MOLECULAR CHARACTERIZATION OF PATHOGENS AND THEIR RESPONSES TO ENVIRONMENTAL FACTORS
Location: Molecular Characterization of Foodborne Pathogens
Title: Molecular Characterization of the Escherichia coli Serogroup O117, O126, and O146 O Antigen Gene Clusters and Development of PCR Assays Targeting Serogroup O117-, O126-, and O146-Specific DNA Sequences
Submitted to: Molecular and Cellular Probes
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 8, 2007
Publication Date: March 14, 2007
Citation: Liu, Y., Debroy, C., Fratamico, P.M. 2007. Molecular Characterization of the Escherichia coli Serogroup O117, O126, and O146 O Antigen Gene Clusters and Development of PCR Assays Targeting Serogroup O117-, O126-, and O146-Specific DNA Sequences. Molecular and Cellular Probes. 21:295-302.
Interpretive Summary: The bacterium, Escherichia coli, causes a variety of diseases in humans and animals, and non-harmful E. coli types, referred to as serogroups, also exist. Traditionally, a procedure called serotyping is used to distinguish among the ca. 165 different E. coli serogroups. This procedure relies on the use of antibodies raised in rabbits against different surface polysaccharides of the bacteria. Serotyping, however, can generally only be performed in specialized laboratories, is labor intensive and may require several days to complete, and one antiserum can react with multiple E. coli serogroups, rendering identification of the specific strain of E. coli difficult. Thus, due to the lack of simple, rapid, and reliable methods for detection and identification of harmful and non-harmful E. coli types, the incidence of disease caused by harmful strains of E. coli may be underestimated, and epidemiological studies are difficult to perform. To develop a more rapid and simple method for detection and typing of the E. coli serogroups O117, O126, and O146, which have caused serious diseases in humans and animals, the DNA sequence of a cluster of genes involved in the production of specific surface polysaccharides of E. coli O117, O126, and O146 was determined. Genes required for biosynthesis of the surface polysaccharides in each of the serogroups were identified, and multiplex polymerase chain reaction (PCR) assays were developed targeting unique DNA sequences in two genes present in each of these serogroups. The PCR assays were serogroup specific when tested against a variety of different bacterial strains. Thus, the use of the E. coli O117-, O126, and O146-specific PCR assays facilitates the ability to identify, detect, and type these bacteria, and potentially can eliminate the use of the labor-intensive serotyping procedure.
The O antigen gene clusters of Escherichia coli serogroups O117, O126, and O146 were sequenced, and eleven, ten, and eleven open reading frames (ORFs) were identified, respectively. Genes required for O antigen sugar biosynthesis, sugar transfer and sugar processing in these O-antigen gene clusters were identified. Multiplex polymerase chain reaction (PCR) assays were developed targeting the wzx and wzy genes present in the O antigen gene cluster of these serogroups. The assays were highly serogroup specific when tested against strains belonging to serogroups that were isolated from food, humans, animals and/or environmental sources, as well as against representative strains belonging to ca. 165 Escherichia coli O serogroups and a number of non-E. coli bacteria. Thus, the results demonstrate that the wzx and wzy gene sequences were specific to E. coli O117, O126, and O146 and can be used as diagnostic markers for rapid identification and detection of these serogroups.