[Influence of Lactobacillus fermentum metabolites on ultrastructure of pathogenic Escherichia coli].

AIM: To study ultrastructural changes of hemolytic and enterohemorrhagic Escherichia coli during interaction with metabolites of Lactobacillus fermentum. MATERIALS AND METHODS: Strains of pathogenic hemolytic (Hly) and enterohemorrhagic Escherichia coli (O157:H7) as well as symbiotic bacteriocinogenic strain Lactobacillus fermentum 97 were used. Inhibition of growth of viable cells was performed by delayed antagonism method. Using electron microscopy, assessment of ultrastructural changes of hemolytic and enterohemorrhagic E. coli under the influence of diffusing in MPC-agar metabolites of lactobacilli. RESULTS: Changes pointing to profound destructive processes in bacterial cells were detected on ultrathin sections. Under the influence of diffusing metabolites of lactobacilli, the following changes were observed: destabilization of cell wall, expansion of periplasmatic space, and emergence of low electron density areas of cytoplasm in polar sections of cells with visualization of floccular material. Emergence of elongated paracrystallic packings and filamentous structures of different length, which deserve special study, was observed in cells of hemolytic E. coli. CONCLUSION: Bacteriocin-like products of lactobacilli during interaction with pathogenic E. coli cause profound destructive changes in the latter which lead to destruction of target cells.

[Interaction of Clostridium difficile with bacterial associations of parietal biofilm in colon of mice].

Using electron microscopy, ultrastructural organization of microbial parietal biofilm in colon of immunodeficient mice line B10-hr(rhy) was studied before and after peroral inoculation with enteropathogenic strain of Clostridium difficile. It was shown that infection leads to dispersion of normal biofilm in various sites and imbalance in natural proportions of different bacterial associations. Also, clear ultrastructural signs of involution of Gram-negative microorganisms were observed. In the remaining areas of the biofilm, density of bacterial population increased. In the same areas massive intrusion of microorganisms in epitheliocytes occurred with their subsequent localization in phagosomes, phagolysosomes and, in some cases, in the cytoplasm of degenerating eukaryotic cells.

[A comparative study of the role of the yadA, invA, and psaA genes in the pathogenicity of Yersinia pseudotuberculosis].

The roles of yadA, invA, and psaA genes introduced into the genetic background of the Y. pseudotuberculosis strain possessing the large p VM82 plasmid in virulence and invasion capacity were studied. Isogenic single mutants as well as double and multiple mutants of these genes were constructed and used. LD50 was used as a measure of virulence and the estimation of the ability to invade mammalian cells and the effect of infection on the weight changes of infected mice were used as additional indicators of pathogenicity. It was shown that the YadA had a major effect on the bacterial virulence when compared with the effects of PsaA and InvA. InvA appears to mediate the main pathway of the cellular invasion. YadA is responsible for the weight loss after infection of mice with sublethal doses of Y. pseudotuberculosis. The effects of YadA on virulence and of InvA on bacterial invasion were independent of the expression of the other genes studied. To our knowledge, this study showed for the first time the direct involvement of YadA in the virulence of Y. pseudotuberculosis in mice. Further pathomorphological studies are required to reveal the differences in the pathogenesis of pseudotuberculosis caused by yadA mutants or yadA+ bacteria of Y. pseudotuberculosis.

[Preparation of T-lymphoblastoid cell lines from patients with T-cell leukemia and asymptomatic carriers of HTLV-I: analysis of the viral genome status]. / Poluchenie T-limfoblastoidnykh kletochnykh linii ot bol'nogo T-kletochnym leikozom i bessimptomnykh nositelei HTLV-I: analiz statusa virusnogo genoma.

Three HTLV-I-infected partially interleukin-2-dependent lymphoid cell lines were derived from a patient with T-cellular leukemia (ATL) from Georgia and a carrier of HTLV-I from Sakhalin. The strains cultured in the presence of 3-5% interleukin-2 were designated as NBK-1, NBK-2, and YE-1, respectively. Immunoblotting analysis showed typical HTLV-I proteins in them except NBK-2 which expressed nontypical proteins p40K and p28-29K. Unexpectedly, leukemic cell fraction ATL/NBK from a patient contained gag proteins p19 (CA), p24 (MA), Pr53, and unidentified protein p29. Southern blot analysis of primary leukemic cells NBK showed one full-length non-defective provirus with restriction sites Sacl in both LTRs. Limited restriction map of the provirus virtually did not differ from previously described HTLV-I prototypes. Although the mechanism of abnormal protein expression remains to be determined, this event can be explained by defective provirus formation in NBK-2 cell line during coculturing of leukemic cells with human umbilical cord blood lymphocytes.

[Clinical and morphological characteristics of migrating erythema in patients with Ixodes tick-borne Lyme disease]. / Kliniko-morfologicheskie osobennosti proiavlenii migriruiushchei éritemy u bol'nykh iksodovym kleshchevym borreliozom.

