Yersinia pestis Surface Antigens in Reception of Specific Bacteriophages.

The significance of Yersinia pestis surface antigens in adhesiveness to specific bacteriophages has been studied with the use of two methodological approaches. It was shown that Ail protein immobilized on the surface of polystyrene microspheres (but not in the solution), can bind both the Pokrovskaya phage and pseudotuberculous diagnostic phage. YapF autotransporter interacted with both phages in a water-soluble form, but YapF bound to polystyrene microspheres interacted only with the Pokrovskaya phage. An assumption was made that Ail and YapF proteins can be the primary receptors providing non-specific reversible binding to the phages used in this work.

Influence of Specific Bacteriophage on the Level of Vesicle Formation and Morphology of Cells of Yersinia pseudotuberculosis.

Incubation of Yersinia pseudotuberculosis cells grown on a solid medium with pseudotuberculous diagnostic bacteriophage for 20 min at 37oC led to a significant decrease in the concentration of both components of the system. This effect was absent when the bacteria were grown in a fluid medium. At the same time, this incubation regimen promoted vesicle formation and typical morphological changes in bacteria grown in both surface and suspension cultures. Co-incubation of the bacteriophage with suspension of vesicles isolated from the suspension culture of Y. pseudotuberculosis grown at 10oC (but not 37oC) led to a decrease in plaque-forming activity of the bacteriophage.

[Effect of lipopolysaccharide O-side chains on the adhesiveness of Yersinia pseudotuberculosis to J774 macrophages as revealed by optical trap].

A method has been developed for the quantitative estimation of the binding force of a model microsphere with a eukaryocyte based on the optical trap in order to study the molecular mechanism of adhesion between an individual bacterium and a host cell. The substantial role of LPS O-side chains in the adhesiveness of Yersinia pseudotuberculosis 1b to J774 macrophages has been revealed with the use of a set of microspheres functionalized with lipopolysaccharide (LPS) preparations and antibodies with different specificities. The results indicate the significance of the O-antigen as a pathogenicity factor of Y. pseudotuberculosis in colonization of a macroorganism. The developed methodical approaches can be applied to the study of molecular mechanisms of the pathogenesis of pseudotuberculosis and other infectious diseases to improve antiepidemic service.

Immunochemical Nature of Receptors of Pseudotuberculosis Diagnostic Bacteriophage.

The effect of treatment of Yersinia pseudotuberculosis cells with antibodies of various specificities on adhesiveness of pseudotuberculosis bacteriophage was analyzed by competitive inhibition technique. Bacteriophage adsorption to bacteria was sterically inhibited by monoclonal antibodies to protein epitopes of Y. pseudotuberculosis outer membrane. These results suggest that receptors of pseudotuberculosis diagnostic bacteriophage are localized on the LPS core of microbial cell.

[IMMUNOCHEMICAL ACTIVITY OF YERSINIA PSEUDOTUBERCULOSIS B-ANTIGEN].

A hybridoma panel producing monoclonal antibodies to immunochemically non-identical antigenic epitopes of the protein nature located in outer membrane of Yersinia pseudotuberculosis was obtained. It was revealed that the previously identified B-antigen protecting laboratory animals from experimental plague was detected using both monoclonal antibodies against mentioned protein determinants and the determinants of lipopolysaccharide O-side chains. The B-antigen is a component of the Y. pseudotuberculosis outer membrane, egested in the form of the vesicles (OMVs) by bacteria cultivated in fluid nutrient medium.

[Study of Yersinia pseudotuberculosis surface antigen epitopes using monoclonal antibodies].

A study of the influence of exogenous factors on the immunochemical activity of the bacterium Yersinia pseudotuberculosis and lipopolysaccharide preparations isolated from bacteria was performed using monoclonal antibodies. It was shown that the hybridomas that were obtained in this work produce antibodies against different and, most likely, species-specific epitopes associated with lipopolysaccharide O side chains. The antibody concentrations produced increased with a decrease in the temperature, at which the bacteria were cultivated. An inhibitory effect of proteinase K, pepsin, and trypsin on the immunochemical activity of bacterial cells, determined using a solid-phase enzyme immunoassay, was demonstrated. Treatment with sodium periodate showed no uniform effect on the reactions between monoclonal antibodies and antigens (lipopolysaccharides and microbial cells), as adjudged by an immunoassay, which is most likely a consequence of the different localization of lipopolysaccharide epitopes recognized by the antibodies from four hybridomas.

