Pathogenetic significance of the psychrophilia of Yersinia pseudotuberculosis

The work presents the data indicating that the temperature of Y. pseudotuberculosis cultivation is very important in regulating the activity of pathogenicity factors, necessary for the initiation of the pathogenic process in the cells of the macroorganism. Low temperature (8-10 degrees C), necessary for the growth of Y. pseudotuberculosis, facilitates the activation of invasive and toxic pathogenicity factors. At a growth temperature of 37 degrees C the inhibition of such necessary attributes of virulence as adhesion and invasion into epithelial cells occurs in Y. pseudotuberculosis, which decreases the capacity of these bacteria for inducing the infectious process. The virulence of Y. pseudotuberculosis population, lost as the result of its cultivation in synthetic culture media at a temperature of 37 degrees C, has been found to be restored at low temperature.     

The entry portals and routes of Yersinia pseudotuberculosis penetration into the warm-blooded organism

This work presents data indicating that Y. pseudotuberculosis population, cultivated at lower temperature (6-8 degrees C), has a high potential of cellular and tissue invasiveness. Pseudotuberculosis has been experimentally shown to start as generalized infection due to the rapid (during 10-15 minutes) penetration of the infective agent through the epithelium of the mouth cavity, the small intestine, the urinary bladder, conjunctiva, pulmonary alveoli and vascular walls into the blood and then into internal organs. The data, obtained in this study, on the penetration of Y. pseudotuberculosis through any mucous surfaces in different species of warm-blooded animals are indicative, to a considerable extent, of the fact that this process occurs due to the action of nonspecific mechanisms.     

Influence of cultivation conditions on spatial structure and functional activity of OmpF-like porin from outer membrane of <Emphasis Type="Italic">Yersinia pseudotuberculosis</Emphasis>

The influence of cultivation conditions of pseudotuberculosis bacteria on the spatial structure and the functional activity of nonspecific OmpF-like porin was studied by means of optical spectroscopy, scanning microcalorimetry, and bilayer lipid membrane technique. With this goal, porin samples isolated from microbial masses grown at different temperatures, nutrient medium densities, and growth phases were characterized. According to CD data, the porin samples under investigation represent beta-sheet proteins. It was found that the protein isolated from the colonial culture of pseudotuberculosis bacteria grown at low temperature has the most compact structure. Using intrinsic protein fluorescence, it was shown that different conditions of pseudotuberculosis bacteria cultivation (temperature, medium, growth phase) led to the changes in spectral properties of porin fluorescence due to the redistribution of the contributions of tyrosine and different classes of tryptophan residues to the total protein emission. Heat inactivation of porin samples was studied using CD spectroscopy, intrinsic protein fluorescence, and scanning microcalorimetry. Spatial features of the porin samples were found to affect their functional activities. Considering all these data, it is possible to correlate the spatial structure and functional activity of porin samples isolated under different cultivation conditions of bacteria and the composition of the outer membrane lipid matrix.     

Pathogenic properties of Yersinia pseudotuberculosis in different biological models

The study of the action of Y. pseudotuberculosis introduced into the cavity of the ligated intestinal loop of a rabbit, into the stomach of 2- to 4-day-old suckling mice or injected intradermally has made it possible to establish the importance of the invasive and toxic activity of this microbe in the pathogenesis of pseudotuberculosis infection. The lesion of the small intestine develops due to the penetration of the bacteria into the mucous membrane and the combined action of the microbial cells inhabiting the stroma, and secreting into the environment of cytotoxins, toxic substances with an enterotropic action or enterotoxins and factors increasing the permeability of the vessels. Y. pseudotuberculosis strains with high and low invasiveness have been isolated.     

Chemotaxis of Yersinia pseudotuberculosis as a mechanism in its search for target tissues of the host organism

Y. pseudotuberculosis cells grown at biologically low temperature have been shown capable of chemotaxis with respect to carbohydrates and amino acids. During cultivation at 36-37 degrees C Y. pseudotuberculosis cells retain this property for 10-15 hours and then lose it. The mechanism of chemotaxis makes it possible for Y. pseudotuberculosis to "find" human and animal tissues and can facilitate the realization of the pathogenicity potential of these bacteria. When administered orally to mice motile bacteria, i. e. those grown at 6-8 degrees C, have been more virulent for the animals than nonmotile ones cultivated at 36-37 degrees C.     

