Peculiarities of the Formation of Antimeningococcus Immunity in Mice Immunized with Fragments of N. meningitidis IgA1 Protease.

We studied immunogenicity of two recombinant proteins FR.9 and FR.11-3 created on the basis of fragments of the primary structure of N. meningitidis IgA1 protease with different molecular weights containing different sets of T and B epitopes. The proteins actively protect animals infected with live virulent culture of meningococci, serogroups A, B, and C. Analysis of CD4+, CD8+, and CD19+ lymphocyte populations in mouse blood showed predominant contribution of different cell populations to the formation of immune response to different proteins. Injection of FR.11-3 protein to animals did no affect the immunoregulatory index, hence, this protein can be used for creation of immunologically safe vaccine preparation.

Serological Analysis of Immunogenic Properties of Recombinant Meningococcus IgA1 Protease-Based Proteins.

Using the genome sequence of IgA1 protease of N. meningitidis of serogroup B, four recombinant proteins of different structure and molecular weight were constructed. These proteins were equal in inducing the formation of specific antibodies to IgA1 protease and had protective properties against meningococci. In the sera of immunized mice, anti-IgA1 protease antibodies were detected by whole-cell ELISA, which indicated the presence of IgA1 protease on the surface of these bacteria. We hypothesized that the protective properties of IgA1 protease-based antigens and IgA1 protease analogs could be realized not only via impairment of bacterium adhesion to the mucosa, but also via suppression of this pathogen in the organism. The presented findings seem promising for using these proteins as the basis for anti-meningococcus vaccine.

[Isolation and determination of activity of IgA1 protease from Neisseria meningitidis].

A method of the isolation and purification of IgAl protease from a culture of Neisseria meningitidis serogroup A has been developed. Three inactivated intermediates of the production of the meningococcal vaccine, a culture liquid, as well as a supernatant and sediment obtained by the precipitation of bacterial cells by cetavlon, served as a starting material. The purity of IgA1 protease was determined by SDS-PAGE. An immunoenzyme assay for determining the IgA1 protease activity has been devised. The yield of the enzyme with a specific activity of 0.5 to 4 million units/mg from 103 g of the cetavlon precipitate (40 l of culture liquid) was about 600 mug. It was shown that IgAl protease isolated from serogroup A meningococcus is capable of protecting experimental animals (mice) infected with meningococcus of serogroup B.

A new approach to establishing the set of phages for typing methicillin-resistant Staphylococcus aureus.

A new approach to using experimental phages for typing methicillin-resistant S. aureus (MRSA) non-sensitive to the phages of International Basic Set (IBS) is described. The collection Includes phage 85, modified on a culture of MRSA, and 5 phages induced from MRSA strains isolated in clinics of Moscow in 1975-76. Firstly, the modified phage selects cultures according to the specific character of its restriction-modification system, then the induced phages differentiate the selected strains into 5 groups (1, 2, 3, 4, 5) based on the specificity of the prophages they contain. Group 1 strains can further be differentiated into 5 subgroups (A, B, C, D, E) by additional phages. Forty-one MRSA strains isolated in 1987-90 in various hospitals of Moscow showing no sensitivity to IBS phages, were lysed by the modified phage, 15 of them belonging to Group 2 and isolated in the traumatological hospital, 26 belonging to Group 1 and were circulating in the burn center. Twenty-three strains of Group 1 appertain to subgroup 1B and were isolated over a 4-year period from the burned surface of patients and from the throat of a medical staff carrier.

Lysogeny of methicillin-resistant Staphylococcus aureus and the role of prophages in transfer of conjugative and non-conjugative plasmids.

The lysogenicity of 49 strains of methicillin-resistant S. aureus (MRSA) isolated in Moscow clinics in the 1970s and '80s was studied by the method of mitomycin C induction. It was found that one strain had phage of serogroup B, 33 strains had serogroup F phages and 15 strains had phages of both serogroups. In the course of genetic crossing on nitrocellulose filters it was demonstrated that serogroups B and F prophages contained in recipient cells 1) increase the frequency of transfer of conjugative plasmid pG873 and 2) mobilize transfer of non-conjugative plasmids pE994 and rms7.

[Sialylation of N-carbohydrate chains of glycoproteins with immobilized trans-sialidase from Trypanosoma cruzi]. / Sialilirovanie N-uglevodnykh tsepei glikoproteinov s pomoshch'iu immobilizovannoi trans-sialidazy Trypanosoma cruzi.

Alpha2,3-sialylation of the lactosamine type N-glycans with trans-sialidase from Trypanosoma cruzi is reported. Trans-sialidase (160 kDa, pI 5.35-5.65) and its catalytic fragment (70 kDa, pI 6.0-6.3) were isolated from T. cruzi cells and immobilized on ConA-Sepharose. The resulting preparation retained activity for several months and was repeatedly used for obtaining mono-, di-, tri-, and tetrasialylated 7-amino-4-metylcoumarine-labeled oligosaccharides with various numbers of antennas and for alpha2,3-sialylation of glycans within glycoproteins and neoglycoconjugates.

