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Journal of Clinical Microbiology, July 2009, p. 2008-2012, Vol. 47, No. 7
0095-1137/09/$08.00+0     doi:10.1128/JCM.02013-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Rapid and Sensitive Detection of Shiga Toxin-Producing Escherichia coli from Nonenriched Stool Specimens by Real-Time PCR in Comparison to Enzyme Immunoassay and Culture{triangledown}

Thomas E. Grys,1,2* Lynne M. Sloan,1 Jon E. Rosenblatt,1,3 and Robin Patel1,3

Division of Clinical Microbiology, Mayo Clinic, Rochester, Minnesota,1 Molecular Microbiology Laboratory, Mayo Medical Laboratories, Andover, Massachusetts,2 Division of Infectious Diseases, Mayo Clinic, Rochester, Minnesota3

Received 17 October 2008/ Returned for modification 12 January 2009/ Accepted 5 May 2009

Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are a frequent cause of food-borne gastroenteritis, hemorrhagic colitis, and hemolytic uremic syndrome. Because antimicrobial agents are generally contraindicated in patients infected with STEC, a sensitive and specific diagnostic test with rapid turnaround is essential. Current culture methods may fail to detect non-O157 STEC. We evaluated a Stx gene real-time PCR assay using hybridization probes and the LightCycler instrument with 204 prospectively collected stool specimens, which were also tested for Stx by enzyme immunoassay (EIA) (ProSpecT STEC; Remel, Lenexa, KS) and by culturing on chromogenic agar (Chromagar O157; BD BBL, Sparks, MD). In addition, 85 archived stool specimens previously tested for Stx (by EIA) and/or E. coli O157:H7 (by culture) were tested by PCR. Sample preparation for PCR included mixing the stool in sterile water and extraction of nucleic acid using the MagNA Pure LC instrument (Roche Diagnostics). The PCR assay had 100% sensitivity and specificity compared to EIA and culture for specimens collected prospectively (4 of 204 specimens were positive) and compared to culture and/or EIA for archival specimens (42 of 85 specimens were positive). Both the EIA and PCR produced positive results from a specimen containing an O103 serotype STEC in the prospective specimens, and the PCR test detected three positive specimens that contained nonviable STEC in the archived specimens. The PCR assay demonstrated 100% sensitivity and specificity compared to EIA and/or culture and more rapid turnaround than either EIA or culture.

* Corresponding author. Mailing address: Mayo Medical Laboratories New England, 160 Dascomb Road, Andover, MA 01810. Phone: (978) 253-6187. Fax: (978) 658-0439. E-mail: grys.thomas{at}mayo.edu

{triangledown} Published ahead of print on 13 May 2009.

Journal of Clinical Microbiology, July 2009, p. 2008-2012, Vol. 47, No. 7
0095-1137/09/$08.00+0     doi:10.1128/JCM.02013-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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