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Shigella is an important human pathogen and is closely related to Escherichia coli. O-antigen is the most variable part of the lipopolysaccharide on the cell surface of Gram-negative bacteria and plays an important role in pathogenicity. The O-antigen gene cluster of S. boydii O1 was sequenced. The putative genes encoding enzymes for rhamnose synthesis, transferases, O-unit flippase, and O-unit polymerase were identified on the basis of homology. The O-antigen gene clusters of S. boydii O1 and E. coli O149, which share the same O-antigen form, were found to have the same genes and organization by adjacent gene PCR assay. Two genes specific for S. boydii O1 and E. coli O149 were identified by PCR screening against E. coli- and Shigella-type strains of the 186 known O-antigen forms and 39 E. coli clinical isolates. A PCR sensitivity of 103 to 104 CFU/mL overnight culture of S. boydii O1 and E. coli O149 was obtained. S. boydii O1 and E. coli O149 were differentiated by PCR using lacZ- and cadA-based primers.

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.

The O antigens found in Salmonella enterica (Se) and Escherichia coli (Ec) show a great deal of diversity, and only three structures are known to be common to both genera. Two of them contain the 3,6-dideoxyheoxse colitose, not found in other serogroups of the two species. The first of these is common to Ec O111 and Se O:35 (sv Adelaide); the other is found in both Ec O55 and Se O:50 (sv Greenside). The genes specific for the synthesis of O antigen are generally located in the rfb gene cluster at map position 45 min in Ec and 42 min in Se. The rfb (O antigen) gene cluster of an Ec O111 strain M92 had been cloned earlier and hybridisation analysis suggested that the rfb clusters of Ec M92 and a Se sv Adelaide strain had been acquired separately by the two species since their divergence. We have now sequenced part of the rfb cluster from Ec M92. We identify two genes of the GDP-colitose pathway, rfbM and rfbK, and show that several other ORFs have similarity to the rfb and cps (capsular polysaccharide) genes. Downstream of this block of genes is an ORF which encodes a protein with predicted transmembrane segments which is presumed to correspond to the rfbX gene. The % G+C values of the Ec M92 rfb sequence are extremely low, indicating that the rfb evolved in a low % G+C species of bacteria before transfer into Ec.

The O antigen of Escherichia coli O111 is identical in structure to that of Salmonella enterica serovar adelaide. Another O-antigen structure, similar to that of E. coli O111 and S. enterica serovar adelaide is found in both E. coli O55 and S. enterica serovar greenside. Both O-antigen structures contain colitose, a 3,6 dideoxyhexose found only rarely in the Enterobacteriaceae. The O-antigen structure is determined by genes generally located in the rfb gene cluster. We cloned the rfb gene cluster from an E. coli O111 strain (M92), and the clone expressed O antigen in both E. coli K-12 and a K-12 strain deleted for rfb. Lipopolysaccharide analysis showed that the O antigen produced by strains containing the cloned DNA is polymerized. The chain length of O antigen was affected by a region outside of rfb but linked to it and present on some of the plasmids containing rfb. The rfb region of M92 was analysed and compared, by DNA hybridization, with that of strains with related O antigens. The possible evolution of the rfb genes in these O antigen groups is discussed.