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Title: Incidence of Escherichia coli O157:H7 and E. coli Virulence Factors in US Bulk Tank Milk as Deteremined by Polymerase Chain Reaction

item Karns, Jeffrey
item Van Kessel, Jo Ann
item Mcclusky, Brian
item Perdue, Michael

Submitted to: Journal of Dairy Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/10/2007
Publication Date: 7/1/2007
Citation: Karns, J.S., Van Kessel, J.S., McClusky, B.J., Perdue, M.L. 2007. Incidence of Escherichia coli O157:H7 and E. coli Virulence Factors in U.S. Bulk Tank Milk as Deteremined by Polymerase Chain Reaction. Journal of Dairy Science. 90:3212-3219.

Interpretive Summary: Severe disease caused by enterohemorrhagic strains of the common bacterium Escherichia coli has been associated with the consumption of raw milk and products made from raw milk. E. coli O157:H7 is the serotype most commonly associated with severe E. coli disease in the US but other strains of E. coli can also cause disease. The specific isolation and identification of disease-causing E. coli is difficult because there are few major differences that separate them from common E. coli. We used assays based upon the polymerase chain reaction (PCR) to detect several genes associated with virulence in disease-causing strains of E. coli in samples of bulk tank milk taken from dairies across the US. The PCR assays indicated that the incidence of O157:H7 strains in raw milk was very low but that there was a somewhat greater possibility of contamination by other disease-causing serotypes.

Technical Abstract: Samples of bulk tank milk from dairies across the United States, taken as part of the National Animal Health Monitoring Dairy 2002 survey, were analyzed for the presence of several virulence factors associated with enterohemorrhagic forms of Escherichia coli (EHEC) using real-time and conventional PCR assays. Samples from 859 farms in 21 states were collected and enriched in EC Medium at 42.5oC to amplify any E. coli present and DNA was isolated from the resulting biomass. The eaeA gene encoding intimin, a virulence factor associated with enteropathogenic (EPEC) and EHEC forms of E. coli, was detected in 199 (23%) of the samples. Seventy-one samples (8.3%) were positive for both eaeA and one or both shiga-like toxin genes (stx1 and stx2), a combination that may be indicative of the presence of EHEC. Samples positive for eaeA were also tested for the presence of the gamma allele of the translocated intimin receptor (gamma-tir), found in EHEC strains of O157:H7, and 61 were found to be positive. Of these 61, 36 also were positive for stx1 and/or stx2, indicating potential contamination with O157:H7. Testing with a commercially available real-time PCR kit indicated that 5 could be contaminated with O157:H7. A multiplex PCR to detect the presence of fliC, rfbE, and hlyA, genes found in O157:H7, reduced to 2 (0.2% of all samples) the number of samples likely to be contaminated with this organism. A strain of O157:H7 (eaeA+, gamma-tir+, stx2+, rfbE+, fliC+, hlyA+) was subsequently isolated from one sample. These results indicate a low incidence of O157:H7 in bulk tank milk but suggest that greater risk from other enteropathogenic and enterohemorrhagic forms of E. coli may exist.