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Aims To develop an oxygen sensor-based method for testing total aerobic viable counts (TVC) in raw meat samples and cattle carcass swabs, which is rapid, simple, affordable, provides good sensitivity and analytical performance and allows on-site use. Methods and Results The test uses the same sample preparation procedure as the established plate counting TVC method for meat samples and carcasses, ISO4833-1:2013. After this liquid samples are transferred into standard 25-ml vials with built-in phosphorescent O-2 sensors and incubated on a block heater with hourly readings of sensor signals with a handheld reader, to determine signal threshold time (TT, hours) for each sample. The method is demonstrated with the quantification of TVC in industrial cuts of raw beef meat (CFU per g) and carcass swabs (CFU per cm(2)). Calibration curves were generated, which give the following analytical equations for calculating the TVC load in unknown samples from measured TT values: TVC [Log(CFU per cm(2))] = 7.83-0.73*TT(h) and TVC [Log(CFU per g)] = 8.74-0.70*TT(h) for the carcass swabs and meat samples respectively. The new tests show good correlation with the ISO methods, with correlation coefficients 0.85 and 0.83 respectively. The testing requires no dilutions, covers the ranges 2-7 Log(CFU per g) for the meat samples and 1-7 Log(CFU per cm(2)) for carcass swabs, and has time to result 1-10 h with faster detection of more contaminated samples. Conclusions The sensor-based testing demonstrates simplicity, high speed, sample throughput and automation. It can provide a straightforward replacement for the conventional TVC tests, which are time consuming, laborious and have time to result of 48-72 h. Significance and Impact of the Study The method(s) can be adopted by the meat industry and research labs, and used to improve microbial quality and safety of meat products and processes.

Escherichia coli (E. coli) are a group of bacteria, which are a natural part of the intestinal flora of warm-blooded animals, including humans. Most E. coli are non-pathogenic and are essential for the normal function of a healthy intestine, however, certain types such as Shiga toxin-producing E. coli (STEC) is a foodborne pathogen that can cause a life-threatening illness. Development of point-of-care (POC), rapid detection devices for E. coli have high importance from a food safety point of view. The most suitable way to distinguish between generic E. coli and STEC is by using nucleic acid-based detection, focusing on the virulence factors. Electrochemical sensors based on nucleic acid recognition have attracted much attention in recent years for use in pathogenic bacteria detection. This review has summarised nucleic acid-based sensors for the detection of generic E. coli and STEC since 2015. First, the sequences of the genes used as recognition probes are discussed and compared to the most recent research regarding the specific detection of general E. coli and STEC. Subsequently, the collected literature regarding nucleic acid-based sensors was described and discussed. The traditional sensors were divided into four categories such as gold, ITO, carbon-based electrodes, and those using magnetic particles. Finally, we summarised the future trends in nucleic acid-based sensor development for E. coli and STEC including some examples of fully integrated devices

Shiga toxin-producing E. coli (STEC) is a food-borne pathogen of significant public health concern, due to the severity of the illness it can cause and a low infection dose. The key targets in molecular-based assays to detect STEC are stx1 or stx2 genes, coding for the ability to produce Shiga T Toxin 1 and 2. The most commonly used molecular detection techniques, such as PCR and real-time PCR, are considerably time-consuming and there is an urgent need for a more rapid screening assay which could be used in agri-food settings. In this work, an amplification-free multiplex electrochemical sensor for the simultaneous detection of stx1 and stx2 genes was developed using a silicon-based chip comprising of six interdigitated gold microelectrodes(IDE) sensors. Two probes complementary to stx1 and stx2 genes were immobilised on a single chip allowing for multiplex detection. The selectivity of the multiplex sensor was confirmed using fluorescence and electrochemistry. The developed assay conditions allowed for an improved limit of detection by three orders of magnitude compared to the previous reports achieving a amplification-free limit of detection of 100 zM (10-19 M) for both, stx1 and stx2 genes. The developed sensor was validated using chromosomal DNA extracted from a bacterial culture containing no virulence genes, stx1 gene only, and both stx1 and stx2. Such multiplex sensor, if combined with on-chip DNA extraction, could revolutionise the point-of-use detection of STEC as well as other pathogens for instance on-farm or in the food industry.

Shiga-toxin producing Escherichia coli (STEC) are zoonotic foodborne pathogens that are capable of causing serious human illness. Ovine ruminants are recognized as an important source of STEC and a notable contributor to contamination within the food industry. This review examined the prevalence of STEC in the ovine food production chain from farm-to-fork, reporting carriage in sheep herds, during abattoir processing, and in raw and ready-to-eat meats and meat products. Factors affecting the prevalence of STEC, including seasonality and animal age, were also examined. A relative prevalence can be obtained by calculating the mean prevalence observed over multiple surveys, weighted by sample number. A relative mean prevalence was obtained for STEC O157 and all STEC serogroups at multiple points along the ovine production chain by using suitable published surveys. A relative mean prevalence (and range) for STEC O157 was calculated: for feces 4.4% (0.2-28.1%), fleece 7.6% (0.8-12.8%), carcass 2.1% (0.2-9.8%), and raw ovine meat 1.9% (0.2-6.3%). For all STEC independent of serotype, a relative mean prevalence was calculated: for feces 33.3% (0.9-90.0%), carcass 58.7% (2.0-81.6%), and raw ovine meat 15.4% (2.7-35.5%). The prevalence of STEC in ovine fleece was reported in only one earlier survey, which recorded a prevalence of 86.2%. Animal age was reported to affect shedding in many surveys, with younger animals typically reported as having a higher prevalence of the pathogen. The prevalence of STEC decreases significantly along the ovine production chain after the application of postharvest interventions. Ovine products pose a small risk of potential STEC contamination to the food supply chain.

