The molecular epidemiology of hepatitis E virus genotype 3 in Canada.

Autochthonous hepatitis E virus (HEV) infection is increasingly reported in industrialized countries and is mostly associated with zoonotic HEV genotype 3 (HEV-3). In this study, we examined the molecular epidemiology of 63 human clinical HEV-3 isolates in Canada between 2014 and 2022. Fifty-five samples were IgM positive, 45 samples were IgG positive and 44 were IgM and IgG positive. The majority of the isolates belong to the subtypes 3a, 3b, and 3j, with high sequence homology to Canadian swine and pork isolates. There were a few isolates that clustered with subtypes 3c, 3e, 3f, 3h, and 3g, and an isolate from chronic infection with a rabbit strain (3ra). Previous studies have demonstrated that the isolates from pork products and swine from Canada belong to subtypes 3a and 3b, therefore, domestic swine HEV is likely responsible for the majority of clinical HEV cases in Canada and further support the hypothesis that swine serve as the main reservoirs for HEV-3 infections. Understanding the associated risk of zoonotic HEV infection requires the establishment of sustainable surveillance strategies at the interface between humans, animals, and the environment within a One-Health framework.

Quantitative Risk Assessment of Salmonella in Breaded Pork Products in China.

Pork products were the most common media of Salmonella in China, breaded pork products as a very popular meat presently, whose Salmonella risk should be drawn to attention. Given that quantitative risk assessment is a more scientific method for risk evaluation, a quantitative risk assessment model of Salmonella in breaded pork products was first constructed from processing to consumption, and was used for assessing the risk and the effective interventions in this study. The data of Salmonella contamination in breaded pork products during processing were obtained from the actual detection data of samples from a representative meat processing plant. With combining the predictive microbial modeling and dose-response relationship, the risk of Salmonella in breaded pork products was charactered, and the probability of Salmonella infection per meal was found to be 5.585 × 10-9. Based on the results of sensitivity analysis, the curing and seasoning process was found to be the key control point for Salmonella contamination during the processing, and consumer behavior was the key control point affecting the probability of Salmonella infection from processing to consumption. The model was also applied for assessing the effectiveness of risk interventions, and among the nine interventions given, control of thawing temperature before cooking such as microwave thawing could reduce the risk of infection by 30.969-fold, while cooking the products thoroughly, Salmonella would not pose a pathogenic hazard to consumers. The model and the assessed results in this study may provide guidance on microbial control in producing process and safety consumption of breaded pork products.

Hepatitis E virus genome detection in commercial pork livers and pork meat products in Germany.

The hepatitis E virus (HEV) is one of the most common causes of hepatitis worldwide. HEV is also widespread in many developed countries, where the number of infections is steadily increasing. In those countries, the virus is transmitted mainly through consumption of undercooked or raw food or through contact with animals. Especially, pigs serve as a main reservoir of HEV. Here, we investigated the prevalence of HEV RNA in pork livers and pork meat products to assess the actual risk of HEV infection through food consumption in Germany. A total of 131 pork products were collected from grocery stores and butcher shops between October 2019 and February 2020 and screened for HEV RNA using nested PCR and subsequent sequencing. Overall, 10% of the samples were positive for HEV, including pork livers (5%), spreadable liver sausages (13%) and liver pâté samples (15%). Sequence analyses indicated that the large majority of HEV strains belonged to subtype HEV-3c, representing the most frequent subtype in Germany. One sample belonged to subtype HEV-3f. Further sequence analysis revealed large sequence variation between the samples; however, most of the mutations identified were synonymous. Although infectivity of the virus was not tested, the results suggest a considerable risk of HEV infection through food consumption. Therefore, preventive measures should be taken according to a One Health approach.

Risk Assessment of Human Toxoplasmosis Associated with the Consumption of Pork Meat in Italy.

