularemia foci, the numbers of isolated cultures and incidence of the vectors and carriers of the agent, as well as taking into consideration the scope of preventive vaccination in the regions, identified were the territories in which epidemic complication are most likely to occur in 2017.

Analyzed are the data on epizootiological and epidemiological surveillance of tularemia foci, situated in the territory of 85 constituent entities of the Russian Federation. Positive findings, obtained in the process of examination of small mammals, Ixodidae ticks, mosquitoes, and environmental objects, using immunological and molecular-genetic methods; isolated tularemia agent cultures; as well as tularemia incidence rates among the population have made provisions for the assessment of circulation and infection activity in the Regions. In period of 2015, 71 cases of human infection with tularemia have been registered. Given is a brief characteristics of activity of the natural tularemia foci and epidemic situation in the territory of the Russian Federation in 2015. Specified are the regions where preventive vaccination is on a low level. Given is the differentiation of RF territories according to the risk of exposure to tularemia infection for 2016.

F. tularensis strain 15/23-1 recA is different from the vaccine strain F. tularensis line 15 NIIEG by lacking one from the two copies of iglC gene and recA gene has reduced its reactogenicity for experimental animals (BALB/c mice and guinea pigs): transient weight reduction is less expressed, as well as leukopenia and thrombocytopenia during the first week after the immunization, compared to vaccine strain. The concentration of gamma-interferon in mice blood serum after 7 days immunized with strain 15/23-1 recA, was 5 times less than in mice immunized with strain 15 NIIEG. The virulence of the modified strain for mice is less than for the strain 15 NIIEG by more than two orders of magnitude. According to histological studies F. tularensis strain 15/23-1 recA stimulates morphologically a more expressed immune rearrangement in the spleen and lymph nodes as compared to F. tularensis strain 15 NIIEG. The difference of strain 15/23-1 recA from vaccine is that it is not detected in the spleens of experimental animals at 21 days after immunization. The level of specific antibodies in sera of immunized mice and guinea pigs were almost identical in both variants and there was no significant difference in the amount of gamma interferon synthesized by splenocytes of mice under specific induction. Immune mice were protected against subcutaneous challenge with virulent strain of tularemia 503 (subsp. holarctica) and protective immunity formed by the strains was lasting more than 180 days.

) as framework for the production of tools for CCHF virus detection and identification in artificially contaminated samples and clinical specimens containing CCHF antigens was proven efficient.

Aptamers are short single-stranded oligonucleotides which are selected via targeted chemical evolution in vitro to bind a molecular target of interest. The aptamer selection technology is designated as SELEX (Systematic evolution of ligands by exponential enrichment). SELEX enables isolation of oligonucleotide aptamers binding a wide range of targets of interest with little respect for their nature and molecular weight. A number of applications of aptamer selection were developed ranging from biosensor technologies to antitumor drug discovery. First aptamer-based pharmaceutical (Macugen) was approved by FDA for clinical use in 2004, and since then more than ten aptamer-based drugs undergo various phases of clinical trials. From the medicinal chemist’s point of view, aptamers represent a new class of molecules suitable for the development of new therapeutics. Due to the stability, relative synthesis simplicity, and development of advanced strategies of target specific molecular selection, aptamers attract increased attention of drug discovery community. Difficulties of the development of next-generation antibiotics basing on the conventional basis of combinatorial chemistry and high-throughput screening have also amplified the interest to aptamer-based therapeutic candidates. The present article reviews the investigations focused on the development of antibacterial aptamers and discusses the potential and current limitations of the use of this type of therapeutic molecules.

Within the frames of the investigation on constructing of an improved live tularemia vaccine with less reactogenicity by the method of allelic replacement using a suicidal plasmid pGM5, the mutant variants have been obtained from the vaccine strain Francisella tularensis 15 NIIEG with a reduced expression of the superoxide dismutase sodB gene. This reduction was a result of the substitution of rare for frequent natural codons in the F e-superoxide dismutase (FeSOD) gene. It was shown that the decrease in the FeSOD production up to 30% of the level in the original strain had no negative effect on the rate of the tularemia cause reproduction. It was shown that the mutant strains were more sensitive to hydrogen peroxide and paraquat as compared to the vacci- he bacteria. All the strains were shown (by Q-PCR) to have at the log phase the elevated content of gene groEL mRNA encoding GroEL chaperone.

