[Repeats in bacterial genomes: evolutionary considerations].

Contemporary data on repeated nucleotide sequences (repeats) in bacterial genomes are discussed. Different classifications and distribution of the repeats in the genomes of bacteria belonging to different species are reviewed. Comparative data on the density of repeats in the genomes of pathogenic and non-pathogenic bacteria, as well as in the genes encoding different functions of microbial cell (genes of stress response, genes controlling DNA metabolism and repair, etc.), are discussed. The role of the repeats in the uptake and loss of genetic material by bacterial genomes is discussed. It is suggested that repeats are responsible not only for the functioning of bacterial genome at the given moment of time, but also for the structures of the implicit genomes, which may appear in the future due to evolutionary significant genomic rearrangements.

[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.

[Molecular genetic methods of typing of the Bartonellae].

The primer systems for the PCR detection of four house-keeping genes of bartonellae in clinical material were developed and tested. The tactics of the species RFLP typing was also developed and tested. The scheme of the species RFLP typing of bartonellae was tested using as an example two strains for the first time isolated in Russia from patients with endocarditis and fever of uncertain origin. The results of the typing were supported by sequencing of the amplicons obtained. According to the sequencing the isolates were attributed to the sub species Bartonella vinsonii, subsp. arupensis. The necessity of molecular epidemiological analysis of bartonelloses in Russia was substantiated.

[Wild small mammals are the reservoir hosts of the Bartonella genus bacteria in the south of Moscow region].

A total of 103 blood samples collected from wild small mammals captured in the Prioksko-Terrasny Reserve on the south of Moscow region were studied to determine the bartonellae prevalence. The examined species were the yellow-necked mice Apodemus flavicollis (35 samples), the European wood mouse Apodemus uralensis (10 samples), the bank vole Clethrionomys glareolus (51 samples), the house mouse Mus musculus (3 samples), the common vole Microtus arvalis (2 samples), and the shrew Sorex araneus (2 samples). Initially, we obtained 76 bacterial Bartonella-like isolates after plating onto the surface of the solid nutrient media. 66 of them were PCR-positive at least for three of four targets, gltA, ftsZ, ribC and 16S RNA. Thus, the percentage of the infection in the studied community was 64%. Subsequent RFLP assay showed that obtained isolates belonged to the Bartonella grahamii and/or B. taylorii species. In 7 cases we found both bartonellae species in one animal. These data were confirmed by direct sequencing of four ftsZ, four ribC and two gltA amplicons. According to our data, there is no any marked host specificity for these bartonellae species. Now we have laid the bartonellae strain collection consisting of 31 isolates. To our knowledge, this is the first investigation of the bartonellae prevalence in wild small mammals performed in Russia. The comparison of our data with those obtained by European researchers and issues of coinfection by different bartonellae species and host specificity are discussed.

Reversions of the two missense and one nonsense trp-mutations induced by UV-light or methyl methanesulfonate in E. coli wild type and its polAi and uvrE502 mutants.

Two missense mutations, trpA58 and trpA78, and one nonsense mutation-trp-ochre, were used to determine the types of base-pair substitution caused by ultra, violet irradiation and methyl methanesulfonate (MMS) in Escherichia coli. UV irradiation of the wild-type bacteria led to the formation of revertants mainly arising as a result of GC yields AT transitions (suppressor revertants of the trpA58 mutant). True revertants of the trp- mutant (arising via transitions of AT pairs) and 5-methyl tryptophan-sensitive (MT-s) Trp+ of the trpA78 mutant (arising via unidentified transversions) occurred at a lower frequency. The polAI mutation did not change the frequency of the UV-induced transitions GC yields AT or that of the substitutions of the AT pairs. The uvrE502 mutation significantly increased the frequency of the UV-induced revertants arising via the transition GC yields AT. Treatment of the wild-type bacteria with MMS resulted in the formation of revertants mainly due to the GC yields AT substitution, and with a lower frequency to the AT yields GC transitions. MMS also induced, with a low frequency, some transversions. The frequency of the MMS-induced GC yields AT transitions was enhanced in the uvrE502 mutant. On the other hand, the uvrE502 mutation eliminated or significantly lowered MMS-induced revertants arising as a result of AT yields GC transitions or transversions.

Cloning and expression in Escherichia coli of the 37-, 14-, and/or 16-kilodalton antigens genes from Rickettsia prowazekii strain E.

