[A case of legionellesis pneumonia verified by isolation of Legionella pneumophila serogroup 1 from bronchoalveolar lavage fluid treated with levofloxacine and tigecycline].

A male patient received non-chemotherapeutic drugs which induced deep neutropenia complicated with sepsis, bilateral pneumonia, acute respiratory insufficiency. Artificial pulmonary ventilation was applied. The examination of bronchoalveolar lavage showed the presence of the culture L. pneumophila (serogroup 1) in a concentration 2 x 10(3) CFU/ml. Antibacterial therapy with levofloxacin in a dose 1000 mg/day was conducted. In a week not only L.pneumophila but also Acinetobacter baumanii was isolated from bronchoalveolar lavage. Tigecyclin was added to levofloxacin treatment. Two air cavities were found in the left lung. The treatment reduced the size of these cavities, infiltrative changes in the lungs and respiratory insufficiency regressed. The patient was discharged from hospital This case is the first case in Russia of L.pneumophila isolation from bronchoalveolar lavage. The case is also characterized by use of tigecycline for treatment of combined legionella and akinetobacterial infection and cavities in the lungs in legionella pneumonia.

[Study of persistence of low virulent strain of Toxoplasma gondii on in vivo model].

AIM: To study virulence of Toxoplasma gondii strain isolated from pathologicoanatomic brain tissue of HIV-infected patients on in vivo model as well as immune response to the pathogen in immunocompetent animals and in animals with cyclophosphamide-induced areactivity. MATERIALS AND METHODS: Groups of immunocompetent and immunodeficient mice were inoculated with cysts of T. gondii obtained from brain tissues of deceased HIV-infected patients. Parasites were detected in different organs and tissues by PCR as well as by parasitological and histological methods. Antibodies to T. gondii belonging to isotypes IgG, IgG1, IgG2, IgA were detected by immunofluorescence assay. RESULTS: Cysts of the pathogen located predominantly in animals' brain, and more rarely - in spleen and liver. Using detection of antibodies of different isotypes, which marked stage of invasion, dynamics of humoral response during persistence of toxoplasms in organism was determined. Analysis of disease pathogenesis as well as peculiarities of immune response to the pathogen in immunocompetent animals and in animals with cyclophosphamide-induced areactivity was performed. CONCLUSION: Characteristics of strains of toxoplasms isolated from material of brain section of patients with HIV infection were studied. Their low virulence and ability to prolonged persistence in organism of mice were demonstrated on in vivo model. Possible association of T. gondii strains' virulence and clinical symptoms with pathogen's genetic polymorphism and its clonal population structure was discussed.

[Effect of the constitutive activity of pathogenicity genes in Listeria monocytogenes].

The effect of the constitutive expression of pathogenicity factors in L. monocytogenes was studied on the model of the intraperitoneal infection of mice. The constitutive expression was due to a single amino acid substitution in the transcriptional factor PfrA, the master regulator of L. monocytogenes virulence genes. The effective lethal dose (LD50) for the strain with the constitutive expression of pathogenicity factors was 3 times lower than for the isogenic wildtype strain. When introduced in equal doses, this Listeria strain led to the death of the animals 1-2 days earlier than the wild-type stain. The study revealed that the constitutive expression of pathogenicity factors resulted in faster dissemination of bacteria in the body of the animal during the first 24 hours, but led to their earlier elimination from the internal organs at later stages of infection.

[Different species of Legionella plasmids and virulence].

Plasmids with a molecular weight of 2.5 to 80 MD were shown to be present in a significant portion of different-type Legionella strains including high-virulence isolates L. pneumophila, serogroup 1, and L. bozemanii detected both in Russia and abroad in different sources. Plasmid-free derivative were obtained from the L. pneumophila strains each carrying only one different-size plasmid DNA. The variants had the same virulence as the original cultures for chicken embryos and guinea pigs, when the latter were infected in the bile sac or through intraperitoneal routes, respectively; their virulence was also similar to that of strains resistant to the normal serum of guinea pigs. Hence, the infection models used by us failed to show any action of plasmids on the virulence of Legionella.