Objective of the study was to carry out comparison between the microorganisms appurtenant to six different species of Bacilli genus as regards mobility of the amplicons obtained using the primer that contained chi-sequence of Bacillus subtilis . Therewith, analyzed were 30 Bacillus anthracis strains and closely related bacilli. Cultures grew on L-agar and L-broth at 37 °C. DNA was isolated from vegetative B. anthracis and Bacillus spp . cultures introducing lysozyme and phenol de-proteinizing. In order to perform single-primer PCR constructed was ChiBs primer - 5’-CTAGGAGCGGG-3’. Single-primer amplification (30 cycles) was carried out at annealing temperature of 47 °C. Comparative analysis of individual electrophoretic amplicon profiles, obtained by means of ChiBs-primer PCR, allows for identification of genetic intraspecific variability and differentiation of B. anthracis strains from the atypical ones and closely related species of bacilli.

Background : Staphylococcus aureus is one of the most important human pathogens and causes over 100 nosological forms of diseases. The lack of data on the spread of S. aureus genetic types specific for different forms of staphylococcal infections in Russia makes it difficult to timely identify and control strains of this epidemiologically dangerous bacterial pathogen. Objective : The aim of the study was to carry out a molecular genetic research of S. aureus isolates obtained during a widespread foodborne illness outbreak among builders at the Pulkovo airport in St. Petersburg in 2013. Methods : The ability of the isolates to produce staphylococcal enterotoxins was studied by immunoenzyme techniques. Gene typing was carried out by sequence-specific primer-based PCR, as well as by sequencing genomic nucleotide sequences of two independent isolates of the pathogen. Results : An enterotoxin A gene in genomes of S. aureus isolates etiologically associated with the outbreak was identified. The production of enterotoxin A by the isolates was shown. According to the complex analysis all isolates producing staphylococcal enterotoxins were identical and constituted the S. aureus strain, sequence-type ST30 and spa-type t2509. The genome of the identified S. aureus strain carried a set of various staphylococcal toxins. The full genome sequence among other techniques revealed high levels of similarity between genomes of the strain under study and well-known reference strain S aureus MRSA 252. Conclusion : The complete molecular genetic study of the S. aureus strain involved into the widespread foodborne illness outbreak was first carried out in Russia, allowing of further using the strain as a Russian reference strain to study potential epidemic outbreaks in the Russian Federation.

Using site-specific mutagenesis constructed were the variants of the vaccine strain F. tularensis subsp. holarctica 15 with the deleted iglC genes. Identified was the fact that genome of the strain 15 contained two copies of iglC gene. Deletion of one of them as well as both had little effect on the cultural-morphological and growth properties of the microbe. At the same time F. tularensis 15 lacking one copy of the iglC gene propagated in mice macrophages several times slower, than the original strain. Inactivation of both of the copies in the chromosome leaded to the emergence of a variant incapable of intracellular reproduction. This capacity in F. tularensis 15/23-2 with two inactivated iglC gene copies was partially recovered after integration of a complementing plasmid. Therewith the data mentioned above testifies to the significance of iglC gene for the process of reproduction in macrophages.

Within the frames of activities attributed to the Reference Center for tularemia monitoring at SRC AMB, genetically identified are 4 isolates of Francisella tularensis , isolated in 2011 in the Altai Territory. These bacteria prove to be virulent for BALB/c mice, DCL being lower than 10 CFU. Using single-primer PCR-typing and MLVA assay distinguished have been the subspecies of the isolates. Three of them refer to the Central Asian subspecies, one – to the Holarctic, the former being isolated in the territory of the Russian Federation for the first time ever.

A model of a controlled conversion of polysaccharide Vi-antigen of S. typhi into zwitterionic antigen is proposed. The immunological properties of modifications of this antigen conjugated to a protein support were studied.

Magnetic immunosorbents (MIS) have been designed using antianthrax monoclonal antibodies immobilized on aluminum silicate—iron oxide matrix activated by sodium perchlorate. MIS allows us to concentrate the analyzed microorganism and to decontaminate culture from concomitant microflora. Diagnostic test systems constructed on the basis of MIS were tested on pure Bacillus anthracis cultures and in the model experiments. The results testify to the high specificity and sensitivity (102–103 spore/ml) of the designed test systems.

