Phenotypic and Genotypic Characterizations of Antimicrobial-Resistant Escherichia coli Isolates from Diverse Retail Meat Samples in North Carolina During 2018-2019.

Surveillance of antimicrobial-resistant pathogens in U.S. retail meats is conducted to identify potential risks of foodborne illness. In this study, we conducted a phenotypic and genotypic analysis of Escherichia coli recovered from a diverse range of retail meat types during 2018-2019 in North Carolina. The investigation was conducted as part of the National Antimicrobial Resistance Monitoring System (NARMS). Retail meat sampling and E. coli isolation were performed in accordance with NARMS retail meat isolation protocols. We used the Sensititre™ broth microdilution system to determine phenotypic resistance to 14 antimicrobial agents and the Illumina next-generation sequencing platform for genotypic resistance profiling. The highest prevalence of E. coli isolates was found in ground turkey (n = 57, 42.9%) and chicken (n = 27, 20.3%), followed by ground beef (n = 25, 18.9%) and pork (n = 24, 18%). The isolates were divided into seven different phylogroups using the Clermont typing tool, with B1 (n = 59, 44.4%) and A (n = 39, 29.3%) being the most dominant, followed by B2 (n = 14, 10.5%), D (n = 7, 5.3%), F (n = 6, 4.5%), E (n = 3, 2.3%), and C (n = 2, 1.5%). Using multilocus sequence typing (MLST), 128 Sequence types (STs) were identified indicating high diversity. Phenotypic and genotypic resistance was observed toward aminoglycosides, sulfonamides, beta-lactams, macrolides, tetracyclines, phenicols, and fluoroquinolones. Ground turkey samples were more resistant to the panel of tested antimicrobials than chicken, beef, or pork (p < 0.05). All isolates were found to be susceptible to meropenem. A high percentage of turkey isolates (n = 16, 28%) were multidrug-resistant (MDR) compared with 18.5% of chicken (n = 5), 8.4% of pork (n = 2), and 8% of beef isolates (n = 2). This study highlights the benefit of surveillance to identify MDR E. coli for epidemiologic tracking and is a comprehensive report of the phenotypic and genotypic characterization of E. coli isolated from retail meats in North Carolina.

Descriptive analysis of the most common types of food safety infractions at ready-to-eat meat processing plants in Ontario, Canada.

Food safety inspections of meat processing plants and abattoirs that process ready-to-eat (RTE) meats have identified a lack of compliance with good manufacturing practices. This study was undertaken to identify common food safety infractions in the RTE meat processing sector in Ontario through an analysis of historical audit records. A total of 376,457 audit item results were evaluated across 912 unique audits of 204 different RTE meat plants. A nearly two-thirds overall item pass rate (64.4%; n = 242,478) was identified. Across all other risk categories, the highest rates of infractions were observed in the "maintenance of premises, equipment and utensils" (56.7%; n = 750). The overall item pass rate was higher in free-standing meat processing plants than abattoirs, while pass rates gradually decreased across the study period. The results of this study have identified key areas for improvement in future inspection, audit and outreach with RTE meat processing plants.

Dairy, Meat, Seafood, and Plant Sources of Saturated Fat: United States, Ages Two Years and Over, 2017-2020.

BACKGROUND: Research suggests that the effects of saturated fat (SF) on health differ depending on its food source. Dairy SF has been associated with lower cardiovascular disease (CVD) risk, whereas meat SF is linked to a higher CVD risk. OBJECTIVES: To estimate the contribution to the total intake of SF of 1) 5 food groups - dairy, meats, seafood, plants, and "other," and 2) the top 10 specific food category sources in the United States population overall and by sociodemographic subgroup. METHODS: The analysis included data from 11,798 participants in the 2017-March 2020 National Health and Nutrition Examination Survey aged 2+ y. Grams of SF from the food sources expressed as a percentage of the total grams of SF consumed, were estimated using the population ratio method. RESULTS: Mean daily intake of SF was 28.1 g [95% confidence interval (CI): 27.6-28.6 g], comprising 11.9% (95% CI: 11.7-12.1%) of total energy intake. Dairy contributed 28.4% of SF, followed by meats (22.1%), plant sources (7.5%), fish and seafood (1.2%), and the remaining foods (41.6%). Youth had higher SF intake from dairy than adults (P < 0.001), whereas non-Hispanic Whites had higher intake than non-Hispanic Blacks (P < 0.001) and Hispanics (P = 0.016). Adults had higher SF intake from meats than youth (P = 0.002), males more than females (P < 0.001), and non-Hispanic Blacks more than non-Hispanic Asians (P = 0.016) and Hispanics (P < 0.001). The top 10 specific sources of SF were unprocessed red meats, sweet bakery products, cured meats, milk, cheese, pizza, unprocessed poultry, Mexican mixed dishes, eggs, and combined fruits and vegetables. CONCLUSIONS: Although dairy contributed ∼30% of SF compared to ∼20% for total meat, the top specific food category source of SF was unprocessed red meats, which ranked in the top 2 food category sources of SF for most subgroups. These findings may be useful for further research to examine the relationship between the different sources of SF and health outcomes.

