Over the three waves of the 7th cholera pandemic, the genome of the pathogen has undergone significant changes, as a result of which, in the last decade, new genetic variants with a high pathogenic and epidemic potential have emerged. The purpose of this study is to investigate the genomic diversity of the cholera agent and the prevalence of new genetic variants in cholera-endemic regions and Russia, and to identify their phylogenetic relations. Materials and methods. To search for new variants of the pathogen, bioinformatic analysis of the whole genome nucleotide sequences of 124 V. cholerae El Tor strains obtained by us earlier and taken from the NCBI Gen Bank and European Nucleotide Archive database was carried out. Phylogenetic relations have been established for 115 different strains isolated in Asia, Africa, Russia and Ukraine during three waves of the pandemic, based on SNP analysis. Results and discussion. Analysis of the genomes of 91 strains isolated in seven endemic countries between 2007 and 2019 showed that, by the set of mutant virulence and epidemicity genes localized in mobile genetic elements (CTXφ prophage, pathogenicity island VPI-1 and pandemicity island VSP-II) and the core region, 74.7% of them are classified as new genetic variants of the pathogen. When analyzing the genomes of toxigenic strains from Russia and Ukraine (1970—2014), five groups were identified that differ in mutations in key genes associated with virulence, epidemic potential, and drug resistance. Among them, imported from endemic regions in recent years (2010—2014), 80.0% are classified as new variants by the spectrum of mutant genes. Conclusion. Analysis of phylogenetic relations based on SNP typing of 115 strains showed the phylogenetic similarity of new variants from Russia and Ukraine with those circulating in Africa and Asia over the past decade. The results obtained are of interest for understanding the molecular mechanisms and dynamics of changes in the most important genetic properties of this pathogen, and can also be used to develop methods for gene diagnostics of new variants of the pathogen.

The aim of the work was to study the prevalence of different types of SXT element with different composition of antibiotic resistance genes among clinical strains of the El Tor cholera pathogen isolated in Russia, Ukraine and cholera-endemic countries in Asia and Africa.Materials and methods. The subject of the study was 27 strains and nucleotide sequences of 77 strains of Vibrio cholerae El Tor available from the NCBI GenBank. The structure of the SXT element and its type were determined using the Mauve and BLAST v.2.9.0 programs. Phylogenetic relations of strains with different types of SXT were identified using Snippy v.4.6.0 and MrBayes v.3.2.7 software. Assessment of strain sensitivity to antibiotics was carried out in accordance with Methodological Regulations 4.2.2495-09.Results and discussion. Two types of SXT element (ICEVchInd5 and ICEVchBan9) have been identified among the studied strains from Russia and Ukraine, which have different composition of antibiotic resistance genes: floR, strAB, sul2, dfrA1 and floR, tetAR, strAB, sul2, dfrA1, respectively. At the same time, the studied strains from Asia and Africa contain five types of SXT: ICEVchInd5, ICEVchBan9, ICEVchBan5, SXTTET, ICEVchInd5ΔVRIII, which differ in size and/or composition of resistance genes. Of these, the last three have not been found in Russia and Ukraine. Due to the high level of genomic diversity of SXT in the population of V. cholerae in endemic regions, there is a risk of importation of cholera pathogen strains with altered resistance to antibiotics into Russia. Phylogenetic relations of 76 strains with different SXT types and different alleles of the ctxB gene encoding the B subunit of cholera toxin have been assessed based on SNP analysis. A close phylogenetic relation between strains with the same type of SXT isolated in Russia and Asian countries has been demonstrated, which confirms the importation of the causative agent of cholera with multiple resistance to antibiotics from this region and the need for constant monitoring of the sensitivity of V. cholerae to antimicrobial drugs.

Objective of the study was to identify the differences in the production of proteins in typical strain and genetically altered variant of V. cholerae O1, biovar El Tor.Materials and methods. Natural strains M1062 (Astrakhan, 1970) and M1509 (Moscow, 2010) were used as model strains in this work. Strains were cultivated on Luria Bertani agar at 37 °C. Electrophoresis was performed in accordance with W.K. Laemmli technique (1970), mass-spectrometric profiling – the method described by A. Shevchenko et al.Results and discussion. Mass-spectrometric scanning of cell lysates of the examined strains showed significant similarity of their proteomes (615 common proteins). The identified differences pertained to high expression of proteins in the strain M1062, participating in biosynthesis of DNA/RNA, included into “purines, pyrimidines, nucleosides” group, as well as regulatory proteins. In M1509 strain, biosynthesis of the proteins responsible for pathogenesis, adaptation under the influence of unfavorable environmental factors, included into “co-factors, vitamins, pigments” group, involved in lipid, carbohydrate, and protein metabolism, cellular processes, as well as proteins-transporters was increased. It has been suggested that the wide dissemination of El Tor genovariants is probably due to enhanced pathogenic and adaptive properties and also to the considerable transformation of cell metabolism.

