Cholera Vibrios of nonO1/nonO139 Serogroups in Etiology of Acute Intestinal Infections: Current Situation in Russia and Around the World

Cholera vibrios of nonO1/nonO139 serogroups (NAG vibrios) are well known as natural inhabitants of aquatic environments and as agents of acute intestinal infections (AII) with different degree of severity. On the background of the current vehement dissemination of the new highly virulent Vibrio cholerae El Tor strains, the moral and economic damage from NAG vibrios is not that evident, but still, not less real and rather significant. In spite of rare association with large outbreaks, NAG vibrios rank in the etiology of AII all over the world and pose a potential threat for the population health in our country. The paper reviews the present-day morbidity as regards NAG infections in various countries, including Russia; as well as the molecular premises of the agents’ pathogenicity and drug resistance. Analysis of the published data has revealed an utter heterogeneity of NAG populations, circulating in certain territories, by reference to the presence/absence of genetic determinants of pathogenicity, persistence, housekeeping and antibiotic resistance factors. Nevertheless, the joint gene-pool of the population includes rather a wide set of genes and can expand due to importations of strains, which differ from resident ones in the genotypes, from other regions. This brings about the danger of emergence of clones with higher pathogenic and, probably, even epidemic potential driven by gene exchange, and the consequences of their occurrence are unpredictable. Therefore, the NAG vibrios demand proper attention from investigators and sanitary-epidemiological institutions of the Russian Federation

Polimeric Immunoglobulin Diagnosticum for Detection of Cholera Toxin and Assessing the Level of Its Production by Vibrios

A marker of the epidemic significance of Vibrio cholerae is their toxigenicity. Therefore, much attention is currently paid to the creation of diagnostic preparations for the detection of cholera toxin and assessment of the level of its production. The volumetric immunosuspension agglomeration reaction, carried out with the help of latex diagnosticums, is an analogue of the indirect hemagglutination reaction, an affordable and technically simple method, since it does not require special equipment and can be used when conducting research in the field. The aim of the study was to design a polymeric immunoglobulin diagnosticum for determining cholera toxin and the level of its production by vibrio strains. Materials and methods. Cholera toxin was obtained from the producer strain Vibrio cholerae Classical 569 B. Rabbit serum to the toxin was obtained according to the method selected by the authors. A polymeric diagnostic immunoglobulin antitoxic drug was obtained through sensitizing immunoglobulins from cholera antitoxic rabbit serum on the surface of polyacrolein microspheres with a size of (1±0.1) μm. Results and discussion. The analytical sensitivity of the developed diagnostic preparation with control cholera toxin is 100 ng/ml. It detects cholera toxin in toxigenic strains of Vibrio cholerae in a titer of 1:16 – 1:512, gives negative results with non-toxigenic strains of V. cholerae O1, V. Cholerae nonO1/nonO139, with samples of heterologous cultures, LPS preparations, liquid nutrient medium used for the cultivation of V. cholerae. Thus, a polymeric immunoglobulin diagnosticum has been constructed to detect and quantify the production of cholera toxin by vibrio strains, and its analytical sensitivity and specificity have been established

Construction of Polymeric Antigenic Diagnosticum Based on <i>Vibrio cholera</i> О1 Lipopolysaccharide

Representatives of the genus Vibrio cholerae differ in the structure of lipopolysaccharide, in particular, its O-polysaccharide chains (O-antigen), which determines the serological specificity of vibrios. Currently, the water-phenolic method is used to obtain the lipopolysaccharide preparation. However, this technique relates to harsh chemical methods, leads to a change in original molecular organization of biopolymer, violating its structure and biological properties. Modern technologies in the development of diagnostic kits for the immunosuspension reaction of volume agglomeration allow for obtaining synthetic carriers with different reaction groups on the particle surface capable to bind antigens/antibodies. The aim of this study was to construct cholera antigenic polymeric diagnostic kit based on the lipopolysaccharide of Vibrio cholerae O1 serogroup. Materials and methods. The lipopolysaccharide was used as a sensitizer obtained through the author's modification of enzymatic purification from the cell membranes of Vibrio cholerae using ultrasonic disintegration. Results and discussion. The resulting sensitin contains small impurities of protein (1.5 %) and nucleic acids (0.1 %). Diagnosticum is characterized by high analytical sensitivity in agglomeration reaction with commercial and experimental rabbit serum to Vibrio cholerae O1 serogroup (1:640 - 1:5120) and analytical specificity (the diagnosticum does not interact with heterologous sera, with serums to pathogens of acute intestinal infections, as well as with sera from healthy donors). A polymeric antigenic cholera diagnosticum designed to detect antibodies to lipopolysaccharide of Vibrio cholerae in the blood serum of patients who were ill, suspected of the disease or vaccinated people has been constructed

Analysis of the Results of Cholera Vibrios Monitoring in Environmental Objects in the Administrative Territories of the Russian Federation Using GIS “CHOLERA 1989–2014”

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

Cholera Vibrios non-O1/non-O139 Isolated in the Process of Epidemiological Monitoring over Rostov-on-Don Water Basins and Drain Sewage System within the Period of 2009–2011

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

Assessment of the Variation Range of Agglutinability in <i>Vibrio cholerae</i> Strains Isolated in the Course of Monitoring Studies

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

Impact of Cultivating Environment on the Terms of Persistence and Certain Properties of Cholera Vibrios

Objective of the study is to investigate the impact of cultivating temperature and medium on the terms of persistence, ctx gene retention, and enzymatic activity of V. cholerae O1 with various toxigenicity.Materials and methods. Utilized were the strains of V. cholerae El Tor: P-5879, P-19613, and also the strain P-19787.Results and conclusions. In the process of studying cholera vibrios El Tor with different genetic characteristics it was determined that the longest terms of persistence (19 days) on mineral substrates at 5 ºC were observed for toxigenic strains, while for non-toxigenic ones it made less than 17 days. At the same time cholera vibrios can persist continuously and even reproduce on mineral substrates under the conditions of subnormal lowered temperatures (not less than 10 °C). Toxigenic strains of Vibrio cholerae, irrespectively of cultivating medium and temperature, retained ctx gene in their genome and maintained enzymatic activity throughout the experiment. Such long-term persistence of cholera vibrios at low temperatures on mineral substrates may be regarded as possibility of preservation of V. cholerae toxigenic strains in case of import by the infected persons or vibrio-carriers