[Cell-free Pseudomonas vaccine: its immunochemical characteristics and immunogenicity]. / Beskletochnaia vaktsina psevdomonas: immunokhimicheskaia kharakteristika i immunogennost'.

Two variants of cell-free protein vaccine have been prepared from the mixture of 4 P. aeruginosa strains, serovars 02, 06, 07 and 011, and from a single P. aeruginosa strain, serovar 02. The preparation contains proteins with molecular weight ranging from 20,000 to 100,000 and the admixture of lipopolysaccharide in negligible amounts (not exceeding 0.08% of dry weight). The vaccine produces no signs of toxicosis in laboratory animals. The vaccine effectively protects mice challenged with P. aeruginosa of different O-serotypes and stimulates the formation of specific protective antibodies in rabbits.

Somatic antigens of Pseudomonas aeruginosa. The structure of the O-specific polysaccharide chain of the lipopolysaccharide from P. aeruginosa O13 (Lányi).

The O-specific polysaccharide, obtained on mild acid degradation of lipopolysaccharide of Pseudomonas aeruginosa O13 (Lányi classification), is built up of trisaccharide repeating units involving 2-acetamidino-2,6-dideoxy-D-glucose (N-acetyl-D-quinovosamine, D-QuiNAc), 2-acetamidino-2,6-dideoxy-L-galactose (L-fucosacetamidine, L-FucAm), and a new sialic-acid-like sugar, 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-L-galacto-nonuloso n ic acid (Sug), and thus contains simultaneously both acidic and basic functions. Cleavage of the polysaccharide with hydrogen fluoride in methanol revealed the high stability of the glycosidic linkage of the ulosonic acid and afforded methyl glycosides of a disaccharide and a trisaccharide. The structures of the new ulosonic acid and acetamidino group were established by analysing the oligosaccharide fragments by 1H, 13C nuclear magnetic resonance spectrometry, as well as on the basis of their chemical conversions: alkaline hydrolysis of the acetamidino group into acetamido group, reductive deamination with lithium borohydride into the ethylamino group and acetylation with acetic anhydride in pyridine accompanied by intramolecular acylation of the acetamidino function by the ulosonic acid to form a six-membered lactam ring. Identification of the oligosaccharide fragments and comparative analysis of the 13C nuclear magnetic resonance spectra of the oligosaccharides and polysaccharide revealed the following structure of the repeating unit: ----3)D-QuiNAcp(alpha 1----3)Sugp(alpha 2----3)L-FucAmp(alpha 1----.

Somatic antigens of Pseudomonas aeruginosa. The structure of O-specific polysaccharide chains of the lipopolysaccharides from P. aeruginosa O5 (Lányi) and immunotype 6 (Fisher).

Lipopolysaccharides were isolated from dry bacterial cells of Pseudomonas aeruginosa O5a,b,c, O5a,b,d, O5a,d (Lányi classification) and immunotype 6 (Fisher classification) by the Westphal procedure. Their polysaccharide chains were built up of trisaccharide repeating units containing D-xylose, 2-acetamido-2,6-dideoxy-D-galactose and a new sialic acid-like sugar, the di-N-acyl derivative of 5,7-diamino-3,5,7,9-tetradeoxy-L-glycero-L-manno-nonulosonic (pseudaminic) acid. Formyl, acetyl and (R)-3-hydroxybutyryl groups were identified as the N-acyl substituents of the last monosaccharide; O5a,b,c and O5a,b,d lipopolysaccharides also contained O-acetyl groups. The glycosidic linkage of pseudaminic acid was extremely labile towards acids, and mild acid degradation of the lipopolysaccharides produced, instead of the O-specific polysaccharides, their trisaccharide fragments with pseudaminic acid at the reducing terminus. Similar degradation of immunotype 6 lipopolysaccharides, followed by oxidation with sodium metaperiodate, resulted in a disaccharide fragment due to destruction of xylose. In contrast the glycosidic linkage of pseudaminic acid proved to be more stable towards treatment with hydrogen fluoride than those of xylose and N-acetylfucosamine. As a result, solvolysis of immunotype 6 lipopolysaccharide with hydrogen fluoride in methanol gave methyl glycosides of a disaccharide and a trisaccharide with pseudaminic acid at the non-reducing terminus. Mild acid hydrolysis of these oligosides afforded free 5-N-acetyl-7-N-formylpseudaminic acid, which was identified by the 1H ande 13C nuclear magnetic resonance data, as well as by the mass spectrum of the corresponding fully methylated aldonic acid. As a result of the identification of all oligosaccharides obtained and comparative analysis of the 13C nuclear magnetic resonance spectra of the oligosaccharides and lipopolysaccharides the following structures were established for the repeating units of the polysaccharide chains of the lipopolysaccharides: (Formula: see text) where D-Xyl = D-xylose, D-FucNAc = 2-acetamido-2,6-dideoxy-D-galactose, Pse5N7NFm = 5-amino-3,5,7,9-tetradeoxy-7-formamido-L-glycero-L-manno-nonulosonic+ ++ acid (7-N-formylpseudaminic acid). All the polysaccharides have an identical carbohydrate skeleton and differ from each other by the acyl substituent at N-5 of pseudaminic acid [acetyl or (R)-3-hydroxybutyryl group] or by the presence or absence of the O-acetyl group at position 4 of N-acetylfucosamine. The data obtained account properly for the O specificity of the studied P. aeruginosa strains.