The effect of the trophic and temperature conditions of cultivation on lipid synthesis by Yersinia pseudotuberculosis

The comparative study of the synthesis lipids in Y. pseudotuberculosis, depending on the conditions of their cultivation (at different temperatures in mineral media and in media, containing organic compounds), has been carried out. As demonstrated in this study, temperature in the main inducing factor, affecting the synthesis of lipids of definite classes and fatty acids, incorporated into these lipids. During the cultivation of Y. pseudotuberculosis in mineral and organic media under the conditions of low temperature their lipid composition remains unchanged, but at 6 degrees C the synthesis of unsaturated fatty acids prevails, while at 37 degrees C saturated fatty acids are mainly synthesized. On mineral media at 37 degrees C bacteria synthesize mostly nonpolar lipids in the form of reserve substances, represented by triglycerides and free fatty acids.     

Enzymatic inactivation of levomycetin and penicillin by cells of the plague and pseudotuberculosis microbes that contain R plasmid depending on cultivation conditions]

The activity of enzymes, inactivating levomycetin and penicillin in the cells of plague and pseudotuberculosis microbes bearing extrachromosomal determinants resistant to a number of antibiotics was studied as dependent on some cultivation parameters: population age, aeration rate and temperature. It was shown that the highest capacity for levomycetin acetylation was characteristic of the cells in the late logarithmic and early stationary growth phages. Accumulation of levomycetin O-acetothers in the incubation medium markedly increased, when the cells were grown under the conditions of intensive aeration. An increase in the cultivation temperature up to 37 degrees C was accompanied by a reliable decrease in the activity of levomycetin acetylase in the transconjugant plague and pseudotuberculosis microbes though no correlation with the resistance levels in the same strains to the above antibiotics was observed. Optimal conditions for penicillinase production were determined. The maximum levels of penicillinase were found in the cells of Y. pestis 556/106 Rn with the episotic resistance type in the early exponential developmental phase under the aeration conditions and the temperature of 28 degrees C.     

Heterogeneity of lipopolysaccharides from Yersinia pseudotuberculosis: chemical characterization of various molecular types

Prior studies have shown some unusual changes in the lipopolysaccharides (LPSs) from Yersinia pseudotuberculosis that occur when the microbe is grown at low temperature; the specific features of these LPSs in comparison with the LPSs from other enteropathogens may be due to unusual thermal adaptation mechanisms. To gain insight into this question, the chemical composition of Y. pseudotuberculosis LPS has been determined. The data indicate that two different S-form LPS species are produced in "cold"-grown bacteria. These have an identical set of bands after SDS-PAGE, similar elution profiles during gel-filtration on a Sephadex G-200 column in the presence of sodium deoxycholate, identical monosaccharide and fatty acid compositions, and similar polymerization degrees, but they have different acylation degree. On the whole, the macromolecularly different LPS populations, varying not only in their smooth or rough nature and hydrophobicity, but also in their localization in the outer membrane and, probably, their interactions with other cell components, are synthesized in "cold"-grown Y. pseudotuberculosis. The biological sense of the heterogeneity and its connection with psychrophilic and pathogenic properties of pseudotuberculosis organisms are discussed.     

Interaction of Yersinia pseudotuberculosis with the epithelium of the small intestine in experimental infection

Some data showing specific ways of interaction of Yersinia pseudotuberculosis with the epithelium of the small intestinal mucosa have been received for the first time by using light microscopy of semithin sections. Comparative study of the lesion of the epithelium of different parts of rabbit small intestine by oral inoculation with Yersinia tuberculosis has expanded the ideas of the infection entry. It has been established for the first time that the most intensive penetration of the microorganisms into the epithelium of the mucous membrane takes place in the duodenum and jejunum. As the infected contents is evacuated via the intestinal tract, the Yersinia pathogenicity decreases, and in the ileum, the invasion of the microorganisms becomes less pronounced. It has been revealed that in pseudotuberculosis, there takes place a repeated dissemination of the infection in the intestinal tract because of decay of the involved enterocytes, which is of pathogenetic importance for the progress and outcome of the infection.     

Some aspects of the immunology of pseudotuberculosis

Integral evaluation of some humoral and cellular immunity factors in patients with pseudotuberculosis (Far-East scarlatinoid fever) with varying severity of its course was carried out. Inadequate immune reaction of human organism in response to infection with pseudotuberculosis was established at the level of both specific (sporadic cases of lacking synthesis or poor synthesis of IgM antibodies, lacking synthesis of IgG antibodies) and non-specific factors (incomplete phagocytic reaction). Phagocytosis is higher in monocytes than in neutrophils during pseudotuberculosis. The opsonizing properties of blood serum, the intensity of which increases in the course of disease, are great importance in freeing the organism from bacteria of pseudotuberculosis. The most rapid and technically simplest is the determination of antibodies in blood serum of the patients by means of the reaction of indirect hemagglutination. For this purpose the authors developed a new, highly sensitive, and specific preparation - the dry erythrocyte antigenic diagnostic agent.     

