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The aim of the work was to study the composition of vesicle preparations (OMV) in Vibrio cholerae strains O1 El Tor (ctxAB– tcpA– ; ctxAB– tcpA+ ) and O139 (ctxAB– tcpA– ) isolated from various sources. Materials and methods. V. cholerae strains O1 and O139 serogroups were used in the work, from which OMV preparations were obtained. The structure of OMV preparations was studied by transmission electron microscopy (TEM). Genome-wide DNA sequencing was performed on the MiSeq platform. The nucleotide and corresponding amino acid sequences of the genes were studied using bioinformatics analysis. Protein profiling was conducted using MALDI-TOF mass spectrometry. Enzymatic activities were detected using appropriate media and substrates. Determination of (surface determinants) lipopolysaccharide (LPS) and outer membrane proteins in the composition of OMV was carried out by enzyme immunoassay (ELISA). The analysis of the spectrum of fatty acids was performed by gas-liquid chromatography mass spectrometry. Results and discussion. OMV preparations have been obtained from non-toxigenic strains of cholera vibrios. TEM has revealed that OMVs are represented by spherical structures of about 40–200 nm. Genome-wide sequencing has shown that fragments of chromosomes containing genes of V. cholerae strains, O1 and O139 serogroups, from which they were isolated are present in the OMV. The bioinformatic analysis revealed differences in the structure of the ompT gene in the initial strains and in OMV obtained from V. cholerae strains O1 and O139 serogroups. The ΔompT gene of the OMV preparation from V. cholerae O139 serogroup was trunked and formed a peptide of 17 amino acid residues (-LENHHQKNREPDKEFPY-). Analysis of the OMV composition has revealed the presence of enzymes (N-acetyl-β-D-glucosaminidase and DNAazes), proteins of outer membranes (ompT and OmpU), LPS, saturated, сis/trans isomers of unsaturated fatty acids and branched fatty acids. Differences in the composition of OMV obtained from non-toxigenic strains of V. cholerae O1 and O139 serogroups have been identified.

Cholera vibrios, isolated from surface water bodies during monitoring activities, can differ from typical ones in terms of agglutinability with diagnostic cholera sera, which makes it difficult to assign them to a specific serogroup. Therefore, the determination of the causes for these deviations through the study of the structure of genetic determinants responsible for the synthesis of O‑antigen (wb* clusters) is considered a relevant task. The aim of this work was to identify wb* clusters in the genomes of R-variant Vibrio cholerae, study their structure, and conduct phylogenetic analysis of the strains. Materials and methods. Full genome sequencing was performed using Illumina MiSeq and MinION platforms. The assembly was conducted de novo using the SPAdes assembler software (v.3.11.1). Manipulation of cluster sequences and data visualization were carried out using the BLAST program from the ncbi-blast-suite package version 2.13.0, Python scripts and packages such as pyGenomeViz and Biopython. The phylogenetic tree was constructed using the roary program (v.3.13.0). Full genome alignment was carried out using the nucmer and promer programs from the MUMmer 4 package version 4.0. Results and discussion. R-variant V. cholerae contains different types of wb* clusters in the genome, with most commonly occurring O23 and O59. A comparison has been made between translated amino acid sequences of wb* regions in R-variant strains and amino acid sequences of reference strains present in the NCBI database. Some of the analyzed clusters completely match with reference strains, while others show high variability. Phylogenetically, the strains primarily group together according to the wb* cluster type, regardless of the S/R phenotype. Classifying R-variant V. cholerae strains based on agglutinability is challenging. Molecular research methods, polymerase chain reaction, in particular, are necessary to determine the pathogenic potential of such strains.

The aim of the work was to determine the pheno- and genotypic features of the aquatic CTX+ strain of Vibrio cholerae isolated in 2023 and a comparative bioinformatic analysis of whole-genome sequencing data.Materials and methods. Whole-genome sequencing was performed on MiSeq (Illumina) and MinIon Oxford Nanopore Technologies (ONT) platforms; hybrid assembly of the whole genome was carried out using the Trycycler algorithm; assembly errors were eliminated by means of the Medaka algorithm and the Pilon program. Dendrogram construction and bioinformatics analysis were carried out with the help of the scipy and Graphviz packages, BioEdit, BLASTN, BLASTP, CARD, ICE Genotyper, and Vector NTI programs. The ability to produce cholera toxin was tested using the GM1ELISA.Results and discussion. The isolated strain was identified as V. cholerae O1 Ogawa, sensitive to most antibiotics. Based on the totality of genetic properties, it was classified as the first genovariant, distinct from the typical El Tor strains only by the presence of ctxB1 gene of classical type instead of ctxB3 of the El Tor type. It has been established that it contains a tandemly duplicated CTX prophage on the small chromosome and a tandem of two copies of RS1 prophage on the large chromosome. Thereat, the rstR gene of the CTX prophage belonged to the classical type, and the RS1 prophage – to the El Tor type. The remaining criteria of epidemic hazard – tcpAelt, rtxA1 and intact VSP-II did not differ from the prototypes. The genome of the strain carries the ICE element VchBan11, which contains the trimethoprim resistance gene dfrA1, and phenotypically the strain is resistant to this antibiotic. Under in vitro conditions, the strain did not produce cholera toxin, as shown by ELISA results. This may be due to the presence of a deletion within the toxR regulatory gene. Strains similar to the 2023 isolate are mainly attributed to the second wave of the seventh pandemic. Currently, they are almost replaced by new genovariants, but occasionally can emerge and even cause diseases. Therefore, their importation into Russian territory potentially pose a threat to public health.

The paper presents an analysis and assessment of risks of complication of epidemiological situation regarding cholera in the Russian Federation and a forecast for 2024. It briefly characterizes the dynamics of the epidemic process around the world during the last decade (2013–2022). The forecast of cholera importation into the Russian Federation, due to activity of the epidemic process in endemic countries, has been confirmed. In 2023, two cases of cholera importation from India were reported. It was established that the toxigenic strains isolated from patients belonged to the currently dominant in the world “post-Haitian” group, including in India and Bangladesh, with the genotype ctxB7 tcpACIRS rtxA4a VSP-IIΔ0495-0512. Furthermore, non-toxigenic Vibrio cholerae O1 strains were isolated from samples of biological material from persons arriving from India, containing preCTX prophage and distinct from the strain (ctxAB– tcpAEТ–) isolated from a patient with acute intestinal infection (Zaporozhye Region, Melitopol). Nontoxigenic NAG vibrios were isolated from 13 patients with acute intestinal infections, which is 1.9 times more than in 2022. Regardless of importations, a strain of V. cholerae O1 (ctxB1+tcpAEТ) of the “pre-Haitian” group was isolated from a water sample of a surface reservoir. 52 non-toxigenic V. сholerae O1 strains were found on the territory of 11 constituent entities of the Russian Federation, which is 18.9 % more than in 2022. In 2024, epidemiological risks of cholera importation from endemic countries to any administrative territory of the Russian Federation remain. The risks of epidemic manifestations of cholera are especially high in the territories of the constituent entities of the Russian Federation which are under ongoing emergency, due to migration processes, and the impact of other risk-forming factors that contribute to an increase in the degree of contact of the population with water bodies contaminated with cholera vibrios. If epidemic foci of cholera are to emerge in a neighboring country, there will be a high probability of spread of this infection in the stated constituent entities of the Russian Federation and in other territories of southern Russia. In the absence of epidemic manifestations of cholera, the possibility of occasional detection of toxigenic O1 serogroup strains in water samples due to unidentified introductions from cholera-affected areas cannot be ruled out.

