Glabridin is an antimicrobial compound which can be extracted from plants, such as liquorice (Glycyrrhiza glabra) roots. Although its activity against foodborne pathogens and spoilage microorganisms has already been reported, the investigation of potential applications as a surface disinfectant is still largely unexplored. Hence, this study evaluated the disinfectant efficacy of glabridin against Listeria monocytogenes. The activity of glabridin was first tested in vitro in a nutrient-rich medium against eight strains of L. monocytogenes, including food isolates and the model strain EGDe. The tested strains showed similar susceptibility with minimal inhibitory and bactericidal concentrations of 12.5 µg/mL and 25 µg/mL, respectively. Subsequently, L. monocytogenes L6, FBR17 and EGDe were selected to assess the efficacy of glabridin against dried cells (according to the European standard EN 13697:2015 + A1:2019) and biofilm cells on stainless steel surfaces. Moreover, the impact of food residual organic matter was investigated using skim milk, cantaloupe and smoked salmon solution as soiling components. Our results showed that applying 200 µg/mL of glabridin resulted in a substantial reduction (>3 log10) of dried and biofilm cells of L. monocytogenes in standard conditions (i.e. low level of residual organic matter). Cantaloupe soiling components slightly reduced the activity of glabridin, while the efficacy of glabridin when tested with salmon and skim milk residuals was substantially affected. Comparative analysis using standardized protein contents provided evidence that the type of food matrices and type of proteins may impact the activity of glabridin as a disinfectant. Overall, this study showed low strain variability for the activity of glabridin against L. monocytogenes and shed light on the possible application of this natural antimicrobial compound as a surface disinfectant.

Nowadays, the discovery of alternative natural antimicrobial substances such as bacteriophages, essential oils, and other physical and chemical agents is developing in the food industry. In this study, nine bacteriophages were isolated from various parts of raw chickens and exhibited lytic activities against L. monocytogenes and various Listeria spp. The characterization of phage vB_LmoS-PLM9 was stable at 4 to 50 °C and pH range from 4 to 10. Phage vB_LmoS-PLM9 had a circular, double-stranded genomic DNA with 38,345 bp having endolysin but no antibiotic resistance or virulence genes. Among the eight essential oils tested at 10 %, cinnamon bark, and cassia oils showed the strongest antilisterial activities. The combined use of phage vB_LmoS-PLM9 and cinnamon oils indicated higher efficiency than single treatments. The combination of phage (MOI of 10) and both cinnamon oils (0.03 %) reduced the viable counts of L. monocytogenes and inhibited the regrowth of resistant cell populations in broth at 30 °C. Furthermore, treatment with the combination of phage (MOI of 100) and cinnamon oil (0.125 %) was effective in milk, especially at 4 °C by reducing the viable count to less than lower limit of detection. These results suggest combining phage and cinnamon oil is a potential approach for controlling L. monocytogenes in milk.

Invasive listeriosis, due to its severe nature in susceptible populations, has been the focus of many quantitative risk assessment (QRA) models aiming to provide a valuable guide in future risk management efforts. A review of the published QRA models of Listeria monocytogenes in seafood was performed, with the objective of appraising the effectiveness of the control strategies at different points along the food chain. It is worth noting, however, that the outcomes of a QRA model are context-specific, and influenced by the country and target population, the assumptions that are employed, and the model architecture itself. Studies containing QRA models were retrieved through a literature search using properly connected keywords on Scopus and PubMed®. All 13 QRA models that were recovered were of short scope, covering, at most, the period from the end of processing to consumption; the majority (85%) focused on smoked or gravad fish. Since the modelled pathways commenced with the packaged product, none of the QRA models addressed cross-contamination events. Many models agreed that keeping the product\'s temperature at 4.0-4.5 °C leads to greater reductions in the final risk of listeriosis than reducing the shelf life by one week and that the effectiveness of both measures can be surpassed by reducing the initial occurrence of L. monocytogenes in the product (at the end of processing). It is, therefore, necessary that future QRA models for RTE seafood contain a processing module that can provide insight into intervention strategies that can retard L. monocytogenes\' growth, such as the use of bacteriocins, ad hoc starter cultures and/or organic acids, and other strategies seeking to reduce cross-contamination at the facilities, such as stringent controls for sanitation procedures. Since risk estimates were shown to be moderately driven by growth kinetic parameters, namely, the exponential growth rate, the minimum temperature for growth, and the maximum population density, further work is needed to reduce uncertainties.