Review of technology and materials for the development of cultured meat.

Cultured meat production technology suggested that can solve the problems of traditional meat production such as inadequate breeding environment, wastewater, methane gas generation, and animal ethics issues. Complementing cultured meat production methods, sales and safety concerns will make the use of cultured meat technology easier. This review contextualizes the commercialization status of cultured meat and the latest technologies and challenges associated with its production. Investigation was conducted on materials and basic cell culture technique for cultured meat culture is presented. The development of optimal cultured meat technology through these studies will be an innovative leap in food technology. The process of obtaining cells from animal muscle, culturing cells, and growing cells into meat are the basic processes of cultured meat production. The substances needed to production of cultured meat were antibiotics, digestive enzymes, basal media, serum or growth factors. Although muscle cells have been produced closer to meat due to the application of scaffolds materials and 3 D printing technology, still a limit to reducing production costs enough to be used as foods. In addition, developing edible materials is also a challenge because the materials used to produce cultured meat are still not suitable for food sources.

Organ meat consumption and risk of non-alcoholic fatty liver disease: the Tianjin Chronic Low-grade Systemic Inflammation and Health cohort study.

Prospective cohort studies linking organ meat consumption and non-alcoholic fatty liver disease (NAFLD) are limited, especially in Asian populations. This study aimed to prospectively investigate the association between organ meat consumption and risk of NAFLD in a general Chinese adult population. This prospective cohort study included a total of 15 568 adults who were free of liver disease, CVD and cancer at baseline. Dietary information was collected at baseline using a validated FFQ. NAFLD was diagnosed by abdominal ultrasound after excluding other causes related to chronic liver disease. Cox proportional regression models were used to assess the association between organ meat consumption and risk of NAFLD. During a median of 4·2 years of follow-up, we identified 3604 incident NAFLD cases. After adjusting for demographic characteristics, lifestyle factors, vegetable, fruit, soft drink, seafood and red meat consumption, the multivariable hazard ratios (95 % CI) for incident NAFLD across consumption of organ meat were 1·00 (reference) for almost never, 1·04 (0·94, 1·15) for tertile 1, 1·08 (0·99, 1·19) for tertile 2 and 1·11 (1·01, 1·22) for tertile 3, respectively (Pfor trend < 0·05). Such association did not differ substantially in the sensitivity analysis. Our study indicates that organ meat consumption was related to a modestly higher risk of NAFLD among Chinese adults. Further investigations are needed to confirm this finding.

Transforming practice chains through ideological objects: How plant-based meats impact consumers' everyday food practices.

This research investigates the impact of the introduction of plant-based meats (PBMs) on consumers' food practices. Based on the results of 21 in-depth interviews with consumers who use PBMs, this research uses practice theory to explore how the adoption of PBMs affects linked food practices and the meanings associated with these practices. We find that consumers adopt PBMs due to either a desire for meaning coherence or for practicality. Subsequently there are social and embodied ripple consequences associated with this adoption, with consumers revising their social food practices, reconfiguring their understandings of health, and re-orienting their relationship to their body. Our findings extend the research on practice theory by examining how the adoption of a new category of ideological objects shapes other linked consumption practices. Practically, our findings provide important insights for dietary, marketing and health practitioners to understand the overall impact of PBM adoption on consumers' dietary patterns and practices, and their perception about health and body.

Comparison of Antibiotic Resistance Profiles of Salmonella Isolates from Retail Meats in Nanchang, China, in Two Periods.