The aim of the work was to identify genotypic features of clonal complexes formed by CTX–VPI+ strains of Vibrio cholerae isolated from water bodies of the Rostov Region in different years, based on bioinformatics analysis of their whole genome sequences (WGSs).Materials and methods. Whole genome sequencing was performed on the MiSeq (Illumina) platform; identification of genetic determinants in WGSs and bioinformatics analysis – by means of BioEdit, BLASTN, BLASTP, CARD, Vector NTI software packages.Results and discussion. The strains were almost identical in each group of 2002-, 2005-, 2007-, 2020-, 2021-, and 2022-isolates and were found at different points of water sampling over a longer or shorter periods of time (from 3 weeks to 2 months). Bioinformatics analysis of WGSs of representative strains divided them into 4 clusters, which can be considered as separate clonal complexes sharing a number of characteristic genetic markers: 1) 2002-isolates containing one SNP in regulatory rpoS gene; 2) 2007-strains carrying tandemly duplicated preCTX prophage; 3) strains isolated in 2020 in Rostov-on-Don and in 2005, during epidemiological complications in the Kamensk district of the Rostov Region, which had a “hybrid” pathogenicity island including the cluster of type 3 secretion system and the nan-nag region of the VPI-2 island, and 9 SNPs in rpoS gene; 4) 2021- and 2022-strains and a single 2018-isolate containing a 9 bp deletion in vchC collagenase gene. Despite the revealed differences, representatives of all clonal complexes possessed sufficient sets of intact determinants of pathogenicity/persistence factors to realize virulent properties and ability to survive in water bodies. Detection of clonal complexes of potentially pathogenic strains in surface water bodies indicates occasional emergence of favorable conditions for their long-term circulation, which emphasizes the importance of constant monitoring of V. cholerae in the territory of Russia.

Determination of Vibrio cholerae affiliation to one or another serogroup may meet some difficulties in cases of atypical agglutination with diagnostic cholera sera. The study of genetic determinants that allows for identifying a serogroup is a relevant task in monitoring of surface water body contamination with cholera vibrios.The aim of the work was to compare the structural organization (quantitative and qualitative gene composition) of LPS clusters in V. cholerae rough variants.Materials and methods. We used Illumina MiSeq for the whole genome sequencing; SPAdes software (version 3.11.1) for de novo assembly; and blastn (v. 2.5.0) for gene searching. GeneMarkS software was deployed for annotation of the genes incorporated in the clusters; nucmer – for searching homologous sites. Visualization of O-LPS clusters was carried out by means of SnapGene Viewer.Results and discussion. Strains of V. cholerae rough variants had diverse gene clusters responsible for O-antigen biosynthesis. We have identified three types of O-LPS clusters with different size and number of genes. Unique DNA sites, common to the whole group of V. cholerae rough variants, have not been detected. Two genes present in all rough strains have been defined, but they are not unique for this group of strains and can be found in representatives of other serogroups. For two types of clusters, a region containing the IS‑element, common with V. cholerae O1, has been revealed.

The aim of the work was to study the structure of the alkyl sulfatase (asu) gene in Vibrio cholerae strains of various serogroups, as well as to compare nucleotide and amino acid sequences of alkyl sulfatases using various methods of bioinformatic analysis.Materials and methods. 483 strains of V. cholerae O1, O139 and nonO1/nonO139 serogroups were employed in the work. The search for the gene, its recurrence, and localization was carried out applying the Blast software. The nucleotide and corresponding amino acid sequences of the gene, as well as its structure, were studied using bioinformatic analysis. Sequencing was performed on the MiSeq (Illumina) platform. The enzymatic activity was detected using a medium, confirming the presence/absence of the gene by PCR in vitro and in silico.Results and discussion. Bioinformatic analysis of the nucleotide and corresponding amino acid sequences of the asu gene has been carried out and its structure investigated. Four functional domains have been identified. In the beta-lactamase domain, a conservative amino acid sequence -HAHADH- has been found in all strains of cholera vibrios, which is part of the Zn2+ binding motif. It has been established that the alkyl sulfatase of cholera vibrios belongs to the family of Zn2+-dependent β-lactamases. Blast analysis has revealed the similarity of nucleotide and amino acid sequences of alkyl sulfatases in representatives of V. cholerae O1 and O139 serogroups (ctxAB+tcpA+) and representatives of the genera Aeromonas and Pseudomonas, which is in the line with the data of 3D modeling of the amino acid sequence structures of the alkyl sulfatase enzyme in these microorganisms. The bioinformatic analysis of nucleotide and amino acid sequences of alkyl sulfatases in cholera vibrios has showed the conservativeness of these sequences in toxigenic strains and the presence of a number of single mutations in the asu gene in atoxigenic ones. The presence or absence of the asu gene has been established by PCR in vitro and in silico and confirmed by the results obtained using the Blast program. It is demonstrated that the presence/absence of the asu gene correlates with the ability/inability of O139 strains to hydrolyze SDS on the medium. These results can be used in studying mechanisms of cholera vibrios adaptation, persistence and pathogenicity.