In this research, a selective, cost-efficient, highly sensitive Ag nanostructure Surface Enhanced Raman Spectroscopy (SERS) sensor was developed as a methodological approach to rapidly detect a targeted-ss-DNA (stx2) in STEC (Shiga toxin-producing Escherichia coli). The Ag nanostructure-based SERS substrate was functionalized by two type of thiols: thiol-ss-DNA for bonding targeted-ss-DNA and 6-Mercapto-1-hexanol (HS(CH2)6OH) for blocking the Ag nanostructure surface. Methylene Blue (MB) was used as a Raman marker to quantify targeted-ss-DNA, as well as a model molecule to characterize the electrodeposited Ag nanostructure SERS substrate. Ag nanostructure SERS substrates showed good sensitivity and repeatability towards MB detection, with a LOD = 0.3158 μM, RSD = 12.48% (at 45 different random points). More importantly, the Ag nanostructure/ss-DNA SERS substrate showed good selectivity towards STEC O157 stx2 targeted DNA, as well as good linearity and sensitivity towards its detection in a buffer solution. A limit of detection of 0.4900 aM in a wide linear range from 1 aM to 100 pM was demonstrated. The SERS sensors were able to identify target DNA ( stx2) in an STEC strain and the study showed proof of principle that SERS substrate has potential as a cost effective, highly selective, highly sensitive DNA and bacteria sensor without the aid of DNA amplification. With further development and validation, this methodological approach has the potential for use in the point-of-use detection for instance on a farm or in the food industry.

Shiga toxin-producing Escherichia coli (STEC) organisms are a diverse group of pathogenic bacteria capable of causing serious human illness, and serogroups O157 and O26 are frequently implicated in human disease. Ruminant hosts are the primary STEC reservoir, and small ruminants are important contributors to STEC transmission. This study investigated the prevalence, serotypes, and shedding dynamics of STEC, including the supershedding of serogroups O157 and O26, in Irish sheep. Recto-anal mucosal swab samples (n = 840) were collected over 24 months from two ovine slaughtering facilities. Samples were plated on selective agars and were quantitatively and qualitatively assessed via real-time PCR (RT-PCR) for Shiga toxin prevalence and serogroup. A subset of STEC isolates (n = 199) were selected for whole-genome sequencing and analyzed in silico. In total, 704/840 (83.8%) swab samples were Shiga toxin positive following RT-PCR screening, and 363/704 (51.6%) animals were subsequently culture positive for STEC. Five animals were shedding STEC O157, and three of these were identified as supershedders. No STEC O26 was isolated. Post hoc statistical analysis showed that younger animals are more likely to harbor STEC and that STEC carriage is most prevalent during the summer months. Following sequencing, 178/199 genomes were confirmed as STEC. Thirty-five different serotypes were identified, 15 of which were not yet reported for sheep. Serotype O91:H14 was the most frequently reported. Eight Shiga toxin gene variants were reported, two stx(1) and six stx(2), and three novel Shiga-toxin subunit combinations were observed. Variant stx(1c) was the most prevalent, while many strains also harbored stx(2b). IMPORTANCE Shiga toxin-producing Escherichia coli (STEC) bacteria are foodborne, zoonotic pathogens of significant public health concern. All STEC organisms harbor stx, a critical virulence determinant, but it is not expressed in most serotypes. Sheep shed the pathogen via fecal excretion and are increasingly recognized as important contributors to the dissemination of STEC. In this study, we have found that there is high prevalence of STEC circulating within sheep and that prevalence is related to animal age and seasonality. Further, sheep harbor a variety of non-O157 STEC, whose prevalence and contribution to human disease have been underinvestigated for many years. A variety of Stx variants were also observed, some of which are of high clinical importance.

Microbial food contamination is a major concern for consumers and food industries. Consumers desire nutritious, safe and “clean label” products, free of synthetic preservatives and food industries and food scientists try to meet their demands by finding natural effective alternatives for food preservation. One of the alternatives to synthetic preservatives is the use of natural anti-microbial agents in the food products and/or in the packaging materials. Meat and processed meat products are characteristic examples of products that are highly perishable; hence natural anti-microbials can be used for extending their shelf-life and enhancing their safety. Despite several examples of the successful application of natural anti-microbial agents in meat products reported in research studies, their commercial use remains limited. This review objective is to present an extensive overview of recent research in the field of natural anti-microbials, covering essential oils, plant extracts, flavonoids, animal-derived compounds, organic acids, bacteriocins and nanoparticles. The anti-microbial mode of action of the agents, in situ studies involving meat products, regulations and, limitations for usage and future perspectives are described. The review concludes that naturally derived anti-microbials can potentially support the meat industry to provide “clean label”, nutritious and safe meat products for consumers.