Salmonella is one of the most important foodborne pathogens. In this article, a total of 160 Salmonella isolates recovered from retail meats in June-July 2018 (before COVID-19 outbreak) and December 2020-April 2021 (after COVID-19 outbreak) in Nanchang, China, were characterized for serotyping, antimicrobial susceptibility, and specific resistance gene screening. The prevalence of Salmonella Typhimurium increased from 5.4% in 2018 to 19.1% in 2021, and Salmonella Enteritidis increased from 3.3% in 2018 to 8.8% in 2021. Compared with those in June-July 2018, Salmonella isolates in December 2020-April 2021 demonstrated a significant increase in resistance to 13 tested antibiotics except for doxycycline and nitrofurantoin (p < 0.05). The Salmonella isolates in December 2020-April 2021 showed a higher presence of plasmid-mediated quinolone resistance genes (qnrA, qnrB, and qnrS), and mutations in the quinolone resistance-determining region (gyrA Asp87Asn, gyrA Asp87Tyr, parC Thr57Ser, and parC Ser80Ile). Whole-genome sequencing was used to analyze four polymyxin B-resistant strains. Some common mutation sites in eptC and micA were found in the four strains. Based on the data in this article, it indicated that antibiotic resistance was facilitated and more gene mutations related to quinolone resistance were developed.

One Health Genomic Analysis of Extended-Spectrum ß-Lactamase‒Producing Salmonella enterica, Canada, 2012‒2016.

Extended-spectrum ß-lactamases (ESBLs) confer resistance to extended-spectrum cephalosporins, a major class of clinical antimicrobial drugs. We used genomic analysis to investigate whether domestic food animals, retail meat, and pets were reservoirs of ESBL-producing Salmonella for human infection in Canada. Of 30,303 Salmonella isolates tested during 2012-2016, we detected 95 ESBL producers. ESBL serotypes and alleles were mostly different between humans (n = 54) and animals/meat (n = 41). Two exceptions were blaSHV-2 and blaCTX-M-1 IncI1 plasmids, which were found in both sources. A subclade of S. enterica serovar Heidelberg isolates carrying the same IncI1-blaSHV-2 plasmid differed by only 1-7 single nucleotide variants. The most common ESBL producer in humans was Salmonella Infantis carrying blaCTX-M-65, which has since emerged in poultry in other countries. There were few instances of similar isolates and plasmids, suggesting that domestic animals and retail meat might have been minor reservoirs of ESBL-producing Salmonella for human infection.

The range and nutrient profile of alternative protein products sold in Australian supermarkets between 2014 and 2021.

The purpose of this study was to investigate changes in the range and nutrient profile of processed alternative protein "convenience" products available in Australia from 2014 to 2021. Product data were extracted from FoodTrack™, an established database of packaged supermarket products. Eligible products were grouped into subcategories: (i) tofu products; (ii) legume products; and (iii) plant-based meats. Nutrient composition was assessed from the products' nutrition information panel. The number of alternative protein products in supermarkets more than doubled between 2014 and 2021 (+130%). On average, products were available for 2.2 years (range 1-7 years). Generally, tofu products had the highest contents of saturated fat and sodium, legume products had the highest contents of carbohydrates, sugar and fibre, and plant-based meats had the highest contents of protein and total fat (per 100 g). This study found large variation in the nutrient composition of these products, highlighting the importance of reformulation and consumer education in the future.

Exploring the prospects of the fifth quarter in the 21st century.

A variable proportion of slaughtered livestock, generally referred to as the fifth quarter, is not part of the edible dressed meat and regarded as animal byproduct. In order for the fifth quarter to play a significant role in the current effort toward a circular bio-based economy, it has to successfully support food security, social inclusivity, environmental sustainability, and a viable economy. The high volume of these low-value streams and their nutrient-dense nature can facilitate their position as a very important candidate to explore within the context of a circular bio-based economy to achieve some of the United Nations Sustainable Development Goals (UN-SDGs). While these sources have been traditionally used for various applications across different cultures and industries, it seems evident that their full potential has not yet been exploited, leaving these products more like an environmental burden rather than valuable resources. With innovation and well-targeted interdisciplinary collaborations, the potential of the fifth quarter can be fully realized. The present review intends to explore these low-value streams, their current utilization, and their potential to tackle the global challenges of increasing protein demands while preventing environmental degradation. Factors that limit widespread applications of the fifth quarter across industries and cultures will also be discussed.

Effects of seasons and parts on volatile N-nitrosamines and their exposure and risk assessment in raw chicken and duck meats.

The N-nitrosamine (NA) concentrations and types in raw chicken and duck meats of different parts and seasons were estimated by headspace solid-phase micro-extraction with gas chromatography-mass spectrometry (HS-SPME-GC-MS). The exposure level and hazard quotient of each detected volatile N-nitrosamine (VNA) were conducted. The selected chicken and duck samples were contaminated by VNAs to some extent. The major types and contents of VNAs in different parts of chicken and duck meats varied seasonally. For chicken samples, the order of the total VNA concentrations was as follows: autumn > spring > winter > summer. For duck samples, the order was changed as follows: winter > autumn > summer > spring (thigh samples) and autumn > spring > winter > summer (breast samples). The estimated exposure levels for adults caused by duck consumption were slightly higher than those by chickens, which was consistent with the tendency in 2-3 years old children. According to the linear regression correlation between the 10% benchmark dose limit (BMDL10) and subtriplicate of median lethal dose (LD50), BMDL10 values of each VNA were calculated. Due to this hypothesis, the risk assessments of each detected VNA and total VNAs posed by consuming chicken and duck meats in Tianjin, China were of low concern. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-021-05195-1.