The tactics of activities of the specialized anti-epidemic team (SAET) of the Rospotrebnadzor on the territory of the Republic of Crimea has been determined by the conditions of emergency situation of sanitary-epidemiological and natural character. The aim of the work was to analyze the features and effectiveness of the functional response of the Rospotrebnadzor SAET to the dynamics of epidemiological risks caused by emergency of combined nature in the Republic of Crimea in 2020–2021. Materials and methods. We used report materials of the Rospotrebnadzor SAET; information materials of the Interregional Rospotrebnadzor Administration in the Republic of Crimea and the Federal city of Sevastopol; “Coronavirus Monitor” service data. Results and discussion. Tactical decisions on the work of the SAET on the territory of the Republic of Crimea had the stage-by-stage nature. In the event of a complication of the epidemiological situation on the new coronavirus infection, a mobile group of the SAET was involved. Together with specialists from the Plague Control Station of the Republic of Crimea, it conducted 211 148 studies of clinical material to detect SARS-CoV-2 RNA using PCR during the period of 2020–2021. In response to the emergence of additional risks caused by the consequences of a dangerous meteorological phenomenon, the group of SAET specialists was reinforced and mobile complex (MС) of the SAET was deployed. During the entire course of work, a prolonged target rotation of personnel was implemented in order to provide for adequate laboratory support for monitoring upon epidemiological indications in accordance with the nature of emerging risks. That tactics of operation made it possible to conduct 18 591 screening studies of clinical material for acute respiratory viral infections (ARVI), natural-focal and acute intestinal infections (AII), as well as the study of 280 water samples for cholera. Introduction of the sequence of operational procedures adapted for work in emergency situations resulted in isolation of 10 Vibrio cholerae O1 ctxA– strains from water bodies. The phased involvement of the mobile group of SAET specialists, and later the MC of the SAET, in combination with the consistent operational target rotation of personnel in order to provide adequate laboratory support for monitoring studies, determined the tactical features of the SAET functioning. Prompt investigation, substantiation of potential and response to realized (actual) epidemiological risks associated with the extraordinary situation made it possible to prevent complications of the epidemiological situation in the territory of the tourist cluster of the Russian Federation.

The aim of the review was to analyze and assess the epidemiological situation on cholera in the world and Russia in 2013–2022 and to make a forecast for 2023. Over the period of 2013–2022, 500 administrative territories affected by cholera were identified in various regions of 71 countries of Asia, Africa and America (the Caribbean region) with formation of 69 endemic foci in 16, 41 and 12 countries, respectively. In 2022, 1 209 301 cases of cholera were registered in 36 countries of the world. The intensity of epidemic process in Asian countries (Syria, Lebanon) increased. Unfavorable epidemiological situation on the African continent persisted. Epidemics and large outbreaks, which began in 2021–2022 due to emergency situations (ES) of social and natural character, continued. The prediction of the stability of the epidemiological situation on cholera in the territories of constituent entities of the Russian Federation (RF), given for 2022, has been confirmed. In 2022, 43 non-toxigenic strains of Vibrio cholerae О1 were isolated from surface water bodies, 8 V. cholerae nonO1/nonO139 strains – from humans. Similarity of those strains with genetically closely related ones isolated in the course of monitoring in previous years in Russia, Donetsk People’s Republic (DPR) and Zaporozhye Region was demonstrated. In 2023, the risks of importation of the infection into RF are retained. It is associated with the intensification of epidemic processes in Asian, African and Caribbean region countries. The extended border with Ukraine, to where importation of cholera from endemic countries is possible, contributes to increased degree of threat. Bioterrorism is also not excluded. The consequences of a possible cholera epidemic complication in DPR, Lugansk People’s Republic, Zaporozhye and Kherson Regions are exacerbated by social emergencies resulting in disruption of infrastructure, interruptions in water supply, etc. In the absence of implementation of the risks of importation of this infection on the territory of RF, a stable epidemiological situation on cholera will be retained. Detection of non-toxigenic V. cholerae O1 strains (including probability of clonal complexes formation), as well as strains of non-O1/non-O139 serogroups, which can be etiological factors of sporadic cases or outbreaks of diarrheal diseases, in surface water bodies is predicted.

The ability of Vibrio cholerae to form biofilms may cause ineffectiveness of cholera treatment and prevention. The aim of the study was to determine the antibiotic sensitivity of V.cholerae in biofilms. Material and methods. Sensitivity to 11 antibacterial agents was determined for biofilms of 10 strains of V.cholerae of different serogroups isolated from humans and from water obtained for 48 hours at 37°C on plastic plates in vials with autoclaved tap water infected with a suspension of 104 V.cholerae microbial cells. For this purpose the plates were washed three times in physiological solution and transferred into penicillin vials with double dilutions of antibacterial agents in liquid nutrient medium (Martin's broth, pH 7.7). After 24 hours of cultivation in the thermostat (37°C), the biofilms were imprinted and 0.1 ml of plankton culture was seeded onto plates with Marten's agar (pH 7.7). After 24 hours in the thermostat (37°C), the biofilms were imprinted and 0.1 ml of plankton culture was sown on plates with Marten's agar (pH 7.7). The result was counted after 24 h, determining the minimum suppressive concentrations of preparations by the presence or absence of V.cholerae growth. Results. The strains studied in biofilm communities, in contrast to the planktonic form, were highly resistant to all antibacterial drugs taken in the study, except for ciprofloxacin and ceftazidime. To improve the effectiveness of treatment of cholera and other infections whose causative agents are cholera vibrio, it is necessary to determine the antibiotic sensitivity of V.cholerae biofilms.

During more than 50 years, a heterogeneous population of Vibrio cholerae non-O1/non-O139 (NAGs) exists in Rostov region of Russia, whose representatives periodically cause acute human intestinal infections; the last case was registered in 2018. In 2022, a sick child was again revealed in Rostov, from whom 2 NAG subcultures were isolated. Aim of the study: bioinformatics analysis of whole genome sequences (WGSs) of the 2022 clinical NAG isolates, genetic determinants of pathogenicity factors and relevant products. Materials and methods. Isolation, identification of cultures, as well as antibiotic sensitivity were carried out according to the MUK 4.2.3745-22. SNP analysis of WGSs obtained on the MiSeq Illumina platform were analyzed by means of designed software; BioEdit 7.2.5, Vector NTI Advance 11 package, CARD database were used to identify and analyze genes and relevant deduced products. Results. The isolated cultures with identical phenotypic features, PCR-genotypes and results of SNP analysis, were identified as a non-toxigenic NAG. The agent showed sensitivity to chloramphenicol, ceftriaxone, streptomycin, gentamicin, nalidixic acid, ciprofloxacin, ampicillin, amikacin and intermediate resistance to tetracycline, doxycycline, furazolidon and co-trimoxazole. WGSs lacked CTX and preCTX prophages, pathogenicity island VPI, thermostable and cholix toxin genes, pandemic islands VSP-I and VSP-II. The determinants of the type 6 secretion system (T6SS) were not presented completely, which evidenced in favor of the loss of its functionality. An intact T3SS cluster was revealed, which was adjacent to the nan-nag region of the VPI-2 pathogenicity island in the absence of the proximal and distal parts of the latter. Other determinants of pathogenicity factors were revealed: RTX cluster responsible for synthesis of MARTX cytotoxin, genes for hemolysin HlyA, cytotonic toxin Cef, several serine and metalloproteases. The nucleotide compositions of most determinants differed from the prototypes, but their putative products preserved characteristic active domains. The adhesive activity might be provided by mannose-insensitive MSHA-like pili, since the msh-cluster included all the genes necessary for their production, as well as flagella, as both clusters responsible for their assembling were 99% identical to the prototype. From antibiotic resistance determinants only -lactamase VarG gene was found. Conclusion. The NAG strain examined here contains a sufficient set of intact virulence-associated genetic determinants, expression of which could contribute to the development of the disease.

