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P2-74 Rapid and Sensitive Detection of Shiga Toxin-producing Escherichia coli in Environmental Samples by Multiplex PCR

Tuesday, July 30, 2013
Exhibit Hall (Charlotte Convention Center)
Shefali Dobhal, Oklahoma State University, Stillwater, OK
Chris Timmons, Oklahoma State University, Stillwater, OK
Guodong Zhang, U.S. Food and Drug Administration, College Park, MD
Charles Rohla, The Samuel Roberts Noble Foundation, Ardmore, OK
Mike Smith, Oklahoma State University, Stillwater, OK
Li Ma, Oklahoma State University, Stillwater, OK
Introduction: Shiga toxin-producing Escherichia coli (STEC) has been associated with numerous foodborne illness outbreaks. A rapid and sensitive detection method is essential for environmental monitoring and outbreak investigations.

Purpose: The objective of this study was to develop a simple and sensitive method for rapid detection of STEC in challenging environmental samples.

Methods: Multiplex PCR targeting eae, stx1 and 2 genes was developed and its sensitivity was evaluated using pure overnight culture of STEC O157:H7 and O26:H11, respectively, and those spiked with background microflora from enriched soil and pecan samples. The effect of additives, Bovine serum albumin (BSA), Polyvinylpyrrolidone (PVP), Polyethylene glycol (PEG) and Gelatin, on PCR sensitivity was also evaluated. 

Results: The developed multiplex PCR method, pre-spin sample matrices in combination with the addition of BSA and PVP in PCR reaction mix, can detect up to 4 CFU/reaction of STEC O157:H7 and 40 CFU/reaction of O26:H11 either as pure culture or in the presence of background microflora from overnight enriched environmental samples of soil or pecan.

Significance: The developed multiplex PCR assay is simple and sensitive with broad applicability for rapid detection of STEC in intricate environmental samples.

Back to: Poster Session 2 - Pathogens, Antimicrobials
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