Listeria monocytogenes is the causative agent of listeriosis, a severe foodborne infection that is increasing significantly in Europe and North America. A correlating factor contributing to the resurgence of listeriosis is the rise in consumption of cold-stored ready-to-eat (RTE) foods. The steady upsurge in disease requires more focused research to control the pathogen, L. monocytogenes. Currently, there is a plethora of diagnostic methods for the causative agent, however, each has limitations, one of which is the inability to correlate results across laboratories. This is a particular hindrance to an outbreak investigation in an age when food is transported widely across the globe. In this study, proteomic approaches were used to search for biomarkers that facilitate rapid characterisation of isolates against a background of differentially expressed proteins. A preamble to this investigation necessitated incorporation of an efficient lysis procedure to release maximum proteins. This was eventually achieved using a Listeria specific enzyme, endolysin, and a disruptive mechanical method. Liquid Chromatography-Mass Spectrometry/Mass Spectrometry (LC-MS/MS) data showed that bead beating and enzymatic lysis were the most efficient methods for analysis of the proteome. Dendrogram lineages, derived from MALD-TOF-MS spectra, strongly correlated with 16S rRNA analyses. Selective protein capture and analysis by MALD-TOF-MS (designated SELDI-TOF-MS) demonstrated considerable intraspecies diversity as revealed by dendrograms which were also visualised by „Heat Maps‟. One-dimensional polyacrylamide gel electrophoresis and LC-MS/MS analysis of seven L. monocytogenes isolates, led to the successful identification of two proteins; a hypothetical protein, designated lwe06778 and a phosphoribosyl-AMP cyclohydrolase which were uniquely present at 4°C. This finding suggests that L. monocytogenes depends on the histidine biosynthesis pathway in order to survive at cold temperatures. It is hypothesised that the addition of inhibitors, specific to both proteins in RTE cold foods may be a useful means for controlling outbreaks of listeriosis in the future.

Listeria monocytogenes is a human and animal pathogen responsible for severe and sometimes fatal infections. Several outbreaks have been associated with contaminated commercial foodstuffs such as raw milk, soft cheese, fresh and frozen milk, poultry, seafood, fruits and vegetable products. Currently, the official method recognized by the Government of Canada for the detection and isolation of L. monocytogenes can take up to six days without confirmation, which can require two more days. An approach based on molecular beacons that fluoresce upon hybridization was developed and tested to detect L. monocytogenes and the genus Listeria in food. Two different beacons were created: one specific to species L. monocytogenes (MG1) and another for the genus Listeria (MG2). Each of these molecular beacons was used with two separate sets of primers: MG1-1f/MG1-2r, MG1-7f/MG1-4r, MG2-2f/MG2-2r and MG2-3ft/MG2-2r. (Abstract shortened by UMI.)

The 16S/23S ribosomal RNA (rRNA) intergenic space (IGS) regions from pathogenic and non-pathogenic species (spp.) of Listeria were characterized by the polymerase chain reaction (PCR) and DNA sequencing. DNA sequencing data for the small rRNA IGS region showed that this IGS was approximately 244 bp in length and was highly homologous (95 to 99 %) in five of the six Listeria spp examined; ie., L. monocytogenes, L. innocua, L. seeligeri, L. welshimeri, and L. ivanovii. A lower degree of homology (91 to 94 %) was detected in the large rRNA IGS region (ca. 494 bp) of these species. The DNA sequence data was used to develop two sets of oligonucleotide primers for PCR-based detection of the members of the genus Listeria. The first set of primers were Listeria genus-specific and, the second set of primers were L. monocytogenes-specific. / xv, 131 leaves ; 29 cm.