Vacuum packaging (VP) is used to reduce exposure of retail meat samples to ambient oxygen (O2) and preserve their quality. A simple sensor system produced from commercial components is described, which allows for non-destructive monitoring of the O2 concentration in VP raw meat samples. Disposable O2 sensor inserts were produced by spotting small aliquots of the cocktail of the Pt–benzoporphyrin dye and polystyrene in ethyl acetate onto pieces of a PVDF membrane and allowing them to air-dry. These sensor dots were placed on top of the beef cuts and vacuum-packed. A handheld reader, FirestinGO2, was used to read nondestructively the sensor phase shift signals (dphi°) and relate them to the O2 levels in packs (kPa or %). The system was validated under industrial settings at a meat processing plant to monitor O2 in VP meat over nine weeks of shelf life storage. The dphi° readings from individual batch-calibrated sensors were converted into the O2 concentration by applying the following calibration equation: O2 (%) = 0.034 * dphi°2 − 3.413 * dphi° + 85.02. In the VP meat samples, the O2 levels were seen to range between 0.12% and 0.27%, with the sensor dphi signals ranging from 44.03° to 56.02°. The DIY sensor system demonstrated ease of use on-site, fast measurement time, high sample throughput, low cost and flexibility.

Response surface methodology was used to develop sensory acceptable, low-salt, shelf-stable frankfurters. A Box–Behnken experimental design assessed the effects of three independent factors: salt replacer (Artisalt™) (0–100%), high-pressure processing (HPP) (0.1–600 MPa) and a mix of organic acids (Inbac™) (0.2–0.4%). Measured responses included: hardness, flavour, saltiness and overall sensory acceptability (OSA) of the frankfurters. The primary factor affecting (P

The objective of this study was to assess the acceptability and consumer (n = 100) appeal of research-optimised low-salt (ROLS) frankfurters or cooked ham manufactured using high-pressure processing (HPP) and organic acids as hurdles and compared to research control and gold standard commercially available products. For frankfurters, consumers preferred the firmness and saltiness of the ROLS and research control frankfurters while the flavour and juiciness of commercial frankfurters was preferred. In terms of overall sensory acceptability (OSA), the ROLS frankfurter was liked just as much as the commercial brand frankfurter. For cooked ham, the appearance and firmness of ROLS and research control cooked ham was preferred while the juiciness of the commercial cooked ham was preferred. Consumers did not find significant differences in flavour, saltiness or OSA and the ROLS cooked ham was liked just as much as the commercial brand cooked ham. Overall, these results indicate that the ROLS processed meat products were just as acceptable or better than the gold standard commercially available products confirming the potential of the use of response surface methodology to optimise salt replacer Artisalt™, HPP and organic acids to manufacture consumer-accepted low-salt processed meat products with enhanced safety and shelf life.

Shiga-toxin producing

Air is recognized as an important source of microbial contamination in food production facilities and has the potential to contaminate the food product causing food safety and spoilage issues for the food industry. Potential for aerial microbial contamination of food can be a particular issue during storage in cold rooms when the food is not packaged and is exposed to contaminated air over a prolonged period. Thus, there are potential benefits for the food industry for an aerial decontamination in cold storage facilities. In this paper, aerial decontamination approaches are reviewed and challenges encountered for their applications are discussed. It is considered that current systems may not be completely effective and environmentally friendly, therefore, it is of great significance to consider the development of nonresidual and verified decontamination technologies for the food industry and, in particular, for the cold storage rooms.

Epidemic and pandemic clones of bacterial pathogens with distinct characteristics continually emerge, replacing those previously dominant through mechanisms that remain poorly characterized. Here, whole-genome-sequencing-powered epidemiology linked horizontal transfer of a virulence gene, sopE, to the emergence and clonal expansion of a new epidemic Salmonella enterica serovar Typhimurium (S. Typhimurium) clone. The sopE gene is sporadically distributed within the genus Salmonella and rare in S . enterica Typhimurium lineages, but was acquired multiple times during clonal expansion of the currently dominant pandemic monophasic S. Typhimurium sequence type (ST) 34 clone. Ancestral state reconstruction and time-scaled phylogenetic analysis indicated that sopE was not present in the common ancestor of the epidemic clade, but later acquisition resulted in increased clonal expansion of sopE-containing clones that was temporally associated with emergence of the epidemic, consistent with increased fitness. The sopE gene was mainly associated with a temperate bacteriophage mTmV, but recombination with other bacteriophage and apparent horizontal gene transfer of the sopE gene cassette resulted in distribution among at least four mobile genetic elements within the monophasic S . enterica Typhimurium ST34 epidemic clade. The mTmV prophage lysogenic transfer to other S. enterica serovars in vitro was limited, but included the common pig-associated S . enterica Derby (S. Derby). This may explain mTmV in S. Derby co-circulating on farms with monophasic S. Typhimurium ST34, highlighting the potential for further transfer of the sopE virulence gene in nature. We conclude that whole-genome epidemiology pinpoints potential drivers of evolutionary and epidemiological dynamics during pathogen emergence, and identifies targets for subsequent research in epidemiology and bacterial pathogenesis.

