Elena Arkadievna Menshikova

The aim of the work was to analyze the epidemiological situation on cholera in the world and in Russia and to make its forecast for 2026. Proactive epidemiological surveillance data showed that over the period of 2016–2025 there was an upward trend in cholera incidence worldwide: 5,474,013 cases were documented in 91 countries. The proportion of patients was distributed among countries as follows: Yemen (53.4 %), Afghanistan (11.7 %), Democratic Republic of the Congo (6.3 %), Nigeria (4.7 %), Somalia (3.2 %), South Sudan (2.5 %), Sudan (2.5 %), and Haiti (2.4 %). In 2025, 42 countries reported 589,571 cases and 8,140 deaths (5.1 % more than in 2024). The increase in cholera cases in 2025, compared to 2024, amounted to 5.12 %. Population migration remained the main risk-forming factor for the spread of cholera to different countries. In the Russian Federation, monitoring of labor migration was introduced with the organization of medical observation of arriving persons for seven days after arrival from cholera affected countries. As a result of monitoring surface water bodies in 2025, 50 non-toxigenic strains of Vibrio cholerae O1 circulating in the territory of the Russian Federation and belonging to different genetic lines were isolated. A decrease in the number of acute intestinal infections was noted from 13 (2024) to 5 (2025), caused by non-toxigenic V. cholerae nonO1/nonO139. Forecast for 2026: the risk of cholera importation persists due to the expected increase in the intensity of labor migration. Registration of acute intestinal infections and extra-intestinal infections caused by non-toxigenic strains of V. cholerae nonO1/nonO139 is probable. Detection of non-toxigenic strains of V. cholerae O1 and their clonal complexes circulating in water bodies will continue. Stable epidemiological situation on cholera in the Russian Federation is predicted for 2026.

The aim of the study was to perform a detailed molecular-genetic characterization of the toxigenic Vibrio cholerae nonO1/nonO139 strain isolated from the Don River in the Rostov Region in 2024. Materials and methods. Whole-genome sequencing was carried out on the MiSeq Illumina and Oxford Nanopore platforms. Primary assembly was performed de novo through the Canu software based on long reads, and subsequent alignment was carried out using short reads with the Pilon program. Vector NTI package, BLASTN, BLASTP, BioEdit, Ugene, abricate (CARD database) and SnapGene Viewer were used in the bioinformatics analysis of genomes. The search for the limits of wb * clusters and comparison of their genes were conducted by means of the author’s program “O-cluster”. The phylogenetic tree was constructed using the roary and FastTree software. Results and discussion. A toxigenic strain of NAG-vibrio has been identified for the first time in Russia. The sequences of both chromosomes of the studied strain have been determined via hybrid genome assembly. Based on the analysis of the O-antigen biosynthesis region, the strain has been classified into serogroup O105 and it shows a high degree of closeness to clinical strains of this serogroup previously isolated in China. Bioinformatic analysis has revealed the presence of tandems of two identical copies of CTX and RS1 prophages on the large chromosome, which are notably distinct from their prototypes. The ctxB gene is represented by the ctxB12 allele, and all RS1 genes, except for rstC , show no similarity to canonical sequences. A VPI-1 island with an alternative tcpA allele, a T3SS cluster within the VPI-3 island, and the cholix toxin gene have been identified, which constitutes a rare combination within a single strain’s genome. The set of intact determinants of other pathogenicity factors is sufficient for the realization of pathogenic potential. The presence of a number of antibiotic resistance determinants in the genome did not always correlate with the phenotypic manifestation of resistance/susceptibility to antimicrobial agents in vitro . The genome structure of the strain and its close relationship to clinical isolates indicate a potential capacity to cause acute intestinal infections. Since there is a possibility of new introductions of such strains into the Russian Federation, their detection in water bodies necessitates the implementation of a set of anti-epidemic measures.

BACKGROUND: The survival of various strains of Vibrio cholerae in water bodies contaminated with heavy metals, particularly cadmium, poses a threat to public health. Of particular interest is the study of the role of biofilms formed by V. cholerae on chitin in this process. АIM: To investigate the role of complex biofilms formed on chitin as a factor in the persistence of toxigenic and non-toxigenic V. cholerae under the toxic effects of cadmium. METHODS: The study used V. cholerae El Tor O1 toxigenic 5342 ctxA+ csh1- strain isolated from a human in 2023 and non-toxigenic 643 ctxA- csh1+ strain isolated from the Matsesta River (Sochi) in 2025, representing a "persistent" genotype repeatedly isolated from the rivers of Sochi. The formation of complex biofilms by two strains was carried out on chitin fragments, and the viability was determined by seeding the corresponding material on Martin's agar. The presence of molecular markers ctxA and csh1 was determined using multiplex real-time polymerase chain reaction (PCR). The effect of cadmium Cd on the formation stage and mature biofilm was studied by adding it to the incubation medium at the appropriate time. RESULTS: Without external influence, both strains were capable of forming a complex biofilm on chitin. The non-toxic strain 643 adhered and formed a biofilm in the presence of 0.01 mM and 0.05 mM cadmium, while the adhesion of the toxic strain was completely inhibited under these conditions. In the case of a mature complex biofilm, both strains retained their viability in the biofilm and supernatant even when exposed to known toxic concentrations of heavy metal (up to 0.6 mM). CONCLUSION: The obtained results indicate that the mature V. cholerae biofilm formed on a chitin substrate ensures the viability of the pathogen when exposed to Cd concentrations that are lethal for planktonic forms. The differences in the response of the toxigenic and non-toxigenic persistent strains of V. cholerae to the toxic effects of Cd during the adhesion stage may reflect the specific features of their ecological adaptation.

