Separate O-grouping schemes for serotyping clinical isolates of Proteus vulgaris and Proteus mirabilis

et al. 2000

A phosphorylated O-specific polysaccharide (O-antigen) was obtained by mild acid degradation of Proteus vulgaris O12 lipopolysaccharide and studied by sugar and methylation analyses, 1 H-, 13 C-and 31 P-NMR spectroscopy, including two-dimensional COSY, TOCSY, NOESY, H-detected 1 H, 13 C and 1 H, 31 P heteronuclear multiple-quantum coherence experiments. It was found that the polysaccharide consists of pentasaccharide repeating units connected via a glycerol phosphate group, and has the following structure:where FucNAc is 2-acetamido-2,6-dideoxygalactose and the degree of O-acetylation at position 4 of GalNAc is < 25%. Immunochemical studies with P. vulgaris O12 O-antiserum suggested that the lipopolysaccharide studied shares common epitopes with the lipopolysaccharide core of P. vulgaris O8 and with the O-antigens of P. penneri strains 8 and 63.Keywords: bacterial polysaccharide structure; glycerol phosphate; lipopolysaccharide; Proteus vulgaris; serological cross-reactivity.Bacteria of the genus Proteus cause urinary tract infections which can lead to severe complications, such as acute or chronic pyelonephritis and formation of bladder and kidney stones. Outer-membrane lipopolysaccharide (LPS) is considered as a potential virulence factor of Proteus [1,2]. The serological O-specificity of bacterial smooth forms is defined by the structure of the polysaccharide chain of LPS (O-antigen). Based on the O-specificity, two species, P. mirabilis and P. vulgaris, have been classified into 60 O-serogroups [3,4], and a number of additional serogroups have been proposed for the third medically important species, P. penneri [5]. Structures of the O-specific polysaccharides of most Proteus serogroups have been determined and correlated to the immunospecificity of the O-antigens [5]. Most polysaccharides (< 80%) appeared to be acidic due to the presence of uronic acids and various noncarbohydrate substituents, including phosphate groups. Numerous serological relationships between Proteus strains from different serogroups have been reported and accounted for by the presence of common epitopes on LPS [5]. Now we report the structure of another phosphorylated O-specific polysaccharide from P. vulgaris O12 LPS and its serological relatedness to LPS of some other Proteus strains. M A T E R I A L S A N D M E T H O D SBacterial strain, isolation and degradation of LPS P. vulgaris O12 (strain PrK 25/57) was from the Czech National Collection of Type Cultures (Institute of Epidemiology and Microbiology, Prague). The bacterium was grown as described [6]. LPS was isolated from dried bacterial cells by phenol/water extraction [7] and purified by treatment with DNAse and RNAse as described [8].Delipidation of LPS (100 mg) was performed with aqueous 2% HOAc at 100 8C until the lipid precipitated. The precipitate was removed by centrifugation (13 000 g, 20 min), and the supernatant was fractionated on a Sephadex G-50 (S) column (Pharmacia) in 0.05 m pyridine acetate pH 4.5, monitored by a Knauer differential refractometer (Germany). A hig...

Section: Serological Studies With Proteus Lpssupporting

confidence: 78%

mentioning

confidence: 99%

Structure of a glycerol teichoic acid‐like O‐specific polysaccharide of Proteus vulgaris O12

Perepelov

Torzewska

et al. 2000

“…Outer membrane lipopolysaccharide (LPS, endotoxin) is considered as a virulence factor of Proteus. Based on the Ospecific polysaccharide chains of LPS (O-antigens), two species, Proteus mirabilis and Proteus vulgaris, were classified into 60 O-serogroups [1,2], and recently two more O-serogroups, O61 and O62, have been proposed for strains of the third species, Proteus penneri [3,4]. Chemical and immunochemical studies of Proteus LPS are important for understanding the molecular basis of the immunospecificity of Proteus strains and for their classification based on structural and serological data.A peculiar feature of the Proteus LPS is that in most Oserogroups the O-antigens are acidic due to the presence of phosphate groups, hexuronic acids, their amides with amino acids, sugar acetals with pyruvic acid or ethers with lactic acid ([3Ϫ9] and references cited therein).…”

mentioning

confidence: 99%

Structural and serological studies on a new acidic O‐specific polysaccharide of Proteus vulgaris O32