Shiga toxigenic Escherichia coli (STEC) are an important group of pathogens and can be transmitted to humans from direct or indirect contact with cattle faeces. This study investigated the shedding of E. coli O157 and O26 in cattle at the time of slaughter and factors associated with super-shedding (SS) animals. Rectoanal mucosal swab (RAMS) samples were collected from cattle (n = 1,317) at three large Irish commercial beef abattoirs over an 18 month period, and metadata were collected at the time of sampling regarding farm of origin, animal age, breed and gender. RAMS swabs were examined for the presence and numbers of E. coli O157 and O26 using a previously developed quantitative real-time PCR protocol. Samples positive by PCR were culturally examined and isolates analysed for the presence of stx subtypes, eae and phylogroup. Any samples with counts >104 CFU/swab of STEC O157 or O26 were deemed to be super-shedders. Overall, 4.18% (55/1,317) of RAMS samples were positive for STEC O157, and 2.13% (28/1,317) were classified as STEC O157 SS. For STEC O26, 0.76% (10/1,317) of cattle were positive for STEC O26, and 0.23% (3/1,317) were classified as super-shedders. Fewer STEC shedders and SS were noted among older animals (>37 months). There was a seasonal trend observed for STEC O157, with the highest prevalence of shedding and SS events in the autumn (August to October). The majority of E. coli O157 (50/55) isolates had stx2 and were eae positive, with no significant difference between SS and low shedders (LS). Interestingly, all STEC O26 (n = 10) were eae negative and had varied stx profiles. This study demonstrates that, while the overall shedding rates are relatively low in cattle at slaughter, among positive animals there is a high level of SS, which may pose a higher risk of cross-contamination during slaughter.

The objective of this study was to develop sensory accepted low-salt cooked ham by the application of response surface methodology (RSM). A Box-Behnken experimental design was used to assess the effects of the independent factors salt replacer (Artisalt™) (0–100%), high pressure treatment (0.1–600 MPa) and a mix of organic acids (Inbac™) (0.2–0.4%) on hardness, flavour, saltiness and overall sensory acceptability (OSA) of the cooked ham. The main factor that affected all response variables was salt replacement. The optimum parameters to maximise salt reduction and produce hams with similar OSA associated with this type of products were Artisalt™ (53%), HPP (535 MPa) and Inbac™ (0.3%) and the cooked ham manufactured using the optimum parameters contained 1.4% total salt which is a 46% reduction compared to control samples which contained 2.6% total salt. Overall, a combination of salt replacer, HPP and organic acids showed great potential for the development of cooked ham with significantly reduced salt content. Industrial relevance Consumer studies have shown that meat consumption is being more and more influenced by health, nutritional and environmental considerations; therefore, companies are constantly searching for new and emerging technologies to reduce salt in meat products and enhance shelf life to reduce food waste. In this study we used a novel approach which showed great potential in salt reduction of ham as the quality and sensory acceptability of the ham were similar and/or better after salt was replaced by 53%. The hurdle approach used in this study is expected to improve the safety and shelf life of the low-salt optimised ham and this confirmatory study is underway.

This study aimed to provide new insights into the epidemiology of Salmonella in pig production, focusing on potential shedding patterns in breeding pigs throughout a full production cycle and the risk of transmission of infection from the sow to her offspring. A longitudinal study was conducted on five farrow-to-finish commercial pig farms. In each herd, shedding of Salmonella in faeces was monitored in breeders through service, gestation and lactation. Swabs of the farrowing room floor and pools of faeces from piglets were collected on two occasions during lactation. Environmental pen swabs were also taken in the weaning and finisher houses. Salmonella isolates were serotyped, tested for antimicrobial resistance (AMR) and typed by Multiple-Locus Variable number tandem repeat Analysis (MLVA). Shedding by breeding pigs was low in all stages of the production cycle; 5% of sows shed at service, the production stage with highest risk of shedding (p

Water misting/showers are used in abattoir lairages to improve meat quality, and to cool and calm pigs after transport and during hot weather. One novel approach, which has not been investigated to date, is to add a disinfectant to the misting water as a means of topically reducing Salmonella on pigs prior to slaughter, thereby potentially controlling this organism in the abattoir. The objective of this study was therefore to evaluate misting with water or with Virkon® S (an approved disinfectant for use in the presence of animals), for their ability to topically reduce Salmonella on high seroprevalence pig herds before stunning and to reduce Enterobacteriaceae. Three experimental groups were investigated: control group (i.e., no misting); water group (misting with cold, 15–17 °C, water, herein referred to as water); and a disinfectant group (misting with 0.5% Virkon® S). As pigs entered the abattoir, each animal was swabbed along its back before being allocated to its experimental group. Each group was randomly assigned to one of 3 lairage pens that were separated by non-trial pens. After 30 min of misting with water or disinfectant, pigs were moved to the stunning area, where each pig was again swabbed, as above. Swabs were analyzed for the presence of Salmonella and enumeration of Enterobacteriaceae. Before misting, Salmonella prevalence on the pigs was 79.0%, 72.1% and 83.6% for the control, water and disinfectant groups, respectively. After misting, Salmonella prevalence increased to 94.3% in the water group; whereas for the disinfectant group, the prevalence increased marginally to 85.9%. No change in Salmonella prevalence was detected for the control group. In line with the Salmonella results, no significant differences were observed in Enterobacteriaceae counts in the control group at either time point (4.37 and 5.01 log10 CFU/cm2, respectively) or in the disinfectant group before and after misting (4.02 and 4.26 log10 CFU/cm2, respectively). However, a 2.3 log10 CFU/cm2 increase in Enterobacteriaceae was recorded for the water group after misting as compared to before misting (p Since misting with water alone increased topical Salmonella contamination on pigs before slaughter, a risk assessment based on known Salmonella data, meat quality and welfare is recommended to determine whether its use is justifiable. On the other hand, the findings from this study suggest that misting with Virkon® S at 0.5% could have a role in topical antisepsis of pigs contaminated with Salmonella prior to slaughter and as such this warrants further investigation.