With a primary focus on Health Canada (HC) and the Canadian Food Inspection Agency (CFIA), this thesis describes the state of microbial related public involvement and risk communication undertakings. The findings show that HC engages with experts to a far greater extent than with the lay public and that HC has not upheld its stated commitment to transparency. Furthermore, both HC’s and the CFIA´s approach to risk communication is overly general, has failed to provide opportunities for dialogue with vulnerable groups and is not rooted in foodborne surveillance data. Public involvement in food safety governance would be improved if HC provided the lay public with a seat on advisory committees and improved its reporting methods. HC and the CFIA could also make improvements by creating opportunities for dialogue between officials and the general public, and by exploring the potential use of alternative risk communication vehicles, such as food labels.

No description available.

With a primary focus on Health Canada (HC) and the Canadian Food Inspection Agency (CFIA), this thesis describes the state of microbial related public involvement and risk communication undertakings. The findings show that HC engages with experts to a far greater extent than with the lay public and that HC has not upheld its stated commitment to transparency. Furthermore, both HC’s and the CFIA´s approach to risk communication is overly general, has failed to provide opportunities for dialogue with vulnerable groups and is not rooted in foodborne surveillance data. Public involvement in food safety governance would be improved if HC provided the lay public with a seat on advisory committees and improved its reporting methods. HC and the CFIA could also make improvements by creating opportunities for dialogue between officials and the general public, and by exploring the potential use of alternative risk communication vehicles, such as food labels.

Cryptic plasmids ranging from 2.0 to 10C kb in size were isolated from 25 out of 122 Listeria monocytogenes strains, and from 7 out of 11 strains of other Listeria species. / Of 2500 clones of a genomic library of L. monocytogenes 81-861 generated in Escherichia coli cells, 5 clones were identified in which $ beta$-hemolytic activity was stably expressed. Testing by intraperitoneal injection showed that these clones were lethal to mice. Restriction mapping of the inserts of the recombinant plasmids showed that, apart from a 650-bp internal Hind III fragment in 2 inserts, there were no other common sites. No homology was demonstrated between the DNAs of the inserts when Southern blots of restriction digests of the 5 plasmids were probed, though homology was demonstrated between the L. monocytogenes listeriolysin O gene and the DNA of one insert. The evidence suggests that at least one additional $ beta$-hemolysin, other than listeriolysin O, exists in this strain of L. monocytogenes, and that it may be a virulence factor. / Using a direct colony hybridization procedure on hydrophobic grid-membrane filters (HGMFs), the inserts of the recombinant plasmids were screened, and a DNA probe specific for L. monocytogenes was identified. After labelling with horseradish peroxidase and colour development of the chromogen, a commercial counter (HGMF Interpreter) was able to detect and count the organism electronically. When the efficacy of the chromogen-labelled DNA probe method on HGMFs was compared with the conventional method for three artificially-inoculated foods, there were no significant differences ($ alpha$ = 0.05) shown in the recovery of L. monocytogenes from the foods.

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No description available.

Various control methods used in the meat and poultry processing environment to mitigate listeriosis were evaluated using a dynamic in-plant Monte Carlo model. These control methods included food contact surface testing, sanitation, post-processing lethality treatment, and product formulation with microbial growth inhibitors. The dynamic in-plant model served as an input into the risk assessment model developed by the FDA and FSIS in 2003 which predicts the number of deaths and illnesses resulting from the use of each control method. The use of growth inhibitors combined with a post-processing lethality step was estimated to save over 200 more lives than the FSIS proposed minimum sampling standard. An analysis of data collected by the National Alliance for Food Safety and Security (NAFSS) found that retail-sliced deli meats have a greater prevalence and concentration of L. monocytogenes than prepackaged deli meats. Cross contamination at the retail level is suspected due to clustering of sample positives by store and the influence of sampling time of day on the prevalence of L. monocytogenes. The comparative risk of Listeria monocytogenes in retail sliced versus prepackaged deli meats was evaluated using a modified version of the 2003 FDA-FSIS risk assessment model which considered slicing location and the use of growth inhibitors. The comparative risk ratio for the number of deaths from retail-sliced versus prepackaged deli meats was found to be 9.1 and retail-sliced product with a growth inhibitor was found to be at greater risk for listeriosis than prepackaged product without growth inhibitor. / Master of Science