P3-41 Using a Repetitive Sequence-based PCR System for Molecular Characterization of Shiga Toxin-producing Escherichia coli

Wednesday, July 31, 2013
Exhibit Hall (Charlotte Convention Center)
Kimberly Anderson, U.S. Food and Drug Administration, Alameda, CA
Shaohua Zhao, U.S. Food and Drug Administration, Laurel, MD
Eileen Liu, U.S. Food and Drug Administration, Alameda, CA
Sunee Himathongkham, U.S. Food and Drug Administration, Rockville, MD
Ruiqing Pamboukian, U.S. Food and Drug Administration, Rockville, MD
Introduction: Shiga toxin-producing Escherichia coli(STEC) are of major public health concern due to their frequent contamination of food products and ability to trigger foodborne illnesses such as diarrhea, hemorrhagic colitis, and life-threatening hemolytic uremic syndrome.

Purpose: In this study, we used the DiversiLab rep-PCR system to generate a digital DNA fingerprint profile for 152 STEC isolates (63-O157 and 89-Non-O157) from human, food, and animal sources. 

Methods: STEC isolates were collected from Michigan State University (n=104), as well as the Center for Veterinary Medicine (n = 48) within the FDA.  Fourteen serogroups were analyzed, including 5 of the “Big 6” E. coliserogroups, using the Kullback-Leibler method for similarity calculation.

Results: Excellent reproducibility of rep-PCR profiles was observed with % values ranging from 98.4%-99.7% and 97%-99.8% for intra-run and inter-run studies, respectively.  A comparison of Non-O157 isolates revealed that 79/89 (88.9%) isolates clustered according to serogroup status, and peak differences among the Non-O157 isolates ranged from 2 peak differences (94.3% similarity) up to 12 peak differences (59.6% similarity).  Furthermore, the dendrogram of STEC serogroups created from rep-PCR profiles mirrors the distinct clonal groups elucidated for STEC strains by other investigators.  For our panel of O157 isolates, 58/63 (92.1%) yielded a DNA banding pattern typical of the O157:H7 rep-PCR pattern type.  Interestingly, 3 of the 5 O157 STEC that did not exhibit an O157:H7 rep-PCR pattern type were isolated from asymptomatic individuals.  Rep-PCR profiles for STEC isolates were also distinct from 12 other bacterial species, which included S. flexneri, L. monocytogenes, V. cholera, K. pneumoniae, and Salmonella enterica.  Together, these results indicate the ability of the DiversiLab rep-PCR system to distinguish between and within O157 and Non-O157 STEC serogroups.

Significance: Rapid PCR-based typing methods, like repetitive sequence-based PCR (rep-PCR), may facilitate the identification of STEC by complimenting the discriminatory power of PFGE.