Toxoplasmosis is a cosmopolitan disease and has a broad range of hosts, including humans and several wild and domestic animals. The human infection is mostly acquired through the consumption of contaminated food and pork meat has been recognized as one of the major sources of transmission. There are, however, certain fundamental differences between countries; therefore, the present study specifically aims to evaluate the exposure of the Italian population to Toxoplasma gondii through the ingestion of several types of pork meat products habitually consumed in Italy and to estimate the annual number of human infections within two subgroups of the population. A quantitative risk assessment model was built for this reason and was enriched with new elements in comparison to other similar risk assessments in order to enhance its accuracy. Sensitivity analysis and two alternative scenarios were implemented to identify the factors that have the highest impact on risk and to simulate different plausible conditions, respectively. The estimated overall average number of new infections per year among adults is 12,513 and 92 for pregnant women. The baseline model showed that almost all these infections are associated with the consumption of fresh meat cuts and preparations (mean risk of infection varied between 4.5 × 10-5 and 5.5 × 10-5 ) and only a small percentage is due to fermented sausages/salami. On the contrary, salt-cured meat products seem to pose minor risk but further investigations are needed to clarify still unclear aspects. Among all the considered variables, cooking temperature and bradyzoites' concentration in muscle impacted most the risk.

An experimental validated in silico model to assess Staphylococcus aureus growth kinetics on different pork products.

AIMS: To gain an understanding of the growth kinetics of Staphylococcus aureus to ensure the safety of pork, and to develop a predictive growth model of Staph. aureus in raw pork, ham and sausage pork under specific storage time and temperature conditions. METHODS AND RESULTS: Growth of Staph. aureus was evaluated at 5-40°C using 5-6 replicates in each sampling time in order to capture experimental variability. Growth curves were fit to Baranyi model to estimate Lag time (λ) and maximum growth rate (µmax). The effect of temperature on λ and µmax was modelled using natural logarithm of λ and square root of µmax. The variability between repetitions was higher at 15, 20 and 40°C than observed at 25, 30 and 35°C. After only 3 and 2 days of storage at 5 and 10°C, respectively, Staph. aureus populations decreased to limit of detection (≤1 log CFU g(-1) ). Staphylococcus aureus populations on ham presented higher µmax in comparison to those grown on raw pork and sausage. Linear regression lines showed that Staph. aureus populations in ham grew faster than those observed in sausage and raw pork. Staph. aureus presented a theoretical minimum growth temperature (Tmin ) depending on the processed pork products, which were different from in raw pork. CONCLUSION: Staphylococcus aureus model predicts faster growth in ham compared to raw pork and sausage. The validation of models showed good predictions, suggesting that the developed models are useful in estimating growth kinetics of Staph. aureus in different pork products. SIGNIFICANCE AND IMPACT OF THIS STUDY: Established here is a data-driven, in silico model on Staph. aureus growth kinetics on different pork products. The model is extensively validated by experiments and simulations and further supported by comparisons to previous modelling efforts in other species.

African swine fever: implications for the Italian pork trade.

In early 2022, the confirmed presence of African swine fever (ASF) circulating in wild boars in mainland Italy and subsequently found in domestic pigs led to several changes regarding the export of pork and pork products to countries outside the European Union (non-EU). The sector suffered the complete and immediate closure of the markets of some countries, often without the measure being communicated in the forms stipulated by international agreements. Indeed, compliance with the current EU regulations does not guarantee the possibility of exporting to non-EU countries. Knowledge of the animal health status requirements of the country (Italy in this case) is essential for food business operators (FBOs) wishing to enter markets outside the EU according to the 'Agreement on the Application of Sanitary and Phytosanitary Measures'. In cases where a sanitary protocol and a model of an official certificate with the importing country exist, the market is officially accessible according to the agreed sanitary requirements. Where no agreement exists, requirements are detailed in the 'import permit' issued to individual FBOs or may be known by directly accessing national regulations through the client/importer. Therefore, the purpose of this work is to briefly outline the conditions imposed by the main non-EU countries for pork products, especially in light of the new epidemiological situation created by the spread of the ASF into a country previously free of the disease.

Changes in Hepatitis E Virus Contamination during the Production of Liver Sausage from Naturally Contaminated Pig Liver and the Potential of Individual Production Parameters to Reduce Hepatitis E Virus Contamination in the Processing Chain.

In this study, changes in hepatitis E virus (HEV) contamination in the production of liver sausage from naturally contaminated pork liver were investigated. Furthermore, the potential effectiveness of individual production parameters in reducing viral loads was measured. When processing moderately contaminated liver (initial Cq-value 29), HEV RNA persisted in the finished sausages, even after heating for 90 min at 75 °C. A matrix-specific standard curve was created using a spiking experiment to accurately quantify HEV RNA in a particularly challenging matrix like liver sausage. Variations in product-specific production parameters, including mincing and heating times, showed some reduction in contamination levels, but even prolonged heating did not render all finished products HEV negative. The persistence of HEV contamination underscores the importance of ongoing monitoring in the pig population and raw materials to enhance food safety measures and reduce the likelihood of transmission through pork consumption. The detection of HEV RNA within all processing stages of pork liver in the production of liver sausage suggests that further research into the risk of infection posed by this detection and vigilance in managing HEV risks in the food chain, particularly in pork products, are required to protect public health.