Listeria monocytogenes has a range of strategies that allow it to persist as biofilms in food processing environments (FPE), making it a pathogen of concern to the food industry. The properties of these biofilms are highly variable among strains, and this significantly affects the risk of food contamination. The present study therefore aims to conduct a proof-of-concept study to cluster strains of L. monocytogenes by risk potential using principal component analysis, a multivariate approach. A set of 22 strains, isolated from food processing environments, were typed by serogrouping and pulsed-field gel electrophoresis, showing a relatively high diversity. They were characterized in terms of several biofilm properties that might pose a potential risk of food contamination. The properties studied were tolerance to benzalkonium chloride (BAC), the structural parameters of biofilms (biomass, surface area, maximum and average thickness, surface to biovolume ratio and roughness coefficient) measured by confocal laser scanning microscopy and (3) transfer of biofilm cells to smoked salmon. The PCA correlation circle revealed that the tolerance of biofilms to BAC was positively correlated with roughness, but negatively with biomass parameters. On the contrary, cell transfers were not related to three-dimensional structural parameters, which suggests the role of other variables yet unexplored. Additionally, hierarchical clustering grouped strains into three different clusters. One of them included the strains with high tolerance to BAC and roughness. Another one consisted of strains with enhanced transfer ability, whereas the third cluster contained those that stood out for the thickness of biofilms. The present study represents a novel and effective way to classify L. monocytogenes strains according to biofilm properties that condition the potential risk of reaching the consumer through food contamination. It would thus allow the selection of strains representative of different worst-case scenarios for future studies in support of QMRA and decision-making analysis.

Testing of foods for low levels of the human pathogen, Listeria monocytogenes (Lm), involves a selective enrichment procedure. A nonpathogenic species of Listeria, L. innocua (Li), is often present in foods and food-manufacturing environments and is an interference organism for Lm detection due to competition during enrichment. The present study investigated whether a novel enrichment strategy incorporating the sugar allose into the secondary enrichment broth (allose method) could improve the detection of Lm from foods when Li is present. First, Canadian food isolates of Listeria spp. were tested to confirm recent reports that lineage II Lm (LII-Lm), but not Li, could metabolize allose. All LII-Lm isolates (n = 81), but not Li (n = 36), possessed the allose genes lmo0734-lmo0739, and could efficiently metabolize allose. Next, smoked salmon was contaminated with mixtures of LII-Lm and Li and tested using different enrichment procedures to compare the ability to recover Lm. Allose broth was more effective than Fraser Broth, with Lm detected in 87% (74 of 85) compared to 59% (50 of 85) of the samples (P < 0.05), following a common preenrichment. When evaluated against a current Health Canada method (MFLP-28), the allose method was more effective, with LII-Lm detected in 88% (57 of 65) compared to 69% (45 of 65) of the samples (P < 0.05). The allose method also remarkably increased the ratio of LII-Lm to Li postenrichment, which improved the ease of obtaining isolated Lm colonies for confirmation tests. Allose may therefore provide a tool for use when the presence of background flora interferes with Lm detection. As this tool is specifically applicable to a subset of Lm, the use of this method modification may provide a working example of tailoring methodology to target the known subtype of the pathogen of interest in an outbreak investigation, or for regular monitoring activities in conjunction with a PCR screen for allose genes on preenrichment cultures.

Microbial pathogens may be present in different types of foods, and hence the development of novel methods to assure consumers\' safeness is of great interest. Molecular methods are known to provide sensitive and rapid results; however, they are typically targeted approaches. In recent years, the advent of non-targeted approaches based on next-generation sequencing (NGS) has emerged as a rational way to proceed. This technology allows for the detection of several pathogens simultaneously. Furthermore, with the same set of data, it is possible to characterize the microorganisms in terms of serotype, virulence, and/ or resistance genes, among other molecular features. In the current study, a novel method for the detection of Listeria monocytogenes based on the \"quasimetagenomics\" approach was developed. Different enrichment media and immunomagnetic separation (IMS) strategies were compared to determine the best approach in terms of L. monocytogenes sequences generated from smoked salmon samples. Finally, the data generated were analyzed with a user-friendly workflow that simultaneously provided the species identification, serotype, and antimicrobial resistance genes. The new method was thoroughly evaluated against a culture-based approach, using smoked salmon inoculated with L. monocytogenes as the matrix of choice. The sequencing method reached a very low limit of detection (LOD50, 1.2 CFU/ 25 g) along with high diagnostic sensitivity and specificity (100%), and a perfect correlation with the culture-based method (Cohen\'s k = 1.00). Overall, the proposed method overcomes all the major limitations reported for the implementation of NGS as a routine food testing technology and paves the way for future developments taking its advantage into consideration.