Somatic antigens of Pseudomonas aeruginosa. The structure of O-specific polysaccharide chains of P. aeruginosa O10 (Lányi) lipopolysaccharides.

Mild acid degradation of lipopolysaccharides from Pseudomonas aeruginosa O10a and O10a,b (Lányi classification) resulted in O-specific polysaccharides built up of trisaccharide repeating units containing 2-acetamido-2,6-dideoxy-D-glucose (N-acetylquinovosamine, DQuiNAc), 2-acetamido-2,6-dideoxy-D-galactose (N-acetylfucosamine, DFucNAc), and 5-acetamido-3,5,7,9-tetradeoxy-7-[(R)-3-hydroxybutyramido] -L-glycero-L-manno-nonulosonic acid. The latter is a di-N-acyl derivative of a new sialic-acid-like sugar which was called by us pseudaminic acid (PseN2). A 3-hydroxybutyric acid residue was also found in natural carbohydrates for the first time. In the O10a,b polysaccharide pseudaminic acid carried an O-acetyl group at position 4. For selective cleavage of the O10a polysaccharide, solvolysis with hydrogen fluoride was employed which, owing to the relatively high stability of the glycosidic linkage of pseudaminic acid, led to the disaccharide with this sugar on the non-reducing terminus. Performing the solvolysis in methanol afforded the methyl glycoside of this disaccharide which proved to be more advantageous for further analysis. Carboxyl-reduction made the glycosidic linkage of pseudaminic acid extremely labile, and mild acid hydrolysis of the carboxyl-reduced 010a polysaccharide afforded the trisaccharide with a ketose derivative on the reducing terminus. Establishing the structure of the oligosaccharide fragments obtained and interpreting the 13C nuclear resonance spectra of the polysaccharides allowed to determine the following structure for their repeating units: (formula: see text) In the polysaccharides the N-acetylquinovosamine residue is attached not to pseudaminic acid itself, but to its N-acyl substituent, 3-hydroxybutyryl group, and thus the monomers are linked via both glycosidic and amidic linkages.

Somatic antigens of Pseudomonas aeruginosa. The structure of O-specific polysaccharide chains of the lipopolysaccharides from P. aeruginosa O1 (Lányi), O3 (Habs), O13 and O14 (Wokatsch), and the serologically related strain NCTC 8505.