The Impact of the Fresh Pork Display Lamps on the Sensory Response of Consumers to Fresh Pork.

Two studies were conducted to investigate the impact of fresh pork display lamps on consumers' sensory responses to pork products. In the first experiment, 63 participants were asked to evaluate pork products with different degrees of freshness under four fresh pork display lights and two LED lights. In the other experiment, 30 subjects participated in the Farnsworth-Munsell 100 hue test under the same lamps, with the aim of showing whether the fresh pork display lamps impaired color discrimination. The results showed that the light source had a significant effect on the color appearance evaluation of the pork products. The ratings for perceived freshness under the fresh pork display lamps were significantly higher than those of ordinary LED lamps, while the color discrimination performance of the subjects under those lamps was poor. It was demonstrated that improper component proportions of the light spectrum could influence observers' assessment of meat quality and weaken the observers' ability to distinguish the freshness level. Through this study, the authors would like to remind lamp users and manufacturers to not only be concerned about the improvement in the color appearance of pork, but also the need for consumers to be aware of the authentic freshness of the pork products.

Development of a Sensitive and Specific Quantitative RT-qPCR Method for the Detection of Hepatitis E Virus Genotype 3 in Porcine Liver and Foodstuff.

As an international and zoonotic cause of hepatitis, hepatitis E virus (HEV) poses a significant risk to public health. However, the frequency of occurrence and the degree of contamination of food of animal origin require further research. The aim of this study was to develop and validate a highly sensitive quantitative RT-qPCR assay for the detection and quantification of HEV contamination in porcine liver and food. The focus was on genotype 3, which is most common as a food contaminant in developed countries and Europe. The selected assay has its target sequence in the open reading frame 1 (ORF1) of the HEV genome and showed good results in inclusivity testing, especially for HEV genotype 3. The developed assay seems to show high efficiency and a low intercept when compared to other assays, while having a comparable limit of detection (LOD). In addition, a standard curve was generated using artificially spiked liver to provide more accurate quantitative results for contamination assessment and tracking in this matrix. Application of the assay to test 67 pig livers from different origins resulted in a positivity rate of 7.5%, which is consistent with the results of numerous other prevalence studies. Quantitative detection of the viral genome in the food chain, particularly in pig livers, is essential for understanding the presence and evolution of HEV contamination and thus ensures consumer safety.

Hepatitis E virus in pork meat products and exposure assessment in Belgium.

Zoonotic hepatitis E virus (HEV) genotype 3 infections are the predominant cause of acute viral hepatitis in Europe, mostly associated with the consumption of HEV contaminated pork meat. In this study we looked at the HEV RNA positivity rate of pork meat products readily available from Belgian supermarkets and evaluated the overall HEV consumer exposure in a Belgian context. Two basic assessments were performed in a 'worst-case' scenario setting: one solely focusing on the contamination level of the product itself (ingredients and processing parameters) and another estimating the overall consumer exposure, taking into account consumption habits in Belgium. Non-thermal-processed ready-to-eat (i.e. ready for consumption without additional cooking step by consumer) pork meat products (e.g. raw dried sausages), had a high estimated HEV contamination level, while thermal-processed ready-to-eat pork meat products (e.g. pork liver pâté) had the highest overall consumer exposure estimates. Following these assessments, pork liver pâtés, raw dried hams and raw dried sausages (n = 54) were purchased from Belgian supermarkets (n = 3) and analyzed for HEV RNA by RT-PCR. In total, 31 % (n = 17) products tested positive. HEV RNA was found in 65 % of the pork liver pâtés, 15 % of raw dried hams and 0 % of raw dried sausages. Phylogenetic analysis of four isolates (all gt3c) from pork liver pâté samples showed similarities with human clinical cases from Germany and Belgium.

Effects of Nitrite and Phosphate Replacements for Clean-Label Ground Pork Products.