A gene bank of Rickettsia prowazekii strain E constructed in the phage vector lambda EMBL4 was screened for antigen production with anti-R. prowazekii serum. One of the immunoreactive clones, grown at 37 degrees C exhibited the expression of at least two antigens of molecular weight (M(r)) 37 kD and 14 kD. Subcloning and further analysis revealed that the antigens (polypeptides) of Mr 37, 14, and/or 16 kD apparently represent structural units of the 138 kD complex antigen. Assembly of the above mentioned polypeptides was found to be thermosensitive as it took place at 30 degrees C but not at 37 degrees C and resulted in an oligomeric structure of M(r) 138 kD. The nucleotide sequence of the gene coding for a precursor of the mature polypeptides of Mr 14 and/or 16 kD was determined.

[Characteristics of transposition of Tn7-like elements determining resistance to trimethoprim and streptomycin]. / Kharakteristika transpozitsii Tn7-podobnykh élementov, opredeliaiushchikh ustoichivost' k trimetoprimu i streptotritsinu.

The transposing elements Tn7, Tn1824, controlling the resistance to trimethoprim and Tn1925, Tn1826, carrying the streptothricin resistance genes were classified as a new transposon family on the basis of their physical structure. The comparative genetic analysis of the frequency, specificity and insertion orientation in different replicons, obtained in independent research systems in this study, demonstrated the identity of transposition characteristics of the transposons. The latter makes it possible to classify them as an independent transposon family. The peculiar feature of the Tn7-like elements family is their RecA-dependent transposition into the chromosome of Escherichia coli stimulated by bacteriophage Plkc transduction of the transposons.

[Deletion of the phage lambda att80 Tn9 genome; the type of intramolecular recombination]. / Deletsii genoma faga lambda att80 Tn9; kharakter vnutrimolekuliarnoi rekombinatsii.

The intramolecular deletion-generating recombination which transforms lambda bacteriophage genomes into the plasmids (named pLS) proved to be site-specific to a certain extent. Using electron microscopy heteroduplex analysis three preferential sites for this recombination were found in seven independent pLS isolates studied. Att-sites were not registered to be involved in the formation of deletions in isolates studied. It was shown that recombination operating in our system was independent of the phage int and bacterial recA genes.

[RecA-independence of the process of amplification caused by the RS-1 sequence of Vibrio cholerae]. / RecA-nezavisimost' protsessa amplifikatsii, obuslovlennogo RS1- posledovatel'nostiami kholernogo vibriona.

The processes of amplification and deamplification of a plasmid DNA segment flanked by direct repeats of RSI-sequence of Vibrio cholerae and carrying the structural genes of cholera toxin inside the recombinant plasmid in E. coli cells have been studied. These processes determined by RSI-sequences are shown to take place independent of the RecA-system of E. coli cells.

[The effect of mutations in recB and recC genes on the precise excision of Tn5 from pNM1 plasmid genome]. / Vliianie mutatsii v genakh recB i recC na tochnoe iskliuchenie Tn5 iz genoma plazmidy pNM1.

The affect of mutations in chromosomal genes determining the realization of RecBC and RecF pathways of recombination in E. coli K12 on the frequency of transposon Tn5 precise excision from the genome of the conjugative plasmid pNM1 has been demonstrated. The pNM1 plasmid is a derivative of R100.1 and differs from the latter in the presence of Tn5 inactivating the tet gene of transposon Tn10.

[The use of the gene hybridization method for the identification of epidemiologically dangerous strains of cholera vibrios]. / Primenenie metoda gennogo zondirovaniia dlia vyiavleniia épidemicheski opasnykh shtammov kholernykh vibrionov.

Using the labeled DNA fragments containing the genes for cholera toxin the strains of cholera vibrios were studied for the presence of cholera toxin genes. Vibrio cholerae strains isolated from natural water reservoirs under the favourable epidemic situation do not contain the cholera toxin genes. The DNA hybridization method was compared with other methods used in research and practical work for estimation of epidemic importance of cholera vibrios.

[Phenotypic features of a strain of Yersinia pseudotuberculosis with the pVM82 plasmid, detected at pseudotuberculosis outbreak foci]. / Fenotipicheskie osobennosti shtamma Yersinia pseudotuberculosis s plazdmidoi pVM82, obnaruzhivaemoi v ochagakh vspyshek psevdotuberkuleza.

