Rapid sample preparation method for PCR-based detection of Escherichia coli O157:H7 in ground beef

doi:10.1046/j.1365-2672.2003.01951.x

If you are seeing this message, you may be experiencing temporary network problems. Please wait a few minutes and refresh the page. If the problem persists, you may wish to report it to your local Network Manager.

It is also possible that your web browser is not configured or not able to display style sheets. In this case, although the visual presentation will be degraded, the site should continue to be functional. We recommend using the latest version of Microsoft or Mozilla web browser to help minimise these problems.

Wiley InterScience

Journal of Applied Microbiology

Volume 95 Issue 1, Pages 129 - 134

Published Online: 16 Jun 2003

The Official Journals of the Society for Applied Microbiology

Go to Society Site

< Previous Abstract | Next Abstract >

Save Article to My Profile Download Citation Request Permissions

Abstract | References | Full Text: HTML, PDF (Size: 110K) | Related Articles | Citation Tracking

Rapid sample preparation method for PCR-based detection of Escherichia coli O157:H7 in ground beef

S. Cui ¹ , C.M. Schroeder ¹*, D.Y. Zhang ² and J. Meng ¹

¹ Department of Nutrition and Food Science, University of Maryland, College Park, MD, ² Department of Pathology, Mount Sinai School of Medicine, New York, NY, USA

Correspondence to J. Meng, Department of Nutrition and Food Science, 0112 Skinner Building, University of Maryland, College Park, MD 20742, USA (e-mail: jm332@umail.umd.edu).

*US Department of Agriculture, Food Safety and Inspection Service, Washington D.C., USA.

KEYWORDS

detection • E. coli O157:H7 • ground beef • PCR

s. cui, c.m. schroeder, d.y. zhang and j. meng. 2003.

Abstract

Aim: To develop an improved, rapid and sensitive sample preparation method for PCR-based detection of Escherichia coli O157:H7 in ground beef.

Methods and Results: Fresh ground beef samples were experimentally inoculated with varying concentrations of E. coli O157:H7. PCR inhibitors were removed and bacterial cells were concentrated by filtration and centrifugation, and lysed using enzymatic digestion and successive freeze/thaw cycles. DNA was purified and concentrated via phenol/chloroform extraction and the Shiga toxin 1 gene (stx1) was amplified using PCR to evaluate the sample preparation method. Without prior enrichment of cells in broth media, the detection limit was 10³ CFU g⁻¹ beef. When a 6 h enrichment step was incorporated, the detection limit was 1 CFU g⁻¹ beef. The total time required from beginning to end of the procedure was 12 h.

Conclusions: The sample preparation method developed here enabled substantially improved sensitivity in the PCR-based detection of E. coli O157:H7 in ground beef, as compared to previous reports.

Significance and Impact of the Study: Superb sensitivity, coupled with quick turn-around time, relative ease of use and cost-effectiveness, makes this a useful method for detecting E. coli O157:H7 in ground beef.

2002/378: received 15 October 2002, revised 25 February 2003 and accepted 5 March 2003

DIGITAL OBJECT IDENTIFIER (DOI)

10.1046/j.1365-2672.2003.01951.x About DOI