Geraldine Duffy

Prevalence and concentration of verocytotoxigenic Escherichia coli, Salmonella enterica and Listeria monocytogenes in the beef production chain: A review

This review examines the prevalence of three important pathogens, verocytotoxigenic Escherichia coli (VTEC), Salmonella enterica and Listeria monocytogenes, in cattle and beef from the farm to the final, ready-to-eat product. Factors affecting prevalence of pathogens in the beef chain, such as the season and cattle rearing method, are examined. Data from many key surveys are summarized in table form. The observed prevalence of pathogens in cattle and beef varies considerably from survey to survey. An indication of relative prevalence of pathogens at different stages can be obtained by calculating average prevalences observed over multiple surveys, weighted by sample number. Based on the data presented in the tables in this review, for E. coli O157 at selected processing stages the mean prevalences (and range of means from individual surveys) are faeces 6.2% (0.0-57%), hides 44% (7.3-76%), chilled carcasses 0.3% (0.0-0.5%), and raw beef products 1.2% (0.0-17%). For Salmonella the mean prevalence data are faeces 2.9% (0.0-5.5%), hides 60% (15-71%), chilled carcasses 1.3% (0.2-6.0%), and raw beef products 3.8% (0.0-7.5%). For L. monocytogenes the mean prevalence data are faeces 19% (4.8-29%), hides 12% (10-13%), and raw beef products 10% (1.6-24%). Seasonal variation was evident in many surveys, faecal prevalences of E. coli O157 and Salmonella generally being higher in the warmer months. The influence of animal type, animal age, feed and housing on pathogen carriage has also been examined. The significance of non-O157 serotypes of VTEC and their detection and classification are discussed.

Antibiotic resistance among Listeria, including Listeria monocytogenes, in retail foods

Aims: In the past eight to 10 years, reports of antibiotic resistance in food‐borne isolates in many countries have increased, and this work examined the susceptibility of 1001 food isolates of Listeria species.

Development of a real-time multiplex PCR assay for the detection of multiple Salmonella serotypes in chicken samples

BackgroundA real-time multiplex PCR assay was developed for the detection of multiple Salmonella serotypes in chicken samples. Poultry-associated serotypes detected in the assay include Enteritidis, Gallinarum, Typhimurium, Kentucky and Dublin. The traditional cultural method according to EN ISO 6579:2002 for the detection of Salmonella in food was performed in parallel. The real-time PCR based method comprised a pre-enrichment step in Buffered Peptone Water (BPW) overnight, followed by a shortened selective enrichment in Rappaport Vasilliadis Soya Broth (RVS) for 6 hours and subsequent DNA extraction.ResultsThe real-time multiplex PCR assay and traditional cultural method showed 100% inclusivity and 100% exclusivity on all strains tested. The real-time multiplex PCR assay was as sensitive as the traditional cultural method in detecting Salmonella in artificially contaminated chicken samples and correctly identified the serotype. Artificially contaminated chicken samples resulted in a detection limit of between 1 and 10 CFU per 25 g sample for both methods. A total of sixty-three naturally contaminated chicken samples were investigated by both methods and relative accuracy, relative sensitivity and relative specificity of the real-time PCR method were determined to be 89, 94 and 87%, respectively. Thirty cultures blind tested were correctly identified by the real-time multiplex PCR method.ConclusionReal-time PCR methodology can contribute to meet the need for rapid identification and detection methods in food testing laboratories.