[Role of lysogenizing phages in the spread of drug-resistance plasmids in a staphylococcal population]. / Rol' lizogeniziruiushchikh fagov v rasprostranenii plazmid lekarstvennoi ustoichivosti v populiatsii stafilokokkov.

The frequency of the transduction of plasmids rms5, rms7, pT127, pC194, pS194 and pUB101 by phages belonging to serological group B (80, 52, 52A, 53, 85, phi 11, S2) in two systems was compared. In system 1 phages for transduction were obtained from plasmid-containing lysogenic donors in the process of induction with mitomycin C; in system 2 phages for transduction were obtained by their multiplication in plasmid-containing nonlysogenic donors. In system 1 the transduction of plasmids rms5, rms7, pT127, pS194 by phage 52A was found to occur with a greater (by 3-5 orders) frequency than in system 2 (the frequency of transduction was 10(-2) to 10(-4), and 10(-6) to 10(-8) respectively). A similar situation was observed with plasmids rms5 and rms7 and phage 52; plasmid pT127 and phage 53; but not observed with plasmids rms5 and rms7 and phages 80, phi 11 and S2; plasmids pC194 and pS194 and phage 53; plasmid pUB101 and phages 52A, 80 and phi 11; plasmids pC194, pS194 and pT127 and phage 85.

[Transfer of drug resistance plasmids from Staphylococcus epidermidis to Staphylococcus aureus in mixed culture]. / Peredacha plasmid lekarstvennoi ustoichinosti ot Staphylococcus epidermidis k Staphylococcus aureus v smeshannykh kul'turakh.

A possibility of transfer of chloramphenicol-resistance and penicillin-resistance plasmids from 4 different donor S. epidermidis strains to 2 S. aureus strains was demonstrated. Chloramphenocol-resistance plasmid was found in S. epidermidis 1065/77 which was not expressed in this strain but could be transferred to and was expressed in Staphylococcus aureus strains. Penicillin-resistance and chloramphenicol-resistance plasmids were transferred simultaneously in 40 per cent of the colonies.

[A new collection of phages for typing methicillin resistant Staphylococcus aureus]. / Novaia kollektsiia fagov dlia tipirovaniia metitsillino-rezistentnykh Staphylococcus aureus.

A new collection of phages for typing methicillin-resistant S. aureus (MRSA) is proposed. The collection includes phage 85 (modified by the MRSA strain), capable of selecting strains with the similar specificity of the restriction-modification system, and 9 MRSA-induced phages. The latter differentiate MRSA strains according to the specificity of prophages present in bacterial cells. The use of this phage collection has permitted the typing of MRSA strains insensitive to the phages of the international collection. Among these cultures an epidemic strain has been detected and the source of its spread in the burn center has been established.

[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.

[Effect of interferon type I on the in vivo persistence of Salmonella typhimurium]. / Vliianie interferona tipa I na persistentsiiu Salmonella typhimurium in vivo.

The influence of type I interferon on the persistence of S. typhimurium in the body of mice has been studied. The injection of the preparation of interferon has been shown to be conductive to the survival of the animals and to reduce the time of Salmonella persistence in the body. The injection of interferon enhances the phagocytic activity of macrophages in the peritoneal exudate of mice.

[Changes in the bactericidal activity of macrophages from mice with various degrees of resistance to Salmonella typhimurium infection as affected by interferon]. / Izmenenie bakteritsidnoi aktivnosti makrofagov myshei s razlichnoi ustoichivost'iu k infektsii Salmonella typhimurium pod deistviem interferona.

The bactericidal activity of mouse macrophages with different sensitivity to Salmonella infection has been studied. The sensitivity of BALB/c mice to S. typhimurium infection is associated with the low bactericidal activity of their macrophages. The introduction of interferon stimulates the bactericidal activity of macrophages sensitive to Salmonella infection of mice, which sharply enhances the resistance of the animals to this infection.

[Interferon enhances the bactericidal activity of peritoneal exudate macrophages against Salmonella typhimurium in mice]. / Interferon povyshaet bakteritsidnuiu aktivnost' makrofagov peritoneal'nogo ékssudata myshei protiv Salmonella typhimurium.

The treatment of macrophages of mouse peritoneal exudate has been found to enhance their bactericidal activity with respect to S. typhimurium. This activation depends on the dose of interferon and the cells/bacteria ratio. The action of interferon is species-specific.

[Epidemiological mechanism for the formation of drug-resistant bacterial populations]. / Epidemiologicheskii mekhanizm formirovaniia lekarstvennoustoichivykh populiatsii bakterii

The concept on the epidemiological mechanism of formation of drug-resistant bacterial populations was substantiated. Investigations showed the epidemiological mechanism of formation of the drug-resistant populations of staphylococci in the purulent-inflammatory foci to be represented in 81.4 per cent of the patients by superinfection with resistant microbes. The latter group of the patients could be divided into two subgroups: in one of the subgroups (34.9 per cent) a reduction of sensitivity in the initial staphylococci was seen in the presence of donor bacteria which later either remained in the focus or were eliminated. In this case it could be supposed that there occurred a preliminary infection with the resistant staphylococci or with the transducing bacteriophages.

