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United States Department of Agriculture

Agricultural Research Service

Title: Methods for Recovering Escherichia Coli O157:h7 from Cattle Fecal, Hide, and Carcass Samples: Sensitivity and Improvements

Authors
item Barkocy-Gallagher, Genevieve - CDC
item Edwards, Kelly - QUINTILES, INC.
item Nou, Xiangwu
item Bosilevac, Joseph
item Arthur, Terrance
item Shackelford, Steven
item Koohmaraie, Mohammad

Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: June 10, 2005
Publication Date: November 20, 2005
Citation: Barkocy-Gallagher, G.A., Edwards, K.K., Nou, X., Bosilevac, J.M., Arthur, T.M., Shackelford, S.D., Koohmaraie, M. 2005. Methods for recovering Escherichia coli O157:H7 from cattle fecal, hide, and carcass samples: Sensitivity and improvements. Journal of Food Protection. 68:2264-2268.

Interpretive Summary: The MRU methods have been used to study the prevalence of E. coli O157:H7 in cattle carcass, hide, and fecal samples. The sensitivity of these methods for recovery of injured cells has now been determined. Ninety-one percent of the carcass samples collected early in the slaughter process tested positive for E. coli O157:H7 when inoculated with five to ten bacterial cells. For hide samples, 100% tested positive for E. coli O157:H7 when inoculated with 30 to 50 bacterial cells. At lower inoculum levels, substituting a commercially available wash buffer for the MRU wash buffer during bacterial concentration steps improved recovery of the target bacteria. When comparing naturally contaminated samples, substituting a buffer with added detergent for the MRU wash buffer also had a positive effect on recovery of E. coli O157:H7 from cattle hide samples. The standard MRU method identified 96% of the fecal samples as positive when inoculated with 300 to 400 bacterial cells per ten grams of cattle feces. The addition of a phosphate buffer to the enrichment broth improved recovery of E. coli O157:H7 from feces. Using the modified enrichment, 92% of the samples were identified as positive when inoculated with 10 to 30 bacterial cells per ten grams of cattle feces. When naturally contaminated fecal samples were split and analyzed, 26% tested positive for E. coli O157:H7 by the MRU method and 39% tested positive using the modified enrichment. The MRU methods are sensitive methods for recovery of E. coli O157:H7 from cattle carcass, hide, and fecal samples. However, these methods can be improved by changing the wash buffer for hide sample analyses and by adding a phosphate buffer to the fecal enrichment broth.

Technical Abstract: The MRU methods have been used to study the prevalence of Escherichia coli O157:H7 in cattle carcass, hide, and fecal samples. Sensitivity of these methods for recovery of injured cells has now been determined. Ninety-one percent of the pre-evisceration carcass samples tested positive for E. coli O157:H7 when inoculated with five to ten CFU. For hide samples, 100% tested positive for E. coli O157:H7 when inoculated with 30 to 50 CFU. At lower inoculum levels, substituting a commercially available wash buffer for phosphate buffered saline (PBS)-Tween 20 wash buffer during immunomagnetic separation improved recovery of the target organism. When comparing naturally contaminated samples, substituting a PBS-CHAPS buffer for PBS-Tween 20 also had a positive effect on recovery of E. coli O157:H7 from hide samples. The standard MRU method identified 96% of the fecal samples as positive when inoculated with 300 to 400 CFU (per 10 g). The addition of a phosphate buffer to the tryptic soy broth enrichment improved recovery of E. coli O157:H7 from feces. Using the modified enrichment, 92% of the samples were identified as positive when inoculated with 10 to 30 CFU (per 10 g). When naturally contaminated fecal samples were split and analyzed, 26% tested positive for E. coli O157:H7 by the MRU method and 39% tested positive using the modified enrichment. Data presented here indicate that the MRU method is highly effective at recovering injured E. coli O157:H7 from beef carcass samples. The MRU method also is effective for recovery of E. coli O157:H7 from hide samples, however incorporating IMS buffer or PBS-CHAPS as the wash buffer improves the ability to correctly identify positive samples (when interference by background microflora is expected). Finally, the MRU method for recovering E. coli O157:H7 from feces is substantially improved by the incorporation of a phosphate buffer in the TSB enrichment broth.

Last Modified: 7/25/2014
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