Lipopolysaccharides from Pseudomonas aeruginosa O1 (Lányi classification), O3 (Habs classification), O13 and O14 (Wokatsch classification), and strain NCTC 8505, which is also related to serogroup O3 (Habs), have structurally similar O-specific polysaccharide chains built up of tetrasaccharide repeating units involving L-rhamnose (Rha), 2-acetamido-2-deoxy-D-glucose (GlcNAc), 2-acetamido-2-deoxy-L-galacturonic acid (GalNAcA), and a di-N-acyl derivative of bacillosamine (BacN): 2,4-diacetamido-2,4,6-trideoxy-D-glucose or 2-acetamido-2,4,6-trideoxy-4-[(S)-3-hydroxybutyramido]-D-glucose. The latter derivative was obtained free by solvolysis with hydrogen fluoride of carboxyl-reduced Habs O3 polysaccharide, and was identified by 1H-nuclear magnetic resonance spectroscopy and by mass spectrometry of the corresponding methylated alditol. Habs O3, Lányi O1, and Wokatsch O14 polysaccharides contained O-acetyl groups. Solvolysis with hydrogen fluoride of the native Habs O3 polysaccharide resulted in selective cleavage of the glycosidic linkages of 6-deoxy sugars to give the trisaccharide fragment involving all three N-acylated amino sugars. Similar solvolysis of NCTC 8505 polysaccharide afforded a mixture of disaccharide and trisaccharide with N,N'-diacetylbacillosamine at the reducing end. Smith degradation of Habs O3 polysaccharide resulted in selective oxidation of rhamnose to give a glycoside of a trisaccharide with glyceraldehyde as the aglycone. Smith degradation of NCTC 8505 polysaccharide was complicated by the formation of the glycoside of a trisaccharide with an aglycone of unknown structure. A trisaccharide with rhamnose at the reducing end was also isolated after Smith degradation of the latter polysaccharide. Analysis of the composition and structure of all oligosaccharides obtained, and detailed examination of the 13C-nuclear magnetic resonance spectra of these oligosaccharides, and of both intact and modified polysaccharides, revealed the following structures of the repeating units. The structure for the NCTC 8505 polysaccharide differs from that proposed previously [Tahara, Y. and Wilkinson, S.G. (1983) Eur. J. Biochem. 134, 299-304] in the configurations assigned to the glycosidic linkages of rhamnose and bacillosamine. The results obtained show the P. aeruginosa strains studied to represent three different O-serotypes in a single O-serogroup (Formula: see text).

[Effect of radiation sterilization on the immunobiological properties of the sorbed protective fraction of Bordetella pertussis]. / Izuchenie vliianiia radiatsionnoi sterilizatsii na immunobiologicheskie svoistva sorbirovannoi protektivnoi fraktsii kokliushnykh mikrobov.

The possibility of using radiation for the sterilization of the dried preparation of B. pertussis adsorbed protective fraction has been studied. The use of gamma irradiation in a dose of 1.5 X 10(3) J/kg has been found to permit obtaining sterile preparations while preserving their initial protective properties.

Immunochemical and immunological study of cell-wall proteins of Pseudomonas aeruginosa.

Crude aqueous extract was obtained from acetone-dried cells of Pseudomonas aeruginosa strain 868 (serogroup O2, Lányi and Bergan's schema) and subjected to ultracentrifugation (105,000 g, 3 h); the lipopolysaccharide (LPS)-containing precipitate was discarded and the supernatant containing water-soluble cell proteins was subjected to further fractionation. From a partially purified aqueous extract two fractions were obtained by step-wise precipitation with ammonium sulphate, namely, F1 (by 50% saturation), and F2 (by 80% saturation). By gel and ion-exchange chromatography from both fractions 9 subfractions were isolated differing in molecular weight, protein content, and LPS contamination. Subfractions 4 and 7 were practically free from LPS, and gave one precipitation line with antisera for strain 868. By immunoelectrophoresis subfraction 4 contained 2 cathodic and 1 anodic, whereas subfraction 7 mainly 1 anodic component. These subfractions were antigenically identical. With ELISA these subfractions were less active as compared to other subfractions, in particular to those of high molecular weight. The anti-subfraction 4 and anti-subfraction 7 sera were found to protect passively mice against intraperitoneal challenge by P. aeruginosa strain 8 (serogroup O2). These data support the authors' opinion that subfraction SF-4 and SF-7 are protective protein antigens (mol wt about 40,000 and 30,000, respectively), that are localized in the outer membrane of P. aeruginosa cell envelope.

Somatic antigens of Pseudomonas aeruginosa. The structure of the O-specific polysaccharide chains of lipopolysaccharides of P. aeruginosa serogroup O4 (Lányi) and related serotype O6 (Habs) and immunotype 1 (Fisher).