Associations of dietary patterns and nutrients with coronary artery calcification and pericardial adiposity in a longitudinal study of adults with and without type 1 diabetes.

PURPOSE: To examine the associations of dietary patterns and nutrients with coronary artery calcification (CAC) and pericardial adiposity (PAT) in adults with and without type 1 diabetes. METHODS: We conducted a six-year longitudinal analysis of data from Coronary Artery Calcification in Type 1 Diabetes study [n = 1255; T1D: n = 563; non-DM: n = 692] collected at baseline, year 3 and year 6. Participants completed a validated food frequency questionnaire, a physical examination, and fasting (12 h overnight fast) biochemical analyses. CAC and PAT were measured using electron beam computed tomography. Dietary patterns were identified using factor analysis. Generalized estimating equations were used to examine associations of dietary patterns and nutrients with CAC and PAT in models adjusted for traditional cardiovascular risks. RESULTS: The 'starchy veggies, meats and alcohol pattern' was associated with significantly increased risk of CAC presence in all adjusted models; an increasing trend was observed with CAC progression. Increasing intake of dietary proteins and total fats were also associated with higher risk of the presence and/or progression of CAC in adjusted models (all p < 0.05). PAT was positively associated with dietary total fats, and inversely associated with dietary intakes of saturated fats, omega-3 fats and fiber in models adjusted for age, sex, total calories, time, diabetes status, systolic blood pressure, serum lipids and physical activity. CONCLUSION: Diets high in total fats and proteins, and in meats (processed and red) and alcohol may increase risks of CAC, while saturated fats, omega-3-fats and fiber may be protective against pericardial adiposity as a risk factor for coronary artery disease. ClinicalTrials.gov Identifier: NCT00005754.

Diet quality and periodontal disease: Results from the oral infections, glucose intolerance and insulin resistance study (ORIGINS).

AIMS: This study examined the cross-sectional association between diet quality and periodontal disease. MATERIALS AND METHODS: In the Oral Infections, Glucose Intolerance and Insulin Resistance Study (ORIGINS), 923 individuals completed the National Cancer Institute's validated Diet History Questionnaire 1, from which the Alternative Healthy Eating Index (AHEI) scores and A Priori Diet Quality Scores (APDQS) were calculated. Mean probing depth (MPD), mean clinical attachment loss (MAL) and % of sites bleeding on probing (%BOP) were derived from full-mouth periodontal exams. Multivariable adjusted linear and logistic regression models assessed the associations between diet quality and MPD, MAL, %BOP, and the odds of periodontitis (defined via the CDC/AAP classification). RESULTS: Alternative Healthy Eating Index and APDQS were not associated with MPD, MAL, or periodontitis. While AHEI was also not associated with %BOP, the APDQS was associated with %BOP (p = .03). Higher nut consumption was related to lower MPD (p = .03) and periodontitis odds (p = .03). Higher red meat consumption was associated with higher MPD (p = .01) and %BOP (p = .05). Higher trans-fatty acid consumption was also associated with increased %BOP (p = .05). CONCLUSION: Overall diet quality scores were not associated with periodontal status. Future studies are necessary to replicate the associations observed in this study to minimize the risk of false discovery.

High-pressure processing of meat: Molecular impacts and industrial applications.

High-pressure processing (HPP) has been the most adopted nonthermal processing technology in the food industry with a current ever-growing implementation, and meat products represent about a quarter of the HPP foods. The intensive research conducted in the last decades has described the molecular impacts of HPP on microorganisms and endogenous meat components such as structural proteins, enzyme activities, myoglobin and meat color chemistry, and lipids, resulting in the characterization of the mechanisms responsible for most of the texture, color, and oxidative changes observed when meat is submitted to HPP. These molecular mechanisms with major effect on the safety and quality of muscle foods are comprehensively reviewed. The understanding of the high pressure-induced molecular impacts has permitted a directed use of the HPP technology, and nowadays, HPP is applied as a cold pasteurization method to inactive vegetative spoilage and pathogenic microorganisms in ready-to-eat cold cuts and to extend shelf life, allowing the reduction of food waste and the gain of market boundaries in a globalized economy. Yet, other applications of HPP have been explored in detail, namely, its use for meat tenderization and for structure formation in the manufacturing of processed meats, though these two practices have scarcely been taken up by industry. This review condenses the most pertinent-related knowledge that can unlock the utilization of these two mainstream transformation processes of meat and facilitate the development of healthier clean label processed meats and a rapid method for achieving sous vide tenderness. Finally, scientific and technological challenges still to be overcome are discussed in order to leverage the development of innovative applications using HPP technology for the future meat industry.