The phenotypic properties conferred to Yersinia pseudotuberculosis cells by the genetical determinants of a 25Md fragment of the plasmid pVM82 coding for the modified cellular immune response in the infected organism. The fragment was shown to determine the conjugative properties of the plasmid, the resistance of bacterial cells to a number of hydrophobic agents and cellular ability to absorb the Congo red dye. The latter confirms the presence of additional structural components in the cell wall of the strain harbouring the plasmid pVM82. The increased resistance of the plasmid-containing strain to bactericidal effect of the blood plasma was demonstrated as compared with the resistance of the strains harbouring the p57 plasmid lacking the 25Md fragment or no plasmid at all.

[Isolation of Escherichia coli K12 mutants with deficient in precise excision of transposons]. / Vydelenie mutantov Escherichia coli K12 s narusheniem protsessa tochnogo iskliucheniia transpozonov.

Plasmid pNM1, the derivative of R100.1, has been constructed by insertion of transposon Tn5 into structural tet genet (Tn10) of the parental plasmid. The frequency of precise excision of Tn5 from plasmidic genome is 10(-5). The high frequency of precise excision obtained in this system permits one, to use it for isolation of mutants having low frequencies of precise excision. Two mutants were isolated in which the frequencies of precise excision of Tn5 were decreased for two orders. The pex1 and pex2 mutations responsible for the effect decrease the precise excision of Tn5 from R100.1 as well as from RP4 genomes.

[Genetic analysis of mutations affecting the synthesis of somatic O-antigen of Vibrio cholerae]. / Geneticheskii analiz mutatsii, vliiaiushchikh na sintez somaticheskogo O-antigena u kholernogo vibriona.

The mutations oagM and oagR affecting the synthesis of somatic O-antigen have been localized on the chromosome of Vibrio cholerae El Jor and classic biotypes by conjugational crosses between different donor and recipient strains. The mutations are localized in the vicinity of the arg marker in both classic and El Tor biotypes of Vibrio cholerae.

[Alternative localization of the determinant of pathogenicity coded by the Yersinia pseudotuberculosis pVM82 plasmid]. / Al'ternativnaia lokalizatsiia determinant patogennosti, kodiruemykh plazmidoi pVM82 Yersinia pseudotuberculosis.

The chromosomal DNA regions in Yersinia pseudotuberculosis strains occur that are homologous to 25 Md DNA segment of the plasmid pVM82 encoding the bacterial capability of immunosuppression. The character of the chromosomal DNA regions dispersion reacting with the 25 Md segment probes is different in epidemiologically hazardous and nonvirulent strains of Yersinia pseudotuberculosis. The specific DNA regions occur as well as identical ones. The suppression of antibody formation to a number of main Yersinia pseudotuberculosis antigens by epidemiologically hazardous strain is demonstrated. The suppression is analogous to the one previously described for Yersinia pseudotuberculosis strains harbouring the plasmid pVM82.

[Genetic control of F' factor production. III. Isolation of a mutant strain of Escherichia coli with reduced frequency and altered specificity of F' plasmid production]. / Izuchenie geneticheskogo kontrolia obrazovaniia plazmid F'. Soobshchenie III. Vydelenie mutantnogo shtamma Escherichia coli so snizhennoi chastotoi i izmennenoi spetsifichnost'iu obrazovaniia plazmid F'.

Isolation and properties of the mutant of HfrH with altered frequency and specificity, of F'-plasmid formation are described. This mutant, designated fpf-5, showed significant reduction in the total yield of transconjugants selected for proximal markers in the cross with the F- recA- recipient. The crosses between the fpf-5 mutant donor and recA-nalA- recipient were sterile. The fpf-5 mutation resulted in the marked changes of the frequency of recombination exchanges involving definite chromosomal regions. The mutation reduced the frequency of the recA-independent recombination in the chromosomal fragment flanked by the leu and proA, sharply decreased the frequency of the recA-dependent and completely blocked recA-independent recombination in the chromosomal regions bordered by the genes proA and gal. Moreover the fpf-5 mutation affected the process of conjugative F'-plasmid formation suggesting effect of the mutation on recombinational exchange between the chromosome and integrated F-plasmid. The data obtained confirmed our earlier conclusion about fundamental role of the Hfr donor cell in the determination of the frequency and specificity of recombination leading to the F'-plasmid formation, and showed that the recombination events initiating F'-formation process occur in the Hfr donor cell.