Acidic O-specific polysaccharides were isolated on mild acidic degradation of lipopolysaccharides of Pseudomonas aeruginosa serotypes O4a,b, O4a,c, O4a,d (Lányi classification) and serologically related to them serotype O6 (Habs classification) and immunotype 1 (Fisher classification). The polysaccharides had identical monosaccharide composition and were built up of L-rhamnose, 2-acetamido-2,6-dideoxy-D-glucose,2-formamido-2-deoxy-D-galacturonic acid and 2-acetamido-2-deoxy-D-galactouronamide residues. The latter two derivatives of D-galactosaminuronic acid were found in nature for the first time. All the polysaccharides, but Lányi serotype O4a,c, contained O-acetyl groups. The polysaccharides were readily de-O-acetylated with aqueous triethylamine and de-N-formylated with dilute hydrochloric acid. De-N-formylated polysaccharide of serotype O4a,c was selectively cleaved with nitrous acid upon 2-amino-2-deoxygalacturonic acid residues to form a tetrasaccharide with a 2,5-anhydrotaluronic acid residue on the reducing end. The tetrasaccharide represented a modified repeating unit of the polysaccharide. Solvolysis of all intact polysaccharides with hydrogen fluoride selectively split the glycosidic linkages of 6-deoxy sugars to give the same trisaccharide, including both derivatives of galactosaminuronic acid and having 2-acetamido-2,6-dideoxyglucose on the reducing end. Structural investigation of the oligosaccharides obtained together with methylation analysis and 13C nuclear magnetic resonance data revealed the following structures of the O-specific polysaccharides: (Formula: see text) An independent confirmation of the structures of the repeating units was obtained as the result of full interpretation of the 13C nuclear magnetic resonance spectra of the intact and modified polymers. Spectral data analysis revealed a number of regularities in the effects of glycosidation connecting their values with the anomeric and absolute configuration of pyranose residues. The data on the structures of the O-specific polysaccharides indicated that each of the five P. aeruginosa strains under study should be considered as an individual O-serotype within one O-serogroup.

Somatic antigens of Pseudomonas aeruginosa. The structure of the O-specific polysaccharide chains of P. aeruginosa O11 (Lányi) lipopolysaccharides.

Lipopolysaccharides were isolated from the phenol layer on aqueous phenol extraction of cells of Pseudomonas aeruginosa O11 (Lányi classification), strains 170021 and 170040. On mild acid degradation of the lipopolysaccharides, with the subsequent gel-filtration on Sephadex G-50, neutral O-specific polysaccharides made up of 6-deoxysugars alone were obtained. Two 2-acetamido-2,6-dideoxy-L-galactose (LFucNAc), 2-acetamido-2,6-dideoxy-D-glucose (DQuiNAc) and L-rhamnose (LRha) residues were found to be the components of the strain 170021 polysaccharide repeating units; those of strain 170040 contained the same monosaccharides, but, instead of 2-acetamido-2,6-dideoxy-D-glucose residue, that of 2-acetamido-2,6-dideoxy-D-galactose (DFucNAc) was present. On the basis of the 13C nuclear magnetic resonance data, methylation analysis and three successive Smith degradations the following structures were determined for the polysaccharide repeating units: strain 170021----2) LRha(alpha 1----3)LFucNAc(alpha 1----3)LFucNAc(alpha 1----3)DQuiNAc(beta 1----; strain 170040,----2)LRha(alpha 1----3)LFucNAc-(alpha 1----3)LFucNAc(alpha 1----3)DFucNAc(beta 1----; differing from one another by configuration of C-4 of 2-acetamido-2,6-dideoxy-D-hexopyranose only.

Vaccines against Pseudomonas aeruginosa infection: 1. Experimental studies.

An experimental study of several types of vaccines against Pseudomonas aeruginosa has been carried out. Pyoimmunogen, a multi-component vaccine consisting mainly of heat-stable proteins (or polypeptides) with a molecular weight of 20 000 to 60 000, was prepared both by laboratory and production techniques using three or five selected strains of P. aeruginosa. The preparations obtained by both methods were found to be of low toxicity and to induce in mice and rats active immunity against challenge with homologous and some heterologous strains of P. aeruginosa, including the toxigenic PA-103 strain. The Pseudomonas vaccine (PV) was prepared from partly purified water-soluble protein antigens (molecular weight ranging from 10 000 to 100 000) obtained from one (mono-PV) or three (tri-PV) selected strains. Mono-PV was prepared by the method of ultrafiltration and tri-PV--by differential centrifugtion. Both types of vaccines proved effective in active mouse protection tests after challenge with homologous as well as with heterologous P. aeruginosa strains. Mono-PV is supposed to induce species-specific immunity against P. aeruginosa infection in mice.

The structure and immunochemical specificity of 0-antigens of 03 serogroup Pseudomonas aeruginosa.