[Immunobiological properties of Yersinia pestis antigens].

The present review contains information concerning immunobiological properties of plague microbe antigens. All of the identified antigens are evaluated in relation to pathogenicity of Yersinia pestis namely a resistance to phagocytosis, toxicity, adhesiveness etc. as well as persistence ability and adaptation to variable environment. In addition, the role of antigens in immunogenicity of living plague microbe for experimental animals is considered. The data concerning mechanisms of antigenic contribution to the development of adaptive immunity are presented.

[Influence of cultivation temperature and Yersinia pestis fra-operon carriage on morphological features of Yersinia pseudotuberculosis].

AIM: Studies of influence of Yersinia pestis fra-operon carriage on morphological properties of Y. pseudotuberculosis recipient strain cells and colonies at different temperature and cultivation time. MATERIALS AND METHODS: Cultures of Y. pseudotuberculosis isogenic variants were grown on solid nutrient medium at 4 - 6, 9 - 11, 26 - 28 and 36 - 38 degrees C. Indirect hemagglutination was used to determine F1 antigen production. Cytorefractometry was used to determine live cell percentage in colonies. Size and dividing cells percentage was evaluated by using phase-contrast microscope "Lyumam-I2". RESULTS: Time period between appearance of micro-colonies and achievement of maximum colony size increased with cultivation temperature decrease. Size of fra+ and fra- variant colony was not significantly different for every temperature regiment used and was significantly lower at the temperature of 36 - 38 degrees C. CONCLUSION: Temperature has a significant influence on population growth, colony size and form.

[Current state of problem of improving tools for plague vaccine prophylaxis].

Literature data on main immunobiological characteristics of 1st generation plague vaccines as well as ways of development of new tools for specific prophylaxis of plague: recombinant live, chemical, antiidiotypic, and DNA vaccines are presented in the review. Their expected advantages and disadvantages, perspectives of development and practical use in system of antiepidemic measures are assessed.

[Influence of cultivation temperature of Yersinia pseudotuberculosis on the immunobiological properties of lipopolysaccharide].

AIM: To study relationship between Yersinia pseudotuberculosis cultivation temperature and immunobiological properties of lypopolysaccharide (LPS). MATERIALS AND METHODS: LPS producing strain--Yersinia pseudotuberculosis 164/84 OX; experimental animal--guinea pigs, rabbits; methods--immunochemical (indirect hemagglutionation assay, RDP), physico-chemical (electrophoresis, gas-liquid chromatography), standard biochemical and statistical methods. RESULTS: It was established that increase of Yersinia pseudotuberculosis cultivation temperature from 10 degrees C to 37 degrees C was associated with attenuation of LPS toxicity in guinea pigs, decrease of its immunochemical activity and contents and degree of acylation of 3OH-14:0 hydoxyacid as well as amount of electrophoretically detected O-side chains. It was assumed that the polysaccharide component plays its role in LPS toxicity. CONCLUSION: Relation between Yersinia pseudotuberculosis cultivation temperature and structural and functional properties of LPS preparations obtained from the culture was determined.

[Biological and physico-chemical properties of Yersinia pseudotuberculosis bacterial culture having the fra-operon Yersinia pestis].

The biological and physico-chemical properties of cultures of two isogenous recombinant variants of Yersinia pseudotuberculosis were studied. The cell genomes of the cultures are distinguished from one another only by the presence or by the absence of the fra-operon, which is a determined attribute of the plague microbe capsule-forming process. The expression of the attribute is amplified by rising the microbial biomass cultivation temperature and stimulates the decrease in the viability of the bacteria and adaptation potential in vitro. In the warm-blooded owner organism the microbes of the capsule-forming recombinant variant are characterized by the greater residual pathogenicity and immunogenic ability to the experimental plague of the laboratory animals as compared to the reference-variant cells. These specific features could be explained by more expressed colonizing ability of the capsule-forming microbes provided by owner cells' stability to the phagocyte process.

[Immunogenicity of B-antigen in experimental plague and pseudotuberculosis].