[The effect of leukinferon on the activity of phagocytosing cells in human salmonellosis]. / Vliianie leikinferona na aktivnost' fagotsitiruiushchikh kletok pri sal'monellezakh u liudei.

The functional activity of phagocytic cells in 52 salmonellosis patients was studied with regard to the following characteristics: percent share of phagocytosis, phagocytic index, nitro blue tetrazolium test results, digestive activity. In patients with the unfavorable course of salmonellosis (the formation of carrier state) disturbances in the bactericidal activity of neutrophils and monocytes were established. For 32 patients leukinferon was included in the complex of etiotropic and pathogenetic treatment. The preparation was introduced in 3 intramuscular injections of 10,000 I. U. at intervals of 48 hours (the course of treatment); 10 days after the last injection this course was repeated. The use of leukinferon restored the normal functioning of phagocytes and the number of T-lymphocytes.

[The mechanism of the resistance of methicillin-resistant Staphylococcus aureus to phages from the International Collection]. / K mekhanizmu ustoichivosti metitsillinrezistentnykh Staphylococcus aureus k fagam Mezhdunarodnogo nabora.

The resistance of methicillin-resistant staphylococci to phage 85 is due to the presence of a certain system restriction modification in microbial cells. The loss of the capacity for restricting phage DNA by the cell as the consequence of the loss of the mec determinant is not accompanied by the loss of its capacity for modifying phage DNA.

[Effect of interferon type I on staphylococcal persistence and indices of body immunoreactivity]. / Vliianie interferona I tipa na persistentsiiu stafilokokkov i nekotorye pokazateli immunoreaktivnosti organizma.

After the subcutaneous injection of type I (alpha) interferon into mice their survival rate in staphylococcal infection greatly increased. At the same time duration of staphylococcal persistence in these animals and the number of persisting staphylococci were found to decrease. After the injection of interferon the splenocytes of the treated animals showed a higher capacity for interferon production. During the whole experiment the characteristics of delayed hypersensitivity in these animals showed a tendency towards normalization in comparison with those in infected mice receiving no interferon.

[Phage-mediated conjugative transfer of plasmids in Staphylococcus aureus]. / Oposredovannyi fagom kon''iugativnyi perenos plazmid u Staphylococcus aureus.

It was shown possible to transfer nonconjugative plasmids during joint cultivation of the donor and recipient cells by transduction and phage-mediated conjugation. In the latter case it was necessary that the phage in the medium was free and the prophage was present in the recipient cells. Differences in the regularities of the transfer of the nonconjugative plasmids mobilized by the conjugative plasmid or phage were observed.

[Effect of prophages on transfer frequency of conjugative plasmid G 873]. / Vliianie profagov na chastotu peredachi kon''iugativnoi plazmidy G 873.

Transfer of the conjugative plasmid G873 on filters and mixed cultivation of the donor and recipient cells in liquid media is described. In the both systems the use of the lysogenic recipient cells (phages of serogroups B and F) in the crossings increased mor than 100-fold the frequency of plasmid transfer. The conjugative transfer of the plasmid in the mixed cultivation system was proved. The conjugative transfer required the presence (while not obligatory) of calcium chloride and was restricted by the serum factors.

[A system of step-by-step differentiation of methicillin-resistant Staphylococcus aureus]. / Sistema poétapnogo differentsirovaniia metitsillinorezistentnykh shtammov Staphylococcus aureus]

One hundred and eighty four strains of methicillin resistant Staphylococcus aureus (MRSA) isolated in pyosurgical and burn departments of Moscow, Minsk, Omsk, Tbilisi, Vologda, Smolensk and Dushanbe were differentiated with using phages of the International Set (BIS), two collections of experimental phages and two-probe fingerprinting. More than 50 per cent of the isolates could not be typed by the BIS phages. The Experimental Collection of the N.F. Gamaleya Research Institute of Epidemiology and Microbiology (Moscow) was more useful in the differentiation in comparison to the Experimental Collection of the London Health Centre. A new approach applied by us to the establishment of our collection i.e. screening of cultures with a definite specificity of the system of the restriction-modification (by a modified phage 85) followed by their differentiation with respect to the specificity of the prophages (by the induced phages with the respective modification specificity) provided reliable and reproducible results. Our collection made it possible to classify the phage type 85 strains which as was confirmed by the fingerprinting data belonged to 3 different genotype variants. The fingerprinting had no advantages over the typing by the phages of the more extended phage set and was recommended for differentiation of the strains (14 per cent) not sensitive to the phages used. A step-by-step scheme for typing MRSA easy for use in clinical and epidemiological laboratories is described.