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.     

Expression of Yersinia pestis antigens encoded by the Ca2+-dependence plasmid

Antigens coded by the Ca2(+)-dependance plasmid were found in the cultural medium, cytoplasm and outer membranes of the three monoplasmid (pCadV) strains of Yersinia pestis with the different basic properties. The presence of 20 mM of Mg2+ at least in the medium is necessary for optimal expression of these proteins. The existence of strain differences in the bacterial cells reaction to temperature, cultivation medium has been demonstrated. No difference in the pCad-dependent proteins was found in Yersinia pestis and the causative agents of pseudotuberculosis, enterocolitis.     

Influence of culture conditions and virulence plasmids on expression of immunoglobulin-binding proteins of Yersinia pseudotuberculosis

The influence of culture conditions and plasmids on immunoglobulin (Ig)-binding activity of two isogenic strains of Yersinia pseudotuberculosis (plasmid-free strain 48(-)82(-) and strain 48(+)82(+) bearing plasmids pYV48 and pVM82) was studied. The highest activity was observed in the bacteria grown on glucose-containing liquid medium in the stationary growth phase. The Ig-binding activity of the bacteria cultured on the liquid medium at pH 6.0 was about 1.5-fold higher than that of the bacteria grown at pH 7.2. Expression of the Ig-binding proteins (IBPs) was most influenced by temperature of cultivation. The IBP biosynthesis was activated in the bacteria grown at 4 degrees C and markedly decreased in those grown at 37 degrees C. The Ig-binding activity of lysates from the bacteria was caused by proteins with molecular weights of 7-20 kD. The activities of the plasmid-free and plasmid-bearing Y. pseudotuberculosis strains (48(-)82(-) and 48(+)82(+), respectively) were analyzed, and the plasmids were shown to have no effect on the IBP expression and biosynthesis, which seemed to be determined by chromosomal genes.     

Diagnostic value of a novel nutrient medium for isolation and cultivation of pathogens causing enteric yersiniosis and pseudotuberculosis

A new nutrient medium for isolation and cultivation of the causative agents of enteric yersiniosis and pseudotuberculosis was found to have advantages over Endo medium in its differentiating and inhibiting properties. This medium permitted the easy differentiation of Yersinia pseudotuberculosis from Y. enterocolitica, as well as from Escherichia coli, Shigella flexneri, Klebsiella pneumoniae, K. rhinoscleromatis, Hafnia, Enterobacter and Citrobacter by color; from Proteus inconstans by swarming. In addition, weakly swarming of P. vulgaris differed by their light bluish color and Pseudomonas aeruginosa, by the brilliance and size of colonies. Endo medium could be used only for differentiation of E. coli from lactose-negative Yersinia colonies, Klebsiella (by mucous growth) and, to a certain extent, all Proteus species (by swarming). The medium under test and the control medium inhibited the growth of Staphylococcus aureus. In contrast to Endo medium, the medium under test partially inhibited the growth of K. rhinoscleromatis and the swarming of P. inconstans. The new medium is now introduced into practice.     

DNA, RNA, and protein biosynthesis in cells of Yersinia pseudotuberculosis at various cultivation temperatures

Y. pseudotuberculosis cells cultivated at temperatures of 37 degrees C and 8 degrees C were found to be capable of incorporating exogenic precursors into DNA, RNA and protein. The linear growth of thymidine incorporation occurred during 8 hours of cultivation at 37 degrees C, then the amount of the incorporated label decreased. At 8 degrees C the level of thymidine incorporation into DNA gradually increased for 80 hours and longer, but not reaching the level of incorporation observed at 37 degrees C. The incorporation of uridine into RNA of Y. pseudotuberculosis cells reached its maximum after 4 hours of cultivation at 37 degrees C, at a lower temperature of cultivation the incorporation of uridine into bacterial cells was almost linear, though slower, and lasted for 20 hours. The content of radioactive alanine in Y. pseudotuberculosis protein increased during 16 hours of cultivation at a high temperature, while at 8 degrees C the growth of the incorporation level lasted for at least 40 hours. For all precursors under study the incorporation rate into the cell biopolymers at the initial stages of cultivation was higher at 37 degrees C, than at a lower temperature.     