We investigated the effects of different phosphate replacements on the quality of ground pork products cured with sodium nitrite or radish powder to determine their potential for achieving clean-label pork products. The experimental design was a 2×5 factorial design. For this purpose, the ground meat mixture was assigned into two groups, depending on nitrite source. Each group was mixed with 0.01% sodium nitrite or 0.4% radish powder together with 0.04% starter culture, and then processed depending on phosphate replacement [with or without 0.5% sodium tripolyphosphate; STPP (+), STPP (-), 0.5% oyster shell calcium (OSC), 0.5% citrus fiber (CF), or 0.5% dried plum powder (DPP)]. All samples were cooked, cooled, and stored until analysis within two days. The nitrite source had no effect on all dependent variables of ground pork products. However, in phosphate replacement treatments, the STPP (+) and OSC treatments had a higher cooking yield than the STPP (-), CF, or DPP treatments. OSC treatment was more effective for lowering total fluid separation compared to STPP (-), CF, or DPP treatments, but had a higher percentage than STPP (+). The STPP (+) treatment did not differ from the OSC or CF treatments for CIE L* and CIE a*. Moreover, no differences were observed in nitrosyl hemochrome content, lipid oxidation, hardness, gumminess, and chewiness between the OSC and STPP (+) treatments. In conclusion, among the phosphate replacements, OSC addition was the most suitable to provide clean-label pork products cured with radish powder as a synthetic nitrite replacer.

Static in vitro digestion methods for assessing essential minerals in processed meat products.

The consumption of processed meat products has increased due to its convenient preparation and characteristics related to taste and consumer acceptance. In this study, we evaluated the content and bioaccessibility of iron (Fe), zinc (Zn), calcium (Ca) and magnesium (Mg) in processed pork and beef products by dialysis assay. In addition, different assays of static in vitro digestion (traditional solubility and INFOGEST) were applied to verify the results on the bioaccessibility of these essential elements. The average content of essential elements (expressed in mg/100 g) were 0.78 ± 0.24 and 1.51 ± 0.51 for Fe; 1.79 ± 0.55 and 3.57 ± 1.42 for Zn; 13.24 ± 9.99 and 20.29 ± 10.45 for Ca and 19.43 ± 2.78 and 26.15 ± 7.33 for Mg, in pork and beef products, respectively. However, we observed high variation between brands and batches. The dialysis assay presented variations in bioaccessibility for all essential elements evaluated in the samples. The results for dialyzable Fe were lower than expected (in vivo assays), probably due to the mechanism simulated in dialysis. This assay was considered inadequate for the simulation of in vitro digestion for meat samples that contain heme-Fe. When comparing the different static in vitro digestion assays employed, the dialysis test showed the lowest bioaccessibility percentages for all essential elements evaluated, when compared with the other in vitro digestion methods. The highest bioaccessible fractions were obtained by applying the INFOGEST protocol. We found that according to the in vitro digestion simulation assay used, the results for bioaccessible percentage displayed high variation. These observations highlight the importance of the assay type used for this purpose. These results did not indicate which is the better assay, but demonstrated the differences of each and their relevance for estimating bioaccessibility of essential elements in meat products.

Survey of hepatitis E virus in pork products and pig stools in Nakhon Pathom Province, Thailand.

BACKGROUND: Hepatitis E virus (HEV) is an important public health threat resulting in more than 3 million symptomatic cases and 70,000 deaths annually. HEV is classified into at least eight genotypes, and five are associated with human infection. Genotypes 1 and 2 primarily affect humans, whereas genotypes 3 and 4 circulate in both humans and swine and are considered zoonotic viruses. Previous studies in Central Thailand have reported human HEV isolates with high similarity to swine strains and high seroprevalence in pigs, suggesting the potential for pig-to-human transmission. OBJECTIVES: This study aimed to detect and analyse HEV in pork products and pig stools collected from local markets and pig farms in Nakhon Pathom Province in Central Thailand. METHODS: A total of 177 pig stool and 214 pork product samples were detected for HEV by using RT-PCR amplification. Next, nucleotide sequencing and phylogenetic analysis were performed. RESULTS: We found one sample of pork products (1/214, 0.5%), which was a pig liver sample (1/51, 2.0%), and 49 HEV-positive samples in pig stools (49/177, 27.7%). Phylogenetic analysis showed that all these HEV sequences belonged to genotype 3, with a high correlation between our samples and HEV from humans and swine was previously reported in Thailand. CONCLUSIONS: This study suggested that the consumption of poorly sanitized or uncooked animal meat or food and frequent exposure to pig stools may be risk factors for HEV infections in humans.

