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P2-181 Evaluation of 16S rRNA Sequencing for the Detection of Salmonella in Cilantro

Monday, July 27, 2015
Exhibit Hall (Oregon Convention Center)
Karen Jarvis , U.S. Food and Drug Administration , Laurel , MD
James White , Oak Ridge Institute for Science and Technology , Oak Ridge , TN
Christopher Grim , U.S. Food and Drug Administration , Laurel , MD
Laura Ewing , U.S. Food and Drug Administration , Laurel , MD
Ninalynn Daquigan , U.S. Food and Drug Administration , Laurel , MD
Junia Jean-Gilles Beaubrun , U.S. Food and Drug Administration , Laurel , MD
Nicole Addy , Oak Ridge Institute for Science and Technology , Oak Ridge , TN
Darcy Hanes , U.S. Food and Drug Administration , Laurel , MD
Introduction: Salmonella enterica is a known cause of foodborne gastroenteritis in the United States and has been associated with outbreaks in fresh produce such as cilantro.  Salmonella detection methods are complex, time consuming, and efforts to decrease the time to detection are needed.  Our previous work indicates that an initial 24-hour non-selective pre-enrichment in Modified Buffered Peptone Water (mBPW) favored the growth of Firmicutes, particularly the Peptostreptococcaceae, as opposed to Enterobacteriaceae.

Purpose: This study evaluates alterations to the FDA Bacteriological Analytical Manual (BAM) method for Salmonella detection to significantly increase the proportional abundance of Enterobacteriaceae; i.e., Salmonella, to that of non-target species, notably Peptostreptococcaceae, in cilantro using16S rRNA sequencing of enrichment cultures.

Methods: Genomic DNA was extracted from broth cultures of cilantro samples processed using the BAM method for the detection of Salmonella in leafy greens. Variable treatments were tested including; the use of two resuscitation broths, mBPW and universal pre-enrichment broth (UP), and shaking (165 rpm) or static incubation at 37°C during the initial 24-hour non-selective resuscitation. 16S rRNA amplicons were generated using primers specific to the V1 - V3 regions and sequenced on the Illumina MiSeq using a 600 cycle reagent kit.

Results: Both pre-enrichment broths performed equally for the enrichment of Enterobacteriaceae (P = 0.882, n = 15). However, shaking the 24-hour mBPW and UP pre-enrichments significantly increased the Enterobacteriaceae by 34.9% (P = 6.61E-06, n = 30) and decreased the Peptostreptococcaceae by 70% (P = 5.62E-04, n = 30). In spiked samples, the effect of shaking was significant for the enrichment of Enterobacteriaceae in general (P = 1.91E-04, n = 13) but significant changes in the abundance of S. enterica were not observed (P = 0.368, n = 13).

Significance: We have shown that aeration via shaking during the 24-hour pre-enrichment of cilantro samples significantly favors the enrichment of Enterobacteriaceae and depresses the growth of Peptostreptococcaceae using 16s rRNA sequencing.

Back to: Poster Session 2 – Laboratory and Detection Methods; Meat, Poultry and Eggs; Dairy and Beverages; Food Law and Regulation; General Microbiology; Modeling and Risk Assessment
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