Verocytoxigenic Escherichia coli in animal faeces, manures and slurries

Animal wastes and effluents from farming operations, including manures and slurries, are frequently applied as fertilizer to land used for crop or silage production and cattle grazing. It is well documented that potentially harmful pathogens including verocytoxigenic Escherichia coli (VTEC) are shed in animal faeces and there is growing concern in many countries about the number of sporadic and outbreak cases of VTEC attributable to direct contact with faecal material either as a result of handling contaminated mud in fields or ingestion of produce grown in contaminated manures or slurries. VTEC has been detected in the faeces of ruminant and non-ruminant farmed animals, wild animals, domestic pets and birds and the pathogen appears to be well adapted to survive in animal faeces and can persist for extended periods ranging from several weeks to many months. Because of this persistence these materials are important as potential vehicles for transmission within herds, farms, the fresh food chain and the wider environment. Appropriate handling of bovine faeces is necessary to control spread of this pathogen and to limit the significant risks of human infection. It may be necessary to hold manure/slurry for extended periods prior to spreading on farmland, or for use in the production of food crops, particularly foods that are to be consumed in the raw or minimally processed state. Alternatively, it may be necessary to apply processes such as composting, heat drying or digestion which can expedite the decline of pathogens including VTEC in manures. However, there is a need for research work to develop economical and practical systems for treatment of manures and slurries. The risk from direct contact with faecal material at farms and petting zoos is also recognized and many public health authorities have put forward measures for strict practices to limit the risk of infection, particularly for young children visiting these environments.

Survival of Escherichia coli O157:H7 during the manufacture of pepperoni.

This study investigated the growth and survival of Escherichia coli O157:H7 during the manufacture of pepperoni to determine whether a 5-log10-unit decline in numbers, as recommended by the U.S. Food Safety and Inspection Service (FSIS), could be achieved. A range of pepperoni formulations with variations in salt (2.5 to 4.8%) and sodium nitrite (100 to 400 ppm) levels, and with pH (4.4 to 5.6) adjusted by manipulation of dextrose concentrations were prepared. The batters produced were inoculated with E. coli O157:H7 380-94 at a level of approximately 6.70 log10 CFU/g; changes in pathogen numbers, pH, titratable acidity, and sodium nitrite concentrations were monitored during fermentation and drying. With the standard commercial formulation (i.e., 2.5% salt, 100 ppm sodium nitrite, pH 4.8) E. coli O157:H7 numbers declined by approximately 0.41 log10 CFU/g during fermentation and a further 0.43 log10 CFU/g during subsequent drying (7 days). A regression equation was fitted to the data which showed significantly (P < 0.001) greater reductions in pathogen numbers in samples with increased salt and sodium nitrite contents and lowered pH. However declines were in all cases less than the target reduction of 5 log10 CFU/g.

Effects of Acid Adaptation, Product pH, and Heating on Survival of Escherichia coli O157:H7 in Pepperoni

ABSTRACT The thermotolerance of E. coli O157:H7 cells (strain 380-94) heated in pepperoni is reported. Information on the pattern of thermal inactivation of E. coli O157:H7 in pepperoni was applied in the development of heating processes designed to reduceE. coli O157:H7 numbers therein by 5 log10units.

A review of quantitative microbial risk assessment in the management of Escherichia coli O157:H7 on beef

Since Escherichia coli O157:H7 first emerged as a food borne pathogen in the mid 1980s, it has been linked to many cases of food poisoning across the world. While multiple sources and routes of transmission for this pathogen are now recognised, beef and beef products remain an important vehicle of the pathogen and continue to be linked to outbreaks across the developed world. Much research has been directed at E. coli O157:H7 transmission, survival and control in the beef chain and this paper presents an overview of current knowledge on this pathogen in the beef chain from primary production through slaughter, processing, distribution, final preparation and cooking. In order to strategically manage E. coli O157:H7 and to devise approaches to reduce the public health risk posed, many national and international groups have applied quantitative risk assessment techniques to model the risk posed by E. coli O157:H7 in beef, particularly in ground/minced beef which is most often linked with infection. This paper reviews these quantitative risk assessments and their application in managing the risk posed by E. coli O157:H7 in beef.