Patients with erythematous Ixodes tick-borne relapsing fever were examined and their skin biopsy specimens were morphologically studied to reveal clinical, immunological and morphological features of erythematous Ixodes tick-borne relapsing fever. Two types of development of erythema migrans were identified. These include 1) a typical type that appears as an area of homogenous hyperemia or that of annular shape and 2) an atypical one that presents as minor vesicles. There were elevated serum immunoglobulins A levels at the height of the disease. Morphologically, at the early stage of the disease, the center of erythema shows disturbances characterized by epidermal dystrophic processes, koilocytosis, subhorny and epidermal vesicles. The dermis displays solid perivascular lymphocytic infiltrates admixed with fibroblasts, fibrocytes, macrophages, plasmocytes, eosinophils, degranulated labrocytes. The interstitium exhibits scanty infiltrates. These changes are less pronounced at the periphery. Electron microscopy shows the structures morphologically similar to those of Borrelia. The late stage (days 15-23) of the disease is marked by insignificant dystrophy and perivascular fibrosis. There were no interstitial infiltrates. By and large, the pattern of clinical and immunological manifestations in patients with erythema migrans correlates with dermal morphological changes.

[Capsule formation in Pasteurella multocida, serovar A]. / Kapsuloobrazovanie u Pasteurella multocida serovarianta A.

In this work the process of capsule formation in P.multocida bovine strain, serovar A, has been studied. The cultures were grown on liquid and solid nutrient media prepared on the basis of Hottinger hydrolysate and synthetic culture medium 199. Extracellular material was detected by the method electron cytochemistry with ruthenium red and polycationic ferritin. As revealed by specific staining and labeling, P.multocida capsule of serovar A was found to contain material of the polysaccharide nature. But the capsular structures of obtained from agar-grown and broth cultures were different. The capsular layer on the surface of cells grown in Hottinger's broth was found to be more pronounced.

[Polymorphism of virus-like particles of retrotransposons in Drosophila and yeast cells]. / Polimorfizm virusopodobnykh chastits retrotranspozonov v kletkakh drozofily i drozhzhei.

Data on the molecular arrangement of viruslike particles (VLPs) of yeast and Drosophila retrotransposons are presented. Two methods for identifying VLPs from specific retrotransposon families have been offered. The first method is based on VLPs fractionation by electrophoresis in agarose gel under strictly controlled conditions. VLPs of the Drosophila melanogaster retrotransposon families copia and gypsy and D. virilis retrotransposon Tv1 were identified by this method. The method based on heterologous induction of retrotransposons in cells of the mutant spt3 strain of Saccharomyces cerevisiae was used to identify VLPs of yeast retrotransposon Tyl and D. melanogaster gypsy retrotransposon.

[The development of a Proteus mirabilis macrocolony on a solid nutrient medium]. / Razvitie makrokolonii Proteus mirabilis na plotnoi pitatel'noi srede.

As revealed in this study, the macrocolony of Proteus mirabilis, formed on solid culture medium, may consist both of the main part and of sporadically appearing dissociating subcolonies, considerably differing one from another. The outer edge of the main part of the macrocolony of swarming cells is represented by bacteria located in three perpendicular directions. The next intermediate area consisting of two layers is represented by bacteria oriented, as a rule, in one direction. The center of the colony is made up of short microbial cells. Between the upper layer and the surface of agar an original subpopulation of microbial cells, forming a separate layer, has been detected; together they determine the planar sandwich-like architectonics of the macrocolony.

Genetic and molecular R-plasmid analysis of Enterobacteriaceae hospital strains at Children's Hospitals of the former USSR.

R-plasmids from Enterobacteriaceae clinical strains, mainly Klebsiella and Serratia, isolated at different neonatal and children's hospitals of different cities of the former USSR for 10 years, were studied for their possible influence on the bacterial host phenotype. Hospital R-plasmids of stable inheritance persisted in hospitals from 2 to 7 years and were disseminated among strains of different genera (Klebsiella, Serratia, Enterobacter) and among different units. The data showed a possibility of long-term molecular rearrangements of R-plasmids in the hospital settings and an acquisition of genetic determinants encoding enterotoxin production. A novel R-plasmid encoding cytotoxicity to HEp-2 cells involved in two nosocomial outbreaks due to K. pneumoniae strains was reported. K. pneumoniae population heterogeneity was evaluated by using the plasmid parameters of strains. Their heterogeneity of a bacterial population was significantly lower during nosocomial outbreaks than in interepidemic periods.

[Conjugative plasmid transfer in Staphylococcus aureus?]. / Kon''iugativnyi perenos plazmid u Staphylococcus aureus?

The microbiological and electron-microscopic study of the transfer of conjugative (pG873) and nonconjugative (rms7 and pE994) plasmids in two systems, on nitrocellulose filters and in a fluid culture medium, was carried out. In both systems the low frequency of the transfer of plasmid pG873 or the absence of the transfer of plasmids rms7 and pE994 were observed when nonlysogenic recipients were used for crossing. The presence of prophage in recipient cells increased the rate of the detection of gentamicin-sensitive transconjugants 100-fold and provided to reveal the transfer rms7 and pE994 plasmids. The electron-microscopic study of specimens with lysogenic recipients revealed a picture which can be interpreted as the fusion of two cells. Such picture was not observed in crossings with a nonlysogenic recipient and in preparations obtained from separate donor and recipient cultures.