Ochratoxin A in Slaughtered Pigs and Pork Products.

Ochratoxin A (OTA) is a mycotoxin that is produced after the growth of several Aspergillus and Penicillium spp. in feeds or foods. OTA has been proved to possess nephrotoxic, hepatotoxic, teratogenic, neurotoxic, genotoxic, carcinogenic and immunotoxic effects in animals and humans. OTA has been classified as possibly carcinogenic to humans (Group 2B) by the IARC in 2016. OTA can be mainly found in animals as a result of indirect transmission from naturally contaminated feed. OTA found in feed can also contaminate pigs and produced pork products. Additionally, the presence of OTA in pork meat products could be derived from the direct growth of OTA-producing fungi or the addition of contaminated materials such as contaminated spices. Studies accomplished in various countries have revealed that pork meat and pork meat products are important sources of chronic dietary exposure to OTA in humans. Various levels of OTA have been found in pork meat from slaughtered pigs in many countries, while OTA levels were particularly high in the blood serum and kidneys of pigs. Pork products made from pig blood or organs such as the kidney or liver have been often found to becontaminated with OTA. The European Union (EU) has established maximum levels (ML) for OTA in a variety of foods since 2006, but not for meat or pork products. However, the establishement of an ML for OTA in pork meat and meat by-products is necessary to protect human health.

Real-Time PCR-Based Methods for Detection of Hepatitis E Virus in Pork Products: A Critical Review.

Standard methods for detection of hepatitis A virus and norovirus in at-risk foodstuffs are available, but currently there is no standard method for detection of hepatitis E virus (HEV) in pork products or other foods that can be contaminated with the virus. Detection assays for HEV are mainly based on nucleic acid amplification, particularly the reverse transcription polymerase chain reaction (RTPCR) in real-time format. RTPCR-based methods can be sensitive and specific, but they require a suite of controls to verify that they have performed correctly. There have been several RTPCR methods developed to detect HEV in pork products, varying in details of sample preparation and RTPCR target sequences. This review critically discusses published HEV detection methods, with emphasis on those that have been successfully used in subsequent studies and surveys. RTPCR assays have been used both qualitatively and quantitatively, although in the latter case the data acquired are only reliable if appropriate assay calibration has been performed. One particular RTPCR assay appears to be ideal for incorporation in a standard method, as it has been demonstrated to be highly specific and sensitive, and an appropriate control and calibration standard is available. The review focuses on the detection of HEV in pork products and similar foodstuffs (e.g., boar). The information may be useful to inform standardisation activities.

A stochastic assessment to quantify the risk of introduction of African swine fever virus to Taiwan via illegal pork products carried by international travellers.

The current study quantified the risk of releasing African swine fever virus (ASFV) into Taiwan from pork products illegally carried by international travellers from 157 countries or territories through six international airports and three international seaports. The association between various factors and the number of pork products detected by the border control authorities was also examined. The risk was estimated with a stochastic process after modelling the number of undetected illegal pork products, probability of pork product detection at international airports and seaports and probability of ASFV contamination of pork products from various countries. The overall annual probability of ASFV release to Taiwan was estimated to be 1 [95% confidence interval (CI): 1-1] under no enhanced mitigation measures. All the median airport-level risks were higher than .921, and four of them reached 1. The total annual risk was .570 (95% CI: .109-.937) for international seaports. The country or territory level risk was estimated to be 1 for Vietnam, China, Hong Kong, the Philippines and South Korea, .999 (95% CI: .628-1) for Macao and .967 (95% CI: .359-1) for Indonesia. After the total number of travellers was factored in, the number of detected illegal pork products was the highest in January and February, and travellers from Vietnam [risk ratio to Japan (RR): 80.45; 95% CI: 58.68-110.3], the Philippines (RR: 37.67; 95% CI: 26.9-52.74) and Cambodia (RR: 28.39; 95% CI: 12.69-63.51) were most likely to bring pork products to Taiwan. Our study indicated a high risk of ASFV introduction through international travellers and also identified the factors associated with the risk. This information can be used as empirical evidence for cost-effective risk mitigation practices.

Estimating the global prevalence of hepatitis E virus in swine and pork products.

