Loss of resistance to ingestion and phagocytic killing by O(-) and K(-) mutants of a uropathogenic Escherichia coli O75:K5 strain.
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To determine the importance of the O75 O antigen and the K5 capsular antigen in resistance to phagocytosis and phagocytic killing, we used previously described O75(-) and K5(-) mutants from an O75(+) K5(+) wild-type uropathogenic Escherichia coli strain in phagocytosis assays with polymorphonuclear leukocytes (PMNs) and monocytes. At a 10-to-1 ratio of bacteria to phagocytes and in the presence of 10% serum, the parental strain GR-12 was resistant to both PMNs and monocytes over a 2-h incubation period. The O75(-) and K5(-) mutants were similar in sensitivity to killing by both PMNs and monocytes, decreasing in viability by 80% in the first hour. Yet, a significant difference in killing between the O75(-) and K5(-) mutants was observed in the first 15 min of incubation. The K5(-) mutant decreased in numbers by almost 60%, while the O75(-) mutant increased in numbers similarly to GR-12 in the first 15 min. The difference in killing was found not to be due to the rate of opsonization. To further determine the mechanism of resistance, a fluorescence assay was used to differentiate attached and internalized bacteria. The K5 capsule hindered the association of both the wild-type strain and the O75(-) mutant in the initial incubation time with PMNs. In conclusion, both the K5 capsule and O75 O antigen play crucial roles in resistance to phagocytosis over time. (+info)
Genetic basis for lipopolysaccharide O-antigen biosynthesis in bordetellae.
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Bordetella bronchiseptica and Bordetella parapertussis express a surface polysaccharide, attached to a lipopolysaccharide, which has been called O antigen. This structure is absent from Bordetella pertussis. We report the identification of a large genetic locus in B. bronchiseptica and B. parapertussis that is required for O-antigen biosynthesis. The locus is replaced by an insertion sequence in B. pertussis, explaining the lack of O-antigen biosynthesis in this species. The DNA sequence of the B. bronchiseptica locus has been determined and the presence of 21 open reading frames has been revealed. We have ascribed putative functions to many of these open reading frames based on database searches. Mutations in the locus in B. bronchiseptica and B. parapertussis prevent O-antigen biosynthesis and provide tools for the study of the role of O antigen in infections caused by these bacteria. (+info)
Serotype 1a O-antigen modification: molecular characterization of the genes involved and their novel organization in the Shigella flexneri chromosome.
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The factors responsible for serotype 1a O-antigen modification in Shigella flexneri were localized to a 5.8-kb chromosomal HindIII fragment of serotype 1a strain Y53. The entire 5.8-kb fragment and regions up- and downstream of it (10.6-kb total) were sequenced. A putative three-gene operon, which showed homology with other serotype conversion genes, was identified and shown to confer serotype 1a O-antigen modification. The serotype conversion genes were flanked on either side by phage DNA. Multiple insertion sequence (IS) elements were located within and upstream of the phage DNA in a composite transposon-like structure. Host DNA homologous to the dsdC and the thrW proA genes was located upstream of the IS elements and downstream of the phage DNA, respectively. The sequence analysis indicates that the organization of the 10.6-kb region of the Y53 chromosome is unique and suggests that the serotype conversion genes were originally brought into the host by a bacteriophage. Several features of this region are also characteristic of pathogenicity islands. (+info)
Structure of the O-polysaccharide of the lipopolysaccharide of Pseudomonas syringae pv. garcae ICMP 8047.
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The composition and structure of the O-polysaccharide of the lipopolysaccharide of Pseudomonas syringae pathovar garcae ICMP 8047 were studied using methylation analyses, Smith degradation, and 1H- and 13C-NMR spectroscopy, including two-dimensional correlation spectroscopy (COSY), total correlation spectroscopy (TOCSY), nuclear Overhauser effect spectroscopy (NOESY), and H-detected 1H,13C heteronuclear multiple-quantum coherence (HMQC) experiments. The polysaccharide was found to contain L-rhamnose and 3-acetamido-3, 6-dideoxy-D-galactose (D-Fuc3NAc) in the ratio 4:1 and to consist of two types of pentasaccharide repeating units. The major (1) and minor (2) repeating units differ from each other only in the position of substitution of one of the rhamnose residues in the main chain. Similar structural heterogeneity has been reported formerly in O-polysaccharides of some other P. syringae strains having a similar monosaccharide composition. A Fuc3NAc residue is attached to the main rhamnan chain as a side chain by a (alpha1-->4) glycosidic linkage; this has not hitherto been described in P. syringae: [figure]. (+info)
The structure of the colony migration factor from pathogenic Proteus mirabilis. A capsular polysaccharide that facilitates swarming.