[Features of expression of the pertussis toxin operon under the control of its own and heterologous promotors in Bordetella bronchiseptica cells]. / Osobennosti ékspressii operona kokliushnogo toksina pod kontrolem sobstvennogo i geterologichnogo promotorov v kletkakh Bordetella bronchiseptica.

Expression of the cloned operon encoding the pertussis toxin synthesis under the control of operons own vir-dependent promoter or vir-independent promoter of Escherichia coli origin was studied. Proteins produced by the recombinant strains have been characterized. The pertussis toxin operon was shown to express under the control of both homologous and heterologous promoters in Bordetella bronchiseptica cells. Use of the lac-promoter increases the yield of produced toxin twofold. Copy number of operon in the cell does not influence the level of toxin production. The synthesized protein can be transported into the culture medium. The biological and physico-chemical properties of the protein are similar to the ones of the natural pertussis toxin. Bordetella bronchiseptica strain producing the toxin with decreased toxic activity has been obtained. Thus, a simple system for cellular expression of the toxin operon was constructed in Bordetella bronchiseptica. It permits one to construct new strains producing nontoxic derivatives of the pertussis toxin for construction of nonreactogenic vaccines.

[The effect of interferon preparations on the ultrastructural organization of Legionella pneumophila]. / Vliianie preparatov interferona na ul'trastrukturnuiu organizatsiiu Legionella pneumophila.

The influence of the preparations of interferon on morphological changes in L. pneumophila on the ultrastructural level has been studied. Disturbances in the ultrastructure of L. pneumophila result from the direct bactericidal action of interferons without any interference of immune mechanisms. These disturbances are manifested by damages in the cell wall, plasma membrane, nuclear and ribosomal apparatuses of microbial cells. Leukinferon exhibits pronounced anti-Legionella activity, both in vitro in a liquid culture medium and in ovo, than reaferon.

[The interaction of Legionella pneumophila with different types of cell cultures and the effect of interferon preparations on this process]. / Vzaimodeistvie Legionella pneumophila s razlichnymi tipami kletochnykhkul'tur i vliianie na étot protsess preparatov interferona.

The interaction of L. pneumophila with lymphoblastoid cell cultures H9 and H9/IIIB and epithelial cell cultures HEp-2 mutual influence have been noted. L. pneumophila penetrates into cells HEp-2 and multiplies there due the so-called "spin phagocytosis". The study of the influence of the preparations of interferon, Leukinferon and Reaferon, on the adhesive capacity of bacteria and their penetration into eukaryotic cells has revealed that the preliminary treatment of both bacteria and cells HEp-2 with the preparations of interferon prior to their infection with Legionella leads to a decrease in the number of microorganisms associated with cells.

Mixed Rickettsia-virus infection in Dermacentor reticulatus imago.

Electron microscopic examination revealed replication and accumulation of Rickettsia sibirica in the fat body of experimentally infected Dermacentor reticulatus ticks. Rickettsia are released from the fat body cells by budding being surrounded with cytoplasm and plasmalemma of the host cell. Eukaryotic cell structures have been detected consisting of lamella layers whirled around the intact rickettsiae. In addition to rickettsia, microorganisms morphologically resembling Francisella tularensis and an orbivirus were found in tick tissues at morphological examination. The morphology of the virus and stages of its morphogenesis are described. Mixed viral and rickettsial infection has been shown to develop in the same ticks and even in the same fat body cells in a very close association.

[The ultrastructure of the interaction of Rickettsia sibirica and R. slovaca with the cells of ixodid ticks]. / Ul'trastruktura vzaimodeistviia Rickettsia sibirica i R. slovaca s kletkami iksodovykh kleshchei.

The ultrastructural aspects of the interaction of R. sibirica and R. slovaca with cells of mites of the species Dermacentor reticulatus, D. marginatus and Ixodes ricinus after their parenteral infection, as well as in the organs of D. marginatus infected naturally in the environment, have been studied. Both rickettsial species have similar morphology in different organs of the vector. These rickettsiae not only multiply, their populations are also partly destroyed in phagolysosomes. The natural mixed infection of R. sibirica and orbivirus in cells of D. reticulatus is described. As shown in this study, both associates pass through the complete ontogenetic cycle of development on the level of the host body and also on the level of an individual cell.

[A surface membrane study of human cell lines with different interferon sensitivities]. / Issledovanie poverkhnostnykh membran v liniiakh chelovecheskikh kletok s razlichnoi chuvstvitel'nost'iu k interferonu.

A comparative study of surface membranes of human cell J-96 and J-48 cultures with different sensitivity to alpha/beta interferon (IF). Reduced sensitivity of J-41 cells to IF-alpha/beta was found to be accompanied by a loss of highly specific receptors for IF-alpha, the lack of changes in the cell surface structures upon treatment with IF-alpha/beta, reduced intensity of cell fusion upon successive treatment with IF and polyethylene glycol. The results are discussed in connection with the observed changes in the activity of superoxide dismutase in J-96 and J-41 cell lines after treatment with IF.