This study investigated several cleaning and disinfection protocols for their ability to eliminate Salmonella and to reduce levels of Enterobacteriaceae, within the lairage pens of a commercial pig abattoir. Eight protocols were evaluated in each of 12 lairage pens at the end of the slaughtering day on 3 occasions (36 pens/protocol): (P1) high-pressure cold water wash (herein referred to as high-pressure wash); (P2) high-pressure wash followed by a quaternary ammonium compound (QAC)-based disinfectant without rinsing; (P3) high-pressure wash followed by a chlorocresol-based disinfectant without rinsing; (P4) high-pressure wash followed by a sodium hydroxide/sodium hypochlorite detergent with rinsing; (P5) P4 followed by P2; (P6) P4 followed by P3; (P7) P5 with drying for 24-48h; and (P8) P6 with drying for 24-48h. Two floor swabs and one wall swab were taken from each lairage pen before and after each protocol was applied, and examined for the presence of Salmonella and enumeration of Enterobacteriaceae. High-pressure washing alone (P1) did not reduce the prevalence of Salmonella in the lairage pens. When high-pressure washing, the probability of detecting Salmonella following application of the chlorocresol-based disinfectant (P3) was lower than with the QAC-based disinfectant, P2 (14.2% versus 34.0%, respectively; p

Salmonella Typhimurium and its monophasic variant S. 4,[5],12:i:- are the dominant serotypes associated with pigs in many countries. We investigated their population structure on nine farms using whole genome sequencing, and their genotypic and phenotypic variation. The population structure revealed the presence of phylogenetically distinct clades consisting of closely related clones of S. Typhimurium or S. 4,[5],12:i:- on each pig farm, that persisted between production cycles. All the S. 4,[5],12:i:- strains carried the Salmonella genomic island-4 (SGI-4), which confers resistance to heavy metals, and half of the strains contained the mTmV prophage, harbouring the sopE virulence gene. Most clonal groups were highly drug resistant due to the presence of multiple antimicrobial resistance (AMR) genes, and two clades exhibited evidence of recent on-farm plasmid-mediated acquisition of additional AMR genes, including an IncHI2 plasmid. Biofilm formation was highly variable but had a strong phylogenetic signature. Strains capable of forming biofilm with the greatest biomass were from the S. 4,[5],12:i:- and S. Typhimurium DT104 clades, the two dominant pandemic clones found over the last 25 years. On-farm microevolution resulted in enhanced biofilm formation in subsequent production cycle.

peer-reviewed The objective of this study was to assess the effect of griddle and steam cooking on the physicochemical and sensory characteristics of high pressure processed (HPP) piri-piri marinated pork chops (MPC). Raw MPC that were HPP at 400 MPa had higher (P

Shiga toxin producing E. coli (STEC) is a foodborne pathogen which causes severe, debilitating, and sometimes fatal, illness. Ireland consistently has one of the highest incidence rates of human STEC infection in Europe. Cattle are one of the primary reservoirs for STEC, excreting the pathogen in their faeces. The amount of pathogen excreted varies greatly, with animals shedding >log104 c.f.u. g−1 faeces being termed ‘super-shedders’. Human infection can occur from faecal contamination of meat, dairy, fresh produce or drinking water. STEC can survive for extended periods in soil, slurry and water, although the exact means is unknown. The objective of this study was to examine phenotypic traits potentially relevant to extended environmental survival in two strain banks: (1) clinical and bovine STEC, in comparison with non-STEC, isolated from the production environment and (2) E. coli O157:H7 of known shedding status. The strain banks were assessed for biofilm-forming abilities and the ability to adhere to the muscle component collagen-I, using a 96-well crystal violet assay, where the absorbance of bound cells indicated biofilm formation levels or adherence to collagen. Extracellular components involved in attachment were assessed using Congo red agar and pellicle formation was also examined. Phenotypic traits potentially related to extended environmental persistence were observed more frequently in non-STEC. However, these traits were also observed in some STEC isolates, showing phenotype is strain dependent indicating a risk for enhanced environmental survival of some STEC isolates. The shedding status of E. coli O157:H7 is not dictated by the investigated characteristics alone.

Salmonella Typhimurium is the second most common cause of foodborne salmonellosis. Phage-mediated horizontal gene transfer contributes to the virulence of S. Typhimurium. An example of a phage-encoded virulence gene is sopE, a T3SS effector, found rarely in Typhimurium and associated with epidemics. The current pandemic and multi-drug resistant monophasic variant of S. Typhimurium (S. 4,[5],12:i:-) acquired sopE in multiple events following the lysogeny of a previously undescribed bacteriophage, mTmV. The current study aimed to further investigate the association of the virulence gene with S. 4,[5],12:i:- and assess its epidemiological impact. To this end, a large collection of clinical S. Typhimurium isolates from the UK have been analysed for the sopE gene and mTmV presence using a phylogenomic approach. While a large proportion of S. 4,[5],12:i:- (41 %) carried the sopE gene, few isolates outside the epidemic clade harboured it. Notably, the mTmV bacteriophage was identified only in S. 4,[5],12:i:-, although laboratory experiments demonstrated that the phage host range is not restricted to it. Nonetheless, we identified the phage in other S. enterica serovars circulating in the same ecological niche of S. 4,[5],12:i:-. In addition, a genomic characterisation of mTmV was performed revealing an unexpected level of phage variation. Finally, we identified a novel phage-like element harbouring the gene. The study revealed the large dissemination and selection of the virulence gene in the current epidemic, which is mobilised by multiple and distinct mobile genetic elements.

