Identification of Escherichia coli O-serogroups by restrictionof the amplified O-antigen gene cluster ( rfb-RFLP)

Abstract

The precise serotyping of clinical Escherichia coli isolates is a crucial step for diagnostic and epidemiological purposes. Epidemiological knowledge associated with serotyping is so important that no alternative method may be considered if it does not correlate with serotyping. Unfortunately, E. coli are difficult to serotype. Genes specifically involved in O-antigen synthesis are clustered in E. coli, Shigella and Salmonella. Published oligonucleotide sequences complementary to JUMPstart and the gnd gene (the conserved flanking sequences upstream and downstream of O-antigen gene clusters, respectively) were used to amplify the O-antigen gene cluster of representative strains of 148 E. coli O-serogroups. A unique amplified fragment was observed for each serogroup (size ranging from 1.7 to 20 kbp). Clearly identifiable and reproducible O-patterns were obtained for the great majority of O-serogroups after MboII digestion of amplified products. The number of bands composing each pattern varied from five to 25. A database was built with the patterns obtained. A total of 147 O-patterns were obtained. Thirteen O-serogroups were subdivided into different O-patterns. However, each of 13 other O-patterns was shared by two or more O-serogroups. O-serogroups of clinical isolates were deduced accurately from O-patterns in all cases, even for some rough or nonagglutinating isolates. The restriction method ( rfb-RFLP) may prove to be better than serotyping since 100% of strains are typable, which is not the case with serotyping.