Cold-smoked salmon is a ready-to-eat seafood product of high commercial importance. The processing and storage steps facilitate the introduction, growth, and persistence of foodborne pathogens and spoilage bacteria. The growth of commensal bacteria during storage and once the product is opened also influence the quality and safety of cold-smoked salmon. Here we investigated the microbial community through targeted 16S rRNA gene and shotgun metagenomic sequencing as means to better understand the interactions among bacteria in cold-smoked salmon. Cold-smoked salmon samples were tested over 30 days of aerobic storage at 4°C and cultured at each time point in a buffered Listeria enrichment broth (BLEB) commonly used to detect Listeria in foods. The microbiomes were composed of Firmicutes and Proteobacteria, namely, Carnobacterium, Brochothrix, Pseudomonas, Serratia, and Psychrobacter. Pseudomonas species were the most diverse species, with 181 taxa identified. In addition, we identified potential homologs to 10 classes of bacteriocins in microbiomes of cold-smoked salmon stored at 4°C and corresponding BLEB culture enrichments. The findings presented here contribute to our understanding of microbiome population dynamics in cold-smoked salmon, including changes in bacterial taxa during aerobic cold storage and after culture enrichment. This may facilitate improvements to pathogen detection and quality preservation of this food.

The aim of this study was to develop a rapid and accurate PMA-qPCR method to quantify viable Brochothrix thermosphacta in cold-smoked salmon. B. thermosphacta is one of the main food spoilage bacteria. Among seafood products, cold-smoked salmon is particularly impacted by B. thermosphacta spoilage. Specific and sensitive tools that detect and quantify this bacterium in food products are very useful. The culture method commonly used to quantify B. thermosphacta is time-consuming and can underestimate cells in a viable but not immediately culturable state. We designed a new PCR primer set from the single-copy rpoC gene. QPCR efficiency and specificity were compared with two other published primer sets targeting the rpoC and rpoB genes. The viability dyes PMA or PMAxx were combined with qPCR and compared with these primer sets on viable and dead B. thermosphacta cells in BHI broth and smoked salmon tissue homogenate (SSTH). The three primer sets displayed similar specificity and efficiency. The efficiency of new designed rpoC qPCR on viable B. thermosphacta cells in SSTH was 103.50%, with a linear determination coefficient (r2) of 0.998 and a limit of detection of 4.04 log CFU/g. Using the three primer sets on viable cells, no significant difference was observed between cells treated or untreated with PMA or PMAxx. When dead cells were used, both viability dyes suppressed DNA amplification. Nevertheless, our results did not highlight any difference between PMAxx and PMA in their efficiency to discriminate viable from unviable B. thermosphacta cells in cold-smoked salmon. Thus, this study presents a rapid, specific and efficient rpoC-PMA-qPCR method validated in cold-smoked salmon to quantify viable B. thermosphacta in foods.

This study investigated the antimicrobial activity of 405 nm light-emitting diode (LED) with and without riboflavin against Listeria monocytogenes in phosphate buffered saline (PBS) and on smoked salmon at different storage temperatures and evaluated its impact on food quality. The results show that riboflavin-mediated LED illumination in PBS 25 °C significantly inactivated L. monocytogenes cells by 6.2 log CFU/mL at 19.2 J/cm2, while illumination alone reduced 1.9 log CFU/mL of L. monocytogenes populations at 57.6 J/cm2. L. monocytogenes populations on illuminated smoked salmon decreased by 1.0-2.2 log CFU/cm2 at 1.27-2.76 kJ/cm2 at 4, 12, and 25 °C, regardless of the presence of riboflavin. Although illumination with and without riboflavin caused the lipid peroxidation and color change in smoked salmon, this study demonstrates the potential of a 405 nm LED to preserve the smoked salmon products, reducing the risk of listeriosis.

Amplicon sequencing approaches have been widely used in food bacterial ecology. However, choices regarding the methodology can bias results. In this study, bacterial communities associated with cold-smoked salmon products and their processing plant surfaces were monitored via sequencing of the V3-V4 region of the 16S rRNA gene. The impact of DNA extraction protocols, sampling methods (swabbing or sponging) and surface materials on bacterial communities were investigated. α and β diversity analyses revealed that DNA extraction methods mainly influence the observed cold-smoked salmon microbiota composition. Moreover, different DNA extraction methods revealed significant differences in observed community richness and evenness. β-Proteobacteria: Photobacterium, Serratia and Firmicutes: Brochothrix, Carnobacterium and Staphylococcus were identified as the dominant genera. Surface microbiota richness, diversity and composition were mainly affected by cleaning and disinfection procedures but not by DNA extraction methods. Surface community richness and evenness appeared higher when sampled by sponging compared to swabbing. β-diversity analyses highlighted that surface topology, cleaning and disinfection and sampling devices seemed to affect the bacterial community composition. The dominant surface bacteria identified were mainly Flavobacteriaceae, β-Proteobacteria and γ-Proteobacteria described as fish spoilers such as Acinetobacter, Pseudomonas and Shewanella. DNA extraction and sampling methods can have an impact on sequencing results and the ecological analysis of bacterial community structures. This study confirmed the importance of methodology standardization and the need for analytical validation before 16S rDNA metabarcoding surveys.