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Swarming by Proteus mirabilis is characterized by cycles of rapid and coordinated population migration across surfaces following differentiation of vegetative cells into elongated hyperflagellated swarm cells. It has been shown that surface colony expansion by the swarm cell population is facilitated by a colony migration factor (Cmf), a capsular polysaccharide (CPS) that also contributes to the uropathogenicity of P. mirabilis (Gygi, D., Rahman, M. M., Lai, H.-C., Carlson, R., Guard-Petter, J., and Hughes, C. (1995) Mol. Microbiol. 17, 1167-1175). In this report, the Cmf-CPS was extracted with hot water, precipitated with ethanol, and further purified by gel permeation chromatography. Its structure was established by glycosyl composition and linkage analyses, and by one- and two-dimensional NMR spectroscopy. The Cmf-CPS is composed of the following tetrasaccharide repeating unit. [see text] (+info)
Value of a single-tube widal test in diagnosis of typhoid fever in Vietnam.
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The diagnostic value of an acute-phase single-tube Widal test for suspected typhoid fever was evaluated with 2,000 Vietnamese patients admitted to an infectious disease referral hospital between 1993 and 1998. Test patients had suspected typhoid fever and a blood culture positive for Salmonella typhi (n= 1,400) or Salmonella paratyphi A (n = 45). Control patients had a febrile illness for which another cause was confirmed (malaria [n = 103], dengue [n = 76], or bacteremia due to another microorganism [n = 156] or tetanus (n = 265). An O-agglutinin titer of >/=100 was found in 18% of the febrile controls and 7% of the tetanus patients. Corresponding values for H agglutinins were 8 and 1%, respectively. The O-agglutinin titer was >/=100 in 83% of the blood culture-positive typhoid fever cases, and the H-agglutinin titer was >/=100 in 67%. The disease prevalence in investigated patients in this hospital was 30.8% (95% confidence interval, 26.8 to 35.1%); at this prevalence, an elevated level of H agglutinins gave better positive predictive values for typhoid fever than did O agglutinins. With a cutoff titer of >/=200 for O agglutinin or >/=100 for H agglutinin, the Widal test would diagnose correctly 74% of the blood culture-positive cases of typhoid fever. However, 14% of the positive results would be false-positive, and 10% of the negative results would be false-negative. The Widal test can be helpful in the laboratory diagnosis of typhoid fever in Vietnam if interpreted with care. (+info)
Structures of the O-specific polysaccharides and a serological cross-reactivity of the lipopolysaccharides of Proteus mirabilis O24 and O29.
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Strains of Proteus mirabilis belonging to serogroups O24 and O29 are frequent in clinical specimens. Anti-P. mirabilis O24 serum cross-reacted with the lipopolysaccharide (LPS) of P. mirabilis O29 and vice versa. The structures of the O-specific polysaccharides (OPSs, O-antigens) of both LPSs were established using sugar analysis and one- and two-dimensional 1H- and 13C-NMR spectroscopy and found to be different. SDS-PAGE and Western immunoblotting suggested that the serological cross-reactivity of the LPSs is due to a common epitope(s) on the core-lipid A moiety, rather than on the OPS. Therefore, the epitope specificity and the structures of the O-antigens studied are unique among Proteus serogroups. (+info)
Structure determination of the O-antigenic polysaccharide from the enteroinvasive Escherichia coli O136.
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The structure of the O-antigen polysaccharide of the lipopolysaccharide from the enteroinvasive Escherichia coli O136 has been elucidated. The composition of the repeating unit was established by sugar and methylation analysis together with 1H and 13C NMR spectroscopy. Two-dimensional nuclear Overhauser effect spectroscopy (NOESY) and heteronuclear multiple-bond correlation experiments were used to deduce the sequence. The absolute configuration for the nonulosonic acid (NonA) could be determined using spin-spin coupling constants, 13C chemical shifts and NOESY. The anomeric configuration of the NonA was determined via vicinal and geminal 13C,1H coupling constants. The structure of the repeating unit of the polysaccharide from E. coli O136 is as follows, in which beta-NonpA is 5,7-diacetamido-3,5,7, 9-tetradeoxy-Lglycero-beta-Lmanno-nonulosonic acid: -->4)-beta-NonpA-(2-->4)-beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1--> (+info)