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The article deals with the results of study targeted to develop polymer diagnostic preparation to identify epidemically significant serogroups Legionella pneumophilia. The preparation combines rate of record (1-5 min) of reaction of paragglutinining preparations with color visualization and demonstrative of reaction of volume agglomeration with polymer diagnosticums. The specially synthesized polymer microspheres were sensibilized with serums enriched with antibodies to lipopolysaccharide of corresponding serovar L. pneumophilia. The derived immunoglobulin diagnostic preparations detect agent of legionellesis in the reaction of slide-agglutination on glass during 1-5 min. The polymer diagnostic preparations provide positive reaction with culture of corresponding serovar and no reaction with other gomologic and geterologic agents of infectious diseases.

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A new immunobiological polymer drug has been designed for the serological identification of hepatitis C. The drug is able to reveal specific antibodies in the sera of patients with hepatitis C, meets the current requirements of diagnostic test systems, and shows a high sensitivity and specificity. It is based on polyacroleinic microspheres; the concentrated cell culture biomass of hepatitis C virus (HCV), which contains an adequate set of viral antigens, is used as sensitin. A new diagnosticum is proposed to be used during primary (screening) laboratory studies based on the serological detection of total antibodies to HCV antigens in the volume agglomeration test. The latter is both one of the alternative methods during serological studies and an additional procedure when a set of diagnostic techniques is used.

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Three diagnostic selective media used for the isolation of Legionella pneumophila were compared. These included BCYEAalpha (Oxoid, a reference medium), BCYEAalpha (Hi Media), and elective legionellosis medium (ELM) developed at the Rostov-on-Don Research Institute for Plague Control. The virulent L. pneumophila strain Philadelphia-1 (LD50 was 10(5) CFU for guinea-pigs) was used a test culture. The susceptibility of the media was determined, by culturing 10(-6) and 10(-7) dilutions of the suspension of the macerated Legionella-infected guinea-pig spleen, as well as the suspension of a culture of this strain (100 CFU) and 6 L. pneumophila cultures freshly isolated from water. The BCYEAalpha (Oxoid) and ELM media demonstrated the similar growth characteristics (chi2 < 0.7; p = 0.05) while the BCYEAalpha (Hi Media) medium showed a low susceptibility. The BCYEAalpha (Oxoid) and ELM media were first found to be successful in detecting Legionella in viable, but nonculturable state, induced by the following factors: 1) starvation in distilled water; 2) exposure to hydroquinone (oxidative shock, and 3) elevated temperature (56 degrees C). The BCYEAalpha (Oxoid) and EML media did not differ either in their ability to suppress extraneous microflora and to maintain stable initial pH under the conditions of incubation of culture plates, as well as in their Na+ concentration (15-19 mmol/l). However, the BCYEAalpha (Oxoid) medium exceeded the ELM one in the growth rate and diameter of Legionella colonies. Two L. pneumophila cultures were isolated in the BCYEAalpha (Oxoid) and EML media used in the field experiment studying 15 water samples from different hot water supply systems. Thus, the conclusion can be drawn that the EML medium is comparable with the reference BCYEAalpha (Oxoid) medium in its susceptibility and ability to detect Legionella in both vegetative and viable, but nonculturable states and is suitable for practical application.

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The paper presents the results of studying the biological properties of Legionella pneumophila strains isolated from environmental objects. Elective legionellosis medium (ELM) has been found to be suitable for the isolation of the causative agent from the starting material and to be as sensitive as CYE (Oxoid company) containing growth and selective additives. Polymerase chain reaction (PCR) with a home-produced commercial test system used to detect L. pneumophila DNA enables identification of the causative agent, including its species. Hyperimmune sera against L. pneumophila 1-7 serogroups used in slide-agglutination and agglutination, as well as a series of co-agglutinating diagnosticums for legionellosis 1-7 serogroups make it possible to identify even the serogroups of L. pneumophilla. Comparative analysis of the virulence of L. pneumophila cultures in vivo and in vitro allows recommendation that practical laboratories should employ a simple NaCl resistance test, which can be used as a guide virulence test.

