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Escherichia coli O86 belongs to the enteropathogenic E. coli (EPEC) group, some strains of which are pathogens of humans, wild birds and farm animals. The O-antigen gene cluster of E. coli O86 was amplified by long-range PCR using primers based on the housekeeping genes galF and gnd, and then sequenced. Genes involved in GDP-Fuc and N-acetyl-galactosamine (GalNAc) synthesis and genes encoding glycosyltransferases, O-unit flippase and O-antigen polymerase were identified on the basis of homology. By screening against 186 E. coli and Shigella-type strains, two genes specific to E. coli O86 were identified. A polymerase chain reaction (PCR) assay, based on the specific O-antigen genes identified here, could be used for the rapid detection of E. coli O86 in environmental and clinical samples. The relationship between E. coli O86 and O127 was also determined by comparing the two O-antigen gene clusters.
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PMID:Characterization of Escherichia coli O86 O-antigen gene cluster and identification of O86-specific genes. 1577 30

O-antigens are highly polymorphic. The genes specifically involved in O-antigen synthesis are generally grouped together on the chromosome as a gene cluster. In Escherichia coli, the O-antigen gene clusters are characteristically located between the housekeeping genes galF and gnd. In this study, the O-antigen gene clusters of E. coli O59 and E. coli O155 were sequenced. The former was found to contain genes for GDP-mannose synthesis, glycosyltransferase genes and the O-antigen polymerase gene (wzy), while the latter contained only glycosyltransferase genes and wzy. O unit flippase genes (wzx) were found immediately downstream of the gnd gene, in the region between the gnd and hisI genes in these two strains. This atypical location of wzx has not been reported before, and furthermore these two genes complemented in trans despite the fact that different O-antigen structures are present in E. coli O59 and O155. A putative acetyltransferase gene was found downstream of wzx in both strains. Comparison of the region between gnd and hisI revealed that the wzx and acetyltransferase genes are closely related between E. coli O59 and O155, indicating that the two gene clusters arose recently from a common ancestor. This work provides further evidence for the O-antigen gene cluster having formed gradually, and selection pressure will eventually bring O-antigen genes into a single cluster. Genes specific for E. coli O59 and O155, respectively, were also identified.
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PMID:Characterization of the Escherichia coli O59 and O155 O-antigen gene clusters: the atypical wzx genes are evolutionary related. 1599 Feb 53

Studies based on the analysis of housekeeping genes indicate that Escherichia coli and all Shigella species, except for Shigella boydii type 13, belong to a single species. This study analysed the phenotypic and genotypic characteristics of 23 E. coli strains isolated in different countries from faecal specimens taken from children with diarrhoea. Strains were identified using the VITEK system and typed with rabbit sera obtained against 186 somatic and 53 flagellar E. coli antigens and against 45 Shigella somatic antigens. Biochemical analysis of these strains showed a typical E. coli profile with a defined reaction against both E. coli O179 and S. boydii 16 somatic antisera. Agglutination assays for flagellar antigens showed a response against H2 in 7 (30 %) strains, H10 in 2 (9 %) strains, H32 in 12 (52 %) strains and H34 in 2 (9 %) strains, demonstrating 4 serotypes associated with this new somatic antigen 64474. A serum against one of these E. coli strains (64474) was prepared. Absorption assays of S. boydii 16 and E. coli 64474 antisera with E. coli O179 antigen removed the agglutination response against this O179 antigen completely, while the agglutination titres against both S. boydii 16 and E. coli 64474 remained the same. Four (17 %) E. coli strains showed antimicrobial resistance to piperacillin only, one (4 %) to piperacillin and trimethoprim/sulfamethoxazole, one (4 %) to ciprofloxacin, nitrofurantoin and piperacillin, and two (9 %) strains were resistant to ciprofloxacin, norfloxacin, ofloxacin, piperacillin and trimethoprim/sulfamethoxazole. With regards to PCR assays, one (4 %) of the strains was positive for Shigella gene ipaH, one (4 %) for ipaA, two (9 %) for ipaB, one (4 %) for ipaD, two (9 %) for sepA and three (13 %) for ospF. The rfb gene cluster in the E. coli strains was analysed by RFLP and compared with the gene cluster obtained from S. boydii 16. The rfb-RFLP patterns for all 23 E. coli strains were similar to those obtained for S. boydii 16. The results from PCR tests to detect rfb genes wzx (encoding O unit flippase) and wzy (encoding polymerase) belonging to a cluster related to the biosynthesis of the S. boydii 16-specific O antigen were positive in 21 (91 %) and 22 (96 %) of the strains, respectively. PCR assays to detect E. coli virulence genes were also performed. These assays detected enterotoxigenic E. coli genes ltA1 in 12 of the strains (52 %), st1a in 4 (17 %), cfa1 in 6 (26 %), cs1 in 1 (4 %), cs3 in 3 (13 %), cs13 in 9 (39 %) and cs14 in 5 (22 %) of the strains. Results from the PFGE analyses confirmed the wide geographical distribution of these strains suggesting that 64474 : H2, 64474 : H10, 64474 : H32 and 64474 : H34 are new serotypes of E. coli strains with a defined virulence capacity, and share a common O antigen with S. boydii 16.
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PMID:New enterovirulent Escherichia coli serogroup 64474 showing antigenic and genotypic relationships to Shigella boydii 16. 2007 11

Based on the O-antigens (O-polysaccharides), one of the most variable cell constituents, 46 O-serogroups have been recognized in the Kauffmann-White serotyping scheme for Salmonella enterica. In this work, the structure of the O-polysaccharide and the genetic organization of the O-antigen gene cluster of S. enterica O56 were investigated. As judged by sugar and methylation analyses, along with NMR spectroscopic data, the O-polysaccharide has a linear tetrasaccharide O-unit, which consists of one residue each of d-ribofuranose, N-acetyl-d-glucosamine, N-acetyl-d-galactosamine, and a novel sugar derivative, 4-(N-acetyl-l-seryl)amino-4,6-dideoxy-d-glucose (d-Qui4NSerAc). The following structure of the O-polysaccharide was established: -->3)-beta-d-Quip4NSerAc-(1-->3)-beta-d-Ribf-(1-->4)-alpha-d-GalpNAc-(1-->3)-alpha-d-GlcpNAc-(1--> The O-antigen gene cluster of S. enterica O56 having 12 open reading frames was found between the housekeeping genes galF and gnd. A comparison with databases and using the O-antigen structure data enabled us to ascribe functions to genes for (i) synthesis of d-GalNAc and d-Qui4NSerAc, (ii) sugar transfer, and (iii) O-antigen processing, including genes for O-unit flippase (Wzx) and O-antigen polymerase (Wzy).
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PMID:Structural and genetic characterization of the O-antigen of Salmonella enterica O56 containing a novel derivative of 4-amino-4,6-dideoxy-D-glucose. 2063 49