Introduction. Vibrio cholerae motility plays a key role in the pathogenesis of cholera, so finding ways to suppress it is a promising approach to combating this infection. Objective: to study the effect of Fluorothiazinone on the motility of V. cholerae. Materials and methods. Flagellar motility was determined in 0.3% agar, with the motility zone measured, in 40 strains V. cholerae El Tor of varying epidemiological significance. The ability of these strains to acquire resistance to Fluorothiazinone was also assessed through repeated subcultures on alkaline agar containing Fluorothiazinone (100 mg/L). Results. A significant decrease in the motility of all studied cultures was observed in the presence of Fluorothiazinone, without the development of resistance. Conclusion. The dose-dependent ability of Fluorothiazinone to reduce the motility of V. cholerae was demonstrated for the first time.

Studies of water bodies in India and Bangladesh have shown that removal of large particles from water reduces the incidence of cholera. Detection of aggregated communities of Vibrio cholerae (biofilms) provided evidence of their role in cholera epidemiology and stimulated investigation of biofilm forms. Summarizing available data in this field is particularly relevant because there are no unified methods for assessing biofilm formation and quantitatively determining the microorganisms comprising biofilms. The work aimed to evaluate methodological approaches to investigating biofilm formation by V. cholerae, focusing on their detection and reproducibility in laboratories of different levels. Models (in vitro, in vivo, and microcosm), as well as methods for biofilm detection, were analyzed. Scopus, Web of Science, MEDLINE, the Russian Science Citation Index, and other sources were used to prepare the review. Data were searched using the following keywords: Vibrio cholerae, биоплёнка (biofilm), методы (methods), микроскопия (microscopy), микрокосмы (microcosms), молекулярно-генетические (molecular genetic). The review included 63 original articles, 3 review articles, and 1 monograph. The review summarizes current concepts regarding the process of V. cholerae biofilm formation. Methods for modeling on abiotic and biotic substrates, including field conditions, approaches to biofilm detection, and the use of in vivo methods to assess infectivity of biofilm cultures, are described. Particular attention is paid to the role of molecular genetic methods. According to the published data, the most effective strategy for studying biofilm formation is an integrated approach combining microbiologic, microscopic, and molecular genetic methods. The review also addresses current challenges and promising research directions in this field. The analysis of current issues and research prospects in V. cholerae biofilms, including recent molecular genetic methods, demonstrates that investigation of biofilms formed by vibrios using a range of methods will allow deeper understanding of the general principles of bacterial coexistence in communities. This, in turn, will contribute to more effective strategies for combating infections, including cholera.

The growth of antibiotic resistance necessitates studying the processes of acquisition and loss of genetic elements responsible for resistance. The aim of the study was to investigate the effect of temperature, biofilm formation, and antibiotics on the efficiency of integrative conjugative element (ICE) transfer in Vibrio cholerae O1 El Tor strains. Material and methods. Conjugative transfer of the ICE element from V. cholerae O1 El Tor strains to Escherichia сoli QD 5003 Rifr and V. cholera O1 El Tor 5879 Nalr cells was carried out in plankton and in biofilms on plastic and chitin at 25–37°C. The presence of ICE was determined by the integrase gene (int). Transconjugants were tested for antibiotic sensitivity and for the presence of resistance genes to tetracyclines (tetR), fluoroquinolones (qnrVC1), trimethoprim (dfrA1), and chloramphenicol (floR). Conjugation was induced by subinhibitory concentrations of ciprofloxacin, doxycycline, trimethoprim/sulfamethoxazole, and streptomycin. Results. Conjugation efficiency was higher in biofilms than in plankton, and lover with decreasing temperature. Streptomycin and trimethoprim/sulfamethoxazole stimulated conjugation in chitinous biofilms. Doxycycline and ciprofloxacin increased conjugation frequency in plankton. Conclusion. Temperature and biofilm formation affect the transfer of antibiotic resistance genes in V. cholerae. In complex biofilm conditions, compared to the planktonic form, there is an increase in the efficiency of conjugation between V. cholerae and other representatives of the Enterobacteriaceae family, which is more pronounced on the biotic substrate (chitin) and at 37°C. Subinhibitory concentrations of antibiotics can both stimulate and suppress the conjugation process in biofilms. It is necessary to solve environmental problems associated with environmental pollution by plastic waste and antibiotics, and to observe dosages when prescribing etiotropic therapy, as well as to search for substances that suppress the transfer of antibiotic resistance genes or promote the elimination of existing mobile genetic elements responsible for antibiotic resistance.

