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P1-26 Evaluating Food Enrichment Methods Using the Illumina MiSeq

Wednesday, May 11, 2016
Megaron Athens International Conference Center
Ninalynn Daquigan, U.S. Food and Drug Administration, Laurel, MD
Christopher Grim, U.S. Food and Drug Administration, Laurel, MD
James White, Resphera Biosciences, Baltimore, MD
Nicole Addy, U.S. Food and Drug Administration, Laurel, MD
Darcy Hanes, U.S. Food and Drug Administration, Laurel, MD
Karen Jarvis, U.S. Food and Drug Administration, Laurel, MD
Introduction: Culture-based methods, critical for identifying pathogens from contaminated foods, vary depending on the commodity type and target pathogen and can take up to one week to complete. This study surveyed enrichments throughout the culture process using 16S rRNA gene sequencing to analyze pathogen recovery and population dynamics, including species missed by traditional culture-based methods.

Purpose: This work uses 16S rRNA gene sequencing on the Illumina MiSeq to evaluate enrichment protocols for the detection of Salmonella enterica and Listeria monocytogenes from food.

Methods: Store-bought cilantro (n=5) and mung bean sprouts (n=4) were inoculated with Salmonella (2-5CFU/25g cilantro) and Listeria (50-100CFU/25g sprouts), aged, and cultured using modified FDA BAM protocols. Genomic DNA, taken from 24 and 48-hour cultures, was prepared for 16S rRNA gene amplification and sequenced in multiplex using the MiSeq platform. High-quality 16S rRNA sequences (normalized to 25,000 reads per sample) were analyzed using Resphera Insight software.

Results: In 24-hour cilantro cultures, we observed an average proportional abundance of 50% for Enterobacteriaceae, 0.2% (49 reads) being Salmonella. Salmonella increased to 37% (9,336 reads) and 93% (23,268 reads) after selective enrichment in Tetrathionate (TT) and Rappaport-Vassiliadis (RV) broths, respectively. Unlike Salmonella, the average proportional abundance of Listeria in 48-hour sprout cultures only reached 0.15% (38 reads). Other families present in 48-hour TT cilantro cultures included Planococcaceae and other Enterobacteriaceae, mainly Lysinibacillus (22%) and Proteus (15%) species, respectively. Streptococcaceae and Enterobacteriaceae families dominated sprout cultures with abundances of 80% (19,883 reads) and 9% (2,366 reads) at 24-hours, respectively, shifting to 52% (13,013 reads) and 28% (7,052 reads) at 48-hours.

Significance: High abundances of nonpathogenic species in cilantro and sprouts demonstrate the need for robust commodity-driven culture methods that favor pathogens such as Salmonella and Listeria. 16S rRNA gene sequencing can provide valuable information to improve detection methods for food safety.

Back to: Poster Session 1 –Antimicrobials, Applied Laboratory Methods, Novel Laboratory Methods, Pathogens and Risk Assessment
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