International audience; Salmonella colonization and infection in production animals such as pigs are a cause for concern from a public health perspective. Variations in susceptibility to natural infection may be influenced by the intestinal microbiota. Using 16S rRNA compositional sequencing, we characterized the fecal microbiome of 15 weaned pigs naturally infected with Salmonella at 18, 33, and 45 days postweaning. Dissimilarities in microbiota composition were analyzed in relation to Salmonella infection status (infected, not infected), serological status, and shedding pattern (nonshedders, single-point shedders, intermittent-persistent shedders). Global microbiota composition was associated with the infection outcome based on serological analysis. Greater richness within the microbiota postweaning was linked to pigs being seronegative at the end of the study at 11 weeks of age. Members of the Clostridia, such as Blautia, Roseburia, and Anaerovibrio, were more abundant and part of the core microbiome in nonshedder pigs. Cellulolytic microbiota (Ruminococcus and Prevotella) were also more abundant in noninfected pigs during the weaning and growing stages. Microbial profiling also revealed that infected pigs had a higher abundance of Lactobacillus and Oscillospira, the latter also being part of the core microbiome of intermittent-persistent shedders. These findings suggest that a lack of microbiome maturation and greater proportions of microorganisms associated with suckling increase susceptibility to infection. In addition, the persistence of Salmonella shedding may be associated with an enrichment of pathobionts such as Anaerobiospirillum. Overall, these results suggest that there may be merit in manipulating certain taxa within the porcine intestinal microbial community to increase disease resistance against Salmonella in pigs. IMPORTANCE Salmonella is a global threat for public health, and pork is one of the main sources of human salmonellosis. However, the complex epidemiology of the infection limits current control strategies aimed at reducing the prevalence of this infection in pigs. The present study analyzes for the first time the impact of the gut microbiota in Salmonella infection in pigs and its shedding pattern in naturally infected growing pigs. Microbiome (16S rRNA amplicon) analysis reveals that maturation of the gut microbiome could be a key consideration with respect to limiting the infection and shedding of Salmonella in pigs. Indeed, seronegative animals had higher richness of the gut microbiota early after weaning, and uninfected pigs had higher abundance of strict anaerobes from the class Clostridia, results which demonstrate that a fast transition from the suckling microbiota to a postweaning microbiota could be crucial with respect to protecting the animals.

Herein we report on the development of an electrochemical sensor for silver ions detection in tap water using anodic sweep voltammetry with in-situ pH control; enabled by closely spaced interdigitated electrode arrays. The in-situ pH control approach allowed the pH of a test solution to be tailored to pH 3 (experimentally determined as the optimal pH) by applying 1.65 V to a protonator electrode with the subsequent production of protons, arising from water electrolysis, dropping the local pH value. Using this approach, an initial proof-of-concept study for silver detection in sodium acetate was undertaken where 1.25 V was applied during deposition (to compensate for oxygen production) and 1.65 V during stripping. Using these conditions, calibration between 0.2 and 10 μM was established with the silver stripping peak ∼0.3 V. The calculated limit of detection was 13 nM. For the final application in tap water, 1.65 V was applied to a protonator electrode for both deposition and stripping of silver. The chloride ions, present in tap water (as a consequence of adding chlorine during the disinfection process) facilitated silver detection and caused the striping peak to shift catholically to ∼0.2 V. The combination of the complexation of silver ions with chloride and in-situ pH control resulted in a linear calibration range between 0.25 and 2 μM in tap water and a calculated limit of detection of 106 nM without the need to add acid or supporting electrolytes.

Pork is an important source of human salmonellosis and low-cost on-farm control measures may provide a useful element in reducing the prevalence of this pathogen in food. This study investigated the effectiveness of dietary supplementation with sodium butyrate administered to finisher pigs for ∼4-weeks prior to slaughter to control Salmonella shedding on highly contaminated farms. Two trials (A and B) were conducted on two commercial pig farms, which had a history of high Salmonella seroprevalence. In both trials, pens (14 pens of 12 pigs/pen in Trial A and 12 pens of 12-17 pigs/pen in Trial B) were randomly assigned to a control (finisher feed without additive) or a treatment group (the same feed with 3kg sodium butyrate/t) for 24-28days, depending on the trial. Faeces were collected from each pig on days 0, 12 and 24/28, and blood, caecal digesta and ileocaecal/mesenteric lymph nodes were collected from the slaughterhouse. Pigs were weighed at the start and end of the trials, feed intake was recorded, and carcass quality parameters were recorded at slaughter. In Trial A, Salmonella shedding was reduced in the treatment compared to the control group at the end of the trial (30% versus 57% probability of detecting Salmonella in faeces, respectively; p0.001). This reflected the serology results, with detection of a lower seroprevalence in the treatment compared to the control group using the 20% optical density cut-off (69.5% versus 89%; p=0.001). However, no effect on faecal shedding or seroprevalance was observed in Trial B, which may be explained by the detection of a concomitant infection with Lawsonia intracellularis. No significant differences in Salmonella recovery rates were observed in the caecal digesta or lymph nodes in either trial. Furthermore, feed intake, weight gain, and feed conversion efficiency (FCE) did not differ between groups (p0.05) in either trial. Numerical improvements in weight gain and FCE were found with sodium butyrate treatment, which gave a cost benefit of €0.04/kg of live-weight gain. Overall, results suggest that strategic feeding of sodium butyrate, at 3kg/t of feed, to finishing pigs for 24-28days prior to slaughter was effective in reducing Salmonella shedding and seroprevalance but perhaps only in the absence of co-infection with other pathogens. However, sodium butyrate supplementation at this rate did not influence intestinal carriage, nor did it reduce seroprevalence to below the cut-off used for the high Salmonella risk category in Ireland (50%), or significantly improve growth performance.