Inhibition of verocytotoxigenic Escherichia coli in model broth and rumen systems by carvacrol and thymol

The antimicrobial activities of thymol and carvacrol were assessed against a selection of verocytotoxigenic Escherichia coli (VTEC) strains (n=11) and other bacterial species and spoilage bacteria (n=7) using a model broth system. The effects of pH, temperature, water activity, sodium chloride concentrations, inoculum size and the presence of competing microflora on the activities of thymol and carvacrol against E. coli O157:H7 strain 380-94 were also determined. The minimum inhibitory and bactericidal concentrations (MIC and MBC, respectively) and numbers of surviving E. coli O157:H7 were determined following incubation. The mean numbers of VTEC surviving exposure to thymol or carvacrol at concentrations of >/=500mug/ml were between 2.0 and 7.8log cfu/ml less than the numbers in the corresponding controls. The susceptibility of E. coli O157:H7 to carvacrol or thymol was found to increase with decreasing storage temperature, water activity, pH and E. coli O157:H7 inoculum size. Sodium chloride (0.5-2.5%) and the presence of a microflora cocktail did not significantly (p>0.05) affect the antimicrobial activities of thymol or carvacrol against E. coli O157:H7. The antimicrobial activity of carvacrol against E. coli O157:H7 was also tested in a model rumen system. A MIC of 500mug/ml carvacrol reduced E. coli O157:H7 inoculated at levels of 10(3) and 10(6)cfu/ml to undetectable levels in the system after 24h incubation. This concentration of carvacrol significantly (p<0.05) decreased the total gas production and volatile fatty acid concentrations in the model rumen assay.

The occurence and initial numbers of Listeria in Irish meat and fish products and the recovery of injured cells from frozen products

A total of 549 samples of meat, fish and poultry products purchased from retail outlets in the Dublin area were examined for the presence of Listeria spp., using a standard recovery method and a new resuscitation method. Listeria spp. were most frequently isolated from frozen beef burgers (97%) and fish fingers (95%). Cooked meats which were prepackaged by the manufacturer were negative for Listeria spp. The pathogen was isolated from 21% of cooked meats which were sold retail unpackaged indicating post-process contamination. Standard recovery methods gave Listeria counts of between 0.7 and log10 5.0 cfu/g on frozen products. Using a resuscitation method, counts were up to log10 2.5 cfu/g higher, indicating the presence of large numbers of injured Listeria cells. The significance of the numbers of Listeria found in the various foods as well as the recovery of injured cells with the new resuscitation method is discussed.

Tracking verocytotoxigenic Escherichia coli O157, O26, O111, O103 and O145 in Irish cattle.

The purpose of this study was to investigate carriage and transfer of verocytotoxigenic Escherichia coli (VTEC) O157, O26, O111, O103 and O145 from faeces and hide to dressed carcasses of Irish cattle as well as establishing the virulence potential of VTEC carried by these cattle. Individual cattle was tracked and faecal samples, hide and carcass (pre-evisceration and post-wash) swabs were analysed for verotoxin (vt1 and vt2) genes using a duplex real-time PCR assay. Positive samples were screened for the five serogroups of interest by real-time PCR. Isolates were recovered from PCR positive samples using immunomagnetic separation and confirmed by latex agglutination and PCR. Isolates were subject to a virulence screen (vt1, vt2, eaeA and hlyA) by PCR. Isolates carrying vt genes were examined by Pulsed-Field Gel Electrophoresis (PFGE). Of the VTEC isolated, E. coli O157 was the most frequently recovered from hide (17.6%), faeces (2.3%) and pre-evisceration/post-wash carcass (0.7%) samples. VTEC O26 was isolated from 0.2% of hide swabs and 1.5% of faeces samples. VTEC O145 was isolated from 0.7% of faeces samples. VTEC O26 and VTEC O145 were not recovered from carcass swabs. Non-VTEC O103 was recovered from all sample types (27.1% hide, 8.5% faeces, 5.5% pre-evisceration carcass, 2.2% post-wash carcass), with 0.2% of hide swabs and 1.0% of faeces samples found to be positive for VTEC O103 isolates. E. coli O111 was not detected in any samples. For the four serogroups recovered, the direct transfer from hide to carcass was not observed. This study shows that while VTEC O157 are being carried by cattle presented for slaughter in Ireland, a number of other verotoxin producing strains are beginning to emerge.