[Biological and genetic characterization of strains of the pathogen of legionellosis, isolated from water on Russian territory]. / Biologicheskie i geneticheskie kharakteristiki shtammov vozvuditelia legionelleza, vydelennykh iz vody na territorii Rossii.

As the result of the study of the spread of Legionella in different regions of Russia, 69 cultures were isolated from different water systems. After serotyping most of these strains (85%) were identified as L. pneumophila, serogroups 1 and 6. 57% of the isolated Legionella strains were shown to be capable of causing fatal infection in guinea pigs, but only 50% of virulent cultures had a high level of virulence. More than a half of the isolated strains (67%) contained plasmid DNA with mol. weights ranging from 2.5 to 80 mD. In some strains of serogroup 1 the identity of plasmids was established by restriction analysis (endonucleases Hind III and Bam HI). The data thus obtained are indicative of the potential epidemic danger of L.pneumophila strains circulating in the environment.

[Isolation and biological characterization of Legionella pneumophila mutants resistant to aminoglycoside antibotics and their protective action]. / Poluchenie, biologicheskaya kharakteristika mutantov Legionella pneumophila k nekotorym aminoglikozidnym antibiotikam i ikh protektivnye svolstva.

Preliminary estimation of virulence in some antibiotic resistant mutants of Legionella pneumophila, Philadelphia 1 in various models of infection revealed its decrease in the mutants resistant to azlocillin, cefotaxime, fluoroquinolone LIB-80, neamine and streptomycin. Detailed investigation of the neamine resistant mutants showed that in relation to streptomycin susceptibility such mutants could be divided into 3 classes: susceptible to streptomycin, resistant to high concentrations of streptomycin and resistant to moderate concentrations of streptomycin. Part of mutants Nea(r)Strr and Nea(r)Strr500 and all mutants Nea(r)Strr100 proved to be less virulent with respect to guinea pigs and chick embryos. The study of the spectinomycin resistant mutants of Legionella spp. did not reveal any changes in the virulence which made it possible to suggest that the influence of the mutations in the ribosomal protein genes determining resistance to streptomycin and neamine on virulence of L. pneumophila was based on the interdependence of the mutant effect on the suppression and the influence on the virulence detected by us in S. flexneri, Y. pseudotuberculosis, L. monocytogenes and F. tularensis. The Legionella mutants Nea(r)Strr100 were characterized by significant protective activity and protected immunized guinea pigs when tested in a model of their aerogenic infection.

[The gentic principles for the design of live Salmonella vaccines]. / Geneticheskie printsipy konstruirovaniia zhivykh sal'monelleznykh vaktsin.

The presently known methods of obtaining Salmonella vaccine strains are characterized, their advantages and drawbacks are noted. Great importance of the genetic safety of Salmonella attenuated strains to be controlled is emphasized, taking into account that they are also used as carrier strains for obtaining hybrid and gene-engineering (vector) vaccines carrying immunogenicity factors of other species of pathogenic microorganisms.

[Characteristics of Listeria monocytogenes strains isolated in Russia and their typing using pulse electrophoresis]. / Kharakteristika shtammov Listeria monocytogenes, vydelennykh v Rossii i ikh tipirovanie s pomoshch'iu pul's-élektroforeza.

On the basis of specially developed scheme for the isolation of Listeria strains comprising 2 enrichment stages and the use of growth inhibitors, 128 L. monocytogenes cultures were isolated from clinical material, foodstuffs and sewage water. Highly virulent L.monocytogenes strains isolated from clinical material belonged to serovar 4b (54%) and 1/2a (38%), while those isolated from foodstuffs and sewage water belonged to 4b (74%). The restriction analysis of the chromosomal DNA of the isolated cultures with the use of restrictase EcoR1 on the basis of pulsed-field gel electrophoresis (PFGE) made it possible to distinguish Listeria strains in accordance with 5 types of restrictograms. The restrictograms of highly virulent L. monocytogenes strains, serovar 4b, belonged to types 1 and 2, while those of L. monocytogenes strains, serovar 1/2a, belonged to types 2 and 3. The comparative use of different methods for typing L. monocytogenes (sero-, phago-, bio- and resistotyping, the analysis of plasmid composition and restriction analysis) revealed that the combination of serotyping and restriction analysis on the basis of PFGE proved to be most promising for the characterization of the isolated L. monocytogenes strains and the assessment of their epidemic importance.