Silver is more often used in consumer products and that in consequence leads to its release into the water. Quick and easy methods for detection of silver ions in water are required. In present work electrochemical-based sensor was developed for detection of silver ions in aqueous solutions. The solution's pH for detection was lowered either with nitric acid or reagent-free using localized pH control. Linear regions for the current peak height against the silver concentration have been found between 100–1000 nM and 250–5000 nM for samples acidified with nitric acid and using localized pH control respectively. In present work the limit of detection was lower than comparing to similar work without the need to modify the electrode.

The objective of this study was to assess the efficacy of a combination of high pressure processing (HPP) and a mix of organic acids Inbac™ as hurdles to extend the shelf life of previously optimised sensory accepted frankfurters and cooked ham with significantly (P

Zoonotic pathogens constitute the major source (60.3%) of emerging infectious diseases. Previous studies have investigated the prevalence of Shiga-toxigenic Escherichia coli (STEC) among wild animal species, but comprehensive data are needed to assess the role that these animals have in the transmission of STEC infections to the human population via faecal contamination of the environment, agri-food or water chain. Due to the nature of these microorganisms in which this human-animal-environment interface plays a relevant role on the disease's dynamics, a "One Health" approach is needed to prevent and control the worldwide spread. The aim of this study was to review the published research on the prevalence of STEC in wildlife. The search was performed using several online databases consisting of three blocks of specific search terms covering pathogen, type of study and population. Two reviewers applied the inclusion and exclusion criteria to screening and eligibility phases. Two hundred and twenty-five abstracts were screened for relevance, and 72 were included for data analysis. Most studies (77.8%) investigated the prevalence of STEC in ruminants and urban birds. Their role in transmitting the pathogen to humans, other animals and the agri-food chain is potentiated by the peculiar biological characteristics in ruminants and improved adaptation of urban birds to urban environments. The popularity of convenience and voluntary response sampling may be due to the lack of human-made boundaries on the wild animal species' habitat and having some samples from hunted-harvested animals. To our knowledge, this is the first comprehensive review on STEC prevalence in wild animal species from studies conducted across the globe. We recommend that future research includes and compares samples from varying origins (i.e., human, animal, environment and food) and applies a "One Health" approach to the emerging challenges that STEC poses to public health. Department of Agriculture, Food and the Marine Embargo of 12 month applies-AC

Consumer demands for high quality, safe, nutritious and convenient meat products has provided important insights into the development of a host of processed meat products including fermented meats. It offers unique perspectives for the application of novel functional microbial cultures and promising emerging technologies in the manufacture of fermented meats. Novel non-thermal and thermal technologies, including high pressure processing, pulsed-UV light, irradiation can be utilized for processing and decontamination of meat products to improve process efficiency, product safety and product quality and composition. Application of novel technologies and culture in the development of functional fermented meat products are discussed.

Escherichia coli O157 and O26 shedding patterns in cattle are known to vary widely. To address gaps in the understanding of the underlying factors which impact on shedding dynamics, sensitive and rapid quantitative methods which can be applied in surveillance studies on cattle are required. Current approaches for enumeration of verocytotoxigenic E. coli (VTEC) in cattle faeces are based on direct plating onto selective agars, most probable number (MPN) or real time PCR applied directly to faecal samples, all of which have limitations in terms of the labour involved or their sensitivity. The objective of this study was to develop a sensitive real time quantitative PCR assay, to quantify O157 and O26 in bovine recto-anal junction (RAJ) swabs. The approach was to target serogroup specific genes rfbE and wzx, and to couple a short enrichment, with the use of a standard calibration curve relating real time PCR cycle threshold (Ct) values against the initial concentration of the pathogen in the sample. Following initial experiments in broth culture, a 5h enrichment in modified tryptone soya broth with novobiocin (20 mg/l) (mTSBn) was found to be optimal, and a linear correlation between inocula (Log10 1 to 6 CFU ml(-1)) and the PCR Ct values for both E. coli O157 (R(2)=0.99, rsd=0.58) and E. coli O26 (R(2)=0.99, rsd=0.44) was confirmed. The developed method was then applied to bovine RAJ swab samples (n=153), which were inoculated with E. coli O157 or O26 (Log10 1 to 7 CFU swab(-1)). Calibration curves yielded correlations for E. coli O157 of R(2)=0.86, rsd=0.72 and for O26 (R(2)=0.88, rsd=0.69). In conclusion, a sensitive method for detection and enumeration of two significant VTEC serogroups in bovine RAJ samples has been developed and validated, and will support studies on the bovine shedding dynamics of these pathogens in cattle.