Introduction. The presence and structure of antiphage systems that contribute to the resistance of cholera vibrios to lytic phages in non-toxigenic strains of Vibrio cholerae O1 biovar El Tor isolated in the Russian Federation and neighboring countries has not been studied. The aim of the study is the detection and analysis of antiphage systems of non-toxigenic strains of V. cholerae O1 biovar El Tor. Materials and methods. The study involved 126 non-toxigenic (ctxAB–tcpA+ and ctxAB–tcpA–) strains of V. cholerae O1 El Tor isolated from 1972 to 2018. DNA sequencing was performed on the MGI DNBSEQ-G50 platform. For bioinformatics analysis, the following programs were used: fastp v. 0.23, unicycler v. 0.4.7, Blast 2.16.0, MEGA X, CRISPRCasty-per and CRISPRCasFinder. Results. Phage-inducible islands of the PLE, BREX and DISARM systems were not detected in the genome of the studied strains. It was found that 80% of ctxAB–tcpA+ strains contain the type I restriction-modification system, while this system was not detected in ctxAB–tcpA– isolates. The genes of the CBASS system were detected in single strains of both groups. In the genome of 35 (32%) studied ctxAB–tcpA– strains isolated in different regions of the Russian Federation and neighboring countries, the presence of the CRISPR–Cas system of class 1 types I (subtypes I-E, I-F, I-C) and III (subtype III-B) was established. The number of spacers in this system varied from 0 to 80 and their sequence was homologous to the protospacer regions of DNA of lytic and temperate phages, transposons, plasmids of V. cholerae, representatives of the genus Vibrio and unrelated bacteria. The presence in a number of strains of spacers homologous to the genetic material of the phage circulating in endemic territories may indicate the imported nature of these strains. Conclusion. The heterogeneity of the studied non-toxigenic strains of V. cholerae O1 El Tor in the presence of antiphage systems was revealed, which expands the information on their genetic organization. In their genome, restriction-modification systems of type I (ctxAB–tcpA+), CBASS (ctxAB–tcpA+ and ctxAB–tcpA–) and CRISPR–Cas class 1 types I (subtypes I-E, I-F, I-C) and III (ctxAB–tcpA–) were identified. The detection of several types and subtypes of the CRISPR–Cas system in the genome of a number of ctxAB–tcpA strains may indicate its repeated acquisition through horizontal transfer.

The isolation of locally adapted rhizobial strains with high symbiotic activity represents an effective strategy for increasing soybean yield under extreme environmental conditions. In this study, seven novel strains were isolated from nodules of soybeans grown in a greenhouse using field soil from the Lower Volga region. Five genomes were assembled into complete circular chromosomes, whereas two strains yielded near-complete chromosomes containing single repeat-mediated junctions. All strains had putative plasmids that were independently validated as circular by long-read mapping and confirmed by the presence of characteristic replication and conjugation-associated genes. Genome sequences of strains were about 11 Mb, and GC contents were 63.1–63.3%. Comparative genome analyses demonstrated that all strains had average nucleotide identity values of 95.4% with Bradyrhizobium japonicum USDA 6^T and 96.3% with Bradyrhizobium barranii 144S4^T, forming a distinct cluster in phylogenetic trees. No significant differences were detected between B. japonicum and B. barranii that would explain the species boundary. Therefore, it is proposed to unite all novel strains into the subspecies Bradyrhizobium japonicum subsp. saratovii subsp. nov., and all other strains of B. japonicum and B. barranii we suggest dividing into four subspecies: Bradyrhizobium japonicum subsp. japonicum subsp. nov., Bradyrhizobium japonicum subsp. barranii comb. nov., Bradyrhizobium japonicum subsp. apii comb. nov., and Bradyrhizobium japonicum subsp. saratovii subsp. nov. The proposed taxonomic framework expands current knowledge of the biodiversity of soybean symbiotic bacteria and contributes to a better understanding of the distribution and the evolution of bacteria Bradyrhizobium spp. in previously unexplored regions.