[The biological properties of Salmonella enteritidis strains isolated from different sources in the period of 1969-1989]. / Biologicheskie svoistva shtammov Salmonella enteritidis, vydelennykh iz razlichnykh istochnikov v period s 1969 po 1989 g.

The study of S. enteritidis cultures of different origin, isolated at the period of 1969-1989, revealed that almost all strains isolated in 1988-1989 belonged to phagovar 1 and carried a plasmid with a molecular weight of 36 MD, while of the cultures isolated in 1969-1984 only one belonged to phagovar 1 and had a plasmid of 36 MD. All S. enteritidis strains under study, irrespective of their origin and time of isolation, were highly virulent for mice, experimentally infected by different routes. S. enteritidis cultures isolated from patients exhibited higher resistance to the bactericidal action of normal guinea pig serum than strains isolated from chickens and from water. S. enteritidis mutants having lost their plasmid of 36 MD were characterized by lower virulence for mice and guinea pigs.

[Species-specific detection of Listeria monocytogenes by directed DNA amplification]. / Vidospetsificheskoe vyiavlenie Listeria monocytogenes metodom napravlennoi amplifikatsii DNK.

A highly sensitive species-specific test system which allows detection Listeria monocytogenes in animal tissues is elaborated using the principle of a polymerase chain reaction. The potentialities of its application were studied in the conditions of experimental infection of animals. It is shown that while the procedure of detection is relatively simple and fast, the test system is as sensitive and specific as traditional microbiological methods.

[The isolation and characteristics of the biological and genetic properties of neamin-resistant mutants of Salmonella abortus-ovis that are promising for vaccine creation]. / Poluchenie i kharakteristika biologicheskikh i geneticheskikh svoistv ustoichivykh k neaminu mutantov Salmonella abortusovis, perspektivnykh dlia sozdaniia vaktsin.

Neamine-resistant mutants were obtained from S. abortus ovis virulent strain. These mutants were divided into three classes according to their sensitivity to streptomycin: mutants completely retaining their sensitivity, mutants sensitive to moderate and high doses of the antibiotic. On the basis of genetic analysis carried out with the use of bacteriophage P22, the Near mutation of class Near 100 Strr 500 mutants was identified as nea B, and the Near mutation of class Near 100 Strs, as nea A. The study showed a decreased virulence of Salmonella transductants that acquired both neamine-resistant mutation of the two classes and streptomycin-resistant mutation. The streptomycin-resistant mutation produced no changes in the virulence of these bacteria. According to the results of experiments on mice, mutants of the two classes under study were found to possess protective activity.

[The transformation of legionellae by plasmid DNA]. / Transformatsiia legionell plazmidnoi DNK.

For the first time the possibility of the genetic transformation of L. pneumophila and L. bozemanii strains with the use of purified DNA of plasmids pUC19, pUC4K, pSC101 and RSF1010-pBR322 was shown. The frequency of transformation varied from 5.2 x 10(-6) to 5.8 x 10(-7), depending on the strain used in the experiment and plasmid DNA. In some of the transformants obtained in this investigation plasmid DNA whose molecular weight was similar to that of the plasmid DNA used for transformation was detected. The relatively stable preservation of plasmids pSC101 and RSF1010 in Legionella strains and the loss of plasmids pUC19, pUC4K and pBR322 in 80% of transformants during storage were shown.

[Characteristics of the interaction of legionellae and Tetrahymena pyriformis]. / Osobennosti vzaimodeistviia legionell i Tetrahymena pyriformis.

L. pneumophila avirulent strains have been shown to lose their capacity for multiplication in T. pyriformis, while the concentration of the virulent strain increases 1000-fold. The loss of the hemolytic activity of L. pneumophila virulent strain leads to the loss of its capacity for multiplication in infusorians.

