Occurrence of 3-Amino-3,6-Dideoxyhexoses in Salmonella and Related Bacteria

Abstract

Several species of Salmonella, Citrobacter, and Arizona were examined for the presence of 3-amino sugars, which were isolated from lipopolysaccharide hydrolysates by cation exchange chromatography and identified by paper and cation exchange chromatography in several systems and by specific colorimetric procedures. 3-Amino-3 ,6-dideoxyglucose was identified in C. freundii 8090, C. freundii 869, S. halle, S. telaviv, S. dakar, S. wandsworth, and S. champaign; 3-amino-3 ,6-dideoxygalactose was found in S. tranoroa and Arizona 24. Evidence suggests that these 3amino sugars occur in lipopolysaccharides as the N-acetyl derivatives. The composition of the lipopolysaccharides containing the 3-amino sugars was determined, and the lipopolysaccharides were compared serologically in order to correlate chemotypes with serotypes. The lipopolysaccharides containing 3-amino-3, 6dideoxyglucose reacted with serogroups 28 or 39; those containing 3-amino-3,6dideoxygalactose were found to react with group 55 antisera. The preparation of a lipopolysaccharide from the phenol phase of the 44% aqueous phenol extraction procedure is also reported. Ashwell and Volk (2) first reported the isolation and characterization of crystalline 3-acetamido-3,6-dideoxy-D-galactose, obtained from a phenol-soluble lipopolysaccharide prepared from Xanthomonas campestris, a member of the Pseudomonadaceae. Raff and Wheat (18) found a different 3-amino-3 ,6-dideoxy sugar in phenol-soluble cell wall lipopolysaccharide of the enterobacterium, Citrobacter freundii ATCC 8090. Jann et al. (Nature, in press) and Muller Seitz (Thesis, University of Freiburg,Germany) independently identified 3-amino-3, 6-dideoxy-D-galactose and 3-amino-3, 6-dideoxyglucose in Escherichia coli serotypes. The present work describes the tentative identification of 3-amino sugars found in the serologically cross-reactive C. freundii 8090, Citrobacter 869, Salmonella dakar, S. telaviv, S. champaign, S. wandsworth, S. tranoroa, and Arizona 24. MATERIALS AND METHODS Growth of cells. C. 1reundii ATCC 8090 was grown in 10-liter lots in 5-gal (18.9-liter) carboys at 25 C for 24 hr with constant aeration. The medium used was as follows, in grams per liter: yeast extract, 3; K2PHO4, 6.6; KH2PO4, 1.7; (NH4)2SO4, 2.5; MgSO4, 0.05; and glucose (autoclaved separately), 10. Cells were harvested with a DeLaval Gyrotest centrifuge,

Carbohydrate Composition of the Phenol-soluble Lipopolysaccharides of Citrobacter freundii

“…In the general application of the 45 % aqueousphenol fractionation procedure which yields phenol and aqueous soluble fractions (28,29), lipopolysaccharides are isolated from the aqueous soluble phase, and the phenol fractions are usually discarded. Recently, however, phenolsoluble lipopolysaccharides have been reported to occur in Xanthomonas campestris (1,11) and in several Citrobacter species (12,17,18; Raff and Wheat, Federation Proc. 25:410, 1966; 26:281, 1967).…”

“…Neither of the X. campestris lipopolysaccharides contained heptose, and the phenol-soluble lipopolysaccharide also lacked 2-keto-deoxyoctonic acid (KDO; 11). The phenol-soluble lipopolysaccharide from C. freundii 8090 has been shown to contain 2-acetamido-2-deoxy-D-glucose and 3-acetamido-3,6-dideoxy-D-glucose (12,17,18). The present report presents results of studies on the preparation and composition of the C. freundii 8090 polysac-1U.S.…”

Immunochemistry of Group A and Inaba C Antigen Factors Constituting the O Antigen of Ol Vibrio cholerae

“…LPS and PS were deacylated according to the method of Caroff et al [14] in 1 M NaOH at 100°C for 2 h; the products were recovered by freeze drying after dialysis against distilled water. The deacylated LPS and PS were N-acetylated according to the method of Luderitz et al [24]; they were Npropionylated or N-butyrylated by incubating in 50 mM sodium bicarbonate pH 8.3 at 25°C for 16 h with active esters of propionic or butyric acid, respectively, prepared according to the method of Seid and Sadoff [25]. The N-propionylated and Nbutyrylated LPS and PS, obtained by freeze drying after dialysis against distilled water, were again treated with the active esters as described above.…”

Immunochemical studies of SalmonellaDakar and SalmonellaTelaviv O-antigens (serogroup O:28)

“…These results confirmed that 4-linked D-galactose was not responsible for O28 1 -antigen specificity and that this subfactor was attributed to the presence of 3-linked (or 3,4-disubstituted) a-D-GalpNAc in the main chains of S. Dakar and S. Telaviv O-polysaccharides. Lüderitz et al (1967) also supposed that the presence of O28 1 was correlated with the presence of N-galactosamine, the presence of O28 2 with ribose, and the presence of O28 3 with rhamnose, but these conclusions were not confirmed by chemical and immunochemical studies. (Table 2).…”

“…A control test should give a negative cross-reaction with bacteria belonging to serogroups C 1 , C 2 , D (12 2 + ), O:30, O:35 and O:39. Literature data on the serological and immunological properties of this serogroup are very limited. Lüderitz et al studied the cross-reactions of S. Dakar and S. Telaviv with Citrobacter freundii 8090 and Citrobacter freundii 869 using unabsorbed and absorbed test sera (Lüderitz et al, 1967;Keleti & Lüderitz, 1971). They observed that Citrobacter freundii 8090 cross-reacts only with sera containing antibodies against 28 3 factor, whereas Citrobacter freundii 869 behaves like S. Dakar, cross-reacting with those sera containing antibodies against factor either 28 1 or 28 3 .…”