The aim of the study was to retrospectively analyze the range of variability of antigenic properties and genotypic characteristics of Vibrio cholerae R-variant strains atypical in terms of agglutinability.Materials and methods. 169 strains of V. cholerae R-variant with atypical agglutinability have been studied using the “AmpliSens® Vibrio cholerae-FL” test-system. The determination of O1 antigen was carried out using the “Ig-V. cholerae О1/О139 – ELISA/dot-ELISA” reagent kit.Results and discussion. A retrospective analysis of the complex of phenoand genotypic characteristics of strains isolated from surface water bodies in the territories of three former Soviet republics and 13 constituent entities of the Russian Federation in the course of 30-year monitoring and identified upon isolation as nontoxigenic V. cholerae R-variant strains has been performed. Upon re-identification, it was found that the strains belong to both epidemically dangerous (3.0 %) and non-dangerous strains (97.0 %). The range of variability was expressed in their distribution into three groups and consisted in retaining of agglutinability only with cholera RO serum in the first group (34.5 % of strains); the loss of this trait, but the acquisition of the ability to agglutinate in different combinations with O1, Ogawa or Inaba sera – in the second (16.7 %); and also in the loss of agglutinability with all diagnostic cholera sera – in the third (48.8 %). The presence of the wbeT gene in the compared V. cholerae classical R-variant strain does not exclude the presence of the genomic region for O1 antigen biosynthesis in other R-strains, possibly in a modified form, which can be clarified in further molecular-genetic studies. Alternatively, such strains are likely to be attributed to V. cholerae nonO1/nonO139. Strains of V. cholerae R-variant with different amounts of surface antigen (optical density range – from 0.088±0.002 to 1.226±0.003) have been identified. The data obtained can be used for monitoring of cholera in laboratories of regional and federal levels.

The aim of the work was to summarize the results of cholera monitoring in 2021, to assess current trends in the development of the epidemic process, and to predict the epidemiological situation in the Russian Federation for 2022. It was established that within the period of 2012–2021, 4117264 cases of cholera with the spread of infection across 83 countries on all continents were registered in the world and there was a downward trend in the incidence in Asia and Africa. The dynamics of monthly morbidity in 2021 was associated with emergencies as factors of epidemiological risk. Epidemics and outbreaks of cholera were documented against the background of COVID-19 pandemic and laid a double burden on healthcare systems. At the same time, based on the overview of the results of cholera monitoring in the constituent entities of the Russian Federation, it was shown that the forecast of epidemic well-being given for 2021 was fully justified. It has been determined that the increase in the number of non-toxigenic strains of Vibrio cholerae O1 (67) isolated from water bodies compared to 2020 (25) is mainly due to the appurtenance of a number of isolates to clonal complexes. The study of phylogenetic relation has demonstrated that the detection of strains with genotypes which were previously identified in the isolates evidences the persistence potential. The identification of strains with new genotypes, which were earlier established in the strains circulating in other territories, pointed at the possibility of the occasional importations. The forecast of the epidemiological situation on cholera in Russia for 2022 is associated with the continuous existence of risks of introduction. If these epidemiological risks are not realized, a favorable epidemiological situation is predicted regarding this infection in the country. It is expected that the detection of epidemiologically insignificant strains of V. cholerae O1 in environmental water bodies, along with their clones and/or clonal complexes, will remain, including strains that may be an etiological factor in sporadic cases or outbreaks of disease.

The aim of the work was to summarize the experience of the Rostov-on-Don Research Anti-Plague Institute in the organizational and functional response aimed at ensuring the implementation of a complex of on-going and scheduled anti-epidemic (preventive) measures at different territorial levels in the context of COVID-19 pandemic. Stage-wise reassignment of scientific and operational subdivisions of the facility was carried out in a timely manner in a flexible mode for a prompt response to the challenges that arose from community transmission of the new coronavirus infection in the territory of the Russian Federation. It allowed an effective distribution of the forces and assets without damage for specialized functioning in the main areas of scientific and practical activities. Realized was an effective interdepartmental interaction with territorial scientific and practical institutions of the Rospotrebnadzor and specialized medical organizations. Practical, methodological and advisory assistance was provided to personnel of specialized medical institutions within the framework of conversing in-patient facilities into hospitals for the treatment of patients with COVID-19. Priority research to study the level of the humoral and cellular immune response to SARS-CoV-2 among the population of the Rostov Region, the etiological spectrum of the causative agents of community-acquired pneumonia associated with COVID-19, full genome sequencing of SARS-CoV-2 isolates was carried out. Specialists of the task forces reinforced human resources of individual laboratory facilities in Moscow city, the Republic of Crimea and the Rostov Region. Complex objective of effective functioning has been implemented: in an operational mode – participation in anti-epidemic (preventive) measures in the face of changes in the dynamics and intensity of the epidemic manifestations of new coronavirus infection; in scheduled mode – the implementation of activities in the main areas of the scientific work of the organization.

The aim of the work was to study surface antigenic determinants of V. cholerae R-variant strains using enzyme immunoassay and a panel of monoclonal antibodies (MAbs).Materials and methods. 60 strains of V. cholerae R-variant isolated from ambient environment objects in the territories of the former USSR and the constituent entities of the Russian Federation over a 30-year period (1988–2019) were investigated in the slide agglutination reaction with cholera diagnostic sera, enzyme immunoassay (ELISA) using the panel of MAbs specific to membrane proteins and a set of reagents “Monoclonal diagnostic immunoglobulins labeled with horseradish peroxidase, dry, for serological identification of V. cholerae O1 and O139 (in vitro) through ELISA and dot-ELISA”.Results and discussion. The analysis of the surface structures of V. cholerae R-variant strains with atypical agglutinability has been carried out applying enzyme immunoassay. It showed that individual strains with different amounts of O-antigen are registered among the studied strains identified at isolation as V. cholerae R-variant (the optical density range is from 0.261±0.002 to 1.312±0.003). Epitopes of specific O-antigen were found in some “conservative” strains (30 %) that are agglutinated only with RO serum, and in several strains (20 %) that do not have the wbeT gene that determines its synthesis, and lost agglutinability with all diagnostic cholera sera, including RO. The protein epitopes recognized by complementary MAbs are represented with varying frequency in the composition of surface antigens of R-vibrios; a decrease in their representation or absence on the cell surface correlates with the modification or loss of R-LPS and is accompanied by a negative agglutination reaction.