The results of the evaluation of the immunogenic properties of B-antigen, earlier identified in the culture fluid of Yersinia pseudotuberculosis submerged culture, with respect to experimental plague and pseudotuberculosis are presented. B-antigen has been shown to produce protective effect in guinea pigs and, probably, hamadryas baboons, but not in white mice infected with the causative agent of plague. Immunizaton with B-antigen protects guinea pigs from primary pneumonic plague caused by both capsule-forming and noncapsular Y. pestis virulent strains. Passive immunization with antibodies to B-antigen induces limitedly pronounced protective effect in guinea pigs and is not effective for white mice with respect to experimental plague. No active or passive protection of white mice or guinea pigs, infected with Y. pseudotuberculosis cultures, has been achieved by the injection of B-antigen or antibodies to it.

[The role of the Yersinia plasmid of calcium-dependency in the formation of the plague immunity].

It was shown that viable variants of the Y. enterocolitica microorganisms containing the plasmid of Ca-dependency (native one or from EV/ I Y. pestis cells) were more immunogenic to plague-infected guinea pigs than isogenic non-plasmid variant. As a result of studying the protective properties of the bacteria Y. enterocolitica sub-cellular fractions it was determined that the largest quantity of immunogen per unit of protein was accumulated in the cultures of bacteria's plasmid variant. The aggregate of the obtained experimental data suggested that there was a non-identified protective antigen (or antigens) for guinea pigs to be encoded by chromosome's DNA. Besides, the Ca-dependency plasmid gene products take part in its excretion and, as the factors of pathogenicity, provide survivability of the Y. enterocolitica microorganisms in the host organism, increasing thus the immunogen accumulation in it.

[Current understanding of the pathogenesis of infectious diseases]. / Sovremennye predstableniia o patogeneze infektsionnykh zabolevanii.

New experimental data in such research fields as molecular biology, biochemistry, genomics and others as well as the changes occurring in the medical-and-epidemiological situation in respect to a number of infectious diseases dictate the necessity to systemize and to revise the appropriate knowledge for the purpose of ensuring a more profound understanding of the processes providing a foundation for the relations within the system of "host--parasite". The article deals with modern aspects of investigating the pathogenesis of infectious diseases, i.e. issues of the genetic and of structural-and-functional organization as well as of regulation of the pathogenetic potential of microorganisms; and mechanism of antimicrobial protection of microorganisms including from the standpoint of evolutionary biology.

[Connection of outer membrane protein composition in Yersinia pestis cells with intrinsic plasmids]. / Sviaz' belkovogo sostava naruzhnykh membran kletok Yersinia pestis s nositel'stvom sobstvennykh plazmid.

A set of isogenic derivatives of Yersinia pestis EV strain was obtained including the variants harbouring the different compositions of Yersinia own plasmids. The protein profiles of outer membranes of the set of strains were defined. The polyacrylamide gel electrophoresis has shown the small 6.1 Md plasmid to code an outer membrane protein with mol mass 29 kDa, different from pesticin I, while the heavy 60.0 Md plasmid encodes the 15-16 kDa polypeptide different from monomers of F1 and T-antigens of plague microbe.

[Effectiveness of revaccinating hamadryas baboons with NISS live dried plague vaccine and fraction I of the plague microbe]. / Effektivnost' revaktsinatsii pavianov gamadrilov chumnoi zhivoi sukhoi vaktsinoi NIIS i fraktsii I chumnogo mikroba.

The effectiveness of some vaccine preparations for the revaccination of hamadryas baboons after their primary immunization with live plague vaccine " NIIS " administered in the form of aerosol was studied. The study was carried out under the conditions of the aerosol challenge of the animals with Y. pestis. The subcutaneous injection of plague vaccine " NIIS " was found to have advantages over its aerosol administration. Revaccination with Y. pestis fraction I, absorbed, was found to be 8 times more effective than the administration of plague vaccine " NIIS " by inhalation and not inferior to the subcutaneous injection of this vaccine. Y. pestis lipopolysaccharide, when injected simultaneously with fraction I, produced an immunosuppressive effect. The development of chemical plague vaccine on the basis of fraction I, intended for revaccination, was shown to have good prospects.

[Experience using fraction I of the plague microbe for revaccinating experimental animals]. / Opyt ispol'zovaniia fraktsii I chumnogo mikroba dlia revaktsinatsii éksperimental'nykh zhivotnykh.

Experiments on guinea pigs have shown that a pronounced revaccination effect develops in the animals receiving the booster injection of fraction I of Pasteurella pestis 1.5--4 months after the primary immunization with live plague vaccine, while the booster injection of live plague vaccine produces a low revaccination effect due to the fact that this vaccine is badly adapted in the body after the primary immunization.