PFGE Standard Operating Procedures for Listeria monocytogenes: Harmonizing the Typing of Food and Clinical Strains in Europe

Abstract

Listeria monocytogenes is a foodborne pathogen responsible for a severe disease known as listeriosis. The European Centre for Disease Prevention and Control (ECDC) coordinates a network of national public health laboratories (NPHLs) in charge of typing clinical strains. In food, it is the European Union Reference Laboratory for L. monocytogenes (EURL Lm), which manages a network of National Reference Laboratories (NRLs). A pulsed-field gel electrophoresis (PFGE) standard operating procedure (EURL SOP) has been used routinely at the EURL Lm since 2007. The EURL Lm has recommended that NRLs use the EURL SOP, whereas the Statens Serum Institut (SSI), under contract for ECDC, requested that NPHLs use Halpins' SOP (HSOP) published in 2010 for the PulseNet USA network. An update of Halpins' SOP (uHSOP) was published in 2013. To facilitate the exchange of profiles among human and food European reference laboratories, it is crucial to ensure that the PFGE profiles obtained with these different SOPs are comparable. The aim here was to compare the EURL SOP with HSOP and uHSOP. The panel comprised 114 well-characterized SSI/EURL strains. All were characterized at the EURL using both the EURL SOP and uHSOP. Seventy of the 114 strains were also characterized at the SSI using HSOP. The EURL SOP and uHSOP produced indistinguishable combined (ApaI/AscI) profiles for the 114 strains tested. The EURL SOP and HSOP produced indistinguishable combined profiles for 69 of the 70 strains tested. One strain displayed for the AscI profile an additional low-intensity band at 184 kbp with HSOP. For this strain, SSI and EUR Lm had already observed the same profile from NPHLs and NRLs. However, this deviation is minor as it accounted for about 1% of all the 114 combined profiles. This study should facilitate the exchange of reproducible PFGE profiles among human and food reference laboratories.

MALDI-TOF MS identification and tracking of food spoilers and food-borne pathogens

“…PFGE is a golden standard for the identification of various food-borne pathogens. The PulseNet database is a surveillance network for food-borne disease outbreaks in the USA [25,26]. PCR is one of the most sensitive methods.…”

An Assessment of Different Genomic Approaches for Inferring Phylogeny of Listeria monocytogenes

“…The European surveillance system of L. monocytogenes from humans, foods, animals, and environments is still widely based on pulsed field gel electrophoresis (PFGE) (EFSA, 2014 ). PFGE was developed in the 1980s and the current PFGE scheme requires restriction by two enzymes using a validated standard protocol (Brosch et al, 1994 ; Michelon et al, 2015 ). PFGE has been extremely useful in Listeria outbreak investigations but its discriminatory power can be suboptimal for source tracking and source attribution (Ribot et al, 2006 ).…”

A Real-Time PCR Screening Assay for Rapid Detection of Listeria Monocytogenes Outbreak Strains

“…The developed strategy selected only 219 out of 587 strains, classified as IIa by molecular serogrouping, which were submitted to SNP analysis and PFGE with priority. As the estimated cost of PFGE per single strain is between 20.9 and 23.3 Euro, depending on the gel size [25], the gain ranged from 7691 and 8574 Euro. Moreover, the rapid response time of the analyses limited economic loss for the food business operators and farms involved.…”