Purpose. A dynamic comparative analysis of the results of identification of Vibrio cholerae (V. cholera) isolates from water and human specimens isolated during the monitoring of cholera throughout the Russian Federation in 2013–2019.Materials and methods. Total 385 strains of V. cholerae O1 serogroup (including one toxigenic V. cholerae O1 El Tor Inaba — Rostov region, 2014) and the R-variant from environment objects were identified in the territory of 21 regions of the Russian Federation.Results. It was found that 91% of the strains were isolated in 7 regions of the Russian Federation: Republic of Kalmykia, Krasnodar Krai, Transbaikal Krai, Khabarovsk Krai, Primorsky Krai, Rostov region and Irkutsk region. 2.3% of strains were atypical in their agglutinability, and 75.3% of strains had atypical phagosensitivity. Phagotypes were identified for 13.2% of strains isolated in 11 administrative territories. Phageotype 11 included 48.8% of the strains isolated mainly in the Transbaikal Krai. Non-toxigenic strains of V. cholerae nonО1/nonO139 (n = 46) were isolated from 43 patients with acute intestinal infections and otitis in 8 regions of Russia. PCRtyping of 377 non-toxigenic V. cholerae strains demonstrated that they belong to 13 clusters and 71 genotypes. Strains with unique genotypes (probable cases of importation) and with the same genotypes repeatedly isolated in different years in one or several territories of the Russian Federation were identified, indicating the prevalence of strains in environment throughout the country.Conclusion. The results allowed us to characterize the epidemiological situation of cholera in Russia (in terms of the environment contamination with V. cholerae O1) as unstable, but not significantly changed over the past 7-year period. These data are important for substantiating the forecast for further development of the epidemiological situation.

Aim of the work – to assess the epidemiological situation on cholera in 2020 and to make a forecast for 2021 based on the monitoring data and analysis of morbidity around the world for the period of 2011–2020. During the period between 2011 and 2020, 4 413 988 cases of cholera were recorded in 97 countries of the world with a general trend towards a decrease in the incidence (coefficient of accuracy of approximation R2 – 0.5705). However, due to the continuing epidemic manifestations of cholera in the endemic countries of Asia, Africa and America, the epidemiological situation on cholera on these continents was characterized as unfavorable in 2020. The emergence of a new “post-Haitian” lineage was observed among epidemically hazardous strains of Vibrio cholerae O1. In 2020, no epidemically dangerous strains of V. cholerae O1, O139 were isolated from humans on the territory of the Russian Federation. 25 non-toxigenic V. cholerae O1 El Tor strains were isolated from environmental objects, eight out of which (ctxA-, tcpA+), according to PCR-INDEL typing and SNP analysis of sequences, belonged to the clonal complex. The results of the analysis of biological properties and phylogenetic relations between the isolated non-toxigenic strains provided the basis for considering the epidemiological situation on cholera in Russia in 2020 as a stable one and a similar forecast of its development in 2021. At the same time, the possibility of importation of this infection from endemic countries cannot be ruled out, as well as the need to carry out a complex of differentiated anti-epidemic (preventive) measures within the framework of the state sanitary-epidemiological surveillance in order to localize and eliminate the imported focus and avoid the spread of the infection.

The goal is to study the features of the biological properties of Vibrio cholerae strains found in environmental objects (EO) on the territory of Rostov-on-Don from 1989 to 2018.Materials and methods. Bacteriological, molecular biological and statistical methods were used to study 73 strains of cholera vibrios O1, O139 and 1702 strains of nonO1/nonO139 serogroups isolated over a 30-year period.Results and discussion. It was found that in Rostovon-Don, 78.0 % of toxigenic strains were isolated from all isolated from environmental protection in Russia, among which 2 were genetically unchanged. The V. cholerae O1 classical strain and 72 V. cholerae El Tor strains were found, of which 70 % belonged to the Ogawa serovar, and 3 % of the nontoxigenic strains were susceptible to the classical phage. A “very weak direct relationship” was statistically revealed between the isolation of V. cholerae O1, O139 and V. cholerae nonO1/nonO139 strains, in which the dominant serogroups (O67, O76, O75, O53, O16) were identified, and 18 % of the strains showed atypical agglutinability. Revealing the biological characteristics of V. cholerae strains contributes to the optimization of monitoring studies.

Aim. Analysis of the phenotypic characteristics and identification of peculiarities of the genotypic organization in non-toxigenic strains of cholera vibrios having different origin, isolated in Russia. Materials and methods. A sample of 548 non-toxigenic strains obtained using the author’s updated GIS “Cholera 1989–2014” was used. PCR genotyping was carried out in accordance with the patented “Method for the identification of non-toxigenic strains of cholera vibrio O1 serogroup using PCR to isolate genetic determinants.” Cluster analysis was performed applying the UPGMA method. The dendrogram was constructed using MEGA 5 software package.Results and discussion. Representative cultural-morphological, serological and biochemical properties of V. cholerae strains have been specified. The variability of the studied strains on the basis of phagolizability has been revealed. Unique phage-types not previously encountered in Russia have been identified. The population of non-toxigenic strains of cholera vibrio O139 serogroup is genetically homogeneous in contrast to V. cholerae O1 El Tor isolates and has identical PCR genotypes. The universality of the PCR genotyping by 14 target genes has been shown to differentiate the studied strains of V. cholerae O1 and O139, as well as to identify disparities among O139 strains isolated in different geographical regions of the country.

Objective of the study was to monitor the spread of cholera in the world, in the CIS countries and in Russia with an assessment of risks and emergencies that contribute to the activation of the epidemic process. Despite the downward trend in the global incidence rate of cholera during the period between 2010 and 2019, epidemics and major outbreaks occurred in 96 countries. WHO has reported 2013 imported cases of cholera to countries in Asia, the Americas, including the Caribbean, Europe, and Australia with Oceania; cholera-endemic administrative territories are identified in 24 countries. According to WHO, cholera burden reduction in Asia and Africa is associated with large-scale vaccination. During epidemiological surveillance of cholera, 705 strains of V. cholerae O1 and O139 serogroups were isolated from surface reservoirs in 26 constituent entities of the Russian Federation, including 10 strains of V. cholerae O1 ctxA+tcpA+, 35 strains of V. cholerae O1 ctxA–tcpA+, 655 strains of V. cholerae O1 ctxA–tcpA– , and five strains of V. cholerae O139 ctxA–tcpA–. Identification of strains with unique, previously unknown INDEL genotypes testifies to their imported nature. The forecast for cholera in the world for 2020, given the proven high degree of epidemic process activation at the expense of social and environmental risks caused by emergencies of different origin, the presence of endemic foci, infection import and other risk factors is unfavorable. For Russia, the forecast for cholera will be determined by the presence of external risks created by the ongoing 7-th pandemic, possible importation of infection to constituent entities of the Russian Federation that differ by the types of epidemic manifestations.