The objective of this study was to model and quantify the level of Listeria monocytogenes in raw milk cheese (RMc) and pasteurized milk cheese (PMc) from farm to fork using a Bayesian inference approach combined with a quantitative risk assessment. The modeling approach included a prediction of contamination arising from the farm environment as well from cross-contamination within the cheese-processing facility through storage and subsequent human exposure. The model predicted a high concentration of L. monocytogenes in contaminated RMc (mean 2.19 log10 CFU/g) compared to PMc (mean -1.73 log10 CFU/g). The mean probability of illness (P1 for low-risk population, LR) and (P2 for high-risk population, HR, e.g., immunocompromised) adult Irish consumers following exposure to contaminated cheese was 7 × 10(-8) (P1 ) and 9 × 10(-4) (P2 ) for RMc and 7 × 10(-10) (P1 ) and 8 × 10(-6) (P2 ) for PMc, respectively. In addition, the model was used to evaluate performance objectives at various stages, namely, the cheese making and ripening stages, and to set a food safety objective at the time of consumption. A scenario analysis predicted various probabilities of L. monocytogenes contamination along the cheese-processing chain for both RMc and PMc. The sensitivity analysis showed the critical factors for both cheeses were the serving size of the cheese, storage time, and temperature at the distribution stage. The developed model will allow food processors and policymakers to identify the possible routes of contamination along the cheese-processing chain and to reduce the risk posed to human health.

This study investigated the thermal inactivation of five Campylobacteraceae species (Campylobacter coli, Campylobacter helveticus, Campylobacter concisus, Campylobacter fetus subsp. fetus and Arcobacter butzleri) at 55 and 60 °C using an immersed coil heating apparatus. The applicability of nine different models, including log linear and Weibull type models, some of which accounted for shoulders and tailing populations at the start or end of the heat treatment, was assessed to describe the thermal inactivation of each species. The Weibull model with tailing effect provided a good fit with low RMSE value (0.07–0.27), AICc (−88.12 to −6.82) and high R2 values (0.97–0.99) depending on the temperature and the species. The Weibull + tail model showed a significantly high D value for Arcobacter butzerli (180.2 s) at 55 °C and a low D value for C. helveticus (23.9 s) at 60 °C, with a high degree of variability in thermal tolerance noted across the species. Moreover, the Weibull + tail model also indicated high thermal sensitivity for C. fetus subsp. fetus at both temperatures investigated. This study highlights the value of assessing various inactivation models to describe the D value to get the model which best describes the survival data. These model predictions enhance knowledge about the survival characteristics and behaviour of fastidious Campylobacteraceae in the food chain.

Among the zoonotic pathogens affecting pigs, Salmonella stands out due to the high number of human cases linked to pork consumption. In the last two decades many countries have put considerable effort into the control of the infection by surveillance and control strategies on farm. Despite this effort, many herds still have a high Salmonella prevalence and they require guidance to address this problem. The present study, using the serological surveillance data of finishing pigs from the Irish National pig Salmonella Control Programme, aimed to highlight factors associated with increased risk or that might mitigate Salmonella occurrence on farm. A questionnaire with 33 questions regarding herd characteristics, management, feeding, biosecurity, and health was completed for 61 individual herds. After the multivariate analysis by linear regression, nine variables were retained in the final model and linked to herd seroprevalence. Home produced-feed linked to the use of meal showed an eight points reduction in Salmonella prevalence compared to purchased feed (p = 0.042). Different biosecurity measures were associated to lower seroprevalence. Changing of footwear from outside to inside the farm decreased seroprevalence nearly 20 units (p = 0.014) and policies not permitting access to the farmyard to feed trucks (p = 0.048) or avoiding the presence of cats on the farm (p = 0.05) were estimated in 10 units less of seroprevalence. In contrast, the lack of perimeter fence increased the chance to have higher seroprevalence in five units (p = 0.05). Finally, intestinal diseases such as swine dysentery (p = 0.044) and E. coli diarrhea (p = 0.1) were estimated to increase Salmonella prevalence in ~20 and 10 units, respectively, demonstrating the importance of controlling other enteric pathogens in an on-farm Salmonella control programme. These results show the usefulness of surveillance data to improve on-farm control and confirm that Salmonella infection in pigs is multi-factorial and the approach to its control should be multifaceted.

The objective of this study was to investigate the growth of Listeria monocytogenes in semi-soft rind washed cheese made from raw and pasteurised milk at different storage temperatures (4, 10 and 15 °C) over a 28 day period simulating storage following ripening. Changes in water activity (aw) and pH in cheeses were also monitored during storage. Response surface models were used to model the interaction of storage temperature and time on aw, pH and L. monocytogenes population. Growth curves were fitted using Baranyi, modified Gompertz and Logistic models at all storage temperatures for both cheeses, and model parameters were statistically analysed. In raw and pasteurised milk cheeses, all models showed a significant (P 0.87. Overall, the L. monocytogenes population increased up to 3 log10 cfu−g−1 for both cheeses during storage following ripening. The fitted models confirmed different L. monocytogenes growth behaviour between raw and pasteurised milk cheeses, which could support the Food Business Operator in predicting growth during storage following ripening.

A quantitative PCR method is described for the detection and quantification of E. coli O157 and O26 in sheep recto-anal junction swabs. The method incorporated a short enrichment step (5 h) and the use of a developed standard calibration curve relating the real time PCR cycle threshold (Ct) values to the initial concentration of pathogen in the sheep sample.