Lipopolysaccharide and outer membranes from the three virulent encapsulated (Cap(+)) strains of three subspecies of Francisella tularensis and their isogenic avirulent capsule-deficient (Cap(-)) mutants were isolated. It was shown that the Cap cells and their outer membranes almost completely consumed the available complement of normal human serum whereas Cap(-) LPS (R-LPS), Cap(+) cells and their components activated the complement less effectively. Absorption of normal human serum with Cap(-) strain dramatically reduced the complement consumption for homologous strain and its surface structures. This reduction reflected the loss of bactericidal antibodies. Addition of antibodies to whole cells of F. tularensis completely restored complement activity. The cross-absorbing experiments demonstrated that Cap(-) cells more effectively deplete bactericidal antibodies than homologous virulent strain. From these results it can be concluded that normal human serum is bactericidal for serum-sensitive Cap(-) F. tularensis strains through the action of complement initiated by the classical complement pathway and serum resistance of virulent strains is not due to absence of targets for bactericidal antibodies, but is due to their low accessibility because of O-side chains of lipopolysaccharide.

The aim of this study was to detect canonical insertion/deletion (INDEL) markers in the genome of Helicobacter pylori and offer INDEL-typing method for differentiation of H. pylori strains. For comparative analysis of the genomes of H. pylori presented in the GenBank database, a local database of nucleotide sequences of 69 H. pylori strains was created. For detecting all INDEL markers with a preset size of 6-20 bp a pairwise comparison of more than 1500 open reading frames (ORF) in the genomes of local database strains was performed. Ten loci containing INDEL markers were founded. The five most variable loci were tested in silico with 21 strains with known geographical origin from the most common populations of hpEurope, hspWAfrica, and hspEAsia. Fifteen individual genotypes with a high diversity index (DI=0.95) were identified. For cluster analysis, the minimal spanning tree (MST) method was used, which demonstrated a clear distribution of clusters according to the geographical origin of the strains tested. INDEL-typing of 21 regional strains from the Astrakhan region was performed in vitro. It was shown that an extensive majority of them belong to the population hpEurope. The findings in this study indicate that the proposed INDEL-typing method almost perfectly reflects the geographical distribution of H. pylori strains determined by the multilocus sequence typing (MLST) method, despite the fact that the primary object of research is completely different genes. Further research is needed to determine the geographical origin of H. pylori strains in Russia.

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Aims: To improve a multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) assay for Helicobacter pylori typing. Materials and Methods: Polymorphic VNTRs were searched by Gene Expert. The distribution and polymorphism of each VNTR locus were analyzed in 18 H. pylori genomes from the NCBI genome database by BLAST and were compared with a collection of 15 clinical H. pylori strains. The MLVA assay was compared with MLST-typing for discriminating H. pylori isolates. Results: Twelve VNTR loci were identified by bioinformatic screening of H. pylori genomes, and five of them were highly polymorphic. Therefore, an MLVA assay composed of five VNTR loci was developed with greatest discriminatory power. Conclusion: MLVA typing is a faster and more standardized method for studying the genetic relatedness of H. pylori isolates. At preliminary stage it is sufficient to use only 3 VNTR loci for the differentiation of H. pylori strains.

Objective of the study was to investigate biological properties and genetic characteristics of tularemia agent strains isolated from natural foci of the Rostov Region in 2020.Materials and methods. Field material from natural foci of the Rostov Region was examined by serological, bacteriological, biological, and molecular-genetic methods. Cultural-morphological, biochemical, antigenic and pathogenic properties of isolated cultures were studied. Protein profles were obtained through MALDI-TOF MS using mass spectrometer Autoflex speed III Bruker Daltonics and Flex Control of Biotyper software. The genetic characteristics of the strains were determined by VNTR and INDEL typing and SNP analysis.Results and discussion. Six strains of tularemia pathogen were isolated from mouse-like rodents using biological method. The investigation of their biological features and data of PCR analysis and INDEL typing with canonical markers showed that all strains are typical representatives of the Francisella tularensis subsp. holarctica biovar EryR. VNTR typing by six genetic loci revealed that all strains belong to four individual genotypes. The strain isolated in 2020 in the Salsky district was identical to the strain which was isolated in the same area in 1989. Based on the whole genome sequencing of two strains, we established that they are closest to the cultures isolated in Turkey (2009, 2012) and Khanty-Mansiysk (2013) by the studied set of SNP markers. Thus, we found that both identical (or closely related) clones of the tularemia agent and new strains with unique genotypes which previously were not described for the Rostov Region can circulate in natural foci of this region for a long period of time.