Effect of blue-green alga (Cyanobacteria) and their exometabolites on formation of resting forms and variability of Yersinia pseudotuberculosis

Research, carried out with the use of bacteriological methods and polymerase chain reaction, revealed that the transformation of Y. pseudotuberculosis, associated with blue-green algae Anabaena variabilis, into resting (noncultivable) forms took shorter time than in soil extract containing no algae. The exometabolites of "old" cultures of these algae sharply accelerated the formation of resting Y. pseudotuberculosis forms. The influence of the algae and the products of their metabolism was manifested far more intensively at 22 degrees C than at 4 degrees C. After passage through infusoria resting Y. pseudotuberculosis forms, preserved in the mucous covering of cyanobacteria, partially reverted into vegetative forms, capable of growing on solid culture media. The revertants essentially differed from the initial vegetative forms by having lower enzymatic activity, agglutinability and cytopathogenicity, as well as by the loss of plasmid p45. The probable role of blue-green algae, widely spread in soils and water reservoirs, in the processes of reversible transformation of Y. pseudotuberculosis vegetative and resting forms, closely connected with seasonal changes of temperature conditions.     

Reduced secretion of YopJ by Yersinia limits in vivo cell death but enhances bacterial virulence

Numerous microbial pathogens modulate or interfere with cell death pathways in cultured cells. However, the precise role of host cell death during in vivo infection remains poorly understood. Macrophages infected by pathogenic species of Yersinia typically undergo an apoptotic cell death. This is due to the activity of a Type III secreted effector protein, designated YopJ in Y. pseudotuberculosis and Y. pestis, and YopP in the closely related Y. enterocolitica. It has recently been reported that Y. enterocolitica YopP shows intrinsically greater capacity for being secreted than Y. pestis YopJ, and that this correlates with enhanced cytotoxicity observed for high virulence serotypes of Y. enterocolitica. The enzymatic activity and secretory capacity of YopP from different Y. enterocolitica serotypes have been shown to be variable. However, the underlying basis for differential secretion of YopJ/YopP, and whether reduced secretion of YopJ by Y. pestis plays a role in pathogenesis during in vivo infection, is not currently known. It has also been reported that similar to macrophages, Y. enterocolitica infection of dendritic cells leads to YopP-dependent cell death. We demonstrate here that in contrast to Y. enterocolitica, Y. pseudotuberculosis infection of bone marrow-derived dendritic cells does not lead to increased cell death. However, death of Y. pseudotuberculosis-infected dendritic cells is enhanced by ectopic expression of YopP in place of YopJ. We further show that polymorphisms at the N-terminus of the YopP/YopJ proteins are responsible for their differential secretion, translocation, and consequent cytotoxicity. Mutation of two amino acids in YopJ markedly enhanced both translocation and cytotoxicity. Surprisingly, expression of YopP or a hypersecreted mutant of YopJ in Y. pseudotuberculosis resulted in its attenuation in oral mouse infection. Complete absence of YopJ also resulted in attenuation of virulence, in accordance with previous observations. These findings suggest that control of cytotoxicity is an important virulence property for Y. pseudotuberculosis, and that intermediate levels of YopJ-mediated cytotoxicity are necessary for maximal systemic virulence of this bacterial pathogen.     

Effect of cultivation temperature on nucleic acid level in bacteria Listeria monocytogenes and Yersinia pseudotuberculosis

The relationship between the multiplication of bacteria, the content of nucleic acid and the specific rate of their growth during their batch cultivation in nutrient broth and mineral medium at temperatures of 37 degrees C and 4-6 degrees C was studied in the causative agents of saprozoonotic infections with L. monocytogenes and Y. pseudotuberculosis used as typical representatives of such bacteria. The content of DNA was shown to remain practically unchanged after the alteration of cultivation temperature and the conditions of nutrition. The linear relationship between the content of RNA and specific growth rate was registered both at 37 degrees C and 4-6 degrees C. However a higher content of RNA at low temperatures was found to correspond to one and the same specific growth rate, which was linked with the additional synthesis of this nucleic acid.     

Acute oral toxicity of Yersinia pseudotuberculosis to fleas: implications for the evolution of vector-borne transmission of plague

Yersinia pestis diverged from Yersinia pseudotuberculosis相似文献   

The Yersinia Ser/Thr protein kinase YpkA/YopO directly interacts with the small GTPases RhoA and Rac-1

Pathogenic bacteria of the genus Yersinia counteract host defense by interfering with eukaryotic signal transduction pathways. YpkA of Yersinia pseudotuberculosis shares significant homology with eukaryotic Ser/Thr protein kinases, is translocated into the host cell and has been shown to be an essential virulence factor in a mouse infection model. In this study, we identify the small GTPases RhoA and Rac-1 as eukaryotic binding partners of YpkA and its homolog YopO of Yersinia enterocolitica. We demonstrate that the interaction is independent of phosphorylation of YpkA and nucleotide loading state of the GTPases. The interaction with RhoA and Rac-1 might provide an important clue to how YpkA interferes with eukaryotic signaling on a molecular level.