[The conjugational transfer of plasmids to Legionella strains]. / Kon''iugatsionnaia peredacha plazmid v shtammy Legionella.

In experiments on conjugation the transfer of a number of R-plasmids having a wide range of hosts, such as plasmids RP1, R68.45, RP4, N3, RK2, S-a, those having a narrow range of hosts, such as plasmid R64, to strains of different Legionella species was shown. The frequency of transfer varied from 3.1 X 10(-3) to 9.4 X 10(-7). The fact that the conjugation transfer was confirmed by the reverse transfer of plasmids from Legionella transconjugates to Escherichia coli strain K12, as well as by the detection of the DNA of the transferred plasmid by means of electrophoresis in agar gel. Plasmid RP1 showed different behavior in transconjugates of various Legionella species after several passages in a medium free of antibiotics. In the Legionella strain under study the unstable preservation of plasmid R64 was observed.

[The use of a molecular DNA probe based on the cloned cytolysin gene for the identification of Legionella]. / Primenenie molekuliarnogo DNK-zonda na osnove klonirovannogo gena tsitolizina dlia identifikatsii legionell.

A molecular DNA probe has been obtained on the basis of recombinant plasmid pNIG carrying the Legionella cytolysin gene. The use of this probe for the identification of different Legionella species and other microorganisms has shown that it may serve as the species-specific marker of L. pneumophila. The probe has been used for the identification of new Legionella-like strains isolated from the environment.

[Plasmid profile of strains of various Legionella species]. / Plazmidnyi profil' shtammov Legionella razlichnykh vidov.

The plasmid profile of Legionella strains of different origin has been studied. 15 out of 32 Legionella cultures belonging to different strains have been found to contain plasmid DNA in an amount of 1 or 2 plasmids, with the exception of L. feelei having 6 plasmids. Only 1 out of 3 Legionella strains isolated in the USSR has been found to possess a plasmid with a molecular weight of about 80 MD. Plasmids with this molecular weight have been found in 13 Legionella strains under study, such plasmids in L. pneumophila strains of serogroup 1 (strains Flint 1 and Albuquerque 1) and serogroup 9 (strain No. 35282) having an exact molecular weight of 82.4 +/- 2.4 MD and being similar in molecular structure, which has been established as the result of their treatment with restrictases Pst 1 and Hind III.

[The action of interferon preparations on strains of Legionella of various serogroups and species]. / Deitsvie preparatov interferona na shtammy legionell razlichnykh serogrupp i vidov.

The action of different preparations of interferon on Legionella strains has been studied in vivo and in vitro. The preparations of leukinferon at a concentration of 500 international units (I.U.) and reaferon at a concentration of 10,000 I.U. have been found to produce an inhibiting effect on Legionella strains in vitro, in a medium with carbon-yeast agar. Leukinferon at a concentration of 125 I.U. suppresses the growth of L. pneumophila also in a liquid medium. The preparation of leukinferon at a minimal concentration of 100 I.U. has been found to suppress the development of lethal infection in chick embryos infected with L. pneumophila strain Philadelphia 1.

[Enterotoxigenicity of Salmonella strains of various origins]. / Enterotoksigennost' shtammov Salmonella razlichnogo proiskhozhdeniia.

The study of the enterotoxigenicity of S. typhimurium with the use of the skin test on rabbits (to detect the delayed permeability factor) has revealed that these strains produce an enterotoxin similar to Escherichia coli thermolabile enterotoxin (TLE). Study of the enterotoxic activity of lysates obtained from 39 S. typhimurium strains and 5 S. dublin strains by sonication has revealed that 87% of S. typhimurium strains and all S. dublin strains produce an enterotoxin similar to E. coli TLE, as demonstrated by all tests used in this investigation, while 59% of S. typhimurium cultures and all S. dublin strains have been positive when tested for the capacity of producing the rapid permeability factor. "Hospital" strains and polyresistant cultures isolated from the environment (phagovar 20) are characterized by a higher rate of producing an enterotoxin similar to E. coli TLE, detected by the tests used in this investigation (90%), than antibiotic-sensitive strains of different origin (78%).