Immobilization of anti-B. anthracis monoclonal antibodies (MAbs) on latex microparticles was studied, the optimal load of these MAbs was determined to be 20 µg for 50 µl of the stock latex suspension. The highest sensitivity of latex agglutination test was observed for 1E6 MAbs. Latex suspensions with immobilized MAbs were lyophilized. Their sensitivity and specificity were shown to be highly competitive with those of the stock liquid latex suspension. Latex diagnosticum for Bacillus anthracis spores detection was constructed on the basis of these lyophilized reagents, developed and approved was the regulatory documentation that included their application instructions and technical specifications. Carried out were inter-laboratory and commission tests of experimental prototypes of the designed latex diagnosticum. These preparations demonstrated high sensitivity (from 1·105 to 2·106 spores/ml and even more) and specificity (absence of cross-reactions with spores of different species of sporogenous bacilli at concentration of 108 spores/ml).

Bacteriophage enzyme preparations exolysin and endolysin were studied. Exolysin (a phage-associated enzyme) was obtained from tail fraction and endolysin from phage-free cytoplasmic fraction of disintegrated Salmonella enteritidis cells. A new method for purification of these enzymes was developed, and their molecular masses were determined. The main catalytic properties of the studied enzymes (pH optimum and specificity to bacterial substrates) were found to be similar. Both enzymes lyse Escherichia coli cells like chicken egg lysozyme, but more efficiently lyse S. enteritidis cells and cannot lyse Micrococcus luteus, a good substrate for chicken egg lysozyme. Similar properties of exolysin and endolysin suggest that these enzymes are structurally similar or even identical.

At present, in compliance with sanitary and epidemiologic regulations in force, equipping of microbiological laboratory with modern engineering and technical systems and equipment is considered to be the important condition ensuring adherence with the biological safety requirements. The abovementioned systems and equipment, together with the rules and techniques of the safe work with pathogenic microorganisms, should provide the set of biological safety measures for the laboratory personnel. Biological safety cabinets of the second and the third classes serve as additional protection means. Considered are peculiarities of the design and benefits of different biosafety cabinets used in BSL 2-3 laboratories, and some of their shortcomings are determined.

It has been established that all anthrax bacteria in an infected organism have the same morphology and fine structure typical for Bacillus anthracis cells. Anthrax bacteria under conditions of a macroorganism form a capsule and slime. A massive bacterial capsule 0.1–0.4 μm thick is characteristic for bacteria of highly virulent and virulent B. anthracis strains. Microbes of avirulent, weakly virulent, and many virulent strains of the anthrax pathogen form a capsule 0.05–0.75 μm thick.

The possibilities of bacteriophage SPZ7 endolysin functioning in the lysis of gram-negative bacteria “from without” were studied. A significant 1.5–3-fold increase in bacteriophage SPZ7 endolysin activity during the lysis of S. enteritidis N60 and E. coli TG1 cells in the presence of high-molecular surfactants, pluronics with a large hydrophobic block, hen’s egg lysozyme, and low concentrations of a peptide antibiotic (colistin), was shown. The developed approach may be promising for improving the efficiency of pharmaceutical bacteriophage enzyme-based antibacterials working against gram-negative microorganisms.

National and foreign literature was analyzed. Considered were nutrient media for plague agent growth, isolation and differentiation. Present state of nutrient media production was highlighted.

We and other authors have recently shown that the pattern of the immune response to components of anthrax, the Bacillus anthracis lethal toxin, is complex. In addition to the neutralizing antibodies, the antitoxin antibody pool contains antibodies enhancing the toxin lethal action. We mapped the epitopes in the protective antigen that are responsible for the induction of both antibody types. In this study, we obtained new data on the cytotoxicity of the B. anthracis lethal toxin toward the J774 A.1 cell line in the presence of monoclonal antibodies to various domains of the protective antigen and the lethal factor. The role of the Fc fragment of immunoglobulins in enhancing the lethal toxin action was shown. These results may serve as a basis for the development of a new generation vaccine for anthrax.

A knockout mutant with a deletion in a quorum sensing system gene qseC was generated from the vaccine strain Francisella tularensis 15 by site-directed mutagenesis. The variant with the inactivated gene qseC differed from the parental strain in growth rate on solid nutrient medium but had the same growth dynamics in liquid nutrient medium. The mutation abolished almost completely the resistance of the vaccine strain to normal rabbit serum and its ability to survive in macrophages; in addition, the strain lost the residual virulence. A significant phenotypic alteration was observed in the lipopolysaccharide of F. tularensis. Particularly, the mutant strain synthesized no noticeable amount of the lipopolysaccharide with the high-molecular-mass O-polysaccharide, presumably as a result of impairing biosynthesis of the repeating unit, namely, a loss of the ability to incorporate a formyl group, an N-acyl substituent of 4-amino-4,6-dideoxy-D-glucose.