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Technical approaches to construction of preparations for serologic diagnostics of Legionella infection were presented in the article; antigenic- and immunoglobulin-based diagnostic kits with known characteristics were developed. Immunogenic properties of protein and lypopolysaccharide antigens, which have diagnostic value, were studied; similarity of protein antigens from 7 serogroups of L. pneumophila was demonstrated. Soluble antigen with known composition was obtained and used for the development of antigen-based polymeric kit for diagnostics of Legionella infection. On the basis of hyperimmune sera, immunoglobulin-based polymeric diagnostic kit and array of coagglutinating diagnostic kits for the mentioned 7 serogroups were developed. Antigen-based polymeric diagnostic kit was recommended for licensure.

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Study showed that El-Tor strains of V. cholerae isolated from different sources produce lipase for hemolysis after cultivation during 24 h on meat-peptone broth independently from their toxigenic and hemolytic abilities. Study of 3- and 4-hours broth cultures of vibrios revealed possibility to differentiate between hemolytic nontoxigenic strains and toxigenic nonhemolytic ones. Using antilipaze diagnostic kit it was possible to differentiate El-Tor vibrios from vibrios of classic biovar basing on lipase production 24 h after cultivation on meat-peptone broth that was evident in El-Tor vibrios but not in classic biovar strains.

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The F1-specific components of Y. pestis capsular antigen, isolated by Baker's method, were shown to differ by their biological activity and the character of action on cell-mediated immunity factors, used in this study. Depending on the method of isolation, antigens could vary in the proportion of their components, which determined the specific features of the total preparations obtained in this investigation. Out of the four components under study having the same antigenic specificity, but different physico-chemical characteristics and genetically determined synthesis, only one component exhibited biological properties dominating in the secreted form of F1, more similar in their composition to protein Caf1. The other three components exhibited immunological activity on the level of unspecific protection in different ways, depending on the model, and influencing the outcome of the interaction "bacteria-macrophage". Lectin-like hemagglutinating and hemolyzing activity was shown by components not related to protein Caf1. The multi-component character of preparations F1 (Baker) and the individual activity of their components should be taken into consideration when using capsular antigen and different methods of its isolation for different aims.

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The possibility of using a heterogeneous, but structurally similar antigen--the commercial preparation of Pseudomonas sp. lipase (Sigma, USA)--for the development of polymer diagnosticum aimed at determination of lipase production in cholera vibrios was shown. The new diagnosticum (antilipase antibodies) on a polymer carrier was used in the serological volume agglomeration test for the detection of hemolytic atoxigenic V. eltor, obtained from environmental, objects, which produced lipase in 80% of cases. The differentiating capacity of the diagnosticum was confirmed on 120 V. eltor cultures isolated from environmental objects. The newly developed diagnosticum makes it possible to determine the lipase activity in cholera vibrios of different biovars and serovars.

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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.

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Conditions of cryostabilization of Yersinia pestis phages preserving their biological properties at very low temperature are studied.

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The present work deals with summarizing the experience obtained by the specialized antiepidemic brigade of the Rostov-on-Don Research Institute for Plague Control in the work on the liquidation of cholera in some regions of Daghestan with a view to discussing the problems of improvement of anticholera measures. The characteristic features of the epidemic process were its explosive character, sparseness of the foci of infection, the prevalence of its transmission through everyday contacts (family contacts and intensive tribal contacts) and essential delays in taking anticholera measures due to sudden appearance of outbreaks, remoteness of small settlements and the lack of manpower and means for carrying out anticholera measures at a given place and time, as well as delays in epidemiological analysis carried out by local health service bodies. Delays in carrying out such measures led to the spread of infection both within settlements and in the whole region and further in the republic. The epidemic process was complicated by the antibiotic resistance of V.cholerae strains circulating on this territory. All these factors formed specific epidemic situation which introduced amendments into the organization of anticholera measures.

