Genotypic analyses of Escherichia coli O157:H7 and O157 nonmotile isolates recovered from beef cattle and carcasses at processing plants in the Midwestern states of the United States

Abstract

Escherichia coli O157:H7 and O157 nonmotile isolates (E. coli O157) previously were recovered from feces, hides, and carcasses at four large Midwestern beef processing plants (R. O. Elder, J. E. Keen, G. R. Siragusa, G. A. Barkocy-Gallagher, M. Koohmaraie, and W. W. Laegreid, Proc. Natl. Acad. Sci. USA 97:2999-3003, 2000). The study implied relationships between cattle infection and carcass contamination within single-source lots as well as between preevisceration and postprocessing carcass contamination, based on prevalence. These relationships now have been verified based on identification of isolates by genomic fingerprinting. E. coli O157 isolates from all positive samples were analyzed by pulsed-field gel electrophoresis of genomic DNA after digestion with XbaI. Seventy-seven individual subtypes (fingerprint patterns) grouping into 47 types were discerned among 343 isolates. Comparison of the fingerprint patterns revealed three clusters of isolates, two of which were closely related to each other. Remarkably, isolates carrying both Shiga toxin genes and nonmotile isolates largely fell into specific clusters. Within lots analyzed, 68.2% of the postharvest (carcass) isolates matched preharvest (animal) isolates. For individual carcasses, 65.3 and 66.7% of the isolates recovered postevisceration and in the cooler, respectively, matched those recovered preevisceration. Multiple isolates were analyzed from some carcass samples and were found to include strains with different genotypes. This study suggests that most E. coli O157 carcass contamination originates from animals within the same lot and not from cross-contamination between lots. In addition, the data demonstrate that most carcass contamination occurs very early during processing.

FIG. 1

Relatedness of E. coli O157 isolates. The dendrogram was generated using the Dice coefficient and UPGMA analysis (see Materials and Methods). The scale at the top of the dendrogram indicates the level of similarity between isolates; types include isolates connected at approximately >95% similarity. One isolate of each distinct genomic pattern (subtype) was included in the dendrogram; thus, a final branch may represent multiple isolates as indicated at the end of each branch. The presence of stx1 is indicated by solid circles; the presence of stx2 is indicated by solid squares. Subtypes that include isolates that carry stx1 and stx2 as well as isolates that carry only stx2 are indicated by open circles and solid squares. The presence of only motile isolates is indicated by filled triangles. The presence of nonmotile and motile isolates is indicated by open triangles. The absence of a symbol indicates the absence of that characteristic from all isolates within the subtype. The numbers of isolates within each subtype that were recovered at each sampling site are indicated. Subtype designations follow the symbols. Abbreviations: St., subtype designations; Preevis., preevisceration; Postevis., postevisceration.

FIG. 2

Genomic fingerprints from passaged isolates. (A) Fingerprints of strain 60AC3 during passaging. The isolate and passaging day are indicated for each lane. The arrow indicates the band that was lost during passaging. (B) Fingerprints of strain 234AB1 during passaging. Size standards are lambda concatemers (Bio-Rad); sizes are given in kilobases. The isolate and passaging day are indicated for each lane. Dashed lines indicate bands lost from the original fingerprint, solid lines indicate bands gained in comparison to the original fingerprint. Bands marked between lanes 1 and 2 pertain to changes in the day 1 pattern, bands marked outside of lanes 1 and 5 pertain to changes between the original fingerprint and the day 3, 5, and 10 fingerprints. BGB, brilliant green bile broth.