[Characteristics of Vibrio cholerae nonO1/nonO139 serogroup strains that caused diseases in population of Rostov region].

AIM: Genotype characteristic and determination of serological properties of Vibrio cholerae nonO1/nonO139 strains that caused diseases in population of Rostov region from 2000 to 2009. MATERIALS AND METHODS: 15 clinical strains of V. cholerae nonO1/nonO139 were studied. Serotyping was performed by using a kit of monospecific typing sera against serogroup 02-084 cholera vibrios obtained from Rostov Research Institute for Plague Control, PCR and VNTR-genotyping--by using specific primers described in scientific publications and constructed by us. RESULTS: Serologic features of strains are very diverse and strains contain various combination of pathogenicity factor genes that seem to be interchangeable. Similar pattern was observed for VNTR-genotyping. Distribution of the examined strains by VNTR-genotyping did not correlate with either PCR-genotyping data or serotyping, or place and time of isolation. CONCLUSION: The data obtained indicates a lack of general source of human infection even in the same location and time period. On the other hand, serological and genotypic features of V. cholerae nonO1/nonO139 may undergo changes in the process of staying in the macro organism or environment due to high plasticity of their genome.

[New nutrient medium for the cultivation and isolation of the plague microbe ChDS-37 as an element of the mobilization reserve of specialized antiepidemic teams of the Russian Inspectorate for the Protection of Consumer Rights and Human Welfare].

A new nutrient medium has been designed to culture and isolate the plague microbe ChDS-37 on the basis of the pancreatic digest of baker's yeast. The results of laboratory tests of the designed medium, by using 10 plague microbe strains and those of approval during the tactical and special training of a specialized antiepidemic team (SAET), suggest that the medium has some advantage over reference media and creates prerequisites for being incorporated into the mobilization reserve of a SAET.

[Study of expression of contact-dependent secretion systems in Vibrio cholerae on the model of Dictyostelium discoideum].

AIM: To detect T3SS and T6SS genes in Vibrio cholerae genomes and assessment of resistance of strains with different genotype characteristics to ingestion by Dictyostelium discoideum amoeba. MATERIALS AND METHODS: One hundred thirty-five toxigenic and non-toxigenic strains of Echolerae O1, O139, and non-O1/non-O139 were studied by PCR and on Dictyostelium discoideum model. RESULTS: T3SS cluster of genes was detected in several non-toxigenic representatives of 01 and non-O1/non-O139 serogroups. All toxigenic vibrios except one strain belonging to non-O1/ non-O139 serogroup lack this cluster. T6SS genes were presented in all studied strains although far from allofthem contained sequence coding ACD-domain of vgrG1 gene product--key effector of T6SS. Susceptibility of vibrios to ingestion by amoeba did not depend from presence of T3SS genes. Only strains with genetic determinants ACD-vgrG1 together with pathogenicity locus VPI were resistant to ingestion, all others were susceptible. CONCLUSION: D. discoideum model is adequate for study of expression of T6SS but not T3SS in V. cholerae. The latter is rather needed for colonization of intestine than for antagonistic activity against protozoa in external environment. Expression of T6SS is characteristic for many non-toxigenic strains of V. cholerae, whereas use of contact-dependent secretion is unlikely necessary for majority of toxigenic strains.

[Comparison of efficacy of tests for differentiation of typical and atypical strains of Yersinia pestis and Yersinia pseudotuberculosis].

AIM: To characterize species specificity of officially recommended tests for differentiation of Yersiniapestis and Yersinia pseudotuberculosis and propose additional tests allowing for more accurate identification. MATERIALS AND METHODS: Natural, laboratory and typical strains oftwo Yersinia species were studied using microbiological, molecular and biochemical methods. For PCR species-specific primers complementary to certain fragments of chromosomal DNA of each species as well as to several plasmid genes of Y. pestis were used. RESULTS: It was shown that such attributes of Y. pestis as form of colonies, fermentation ofrhamnose, melibiose and urea, susceptibility to diagnostic phages, nutritional requirements could be lost in pestis bacterial species or detected in pseudotuberculosis species. Such attribute as mobility as well as positive result of CoA-reaction on fraction V antigen are more reliable. CONCLUSION: Guaranteed differentiation of typical and changed according to differential tests strains is provided only by PCR-analysis with primers vlml2for/ISrev216 and JS respectively, which are homologous to certain chromosome fragments of one of two Yersinia species.