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Wide circulation of antibiotic-resistant Vibrio cholerae strains again gives prominence to the problem of etiotropic therapy. The results of the treatment of 428 persons infected with V.cholerae (237 cholera patients and 191 Vibrio carriers) in different regions of Daghestan during the outbreak of epidemic in 1994 are presented. The main criterion of the effectiveness of antibacterial therapy was the determination of the percentage of bacterial relapses. The sensitivity of 118 V.cholerae strains to different antibacterial preparations was studied by the method of serial dilutions. After the clinical use of chloramphenicol 29.7% of bacterial relapses were registered, the in vitro resistance of V. cholerae being 32-64 mkg/ml. After the use of tetracycline 16.5% bacterial relapses were registered with in vitro resistance being the same. The use of the combination of these preparations gave 15% of bacterial relapses. Furazolidone gave 4.3% of bacteria relapses, while after the use of ciprofloxacin 2.8% of bacterial relapses were registered with in vitro sensitivity equal to 0.25-0.5 mkg/ml. Ciprofloxacin was recommended for the treatment of cholera patients and furazolidone, for the treatment of Vibrio carriers.

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Retrospective analysis of epidemic cholera manifestations was made in Daghestan using the data of operative epidemic analysis of the break in 1994. Unexpected prolongation of epidemic process of cholera for Daghestan, which was imported by pilgrims from Southern-Western Asia, has been shown using climate-geographical social-demographical and sanitary-hygienic peculiarities. Common laws of development of epidemic complications were demonstrated, as well as the main ways of infection transmission of great number of Daghestan settlements in epidemic process. The importance of antiepidemic means and significant role of created specialized antiepidemic groups have been emphasized in rapid carrying out of means in infection focus, including massive investigation of people in settlements.

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The acidic pH of water of surface water reservoirs in Izberbash and two adjoining regions, including sea water, seems to be unfavorable for the prolonged preservation of Vibrio cholerae eltor, but additional ecological investigations are necessary to study the possibility for infection to take root at this territory. Water from the Zam-Zam spring, if contaminated with V. cholerae, may serve as a transmission factor, but the duration of its action is limited by the survival term of V. cholerae. The water route of transmission did not play any essential role in the spread of cholera in the central regions of Daghestan.

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Influence of high hydrostatic pressure (2600-4300 atm) on the shape of human red blood cells was studied. An irreversible change of the shape was observed, which was similar to that seen in the case of a shift stress. The model modification dynamics of the erythrocyte shape was proposed. Activation energy and activation volume of the defects resulting in shape changes were calculated.

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The immunogenicity of Y. pestis strain EV, grown in yeast-casein medium, yeast medium with Hottinger digest and yeast medium with sunflower-seed protein at 28 degrees C and 37 degrees C, for guinea pigs and white mice has been studied. As revealed in this study, these media ensure the formation of highly immunogenic populations of Y. pestis strain EV and, therefore, can be used for growing Y. pestis vaccine strains. Considerable fluctuations in the content of such highly protective antigen as fraction 1 do not affect the immunogenicity of live cultures of Y. pestis strain EV. This is due to the leveling of differences in the content of this antigen in the process of the multiplication of these bacteria in laboratory animals.

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The content of fraction 1 and VW-antigens in Y. pestis cultures grown in different media (yeast-casein medium, yeast medium with Hottinger digest, and yeast medium with sunflower-seed protein) was studied over the course of their growth by means of the antibody neutralization and microprecipitation in agar tests. The media under study were not inferior to the casein sulfuric hydrolysate-based medium used for control in their capacity for ensuring the synthesis of VW-antigens. The maximum accumulation of fraction 1 was observed in yeast medium with sunflower-seed protein. In all media the maximum content of fraction 1 was registered on day 3 of cultivation, and the maximum accumulation of VW-antigens on days 8-9 of incubation at 37 degrees C. The data obtained in this study make it possible to regard fraction 1 and VW-antigens as the secondary metabolites of Y. pestis.

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