P-061. Phenotypic and Genotypic Characterization of Escherichia coli O157:H7 Isolates from 2006 Outbreaks Linked to Fresh Produce

B. Li, S. Gebru, G. L. George, D. Roberson, D. W. Lacher, M. L. Kotewicz, J. E. LeClerc, T. A. Cebula;
US FDA, Laurel, MD.

Background: The frequency of foodborne outbreaks of Escherichia coli O157:H7 associated with the consumption of fresh produce has increased in recent years. In the fall of 2006, three major outbreaks occurred that were linked to contaminated spinach or lettuce. Preliminary to in-depth analyses of isolates by DNA microarray, optical mapping, and phenotypic microarray, we carried out phenotypic and genotypic characterization of foodborne and clinical samples from these outbreaks. Methods: Isolates of E. coli O157:H7 from outbreaks linked to spinach (Sept.-Oct., 2006) and to lettuce in Taco Bell (Nov.-Dec., 2006) and Taco John (Dec., 2006) restaurants were obtained from State laboratories. A total of 276 clinical and 19 spinach-derived isolates were examined. Microbiological analyses comprised plating assays on sorbitol-MacConkey, MUG, and tellurite agars. Molecular analyses included conventional PCR assays for virulence factors and uidA as well as a real-time PCR assay for ORFs specific to E. coli O157:H7; and a multi-locus variable number tandem repeat analysis (MLVA). Results: Most (91.9%) of the isolates were sorbitol-negative, β-glucuronidase-negative, tellurite-resistant, stx1-, stx2+, eae+, and ehxA+ strains. The isolates linked to lettuce from Taco John were stx1+ and stx2+. A seven-locus MLVA assay showed a total of at least 15 patterns from the three outbreaks. Two samples were mixed cultures, each containing the typical outbreak isolate and an additional strain. Multilocus sequence typing was used to determine the phylogenetic relationships of the atypical isolates to other types of pathogenic E. coli. Conclusions: The results demonstrated the usefulness of conventional phenotypic and genotypic characterization of outbreak samples carried out prior to the more expensive and time-consuming characterization of isolates required in trace-back studies. The use of a seven-locus MLVA assay, however, potentially overvalues the diversity of strains present within outbreaks, limiting its usefulness for establishing attribution in molecular epidemiology or microbial forensics investigations.