Analysis of cholera incidence for the period of 2009–2018 was performed. The upward tendency in the morbidity rate dynamics around the world (compared to 2009) with an average annual growth rate of 5,352 % was revealed. For the first time during the 7th pandemic caused by V. cholerae O1 El Tor, WHO reported 1227391 cases of cholera world-wide in 2017, out of which 1032481 (84.1%) were registered in Yemen, where the war continues and one of the largest epidemics in the world. There have been cross-border epidemiological complications in several African countries. Endemic foci continue to exist and spread in Asia, Africa and the Caribbean. Under the epidemiological surveillance in Russia, 744 strains of V. cholerae El Tor – ctxA– tcpA–, ctxA– tcpA+ and V. cholerae О139 ctxA– and tcpA– were isolated from the surface water bodies, as well as single strains of El Tor ctxA+ tcpA+. As a result of INDEL- and PCR-genotyping, the isolation of strains with identical genotypes and new ones was established. To make the prognosis for 2019, the risk of activation (continuation) of the cholera epidemic process in the world was assessed, taking into account emergencies of different origin and risk factors. The cholera forecast at the global level and in Russia for 2019 is unfavorable.

The trend of morbidity reduction at an average annual rate of 1.113 % in 2017 (in comparison to 2008) continues. The tendency of intensive monthly cholera incidence growth around the world in 2017 in relation to 2015 and 2016 is established. 25 countries with 70 endemic areas and 11734 imported cases of cholera onto different continents with predominance of interstate carrying of infection in Asia were identified. In Russia in 2008, 2010, 2012 and 2014 cholera importations from India have been registered. V. cholerae O1 biovar El Tor ctxA–tcpA–, ctxA–tcpA+, V. cholerae O139 ctxA– and tcpA– and singular V. cholerae O1 biovar El tor ctxA+tcpA+ strains were isolated from surface water bodies. INDEL typing of 242 V. cholerae O1 strains, isolated from environmental objects (2014–2017), provided the data which confirmed their belonging to common cluster of V. cholerae O1 ctxA–tcpA+. Along with genetic diversity of V. cholerae O1 ctxA–tcpA–, several strains of the same or closely related genotypes in some regions were isolated every year. For the first time ever the algorithm for risk of epidemic process activation assessment in case of cholera in Russia, taking into account the various factors (conditions), their gradation and expert scores, has been developed for the prediction. The forecast for cholera at the global level and in Russia for 2018 is unfavorable.

Objective of the work was to compare the results of monitoring of Vibrio cholerae strains isolated in the territories of federal districts, their constituent entities and from certain water bodies between 2006–2016. Based on the methodology, using updated geoinformation system “Cholera 1989–2014”, a comparative study of the dynamics of isolation and biological properties of 586 Vibrio cholerae O1, O139 strains and R-variants, collected from the surface water bodies in the administrative territories of Russia, was carried out. Eventually it is established that Vibrio cholerae strains of various serogroups are found in the territory of all federal districts, but not in all the entities included in them. It is shown that the greatest number of isolated strains was registered in the Southern Federal District. The overwhelming number of Vibrio cholerae strains, isolated in the country, were non-toxigenic. It was noted that against the background of epidemiological welfare, the detection of single epidemiologically significant strains continued (Rostov Region).ctxA–tcpA+ strains were found in the Southern, Far Eastern and North-Western Federal Districts. It is established that until 2013 in five constituent entities strains of Vibrio cholerae O1 and O139 in water bodies were not detected. Non-toxigenic strains of Vibrio cholerae O1 El Tor Ogawa serovar prevailed in the territories of the Southern and Ural, and Inaba serovar – in the remaining Federal Districts.The most frequently isolated were non-toxigenic Vibrio cholerae O1 strains with phagotype 15, while isolates with phagotypes 4, 5, 10, 14 and 17 were traced only in the Southern Federal District. The data obtained formed the basis of the approach that contributes to a comprehensive assessment of the epidemiological situation on cholera in the territory of Russia and highlighted the prospects for using GIS to improve the effectiveness of cholera vibrio monitoring in surface water bodies.

Objective of the study was to develop the methodology for identification of administratively independent areas under potential risk of “external” epidemiological threat realization, by the example of cholera.Materials and methods. Analysis was conducted using free software package with open source code (QGIS 2.8 and GRASS GIS 7.0) on the basis of the data received from Rosgranitsa and the Federal State Statistics Service. The construction of risk cartogram was performed on the base of Euclidean distance and estimation of nuclear density.Results and conclusions. In accordance with the obtained results, the GIS containing information about the checkpoints on the Russian border, settlements, roads and railway lines was worked out. The method of identification of risk areas due to importation of infectious diseases based on the cholera model has been developed, and the total area of such territories was less than 1 % of the total area of the country. It was found that in some cases the risk area is located at a certain distance from the checkpoint, but the existence of checkpoint does not lead to the formation of risk areas. The developed GIS is available on the geo-information portal of FGHI Rostov-on-Don Research Anti-Plague Institute of Rospotrebnadzor.

Analysis of epidemiological situation on cholera around the world has demonstrated downward trend in morbidity rates in 2016; annual average decline being 3.907 % (by reference to 2007). Comparative assessment of monthly incidence in 2015 and 2016 shows perennial registration of cholera cases in a number of African countries and in the Caribbean with up-rises in different seasons. Establishment of epidemic foci is on-going. Between 2007 and 2016, 1733 imported cases of cholera took place across the globe. Interstate import dominated in Asia. For forecast development, for the first time ever, risk assessment of epidemic process activation has been performed, taking into consideration emergency situations of varying origin, social and natural environments, using expert evaluation scores. Epidemic manifestations of cholera in the Russian Federation were characterized by import of the infection in 2008, 2010, 2012, and 2014; isolation of V. cholerae O1 biovar El Tor ctxA– tcpA–, ctxA+ and V. cholerae O139 ctxA– and tcpA– from surface water bodies, as well as individual V. cholerae O1 biovar El Tor сtхA+ tсра+. Worked out has been the algorithm for epidemiological investigation of causes and conditions of surface water body contamination with cholera vibrios. Cholera forecast worldwide and in Russia for the year 2017 remains unfavorable.