Introduction. The evolutionary association of Vibrio cholerae with chitin provided resistance to stress and protection from predators. The most important mechanism that provided V. cholerae with the effectiveness of association with chitin is biofilm formation. The ability to form a biofilm in V. cholerae depends on the presence of a factor, toxin-corrected adhesion pili (TCP), which are synthesized by the tcp A-F genes. One of the key methods for studying biofilms is microscopy. It allows one to visualize the structural elements and study various parameters of biofilms and the effects of various factors on them. Aim. To determine the epidemiological significance of the biofilm-forming ability of toxigenic strains by their morphological characteristics on chitin-containing substrates. Study of structural differences in biofilms of Vibrio cholerae tcpA+– and tcpA– strains on chitin-containing substrates. Results. It has been shown that Vibrio cholerae tcpA+– and tcpA– strains are able to form biofilms on the surface of chitin-containing substrates. The intensity of biofilm formation is more pronounced in tcpA+ strains, because V. cholerae ctxA+ tcpA+ cells in the biofilm are predominantly singly located and the surface of the chitinous exoskeleton with which they are in contact is intact, V. cholerae ctxA– tcpA– cells form chains in the biofilm, which indicates division processes, and scattered chitin of the endocuticle indicates activity of metabolic processes. Conclusion. The strains of V. cholerae used in the work, regardless of the presence or absence of the ctx and tcp genes, form bioplecs on a chitin substrate. The indicator of biofilm formation in terms of the thickness of the biofilm matrix is higher in V. cholerae ctxA+ tcpA+ , in terms of the degree of degradation of the chitin substrate it is higher in V. cholerae ctxA– tcpA– .

Introduction. Evaluation of antibiotic sensitivity of biofilms formed by Vibrio cholerae can help in the search for effective drugs to combat cholera. The aim of the work is to evaluate the effectiveness of antibacterial drugs against V. cholerae cells being a part of a polymicrobial biofilm formed on various substrates. Materials and methods. Mono- and polymicrobial biofilms were simulated in vitro on chitin and plastic in flasks containing tap autoclaved water contaminated with a suspension of 104 microbial cells of three strains of V. cholerae O1 El Tor separately or in combination with Escherichia coli QD 5003. Results. When being a part of mono- or polymicrobial bacterial communities formed by the studied strains on chitin or plastic, the V. cholerae and E. coli cells were less susceptible to the action of antibacterial drugs. Conclusion. The increased antibiotic resistance of V. cholerae biofilms formed on biotic and abiotic surfaces highlights the danger of their spread and preservation in the environment, creates additional problems regarding the use of antibiotics and requires the development of alternative strategies to reduce resistance.

BACKGROUND: In the summer-autumn period, the peak of phytoplankton reproduction and flowering occurs and results in the deterioration of the quality of river water is noted. At the same time, cases of gastrointestinal diseases in humans are recorded, and epidemic complications of cholera may occur in endemic areas. V. cholerae form biofilm on the surface of chitinous hydrobionts and plastic components can lead to the spread of V. cholerae, possibly explaining the autochthonous mechanism of their existence in water bodies. AIM: This study determines the duration of Vibrio cholerae preservation on biotic (chitin) and abiotic (plastic) substrates in the presence of green unicellular algae with a change in cultivation temperature under experimental conditions. MATERIALS AND METHODS: This study used bacteriological and molecular genetic methods to achieve its goal. RESULTS: The preservation of toxigenic and non-toxigenic strains of V. cholerae O1 El Tor and V. cholerae O139 (ctxАВ+tcpА+csh1 and ctxАtcpАcsh1+) for six months, including three months at a low temperature simulating the autumn-winter period, as part of biofilms in microcosms, where one component is green microalgae. We noted an excess of the concentration of V. cholerae O1 and O139 serogroups by two orders of magnitude in samples where one of the components is chitin and the reproduction of green microalgae in the presence of a chitin substrate. This is probably one stage in the food chain in the ecology of water bodies and, accordingly, can be a reservoir for biofilm forms of cholera vibrios. Preservation of V. cholerae O1 El Tor non-toxigenic strain in a viable state in biofilm samples on plastic at decreased temperatures 82C for three months is possibly due to the presence of a cold shock gene in its genome. CONCLUSION: Without a substrate for adhesion, colonization, and biofilm formation, vibrios are incapable of long-term persistence at low temperatures.