Growth Potential of Listeria monocytogenes in Three Different Salmon Products.
ABSTRACT: Cold smoked salmon and sushi salmon have been implicated in outbreaks of listeriosis. We performed challenge tests and a durability study with Listeria monocytogenes on different salmon products to determine the growth potential of this important food-borne pathogen. Data from the challenge test showed a significant growth potential of L. monocytogenes on all of the tested salmon products, with faster growth in sushi salmon than in cold smoked salmon. In identical products that were naturally contaminated at low levels, the durability study did not confirm a high growth potential, possibly due to interactions with competing microflora. The injection of sodium lactate (NaL) at a high concentration (30%) into cold smoked salmon significantly reduced the growth potential of L. monocytogenes. In addition to good manufacturing practices, the injection of higher concentrations of NaL may therefore be a useful additional hurdle to prevent growth of L. monocytogenes to high numbers in the tested salmon products.
Project description:Listeria monocytogenes is an important foodborne pathogen and a major cause of death associated with bacterial foodborne infections. Control of L. monocytogenes on most ready-to-eat (RTE) foods remains a challenge. The potential use of ?-phenylethylamine (PEA) as an organic antimicrobial against L. monocytogenes was evaluated in an effort to develop a new intervention for its control. Using a collection of 62 clinical and food-related isolates we determined the minimum inhibitory concentration (MIC) of PEA against L. monocytogenes in different broth and agar media. Bologna type sausage (lyoner) and smoked salmon were used as food model systems to validate the in vitro findings. PEA had a growth inhibitory and bactericidal effect against L. monocytogenes both in in vitro experiments as well as on lyoner and smoked salmon. The MIC's ranged from 8 to 12.5 mg/mL. Furthermore, PEA also inhibited L. monocytogenes biofilm formation. Based on good manufacturing practices as a prerequisite, the application of PEA to RTE products might be an additional hurdle to limit L. monocytogenes growth thereby increasing food safety.
Project description:As the preparation of salads involves extensive handling and the use of uncooked ingredients, they are particularly vulnerable to microbial contamination. This study aimed to determine the microbial safety and quality of pre-packed salads and salad bar ingredients sold in Singapore, so as to identify public health risks that could arise from consuming salads and to determine areas for improvement in the management of food safety.The most frequently encountered organism in pre-packed salad samples was B. cereus, particularly in pasta salads (33.3%, 10/30). The most commonly detected organism in salad bar ingredients was L. monocytogenes, in particular seafood ingredients (44.1%, 15/34), largely due to contaminated smoked salmon. Further investigation showed that 21.6% (37/171) of the pre-packed smoked salmon sold in supermarkets contained L. monocytogenes. Significantly higher prevalence of L. monocytogenes and higher Standard Plate Count were detected in smoked salmon at salad bars compared to pre-packed smoked salmon in supermarkets, which suggested multiplication of the organism as the products move down the supply chain. Further molecular analysis revealed that L. monocytogenes Sequence Type (ST) 2 and ST87 were present in a particular brand of pre-packed salmon products over a 4-year period, implying a potential persistent contamination problem at the manufacturing level.Our findings highlighted a need to improve manufacturing and retail hygiene processes as well as to educate vulnerable populations to avoid consuming food prone to L. monocytogenes contamination.
Project description:Protocols for the specific detection of Listeria monocytogenes in cold-smoked salmon were developed. PCR was used as the method of detection. Inhibitors of PCR present in the food samples were removed by ether extraction or column purification, or their effect was overcome by the use of Tween 20 as an enhancer. These protocols are many times more rapid than conventional detection methodologies and also have the potential for automation.
Project description:In this study, RNA-seq was used to compare the transcriptomes of L. monocytogenes strain H7858 grown on cold smoked salmon (CSS) and in modified brain heart infusion broth (MBHIB, water-phase salt 4.65%, pH 6.1) at 7oC. RNA-seq was performed on H7858 RNA samples representing four independent biological replicates of growth experiments that involved growth of H7858 on CSS or in MBHIB. Samples for RNA preparation were collected when H7858 was grown to late log phase on these two matrices. Indexed and purified cDNA libraries (8 libraries including 4 replicates for each CSS and MBHIB) were loaded together onto an independent flow cell without any other samples; sequencing was carried out by running Hiseq 2000 (single-end, 100-bp per read). Reads alignment was carried out using the Burrows-Wheeler Aligner (BWA). Differential expression of genes in different strains was statistically assessed using the BaySeq method. Significant differential transcription of 149 genes in H7858 was observed between these two treatments including 88 genes and 61 genes up- and downregulated, respectively, in H7858 grown on CSS compared to in MBHIB. Our results show that genes involved in cobalamin biosynthesis, ethanolamine and 1,2-propanediol utilization, and specific carbohydrate transport systems have significantly higher transcript level in H7858 grown on CSS compared to in MBHIB. PrfA-dependent gene enrichment analysis showed that, genes regulated by PrfA were overrepresented among L. monocytogenes genes with higher transcript level on CSS than in MBHIB. Transcriptome profiles of L. monocytogenes grown on cold smoked salmon and in modified BHI broth at 7oC were generated by deep sequencing, in quadruplicate, using Illumina Hiseq 2000 (100 bp per read, single-end).
