Listeria monocytogenes in pork industry
Several deadly outbreaks of listeriosis occurred in Europe and North America over the last decades. Pork products are often involved in these clustered cases. The Health Canada Policy on Listeria monocytogenes (Listeria policy, 2011) focuses on environmental contamination in ready to eat products “step” in the food production chain. However, there is a clear lack of information about this pathogen upstream the production : slaughter and cutting plants. After the updated Listeria policy (2011), Health Canada introduced new regulations, thus, manufacturers must deal with more frequent detections, particularly from environmental origin. Then the question arises of equivalence of these L. monocytogenes strains detected in terms of risk to the consumer. At previous steps of meat production (slaughtering, cutting), abundant environmental flora is thought to be unfavorable to Listeria monocytogenes detection. First of all, it appears necessary to validate bacteriological methods recommendations of Health Canada. Then detection and suitable typing of strains will allow a temporal and spatial analysis of the evolution of residual contamination of Listeria monocytogenes in facilities. The integration of this information in surveillance of human listeriosis helps to evaluate the needs in terms of risk reduction or characterization of environmental strains.
From slaughter to cutting facilities we suggested that a reorganization of L. monocytogenes populations (decreased of environmental strains diversity), by a selection, occurs. Better characterization (biofilm forming ability, virulence) of these isolates would facilitate management measures by manufacturers.
Identify sites at risk of residual contamination by L. monocytogenes in pig slaughter / cutting plants. Obtain a collection of strains, precisely identify these isolates (molecular typing) to describe the changes in strain populations in and between plants. Compare biofilm forming properties of strains to contribute to the definition of resident and transient strains and their associated virulence properties.
Exhaustive sampling of 4 plants (156 samples each time), focused on the detection of L. monocytogenes (version ISO / TC 34 / SC 9 June 2012) (4 visits per plant in one year). The 2496 analyzes will be conducted according to the protocol MFH-PB-30 with few modifications (using Compass). The comparison of Listeria detection methods (Health Canada methods and fast BAX Listeria spp. detection), particularly for “primary production” samples, will be done from some of these samples.
Isolates characterization: serogrouping by multiplex PCR, ribotyping (EcoRI),MVLST-typing and PFGE (protocol C-enternet, ApaI AscI, DICE / UGPMA using BioNumerics (Applied Math).
Characterization of potential virulence of the isolates: the inlA gene (2400pb) will be sequenced, and SNPs analyzed for premature stop codon and amino acid deletions detection. It will be conducted on 30% of the isolates of the collection, (particularly on isolates of the same pulsotype and found at the start and at the end of the slaughtering/cutting process). Biofilm forming ability of the strains: the formation of biofilm in static conditions (crystal violet staining, confocal microcopy) and analysis of dynamic biofilm formation (Bio flow fluxion platform Biosciences Inc.) will be done. Detection / expression (PCR, Q-PCR, RT-PCR ) of bapL (gene coding for biofilm associate protein) will be done, as well as analyses of quorum sensing (AI-2) and the Agr (regulatory system) in opposite strains.