Using the method of phenol-aqueous extraction, lipopolysaccharides (LPS) were isolated from 5 strains (subgroups) belonging to 03 serogroup of P. aeruginosa (4). Specific polysaccharides were isolated from the LPS by means of acid hydrolysis. It has been established that the polysaccharides determining O-antigenic specificity have a uniform structure. They consist of repeated trisaccharide links comprising: 2,3(1-acetyl-2-methyl-2-imidazolino-5,4)-2,3-dideoxy-D-mannuronic acid (Im) 2,3-diacetamido-2,3-dideoxy uronic acid with -D-manno-(M) or -L-gulo-(G) configuration and 2-acetamido-2-deoxy-D-fucose (F) with alpha or beta configuration of the glycosidic bond. The structure of 0/3a/3d, 3f polysaccharide has not been definitely cleared up. Serological analysis using passive haemagglutination reaction (PHAR) testifies to the presence of antigenic cross activity of all the five LPS. Antigenic specificity of LPS of the individual subgroups was revealed in passive haemagglutination inhibition reaction (PHAIR). Partial cross activity was clearly demonstrated in immunoprecipitation experiments in the five subgroups. Serological properties of the LPS of P. aeruginosa 03 subgroup essentially correlate with the structure of their polysaccharide chains determining O-antigenic specificity.

[Determination of in vivo and in vitro toxin activity. I. A new method of determining in vitro activity]. / Opredelenie aktivnosti toksinov in vivo i in vitro. Soobshchenie I. Novyi metod opredeleniia aktivnosti in vitro.

The potency of 5 toxins of different microbial species has been studied by common in vivo methods and by the in vitro method based on measuring the relative test-microbe growth inhibiting units 50. The possibility of using this method for the determination of toxicity in vitro with a view of studying the potency of diphtheria and gas-gangrene exotoxins, as well as that of Pseudomonas aeruginosa exotoxin, is substantiated.

Somatic antigens of Pseudomonas aeruginosa. The structure of O-specific polysaccharide chains of P. aeruginosa O:3(a),c and O:3a,d,e lipopolysaccharides.

On mild acid degradation of Pseudomonas aeruginosa O:3(a),c, O:3a,d,e and O:3(a),d,f lipopolysaccharides O-specific polysaccharides were isolated. The trisaccharide repeating units of O:3(a),c and O:3a,d,e polysaccharides contained 2-acetamido-2,6-dideoxy-D-galactose and 2,3-(1-acetyl-2-methyl-2-imidazolino-5, 4)-2,3-dideoxy-D-mannuronic acid, which were identified previously as the constituents of P. aeruginosa O:3a,b and O:3a,d O-specific polysaccharides, as well as 2,3-diacetamido-2, 3-dideoxyl-L-guluronic acid, which has never before been found in nature. The last monosaccharide was identified without being isolated in the free state, by means of 13C nuclear magnetic resonance spectroscopy. The structures of O:3(a),c and O:3a,d,e polysaccharides were established by selective hydrogen fluoride solvolysis followed by modification of the trisaccharides obtained and analysis of the 13C nuclear magnetic resonance spectra at each modification stage. The polysaccharides possessed similar structures of repeating units differing from each other in the anomeric configuration of the N-acetylfucosamine residue only: leads to 4)DManImU-(beta 1 leads to 4) LGul(NAc)2U(alpha 1 leads to 3)DFucNAc(beta 1-and leads to 4)DManImU(beta 1 leads to 4)LGul(NAc)2U(alpha 1 leads to 3)DFucNAc(alpha 1-, where DManImU = 2,3-(1-acetyl-2-methyl-2-imidazolino-5,4)-2,3-dideoxy-D-mannuronic acid, LGul(NAc)2U = 2,3-diacetamido-2,3-dideoxy-L-guluronic acid, DFucNAc = 2-acetamido-2,6-dideoxy-D-galactose. The same components were detected in the O:3(a),d,f polysaccharide but not in the same stoichiometric ratio; this polysaccharide possessed no regular structure. The immunochemical study of lipopolysaccharides of all five P. aeruginosa O:3 serotypes showed a definite interrelation between their serological properties and structures of the corresponding O-specific polysaccharides.