Babicka

Грачев

et al. 1998

The following structure of the O-specific polysaccharide chain (O-antigen) of the Proteus vulgaris O32 lipopolysaccharide (LPS) was established by 1 H-NMR and 13 C-NMR spectroscopy, including twodimensional NOESY and H-detected 1 H, 13 C heteronuclear multiple-quantum coherence (HMQC) experiments:In addition, an O-acetyl group was detected, which, most probably, is located at position 3 of a part of Rhap I residues. Serological studies, using rabbit polyclonal anti-(P. vulgaris O32) serum, homologous and heterologous Proteus O-antigens and related artificial antigens, revealed the importance of an A-D-GalA-associated epitope in manifesting the immunospecificity of P. vulgaris O32 and substantiated serological relationships between the O-antigen studied and those of some other Proteus strains.Keywords : Proteus vulgaris ; lipopolysaccharide; O-antigen; bacterial polysaccharide structure; serological specificity.Bacteria of the genus Proteus cause mainly wound and urinary tract infections, the latter sometimes leading to acute or chronic pyelonephritis and formation of bladder and kidney stones. Outer membrane lipopolysaccharide (LPS, endotoxin) is considered as a virulence factor of Proteus. Based on the Ospecific polysaccharide chains of LPS (O-antigens), two species, Proteus mirabilis and Proteus vulgaris, were classified into 60 O-serogroups [1, 2], and recently two more O-serogroups, O61 and O62, have been proposed for strains of the third species, Proteus penneri [3,4]. Chemical and immunochemical studies of Proteus LPS are important for understanding the molecular basis of the immunospecificity of Proteus strains and for their classification based on structural and serological data.A peculiar feature of the Proteus LPS is that in most Oserogroups the O-antigens are acidic due to the presence of phosphate groups, hexuronic acids, their amides with amino acids, sugar acetals with pyruvic acid or ethers with lactic acid ([3Ϫ9] and references cited therein). The role of acidic polysaccharides in the pathogenicity of Proteus and, particularly, in urinary tract infections was discussed [10]. The acidic character of Proteus polysaccharides enables bacteria to bind metal cations, Mg 2ϩ and Ca 2ϩ , via electrostatic interaction. This may enhance the formation of struvite and carbonate apatite stones in urinary tract under alkaline conditions in the presence of ammonia, one of the products of urea decomposition caused by Proteus bacteria [11]. Recently, the importance of an acidic polysaccharide rich in galacturonic acid and galactosamine was demonstrated for migration of P. mirabilis swarm cells by reduction of surface friction [12].A few structures of P. vulgaris O-antigens have been elucidated, some of them having been found neutral [7,13] and the others acidic [7Ϫ9]. Now, we report the structure of a new acidic O-specific polysaccharide of P. vulgaris O32 containing D-galacturonic acid. MATERIALS AND METHODSBacterial strains, growth, isolation and degradation of lipopolysaccharide. P. vulgaris O32 (strain PrK 57/...

“…According to the serological classification based on the specificity of the cultures as described [10]. Crude lipopolysaccharide preparations, obtained after extraction of bacterial mass with a hot phe-outer-membrane lipopolysaccharide (LPS, O-antigen), Proteus mirabilis and Proteus vulgaris strains are divided into 60 O-nol/water mixture [11], were purified by treatment with cold serogroups [1,2]. aqueous 50% CCl 3CO2H followed by dialysis of the superna-Recently the name Proteus penneri has been proposed for tant.…”

mentioning

confidence: 99%

Structure and cross‐reactivity of the O‐specific polysaccharide of Proteus penneri strain 26, another neutral Proteus O‐antigen containing 2‐acetamido‐2,6‐dideoxy‐ L‐glucose (N‐acetyl‐ L‐quinovosamine)

Arbatsky

Widmalm

et al. 1998

A neutral O-specific polysaccharide obtained from the lipopolysaccharide of Proteus penneri strain 26 was studied using sugar analysis and 1 H and 13 C NMR spectroscopy, including two-dimensional NMR techniques. The following structure of the trisaccharide repeating unit was established: →6)-A-DGlcpNAc-(1→3)-A-L-QuipNAc-(1→3)-A-D-GlcpNAc-(1→ where L-QuiNAc is 2-acetamido-2,6-dideoxy-L-glucose (N-acetyl-L-quinovosamine). Cross-reactivity of the Proteus penneri 26 anti-O serum with other strains of P. penneri isolated in Poland and USA and one strain of P. vulgaris is discussed.Keywords : Proteus penneri; O-antigen ; bacterial polysaccharide; lipopolysaccharide ; 2-acetamido-2,6-dideoxy-L-glucose.P. penneri strain 62 was isolated in Poland and P. vulgaris Bacteria of the genus Proteus are a common cause of urinary tract infections which may result in severe complications, such OX2 was purchased from the Czechoslovak National Collection of Type Cultures. Dry bacteria were obtained from aerated liquid as pyelonephritis and formation of kidney stones. According to the serological classification based on the specificity of the cultures as described [10]. Crude lipopolysaccharide preparations, obtained after extraction of bacterial mass with a hot pheouter-membrane lipopolysaccharide (LPS, O-antigen), Proteus mirabilis and Proteus vulgaris strains are divided into 60 O-nol/water mixture [11], were purified by treatment with cold serogroups [1,2]. aqueous 50% CCl 3CO2H followed by dialysis of the supernaRecently the name Proteus penneri has been proposed for tant. Alkali-treated LPSs (LPSs-OH) were prepared by saponifistrains formerly described as Proteus vulgaris biogroup I [3, 4]. cation of LPSs with 0.25 M sodium hydroxide (56°C, 2 h), folThese strains have not been serologically classified and chemical lowed by precipitation with ethanol. LPS was treated with aqueand immunochemical studies of their outer-membrane lipopoly-ous 1% acetic acid at 100°C for 2 h. Products were fractionated saccharides are being undertaken [5Ϫ9] (and references cited by gel-permeation chromatography on a colunm (3ϫ65 cm) of therein) in order to create the basis for such a classification. As Sephadex G-50 in 0.03 M pyridinium acetate pH 5.4 as eluent, a result, the new, separate serogroups O61 and O62 have been monitored using a Knauer differential refractometer, to give a identified which include strains with the O-antigen structures high-molecular-mass O-specific polysaccharide. most widespread among P. penneri strains [6,9]. In continuation Rabbit antiserum. Rabbits were immunized with heatof these studies, we now report the structure of the O-specific killed bacteria of P. penneri 26 as follows. Animals received polysaccharide and cross-reactivity of the P. penneri strain 26. intravenously 50, 100, 100 and 200 µg of heat-killed bacteria in 0.5 ml NaCl/P i (0.015 M Na 2 HPO 4 , 0.15 M NaCl, pH 7.2) on day 0, 4, 7 and 11, respectively. At day 16 blood (20 ml) were MATERIALS AND METHODS obtained from the ear vein (IgM-rich antiserum). The...