Zoonotic transmission of hepatitis E virus (HEV), in particular the genotype (GT) 3 and GT4 strains, constitutes a major one health issue. Swine serves as an important reservoir and the processed pork products essentially contribute to foodborne transmission. This study comprehensively estimated HEV prevalence in domestic pigs, wild boars, and pork products. At global level, we found nearly 60% domestic pigs and 27% wild boars have ever encountered HEV infection based seroprevalence rate. Nearly 13% domestic and 9.5% wild swine are actively infected based on HEV RNA positivity. Importantly, about 10% of commercial pork products are HEV RNA positive, although available data are limited in this respect. Our results indicate the high prevalence rate of HEV infection in pigs and widespread contamination in pork products, although there are substantial variations at regional and country levels. These findings are important for better understanding the global epidemiology and clinical burden of HEV infection in human population related to zoonotic transmission.

Excessive copper in feed not merely undermines animal health but affects food safety.

BACKGROUND: Blackened intestines in slaughtered pigs have been commonly observed in China in recent years. However, no cause has been reported. OBJECTIVES: We attempted to determine whether the blackening of the pig intestine was related to an excess of copper (Cu) in their feed. METHODS: In this study, we observed and collected porcine intestines in small- and large-scale pig slaughterhouses in Shandong province from May to October 2018. Twelve types of metal ions were detected in the black intestinal samples. RESULTS: The Cu level in the intestine samples was mostly higher than the Chinese national limit for food. Further study showed that Cu supplementation in most commercial porcine feed also exceeded the national standard. An animal model (mouse) that could mimic the intestinal blackening in pigs was established. Compared to control mice, Cu accumulated in the liver and intestines of mice fed an excessive Cu level, confirming the excessive Cu in the feed may be considered the major cause of blackened porcine intestines. Microscopic examination revealed that black intestines had many particles containing Cu in the lamina propria of the intestinal mucosa, and the intestinal mucosal epithelial cells showed degeneration and necrosis. CONCLUSIONS: In conclusion, overuse of Cu in animal feed can lead to animal poisoning and Cu accumulation in animal products. Such overuse not only harms the health of livestock but can also affect public health.

Droplet digital polymerase chain reaction assay for identifying and quantifying pork products.

Halal products are growing in consumer markets worldwide, and pork meat is classified as non-halal. Manufacturers of processed foods and products must ensure that their products follow Islamic dietary law because pork is prohibited for Muslims. Droplet digital polymerase chain reaction (PCR) (ddPCR) is a novel method for identifying pig species and quantifying pork products. This experiment aimed to investigate pork species and establish the proportion of pork in meat products using the mitochondrial cytochrome b gene (CYTB). The study found that the correlation coefficient between the meat weight and DNA concentration of pork was 0.997, and the correlation coefficient between the DNA concentration and the target DNA copy number of pork was 0.998. The accuracy of the ddPCR assay was verified using a sample of a known proportion of pork, and it was revealed that this method is highly precise in quantifying pork products. Nine products contained an undeclared meat proportion (90%). The limit of detection for pork was 0.0001 ng. The analysis indicated that the ddPCR assay has high accuracy and sensitivity for quantifying pork products. Therefore, the predictive model can be used in routine laboratories for quality assurance of halal food products.

Prevalence, Molecular Characterization and Antimicrobial Susceptibility of Clostridioides difficile Isolated from Pig Carcasses and Pork Products in Central Italy.

In the last decade, the incidence and severity of Clostridioides difficile infections (CDIs) in humans have been increasing and community-associated infections have been described. For these reasons, the interest in C. difficile in food and in food animals has increased, suggesting other possible sources of C. difficile acquisition. This study evaluated the presence of C. difficile on pig carcasses at the slaughterhouse and in pork products in Central Italy. The contamination rate on pig carcasses was 4/179 (2.3%). Regarding food samples, a total of 216 pork products were tested (74 raw meat preparations and 142 ready-to-eat food samples made by cured raw meat). The real-time PCR screening was positive for 1/74 raw meat preparation (1.35%) and for 1/142 ready-to-eat food samples (0.7%) C. difficile was isolated only from the raw meat preparation (pork sausage). All the isolated strains were toxigenic and susceptible to all the tested antibiotics. Strains isolated from carcass samples displayed A+B+CDTa+CDTb+ profile, were toxinotype IV and belonged to the same ribotype arbitrary named TV93, while the one isolated from food samples displayed A+B+CDTa-CDTb- profile and it was not possible to determine ribotype and toxinotype, because it was lost after freeze storage. It was concluded that the prevalence of C. difficile in the pork supply chain is very low.