The main pathogenic factor of Bacillus anthracis is a three-component toxin encoded by the pagA, lef, and cya genes, which are located on the pXO1 plasmid. The atxA gene, which encodes the primary regulator of pathogenicity factor expression, is located on the same plasmid. In this work, we evaluated the polymorphism of the pagA, lef, cya, and atxA genes for 85 B. anthracis strains from different evolutionary lineages and canSNP groups. We have found a strong correlation of 19 genotypes with the main evolutionary lineages, but the correlation with the canSNP group of the strain was not as strong. We have detected several genetic markers indicating the geographical origin of the strains, for example, their source from the steppe zone of the former USSR. We also found that strains of the B.Br.001/002 group caused an anthrax epidemic in Russia in 2016 and strains isolated during paleontological excavations in the Russian Arctic have the same genotype as the strains of the B.Br.CNEVA group circulating in Central Europe. This data could testify in favor of the genetic relationship of these two groups of strains and hypothesize the ways of distribution of their ancestral forms between Europe and the Arctic.

Objective – genotyping by multilocus sequence-typing (MLST) and phylogenetic analysis of 40 Bacillus anthracis strains isolated in Russia and neighboring countries.Materials and methods. In this study, the sequences of seven housekeeping genes of B. anthracis strains were assembled based on the data of a whole genome new generation sequencing, after which the identified mutations and their coordinates were described. The obtained sequences were used for genotyping of the investigated sample using the MLST method. The results are compared with the data presented in PubMLST database. A phylogenetic analysis was performed for the in silico fused sequences of the seven loci of the identified sequence types. The MEGA 7.0 software package was used to build the dendrograms.Results and discussion. Two sequence types (ST) have been found in the examined sample: 35 strains belong to ST-1, and five strains that differed by one common mutation at the glpF locus – to ST-3 (according to PubMLST coding), which emphasizes the genetic separation of this group of strains. One strain has a unique mutation in the gmk gene located outside the region used for MLST.

Bacillus anthracis is a Gram-positive spore-forming bacterium capable of causing anthrax, a particularly dangerous disease of warm-blooded animals. The indication of B. anthracis is often complicated by the hight genetic similarity between B. anthracis and some closely related Bacillus species. In present work, we propose a set of oligonucleotide primers and Taq-man probes intended for simultaneous detection of three hight-specific for B. anthracis chromosomal loci in multiplex PCR. The proposed set of oligonucleotides makes it possible to identify B. anthracis with a high degree of certainty without regard to the plasmid profile of the strain and to differentiate B. anthracis from closely related microorganisms, even those with atypical genetic properties. Key words: Bacillus anthracis, PCR, species diagnostics

Climate change brings new risks of emergence of especially dangerous diseases. The paper reports the possibility of assessing the pathogenic potential of bacteria as demonstrated by studying the allelic polymorphism of anthrax bacterium pathogenicity factor genes, which is a prerequisite for assessing the associated microbiological risks. The allelic polymorphism of the capBCADE operon (capB, capC, capA, capD, and capE genes) encoding the capsule biosynthesis proteins of Bacillus anthracis, and the acpA and acpB genes encoding the expression regulators of this operon have been studied for the first time. A number of single nucleotide polymorphisms (SNPs) were described in the strains of the studied sample, including 5 SNPs in the capB gene, 3 in capC, 4 in capA, 14 in capD, 2 in capE, and 15 in acpB, as well as 7 SNPs and one insertion in the acpA gene. As a result, the sample has been divided into sequence types for each gene and 17 genotypes, which are combinations of the identified sequence types. In silico translation of the detected alleles of the studied genes revealed three isoforms of the CapB and CapA proteins, two isoforms of the CapC and CapE proteins, six isoforms of the CapD protein, five isoforms of the AcpA protein, and four isoforms in the AcpB protein. It has been demonstrated that the SNP in the 351A → G position of capC is a marker of A.Br.Aust94 group strains. Based on the results, A.Br.Vollum group strains were divided into two subgroups. The strains in the evolutionary lines B and C differed from the line A strains by the presence of an 853G → A SNP in the acpA gene. In addition, a previously unknown variable number tandem repeat (VNTR), has been found in the acpA gene and the possibility of using it for differentiating and genotyping of B. anthracis strains has been demonstrated.

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