Detection, Occurrence, and Characterization of Escherichia coli O157:H7 from Raw Ewe's Milk in Spain

A study was carried out in the Castilla y León region of Spain to investigate the presence of Escherichia coli O157:H7 in raw ewe's milk samples collected from several cheese factories during 1 year. All specimens were examined for E. coli O157:H7 by selective enrichment at 41.5 ± 1.0°C, after both 6 and 22 h of incubation, and then immunomagnetically separated and plated on cefixime–potassium tellurite–sorbitol MacConkey agar. No growth was obtained in the enrichment broth after a 6-h incubation. Presumptive colonies obtained after 22 h of incubation were screened by a multiplex PCR assay for the presence of rfb_O157 and fliC_H7 genes. Of all the ewe's milk samples studied, three were positive for E. coli O157:H7. The E. coli O157:H7 strains that were positive for the rfb_O157 and fliC_H7 genes were then analyzed by multiplex PCR for the presence of virulence genes (stx₁, stx₂, ehxA, and eaeA). All E. coli O157:H7 isolates were Shiga toxigenic and harbored additional genes related to virulence (ehxA and eaeA). The predominant Stx toxin type was stx₂. These results demonstrate that raw ewe's milk used in cheesemaking may be sporadically contaminated with E. coli O157:H7 strains that are potentially pathogenic for humans.