[Nonharsh methods of isolation of antigenic complexes of Klebsiella pneumoniae, and analysis of their protective activity]. / Izuchenie antigennykh kompleksov Klebsiella pneumoniae shchadiashchimi metodami i izuchenie ikh protektivnoi aktivnosti.

Protective antigenic complexes capable of protecting mice from septic Klebsiella infection can be obtained from Klebsiella pneumoniae. The preparations obtained by the method of aqueous extraction and by the action of hydroxyl amine and autolysis have been found to possess approximately the same protective activity and to protect mice from infection with Klebsiella pneumoniae strains belonging to both homologous and heterologous serotypes.

Somatic antigens of Pseudomonas aeruginosa. The structure of O-specific polysaccharide chains of P. aeruginosa O:3a, b and O:3a, d lipopolysaccharides.

On mild acid degradation of Pseudomonas aeruginosa O:3a,b and O:3a,d lipopolysaccharides O-specific polysaccharides were isolated. Both polysaccharides were found to contain 2-acetamido-2,6-dideoxy-D-galactose, identified as fucosamine hydrochloride formed after hydrolysis with a very low yield. The other two components of the trisaccharide repeating unit, 2,3-diacetamido-2,3-dideoxy-D-mannuronic acid and 2,3-(1-acetyl-2-methyl-2-imidazolino-5,4)-2,3-dideoxy-D-mannuronic acid, were identified without isolation in their free state directly in the course of structural investigation of the polysaccharides. Both these monosaccharides have never before been found in nature. Solvolysis of either O:3a,b or O:3a,d polysaccharides with liquid hydrogen fluoride resulted in the formation of the same trisaccharide, N-acetylfucosamine residue being the reducing end. The structure of this trisaccharide, which is the repeating unit of both polysaccharides, was deduced from the results of successive chemical modifications and 13C-nuclear magnetic resonance spectra recorded for every oligosaccharide formed. As a result, the acidic diaminosugars were converted into 2,3-diacetamido-2,3-dideoxy-D-mannose indistinguishable from authentic sample. The O-specific polysaccharides O:3a,b and O:3a,d differed in the configuration of the glycosidic bond of N-acetylfucosamine residue only and had the following structures: leads to 4)DManImU(beta 1 leads to 4)DMan(NAc)2U (beta 1 leads to 3)DFucNAc(beta 1- leads to 4)DManImU(beta 1 leads to 4)DMan(NAc)2U (beta 1 leads to 3)DFucNAc(alpha 1- where DManImU = 2.3-(1-acetyl-2-methyl-2-imidazolino-5,4)-2, 3-dideoxy-D-mannuronic acid, DMan(NAc)2U = 2,3-diacetamido-2,3-dideoxy-D-mannuronic acid, DFucNAc = 2-acetamido-2,6-dideoxy-D-galactose. The structures established were in agreement with optical rotations and assignments of all the signals in the 13C-nuclear magnetic resonance spectra of the polysaccharides.

[Elaboration of a method of stabilizing the protective fraction of pertussis microbes]. / Razrabotka metoda stabilizatsii protektivnoi fraktsii kokliushnykh mikrobov.

A method for stabilizing adsorbed preparations of the protective fraction of B. pertussis has been developed; according to this method, a colloid suspension of protective protein in phosphate buffer is obtained, the protein is then adsorbed on aluminium hydroxide gel and lyophilized with 10% of sucrose. If stored at 4 degrees C, these dried preparations have been found to retain their immunogenicity for 1 year.

[Cell-free Pseudomonas vaccine. IV. Laboratory trials of the effectiveness of experimental Pseudomonas vaccines]. / Beskletochnaia psevdomonas-vaktsina. Soobshchenie IV. Laboratornye ispytaniia éffektivnosti éksperimental'nykh psevdomonas-vaktsin.

Partially purified water-soluble cellular protein antigens have been obtained from 2 newly isolated P. aeruginosa strains and 1 museum P. aeruginosa strain, belonging to immunotypes 2, 3 and 7, by the method of preparative ultracentrifugation. Such trivalent P. aeruginosa vaccine (PV) has proved to be effective in direct and cross experiments of the active protection of mice. The method of ultrafiltration has been used to prepare monovalent PV from a newly isolated strain belonging to immunotype 3/7. This monovalent PV has been found to stimulate immunity to infection with homologous or heterologous P. aeruginosa strains in mice. The comparison of the results obtained in the study of PV prepared by the methods of ultracentrifugation and ultrafiltration suggests that both these methods for the isolation and purification of P. aeruginosa protective protein antigens are equally effective.