Aim. Application of the authors’ GIS «Cholera 1989-2014» for systematization of atoxigenic strains of serogroup 01 cholera vibrios (ctxAB-tcpA-, ctxAB-tcpA+), isolated from aquatic objects of the environment by pheno- and genotype. Materials and methods. A sample of 304 Vibrio cholerae 01 strains was studied. Isolation of 39 genes related to pathogenicity was carried out. Discrimination ability of a set of genes was determined by Simpson formula. Cluster analysis was carried out by UPGMA method. Results. Analysis of multi-year data on aquatic V. cholerae 01 strains in country’s subject was carried out using GIS. A possibility of systematization of phenotypes of the isolated strains by defined parameters was shown. An experimental program for detection of presence/lack of various genes and their combinations forgenotyping was developed. Conclusion. GIS was established to allow to carry out analysis of phenotypes by defined parameters, as well as implement approximate systematization of genotypes of atoxigenic strains of cholera vibrios 01 by optimally sufficient detection of 14 genes.

Objective of the study is the retrospective evaluation of epidemic cholera manifestations in the Republic of Dagestan in 1994, taking into account VNTR-typing and studies of genome peculiarities in the isolated V. cholerae O1, Biovar El Tor. Materials and methods. Utilized have been the data on the infection rates in different periods of the epidemic. The strains have been investigated using VNTR-analysis and whole genome sequencing. Results and conclusions. Heterogeneity of the agent population has been established on the basis of the circulating VNTR-genotypes. The whole-genome sequencing has revealed the presence of B-subunit allele ctxB1 in CTXφ prophage of El Tor V. cholerae O1 strains. The strains isolated in the early and the following stages of epidemic development have been found to be closely related to strains from India, Bangladesh, and Nepal. Some strains possessed a cluster of multiple antibiotic resistance genes, SXTET: floR, strA, strB, sul2 – of “Indian” type – SXT-ICE-Ind, as well as ICE mobile elements, containing determinants of resistance to antibiotics of tetracycline group – tetA. Phylogenetic relatedness of the Dagestan strains to the strains from Asia indicates to the origin of the strains, as well as to the independent cholera importations that took place at different stages of the epidemic. The role of genetically altered V. cholerae O1 El Tor variants with identified peculiarities on molecular-genetic level, as a system-forming factor in the emergence and development of epidemic in the Dagestan Republic, was directly linked to the social conditions.

In the course of assessment of epidemiological situation around the world in 2015, established has been the tendency to incidence rate reduction, with the annual average tempo - 10.225 % (in reference to 2006). For the first time ever, defined has been equivalent epidemiological year in case of cholera within an interval between 2014-2015.The key epidemiological risks of infection transmission globally are still inter-state (Africa, Asia) and extra-continental imports of cholera from the Caribbean (Cuba) and Asia into America (Canada) and Europe (Great Britain, Italy, Belgium, and Germany), which took place in 2015 too. Identified have been 21 endemic administrative territories in eight countries of Africa, America, the Caribbean, and Asia. Epidemiological complications are mainly caused by V. cholerae O1, biovar El Tor, producing cholera toxin of classical type, as well as by V. cholerae O1, classical biovar, and V. cholerae O139 serogroup. Epidemic manifestations of cholera in the Russian Federation are characterized by import of the infection by citizens returning from abroad; the isolation from surface water bodies of V. cholerae O1, biovar El Tor ctxA- tcpA-, ctxA-tcpA+ and V. cholerae O139 ctxA-and tcpA-, as well as of single V. cholerae O1, biovar El Tor ctxA+tcpA+. Forecast on cholera world-wide and in Russia for 2016 remains unfavorable.

Aim. Comparative study of antibiotics resistance and VNTR-typing of Vibrio cholerae non O1/ non O139 strains, isolated on the territory of Rostov region in 2014. Materials and methods. Antibioticogramms of strains were determined by serial dilution method in dense nutrient medium according to MG 4.2.2495-09 (2009). Pheno-, sero- and VNTR-typing was carried out by conventional methods. Results. The studied strains belonged to V. cholerae species, did not agglutinate with O1 and O139 sera, were atoxigenic, hemolysis-positive, did not contain genes of cholera toxin and toxin-coregulating pili ofadhesion, contained genes ofhemagglutinin/protease, protease PrtV, collagenase, cytotonic factor Cef, outer membrane protein OmpW, tol- and vps-clusters, regulatory genes toxR and hapR. Antibioticogramms of the strains have shown the presence of cultures, resistant to ampicillin, ceftazidime, furazolidone, trimethoprim/sulfamethoxazole with intermediate resistance to streptomycin, kanamycin, gentamycin, amikacin, netilmicin. Approximately 20% of isolates had multiple drug resistance. Data of VNTR- and geno-typing confirmed a possibility of water transmission route of the infection. Conclusion. Execution of monitoring of cultures from environmental samples is necessary for timely detection of genetic characteristics, antibiotics resistance.

Carried out has been whole-genome sequencing of the Vibrio cholerae 81 El Tor Inaba strain DNA, isolated from Temernik River waters in the territory of Rostov-on-Don in July, 2014. Identified is the presence of hybrid CTX prophage, which contains ctxB gene of the classical type ( ctxB1 allele), as well as rstR gene El Tor type, tcpA gene with the mutations in coding and promoter regions ( tcpETCIRS allele), the null-mutation in rtxA gene, and extended deletion inside the pandemicity island VSP-II. All the stated above peculiarities are characteristic of cholera vibrio genovariants, which have an enhanced epidemic potential and have recently replaced the typical strains of this biovar.

Cholera epidemiological situation around the world (2005-2014) has been assessed. Distribution of infection in the territory of African, Asian, and American countries, as well as in the Caribbean Region has been shown. Interstate and inter-continental importations of cholera in Europe, Australia, and America, including USA and Canada have taken place during this period. Epidemic process is spatially disseminated (with involvement and affection of new countries and administrative territories) and temporally chronic (America, the Caribbean Region; Africa) due to occurring of natural or social emergency situations. Alongside epidemics and outbreaks of the disease, caused by genetically altered variants of V. cholerae O1 El Tor and strains with multiple drug resistance, outbreaks with isolation of clinical strains of V. cholerae O139 serogroup take place in the Southeast Asia (China) on an annual basis. The forecast for 2015, as regards cholera in the world, stays unfavorable, which in its turn allows for the possibility to import this infection in the territory of the Russian Federation.

The Rostov-on-Don Anti-Plague Research Institute interacts with a number of health and scientific institutions of the Federal Service on Consumer Rights Protection and Human Welfare Supervision. The basis of this interaction is formed by the activity on execution of existing and development of new regulatory guidance documents; the work of the Reference Center for cholera monitoring; monitoring of the state of natural focal infections in the Rostov Region; the assessment of anti-epidemic preparedness of health care centres; professional training; guidance and practical assistance to comply with biological safety requirements; as well as the fulfillment of relevant scientific research themes.