[Approaches to species-specific typing of double mixed cultures comprising pseudotuberculosis bacteria and atypical plague bacillus strains].

The species relevance of atypical Yersinia strains was determined by various microbiological, immunological, and genetic (including polymerase chain reaction) tests. These strains were shown to represent mixed cultures of Y. pseudotuberculosis serovariant O1b and Y. pestis var antiqua. Identification-resistant cells with atypical properties and plasmid segregation were found in the populations of Y. pestis strains. Analysis of different diagnostic tests revealed the most reliable ones selected for the identification of atypical Y. pestis strains with unstable genome.

[Primary characterization of the properties and diagnostic value of the antigenic complex "fraction V" in a plague microbe].

The experimentally obtained antigenic complex isolated by extraction in the gradient surfactants from live and acetone-dried bacteria of the capsule-free vaccine strain EV76 of a plague microbe that had lost its ability to synthesize the diagnostic species-specific capsular antigen F1 was investigated. The antigenic complex fraction V (FV) was obtained after the fifth stage of extraction at a concentration of 1.28% of surfactants and after additional purification. The thermostable FV was found to consist mainly of protein. The protein having a molecular mass of about 43 kD predominates in the fraction. The latter is nontoxic for albino mice and antigenic. It forms a precipitate with commercial antiplague serum antibodies. FV antigenic sensitization of tanned sheep red blood cells gave rise to a diagnostic agent that specifically reacted with an antiplague serum rather than with heterologous sera against enterobacteria. The sera immunized with FV specifically reacted in the JDJFR with all the strains of the pathogen of plague irrespective of the temperature of their cultivation, including "fraction-free", which did not interact with a diagnosticum on F1. The animal sera immunized with capsule-free plague microbial strain reacted only with a FV-erythrocytic diagnosticum and they did not interact with F1 antigen-sensitized red blood cells. The erythrocytic FV diagnosticum was tested in ABNR with 130 typical and atypical plague microbial strains and with 133 strains of heterologous bacteria of different species of the family Enterobacteriaceae. The FV diagnosticum identified all the variants of a plague microbe, while the F1 diagnosticum revealed only its capsular variants. Among the heterologous bacteria, some strains of the closely related pathogen of pseudotuberculosis in those who were in the R form, rather than S form, positively reacted. The use of FV identified 2 groups of hybridomas obtained after immunization of albino mice with the capsule-free variant of a plague microbe. Some hybridomas reacted only with plague bacteria while others did with two above pathogens. The authors substantiate the expediency of using FV, its components, and obtained monoclonal plague pathogen antibodies to improve antiplague diagnosticums with an activity spectrum that exceeds that of the existing commercial F1 antigen-based diagnosticums. They also discuss the lines of further studies.

[Detoxifying effect of antibiotics and their effectiveness in experimental plague at the stage of explicit intoxication]. / Detoksitsiruiushchee deistvie antibiotikov i ikh éffektivnost' pri éksperimental'noi chume na stadii vyrazhennoi intoksikatsii.

The effect of antibiotics such as amikacin, rifampicin, doxycycline, polymyxin B and cefotaxime on the toxins of the plague microbe (lipopolysaccharide + fraction II according to Beiker) was studied in vitro and in vivo. The study on the antibiotic neutralization of plague toxins revealed that only polymyxin had toxin neutralizing capacity which depended on the dose. Investigation of the polymyxin effect at various stages of plague infection showed that when polymyxin in a dose of 1250 units and a mixture of plague toxins in lethal doses were administered simultaneously to albino mice, the positive effect amounted to 100 per cent. When the antibiotic was administered 30 or 60 minutes later, the antibiotic efficacy proved to be lower by 90 or 76.6 per cent, respectively. The intoxication in later periods (in 90-120 minutes) resulted in a decrease in animal survival up to 40-15 per cent. It was demonstrated on the model of the plague infection in albino mice that the use of amikacin, cefotaxime, rifampicin or doxycycline during polymyxin therapy at the stage of marked generalization of the infection provided a significant increase in the animal survival (60 to 80 per cent) as compared to that after the use of the same drugs alone (0 to 20 per cent).