P1-47 Comparison of Two Selective Media and Validation of Conventional and Real-time PCR Assays for the Detection of Listeria monocytogenes in Food Matrices

Monday, August 4, 2014
Exhibit Hall D (Indiana Convention Center)
Dong-Hyeon Kim, Konkuk University, Seoul, South Korea
Jung-Whan Chon, Konkuk University, Seoul, South Korea
Dasom Choi, Konkuk University, Seoul, South Korea
Young-Ji Kim, Konkuk University, Seoul, South Korea
Hong-Seok Kim, Konkuk University, Seoul, South Korea
Jin San Moon, Animal, Plant and Fisheries Quarantine and Inspection Agency, Anyang, South Korea
Kun-Ho Seo, Konkuk University, Seoul, South Korea
Introduction: Listeria monocytogenes is a bacterial foodborne pathogen responsible for listeriosis. Most countries have a zero-tolerance policy toward the presence of L. monocytogenes in ready-to-eat (RTE) foods owing to the possible severe consequences. Various methodologies, including conventional culture, molecular biological, biochemical and immunological techniques, have been implemented for the rapid and specific detection of L. monocytogenes. However, not all methods are well suited for routine use.

Purpose: In this study, we compared four detection methods (two standard selective media, conventional PCR and real-time PCR) most commonly used for the detection of Listeria monocytogenes in foods and identified false-positive colonies hindering the specific detection of L. monocytogenes on standard media.

Methods: L. monocytogenes was artificially inoculated into various food samples to generate partial positives and partial negatives. The inoculated samples were pre-enriched in Listeria enrichment broth followed by secondary enrichment in Fraser broth. Secondary enrichments were streaked onto Oxford and polymyxin-acriflavine-LiCl-ceftazidime-aesculin-mannitol (PALCAM) agar. After incubation, suspected colonies were biochemically identified using Vitek 2 system. In parallel, conventional PCR and real-time PCR were conducted with genomic DNA extracted from the secondary enrichment broth.

Results: Among all the methods, real-time PCR exhibited statistically excellent detection sensitivity (P < 0.05). Conventional culture methods performed poorly in the case of food with high levels of background microflora, generating numerous false-negative results. Although the detection of L. monocytogenes in fresh-cut vegetables using current culture methods was hindered only by Listeria innocua, various background microflora such as Listeria innocua, Listeria welshimeri, Listeria grayi, and Enterococcus faecalis appeared as presumptive positive colonies in raw beef, indicating a need to improve these methods.

Significance: We suggest that real-time PCR be used for early screening of L. monocytogenes in food samples, especially those with high levels of background microflora, thus complementing standard culture methodologies.