[Genetic control of the RecF recombination pathway. II. Effect of recL- and uvrE- mutations]. / Izuchenie geneticheskogo kontrolia RecF-puti rekombinatsii. Soobshchenie II. Effekt mutatsii recL- i uvrE-

It was found earlier that the effect of the uvrE- mutation on the yield of recombinants formed via RecF pathway of recombination was mainly due to the decreased viability of the recB-C-sbcB-uvrE- recipient. The results of the present paper show the uvrE- and recL- mutations reduce the recipient ability of the recB-C- sbcB- strains to the same extent. We were unable to find any evidence for complementation between the recL- and uvrE- mutations in transient as well as in stable uvrE-/recL- merozygotes. The recL152 as well as the uvrE502 mutation is lethal for the strains deficient in DNA polymerase I. Both recL- and uvrE- mutations reduce in the recB-C- sbcB- strains the probability of inheritance of the proximal non-selective donor markers as well as selective markers located close to the leading end of the donor chromosome. The former effect however could not be completely due to the latter.

[IS-elements and their role in genetic recombination]. / IS-élementy i ikh rol'v geneticheskoi rekombinatsii.

The data concerning the biological functions and properties of short specific polynucleotide sequences (so called insertion sequences--IS) are reviewed. IS elements integrated in a genome can lead to strongly polar mutations in Escherichia coli, its bacteriophages and plasmids, while some IS (IS2) being integrated in inverted orientation turn on the gene activity. Several copies of the IS elements are present in the E. coli chromosome. A characteristic feature of IS is their ability to recA-independent migration along the bacterial chromosome. Possible mechanisms of IS integration are discussed. IS elements play the key role in the majority of recA-independent recombinational events: F-prime and partially Hfr-formation, plasmid recombination and dissociation, some cases of deletion formation etc. IS elements participate in recombination in the form of direct or inverted repeats. Direct repeats probably determine the processes of dissociation of the complete multicomponent R-factors and other plasmids. Inverted repeats (some of them are palindromes) are responsible for the migration of several drug-resistance determinants called transposons. Possible mechanisms of IS-dependent and probably IS-controlled recombination are discussed.

[Initial characteristics and isolation of bacterial mutants with altered ability to transpose Tn9]. / Metod vydeleniia i pervichnaia kharakteristika bakterial'nykh mutantov s izmenennoi sposobnost'iu k transpozitsii Tn9.

The method allowing the induction of bacterial mutations affecting Tn9 transposition from the bacteriophage genome to the Escherichia coli chromosome is described. Neither impaired ability of cells to adsorb bacteriophages, nor phage DNA degradation in the mutant cells were observed in the transposition-defective mutants isolated by the method. This led us to the conclusion that the isolated mutants were indeed defective in the transposition of Tn9.

[Genetic control of plasmid F' formation. II. Role of recipient recA- in the process of plasmid F' formation]. / Izuchenie geneticheskogo kontrolia obrazovaniia plazmid F'. Soobshchenie II. Rol' retsipienta recA- v protsesse obrazovaniia plazmid F'.

We have chosen the process of F'-plasmid formation as a model for a study of the illegitimate, recA-independent recombination. Our genetic data showed that F'-formation was under genetic control of the Hfr donor cells. To determine the role of recipient cells in the process of F-prime formation we compared the pattern of transconjugant formation in the crosses of Hfr and F' donors with the recA-nalA+ and recA-nalA- recipients, nalA- mutation in the recipient lead to the sharp decrease of the total yield of transconjugants selected for proximal markers and decrease the fraction of conjugative plasmids in progeny. The plasmids inherited in the nalA- recipient in HfrxF- crosses possess only proximal selected markers while the appropriate distribution of proximal and more distal markers took place in the nalA+ recipient. The homogeneity for the sensitivity of nalA- progeny to the male specific phages in comparison with the nalA+ transconjugants was also observed. The data obtained show that the nalA- mutation has a strong influence on the process of final inheritance and probably formation of F'-plasmids in the recipient cells. The extent of the influence of the nalA- mutation on the pattern of F'-plasmid inheritance was dependent upon the primary F' structure formed (or existed) in the donor cell.