Verification of a Commercial Real-Time PCR Listeria monocytogenes and Listeria species Detection Method on Individually Quick-Frozen Vegetable Matrices
Erica Miller 1, Christopher Crowe 2, and J. David Legan 3
Eurofins Microbiology Laboratories, Inc., 1 Louisville, KY, 2 Salinas, CA, 3 Madison, WI
ABSTRACT
Introduction
With increasing concerns and rising numbers of product recalls associated with pathogens in produce, most major PCR testing platforms are validated for detection of Listeria in fresh produce. However, validations for other produce matrices are sparse, particularly for frozen matrices which can severely reduce initial temperatures of enrichment broths.
Purpose
To verify the ability to detect Listeria monocytogenes and Listeria species in frozen vegetable matrices utilizing real-time PCR.
Methods
Twenty-three different frozen vegetable matrices were tested, including cauliflower, carrots, various vegetable medleys, and chopped spinach: each with seven replicates (125g) inoculated with Listeria monocytogenes at ca. 7.5 cfu/sample. All samples were enriched with 1,125 mL of BACGro Ultra Listeria Broth™ (BULB) and incubated at 37°C for 21 ± 3 hours. Following incubation, samples were tested for Listeria monocytogenes and Listeria spp. using BACGene™ real-time PCR kits according to the manufacturer’s instruction. All proprietary media and tests kits were supplied by Gold Standard Diagnostics™.
Results
All seven replicates returned a presumptive positive PCR result for Listeria monocytogenes and Listeria spp. positive PCR results. The FDA microbiological method validation guidelines require 7/7 (or 19/20) presumptive results to consider a method extended to a new matrix, with no statistical analysis necessary. This standard was met 46 times in this study for both Listeria spp. and Listeria monocytogenes.
Significance
These data demonstrate that this method can reliably detect Listeria monocytogenes and Listeria spp. in a wide range of frozen vegetable matrices.
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