T4-08 Digging Deep:  Making the Case for Molecular Based Detection with Real-world Performance and Discrepant Evaluation

Monday, July 10, 2017: 3:45 PM
Room 16 (Tampa Convention Center)
William Chaney , Roka Bioscience , San Diego , CA
Sarah Verver , Roka Bioscience , San Diego , CA
Janelle Lauffer , Roka Bioscience , San Diego , CA
Cambria Berry , Roka Bioscience , San Diego , CA
Ted Andrew , Roka Bioscience , Warren , NJ
Mary Duseau , Roka Bioscience , Warren , NJ
Introduction: Alternative molecular based detection technologies are rapidly evolving, yet results from such methods are often questioned or dismissed based upon follow up with imperfect culture based methodologies.

Purpose: This study evaluated the performance of the Atlas® Listeria Environmental (LE) Detection Assay for Listeria spp. detection in real-world environmental samples with investigation of initially discrepant results.

Methods: Environmental sponge swabs (n≥700) were collected from zones 3 and 4 of multiple processing environments. Swab samples were enriched with 90 ml of Actero Listeria Enrichment Media for 24 hours at 35°C prior to assaying with the LE assay using two analytical volumes, 200 µl and 2,000 µl. Screen positive results were initially struck to Modified Oxford Agar and transferred to Fraser secondary enrichment with subsequent plating to MOX and Listeria Chromogenic agars. Absence of typical morphology resulted in further analytical and culture analyses including PCR, IMS, and Filtration methodologies. Purified isolates were identified with biochemical or sequencing methodologies.

Results: Of n=750 samples, the LE assay reported 24 presumptive results for both the 200 and 2,000 µL analytical volumes. Of the 24 screen positive samples, Listeria spp. was isolated from 23 with standard or follow up culture analyses. Two discrepant samples were resolved utilizing a modified sample preparation for removal of free nucleic acid. In two cases, a biochemical method failed to identify typical isolates, which were identified as L. monocytogenes and L. innocua by 16S based identification. Other samples required extensive culture attempts to obtain isolates for microbial identification, which would be impractical under current industry testing scenarios and turnaround times.

Significance: These real-world data supported the molecular based detection of Listeria spp. while highlighting potential pitfalls and shortcomings of downstream confirmation processes.