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P3-50 Evaluation of the FDA BAM Method for Detection of Low Levels of E. coli O157:H7 in Artificially Inoculated Alfalfa Seeds

Tuesday, July 28, 2015
Hall B (Oregon Convention Center)
Tong-Jen Fu , U.S. Food and Drug Administration , Bedford Park , IL
Qing Li , Illinois Institute of Technology , Bedford Park , IL
Shuoqiu Song , Illinois Institute of Technology , Bedford Park , IL
Ken Yoshitomi , U.S. Food and Drug Administration, Office of Regulatory Affairs, Office of Regulatory Science , Rockville , MD
Karen Jinneman , U.S. Food and Drug Administration, Pacific Regional Laboratory Northwest , Bothell , WA
Peter Feng , U.S. Food and Drug Administration , College Park , MD
Introduction: Contaminated sprouts have been linked to numerous illness outbreaks. Seeds are often identified as the contamination source via epidemiological means. Analytical methods capable of identifying causative pathogens in contaminated seeds are needed to support regulatory actions.  

Purpose:  Evaluate the BAM method for detection of E. coli O157:H7 in inoculated alfalfa seeds. The sensitivity of culture method vs. qPCR for screening contaminated samples was compared. 

Methods: Test seeds spiked with different levels (0.1, 1, or 10 % w/w) of inoculated seeds (containing ~1 log CFU/g of E. coli O157:H7) were divided into 25-g aliquots. Each aliquot was mixed with 225 ml of pre-enrichment media (mBPWp) and incubated according to the BAM. Presence of the pathogen was determined either by culture method (streaking or spread plating on TC-SMAC or R&F E. coli O157:H7 agar) or by qPCR using SmartCycler or ABI-7500. 

Results: Culture method and qPCR showed similar sensitivity. Both detected E. coli O157:H7 in all samples spiked at a 1% level (E. coli O157:H7 concentration of -1 log MPN/g). At a 0.1% spiking level (E. coli O157:H7 level of -2 log MPN/g), presence of the pathogen was indicated in 9/20 and 8/20 samples using culture method and ABI-7500 test, respectively. In seeds spiked at a 0.01% level (E. coli O157:H7 level < -2.52 log MPN/g), the pathogen was detected in 1/20 samples using culture method but was not detected by ABI-7500. The sensitivity of the culture method can be significantly improved by incorporating immunomagnetic separation (IMS). With IMS, the number of positive samples increased from 9/20 to 18/20 and from 1/20 to 8/20 in seeds spiked at 0.1% and 0.01% level, respectively.

Significance: The BAM method performed well for detection of low levels of E. coli O157:H7 in alfalfa seeds and will be useful in supporting outbreak investigations and other surveillance activities.

Back to: Poster Session 3 – Laboratory and Detection Methods, Sanitation; Antimicrobials; Produce; Epidemiology
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