Project description:The foodborne pathogen Listeria monocytogenes is able to survive and grow in ready-to-eat foods, in which it is likely to experience a number of environmental stresses due to refrigerated storage and the physicochemical properties of the food. Little is known about the specific molecular mechanisms underlying survival and growth of L. monocytogenes under different complex conditions on/in specific food matrices. Transcriptome sequencing (RNA-seq) was used to understand the transcriptional landscape of L. monocytogenes strain H7858 grown on cold smoked salmon (CSS; water phase salt, 4.65%; pH 6.1) relative to that in modified brain heart infusion broth (MBHIB; water phase salt, 4.65%; pH 6.1) at 7°C. Significant differential transcription of 149 genes was observed (false-discovery rate [FDR], <0.05; fold change, ?2.5), and 88 and 61 genes were up- and downregulated, respectively, in H7858 grown on CSS relative to the genes in H7858 grown in MBHIB. In spite of these differences in transcriptomes under these two conditions, growth parameters for L. monocytogenes were not significantly different between CSS and MBHIB, indicating that the transcriptomic differences reflect how L. monocytogenes is able to facilitate growth under these different conditions. Differential expression analysis and Gene Ontology enrichment analysis indicated that genes encoding proteins involved in cobalamin biosynthesis as well as ethanolamine and 1,2-propanediol utilization have significantly higher transcript levels in H7858 grown on CSS than in that grown in MBHIB. Our data identify specific transcriptional profiles of L. monocytogenes growing on vacuum-packaged CSS, which may provide targets for the development of novel and improved strategies to control L. monocytogenes growth on this ready-to-eat food.
Project description:Vacuum-packaged cold-salted and cold-smoked fish products are considered typical vehicles for Listeria monocytogenes, the causative agent of the food-borne disease listeriosis, which is increasingly prevalent in the European Union. Efficacy of both the fish processing plant self-checking system and official food control conducted by authorities are crucial for L. monocytogenes prevention in the processing of these risky products. However, the impact of official control on L. monocytogenes prevention in the processing of fish products has not been extensively studied. We investigated the occurrence, control measures, and correction of non-compliances predisposing to L. monocytogenes in Finnish fish processing plants. The following features were associated with L. monocytogenes occurrence: (a) frequency of non-compliances concerning processing machinery, (b) recurrence of non-compliances, and (c) frequency of non-compliances for which official control measures were requested by inspecting authorities. Official control of fish processing plants had focused on risky areas, but non-compliances were common and their correction exhibited deficiencies. We conclude that L. monocytogenes prevention in fish processing can be enhanced by strengthening official food control measures and processing plant compliance. In particular, timely correction of all food safety violations must be improved.
Project description:The complete genome sequence of the persistent Listeria monocytogenes strain R479a isolated from smoked salmon in Denmark and belonging to lineage II, serovar 1/2a, and multilocus sequence type 8 (ST8) is presented here.
Project description:Listeria monocytogenes is a Gram-positive, foodborne pathogen responsible for approximately 28% of all food-related deaths each year in the United States. L. monocytogenes infections are linked to the consumption of minimally processed ready-to-eat (RTE) products such as cheese, deli meats, and cold-smoked finfish products. L. monocytogenes is resistant to stresses commonly encountered in the food-processing environment, including low pH, high salinity, oxygen content, and various temperatures. The purpose of this study was to determine if cells habituated at low temperatures would result in cross-protective effects against osmotic stress. We found that cells exposed to refrigerated temperatures prior to a mild salt stress treatment had increased survival in NaCl concentrations of 3%. Additionally, the longer the cells were pre-exposed to cold temperatures, the greater the increase in survival in 3% NaCl. A proteomics analysis was performed in triplicate in order to elucidate mechanisms involved in cold-stress induced cross protection against osmotic stress. Proteins involved in maintenance of the cell wall and cellular processes, such as penicillin binding proteins and osmolyte transporters, and processes involving amino acid metabolism, such as osmolyte synthesis, transport, and lipid biosynthesis, had the greatest increase in expression when cells were exposed to cold temperatures prior to salt. By gaining a better understanding of how this pathogen adapts physiologically to various environmental conditions, improvements can be made in detection and mitigation strategies.
Project description:We describe a novel quantitative real-time (Q)-PCR assay for Listeria monocytogenes based on the coamplification of a target hly gene fragment and an internal amplification control (IAC). The IAC is a chimeric double-stranded DNA containing a fragment of the rapeseed BnACCg8 gene flanked by the hly-specific target sequences. This IAC is detected using a second TaqMan probe labeled with a different fluorophore, enabling the simultaneous monitoring of the hly and IAC signals. The hly-IAC assay had a specificity and sensitivity of 100%, as assessed using 49 L. monocytogenes isolates of different serotypes and 96 strains of nontarget bacteria, including 51 Listeria isolates. The detection and quantification limits were 8 and 30 genome equivalents, and the coefficients for PCR linearity (R2) and efficiency (E) were 0.997 and 0.80, respectively. We tested the performance of the hly-IAC Q-PCR assay using various broth media and food matrices. Fraser and half-Fraser media, raw pork, and raw or cold-smoked salmon were strongly PCR-inhibitory. This Q-PCR assay for L. monocytogenes, the first incorporating an IAC to be described for quantitative detection of a food-borne pathogen, is a simple and robust tool facilitating the identification of false negatives or underestimations of contamination loads due to PCR failure.
Project description:In August 2017, an outbreak of six listeriosis cases in Denmark was traced to cold-smoked salmon, using epidemiological investigations and whole-genome sequencing (WGS) analyses. Exchange of genome sequences allowed identification in France of a food isolate from a salmon-derived product and a human isolate from 2016 within the same cgMLST cluster as the Danish isolates (L2-SL8-ST8-CT771). The salmon product came from a third European Union country. WGS can rapidly link human cases and food isolates across Europe.