V. cholerae O1 El Tor cultures, serovars Ogawa and Inaba, from surface waters of the reservoirs, which are different in epidemic manifestations of the disease.

Identified are the peculiarities of biological properties and origin of cholera vibrio O1 serogroup strains isolated from the Rostov-on-Don water bodies in the course of surveillance in 2008–2012. 41 atoxigenic V. cholerae O1 strains have been isolated from 767 water samples and investigated. Stable tendency of isolation of increased numbers of atoxigenic V. cholerae O1 strains over time has been demonstrated. In addition, in the strains under study detected have been the genes of additional pathogenicity factors, by PCR genotyping using specific primers for 45 nucleotide sequences associated with V. cholerae pathogenicity. The strains have also been classified according to 19 VNTR-genotypes grouped into 6 clusters based on VNTR-typing using exclusive copyright locus-specific primers. Discovered have been the strains with similar genotypes though isolated at different points both throughout the year and over the period of several years. These ones have probably been imported. They are characterized by a capacity to persist in ambient water bodies for a certain period of time.

Within the last decade increase in cholera morbidity rate in the world has been revealed (with due consideration to linear trend). For 2013 maintenance of the tendency is forecasted. Epidemics, outbreaks and importations of the infection have occurred in 109 countries around Asia, Africa, America, Europe, Australia and Oceania. Nosoarea has been expanding owing to inclusion of new administrative territories and countries into the epidemic process, some cases being registered for the first time during the seventh pandemic. Apart from the endemic territories in India and Bangladesh, formation of endemic foci in twelve African countries has been exposed, as well as continued dissemination of epidemic cholera, consecutive in time and spatially stepwise. These facts testify of the follow-up of the seventh pandemic. Moreover, V. cholerae O1 El Tor strains with classical ctx-B allele have been spreading worldwide, which coupled with factors mentioned above predetermines unfavorable cholera forecast on the global level.

Studied is the anti-cholera activity of 40 strains from different species of probiotic bacteria according to the main qualitative characteristics, which are determined on the basis of quantitative data. Established are the selection criteria for probiotic bacteria to be used for cholera treatment: antagonistic activity against V. cholerae – high or moderate, adhesion – only moderate, acid forming activity – high or moderate. The preventive and medical efficiency of 5 selected strains of lactobacilli and 1 strain of bifidobacterium is demonstrated on the experimental model of cholera. Underlined is the prospective use of probiotics as additional preparations of choice against cholera infection.

The prognosis for cholera on 2011 proved true: the tendency of morbidity growth remained in the last decade (2002-2011) worldwide. Imported cholera cases caused large outbreaks and epidemics in the Caribbean countries and Asia. Formation of endemic foci in a number of countries of Asia and Africa took place, where favorable conditions for epidemics development (environmental and social conditions, military conflicts, economical and political instability) existed. Wide spread of genetically altered V. cholerae eltor and V. cholerae O139 variants with epidemic and pandemic potential was observed. All the events mentioned above predetermine the unfavorable prognosis for cholera on a global level.

Studied were morphological, physiological and ultrastructural changes in the cell culture L-929 and in the intestine of suckling mice in the presence of Cef (CHO cell elongating factor) of Vibrio cholerae. Registered were cellular elongation, cytoplasm vacuolization, an increase in the number of lysosomes, retraction and nuclei deformation in the L-929 cell culture. Statistical significance test confirmed that in vivo Cef caused the accumulation of fluid in suckling mice. Electron microscopy of the epithelial cells of small intestine revealed vacuolization and myelin-like structures formation in cytoplasm and nuclei, swelling of mitochondria, clarification of their matrix and destruction of cristae. Cristae were filled with mucus. The involvement of mast cells caused the increasing of capillary permeability which enhanced dehydration. These changes confirmed Cef participation in manifestation of Vibrio cholerae pathogenic properties.

Carried out is approbation of the new nutrient media set for cholera diagnostics on the basis of bakers' yeast pancreatic digest within the frames of the special tactical training exercises for specialized anti-epidemic team (SAET). It is demonstrated that the developed nutrient media set compare favourably with its analogues, commonly used for practical procedures; and its constituent media have a potential to be included into the SAET mobilization reserve.

Previously developed selective differential medium for V. cholerae growth was modernized. The modernized medium called SDMV-M was shown to possess the required sensitivity and germination index. The growth of E. coli was entirely inhibited, that of P. vulgaris was inhibited considerably. The medium possessed good differentiating ability: orange V. cholerae colonies were clearly distinguished from concomitant microorganisms. In the laboratory trials SDMV-M demonstrated some advantages as regards V. cholerae isolation from contaminated faeces in comparison with the reference medium TCBS.

Demonstrated are the results of microbiological monitoring over the ship’s ballast water at the sea ports of the Rostov region. Analyzed for the presence of Vibrio cholerae are the samples of the ship’s ballast water collected within the period of May-September 2010-2012, a total of 179 samples collected from 127 ships. Among 30 V. cholerae non O1/ non O139 strains isolated in 2010-2011 only 13 have been identified serologically. Application of MALDI-TOF mass spectrometry in 2012 has enhanced informational capacity of the investigations. It has expanded the range and increased the frequency of bacteriological findings. The detection of vibrios implies possibility of V. cholerae importation into the aquatic area of Azov Sea.

Studied have been biological properties of cholera vibrios non-O1/non-O139 circulating in the Rostov-on-Don surface waters and drain sewage system within the period of 2009–2011. Overwhelming majority of the isolates have been classified as typical ones in terms of their phenotype. However 90 strains (21 %) out of total 280 have been identified as atypical ones against agglu­tina­bility in the slide-agglutination and full-scale agglutination reaction to diagnostic cholera sera. Put forward are recommendations on performance of the differentiation between V. cholerae non-O1/non-O139 and V. cholerae O1/O139. Specified are predominant sero­groups of the strains which turned to be unchanged within the stated period (O16, O53, O76, and O67). Revealed is the strain genotype variability and low probability of virulence manifestations. Demonstrated is the futility of the blind gene- and serological typing of aquatic strains.

Studied are Vibrio cholerae of different serogroups on the presence of cef (CHO cell elongating factor) gene and activity against tweens and tributyrin using HDS-agar, prepared on the basis of bakery yeast pancreatic digest. Determined is the fact that all cef-positive strains hydrolyze tweens 20, 40, 60, 80, 85 and all but toxigenic V.cholerae O139, hydrolyze tributyrin. In contrast, non-toxigenic cef-negative strains of O139 serogroup, are active only against the latter. Apparently, the tweenase activity of Vibrio cholerae is provided, partly, by Cef, and the ability to hydrolyze tributyrin is the result of combined activity of Cef and other ferments. Shown is the efficiency of HDS-agar application to determine these characteristics.