[Immunostimulating activity of antigenic complexes isolated from various Staphylococcus strains]. / Immunostimuliruiushchaia aktivnost' antigennogo kompleksa stafilokokka, prigotovlennogo iz raznykh shtammov.

The study of the immunostimulating potency of the staphylococcal antigenic complex obtained from different strains by aqueous extraction indicated that the preparations obtained from highly virulent strains showed the lowest potency after challenge with both homologous and heterologous strains. The preparations obtained from strains with low virulence stimulated resistance to highly virulent strains to a greater extent than the preparations from the same virulent strain which was used for challenge. The necessity of revising the requirements to strains used for the production of antibacterial staphylococcal preparations is substantiated.

Somatic antigens of Pseudomonas aeruginosa. The structure of the O-specific polysaccharide chains of Ps.aeruginosa O:2 (Lanyi) lipopolysaccharides.

Structural and immunochemical studies have been performed on the O-specific antigens of two serologically related types of Pseudomonas aeruginosa O:2a,b and O:2a,c (Lanyi's classification). The O-specific polysaccharide chains of the two serotypes were shown to be acidic polysaccharides composed of repeating trisaccharide units consisting of L-rhamnose, N-acetyl-D-quinovosamine and N-acetyl-L-galactosaminuronic acid residues. Based on 1H and 13C nuclear magnetic resonance spectroscopy, data of methylation analysis and selective solvolysis with hydrogen fluoride, the repeating unit of the O:2a,c O-specific polysaccharide was assigned the following structure: leads to 4)LGalNAcA(alpha1 leads to 3)DQuiNAc(alpha1 leads to 3) LRha(alpha1, where GalNAcA = N-acetylgalactosaminuronic acid, QuiNAc = N-acetylquinovosamine and Rha = rhamnose. The O:2a,b O-specific polysaccharide had the same structure of the trisaccharide repeating unit but was distinguished by the presence of O-acetyl groups on 70-80% of the rhamnose residues in position 2. The O-acetyl groups were located both by methylation with methyltrifluoromethane sulphonate and by comparison of the 13C nuclear magnetic resonance spectra of the native and O-deacetylated polysaccharides. Serological studies revealed the determinant role of the O-acetyl groups in the O-antigen O:2a,b and suggested the O-factor 2b to be related to the 2-O-acetyl-L-rhamnose residue, whereas the O-factor 2c was probably determined by the nonacetylated rhamnose residue. The dominant moiety of the determinant 2a common to the two antigens was obviously presented by the N-acetyl-L-galactosaminuronic acid residue.

Somatic Antigens of Pseudomonas aeruginosa. The structure of the polysaccharide chain of Ps.aeruginosa O:6 (Lanyi) lipopolysaccharide.

An O-antigenic lipopolysaccharide, manifesting a high serological activity in the reaction of passive haemagglutination and its inhibition, was isolated from Pseudomonas aeruginosa O:6 cells (Lanyi). Splitting-off of the lipid component by mild acid hydrolysis resulted in the polysaccharide deprived of O-specific activity. The polysaccharide chain of the lipopolysaccharide was made up exclusively of the residues of amino sugars, among which have been identified: 2-acetamido-2-deoxy-D-galactose (GalNAc), 2-acetamido-2,6-dideoxy-D-glucose (QuiNAc), 2-acetamido-2,6-dideoxy-D-galactose (FucNAc) as well as 2,3-diacetamido-2,3-dideoxy-D-glucuronic acid [Glc(NAc)2A] found naturally for the first time. The principal methods of establishing the polysaccharide structure were 13C nuclear magnetic resonance, methylation analysis and solvolysis with hydrogen fluoride. Depending on solvolysis conditions, a disaccharide or a trisaccharide containing the diacetamidouronic acid residue was formed. From the results obtained it followed that the polysaccharide chain of the O-antigenic Ps. aeruginosa O:6 lipopolysaccharide represents an acidic hexasaminoglycan constructed of the repeating tetrasaccharide units of the following structure: leads to 4)DGalNAc(alpha 1 leads to 4)DGlc(NAc)2A(beta 1 leads to 3)DFucNAc(alpha 1 leads to 3)DQuiNAc(alpha 1 leads to.