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United States Department of Agriculture

Agricultural Research Service

Research Project: MOLECULAR CHARACTERIZATION OF PATHOGENS AND THEIR RESPONSES TO ENVIRONMENTAL FACTORS

Location: Molecular Characterization of Foodborne Pathogens

Title: Comparison of the H7 latex agglutination test with a fliCh7 real-time PCR assay for confirmation of the H type of Escherichia coli O157:H7

Authors
item Narang, Neelam
item Fratamico, Pina
item Tilman, Glenn -
item Pupedis, Kitty -
item Cray Jr, William

Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 11, 2009
Publication Date: October 1, 2009
Citation: Narang, N., Fratamico, P.M., Tilman, G., Pupedis, K., Cray Jr, W.C. 2009. Comparison of the H7 latex agglutination test with a fliCh7 real-time PCR assay for confirmation of the H type of Escherichia coli O157:H7. Journal of Food Protection. 72(10):2195-2197.

Interpretive Summary: Escherichia coli O157:H7 is an important food-borne pathogenic bacterium that causes serious diseases, including hemorrhagic colitis and hemolytic uremic syndrome. Rapid detection and accurate identification of this pathogen is critical to prevent contaminated food from reaching the consumer. Positive identification of E. coli O157:H7 isolated from food is made using biochemical tests to identify the bacterium as E. coli and by a test known as latex agglutination using antibodies that react with the H7 component of the bacterium. However, some E. coli O157:H7 strains isolated from food give a false negative reaction for the H7 component, and thus, these strains cannot be identified as O157:H7. Therefore, a more accurate and reliable test to identify H7 is needed. A commonly used H7 latex agglutination test was compared with a polymerase chain reaction assay (PCR), which targets the gene (fliCh7) that is responsible for production of the H7 component for identification of H7 in E. coli O157:H7 strains. The PCR assay showed 100% agreement with the H7 status reported for reference strains and E. coli O157:H7 meat isolates; however, 42% (42/100) of the E. coli O157:H7 meat isolates were not identified as H7 by the latex agglutination test. In summary, the PCR test described in this paper, which targets the E. coli fliCh7 gene, can be used to rapidly identify strains that possess the H7 component and to confirm E. coli O157:H7 isolates that show a negative H7 agglutination reaction. Accurate identification of E. coli O157:H7 strains recovered from food will prevent food contaminated with this pathogen from reaching the consumer.

Technical Abstract: Escherichia coli O157:H7 is a food-borne pathogen that causes hemorrhagic colitis and hemolytic uremic syndrome. Positive identification of E. coli O157:H7 is made using biochemical tests and latex agglutination using specific antisera. However, under certain conditions, some E. coli O157:H7 isolates can appear to be non-reactive with H7 antisera and may require multiple passages on motility medium to restore H7 antigenicity. In the present study, we compared latex agglutination with a real-time PCR test to detect the presence of the H7 antigen or the fliCh7 gene, respectively, in E. coli O157 isolates. One hundred and twenty-six E. coli strains were tested including reference strains and strains isolated from meat. Four non-E. coli strains were also tested. All strains were analyzed in parallel by a commercial latex agglutination test and by real-time PCR using primers and a probe targeting the fliCh7 gene. The real-time PCR assay targeting the E. coli fliCh7 gene showed 100% agreement with the H7 status reported for reference strains and E. coli O157:H7 meat isolates. The latex agglutination test results agreed with the H7 status reported for the E. coli O157:H7 reference strains and non-O157, H7 strains, except for one, E. coli O117:H7; however, 42% (42/100) of the E. coli O157:H7 meat isolates tested negative for the H7 antigen by latex agglutination. Therefore, the real-time PCR test can be used to confirm E. coli O157:H7 strains that show negative H7 agglutination test results.

Last Modified: 7/23/2014