Characterization and Source Investigation of Multidrug-Resistant Salmonella Anatum from a Sustained Outbreak, Taiwan.

An ongoing outbreak of multidrug-resistant Salmonella enterica serovar Anatum began in Taiwan in 2015. Pork and poultry were identified as vehicles for transmission. Contaminated meat contributed to the high rate of infections among children. Nearly identical Salmonella Anatum strains have been identified in the United Kingdom, the United States, and the Philippines.

Development of a multiplex real-time PCR to differentiate the four major Listeria monocytogenes serotypes in isolates from meat processing plants.

Listeria monocytogenes is an important foodborne pathogen, causative agent of listeriosis. The epidemiology and persistence of this bacterium in meat processing plants may be related to its serotype, so it is of utmost importance to carry out a correct differentiation of L. monocytogenes serotypes. The objective of this study was to develop a unique quadruplex real-time quantitative PCR (qPCR) method able to differentiate the four most predominant and worrying L. monocytogenes serotypes (1/2a, 1/2b, 1/2c and 4b) in isolates from meat processing plants and ready-to-eat (RTE) dry-cured meat products. The design of specific primers and probes was based on the lmo0737, lmo0308, ORFC (locus genomically equivalent to gltA-gltB) and ORF2110 genes. A qPCR based on a fragment of the 16S rRNA gene was used to ensure the amplification of Listeria spp. genomic DNA. The standard curves showed efficiency values ranging between 92.3% and 105.8% and, R2 values > 0.98. The specificity of the method was also confirmed by the comparison of the results with those obtained by a previously reported conventional multiplex PCR. In addition, none of the strains which were not ascribed to L. monocytogenes amplified any of the target genes related to the four major serotypes of this pathogenic species. The qPCR, therefore, provides a sensitive, specific and rapid tool for identifying the L. monocytogenes serotypes 1/2a, 1/2b, 1/2c and 4b. This method could be very useful for identifying sources of L. monocytogenes contamination in the meat industry or for epidemiological monitoring of persistent strains throughout the processing of RTE meat products.

Low prevalence of viable Toxoplasma gondii in fresh, unfrozen American pasture-raised pork and lamb from retail meat stores in the United States.

In a national survey of fresh, unfrozen, American pasture-raised lamb and pork, the prevalence of viable Toxoplasma gondii was determined in 1500 samples selected by random multistage sampling (750 pork, 750 lamb) obtained from 250 retail meat stores from 10 major geographic areas in the USA. Each sample consisted of a minimum of 500g of meat purchased from the retail meat case. To detect viable T. gondii, 50g meat samples of each of 1500 samples were bioassayed in mice. Viable T. gondii was isolated from 2 of 750 lamb samples (unweighted: 0.19%, 0.00-0.46%; weighted: 0.04%, 0.00-0.11%) and 1 of 750 pork samples (unweighted: 0.12%, 0.00-0.37%; weighted: 0.18%, 0.00-0.53%) samples. Overall, the prevalence of viable T. gondii in these retail meats was very low. Nevertheless, consumers, especially pregnant women, should be aware that they can acquire T. gondii infection from ingestion of undercooked meat. Cooking meat to an internal temperature of 66°C kills T. gondii.

Prevalence and Antimicrobial Resistance of Enterococci Isolated from Retail Meats in the United States, 2002 to 2014.

Bacteria of the genus Enterococcus are important human pathogens that are frequently resistant to a number of clinically important antibiotics. They are also used as markers of animal fecal contamination of human foods and are employed as sentinel organisms for tracking trends in resistance to antimicrobials with Gram-positive activity. As part of the National Antimicrobial Resistance Monitoring System (NARMS), we evaluated several retail meat commodities for the presence of enterococci from 2002 to 2014, and we found 92.0% to be contaminated. The majority of isolates were either Enterococcus faecalis (64.0%) or Enterococcus faecium (28.6%), and the antimicrobial resistance of each isolate was assessed by broth microdilution. The resistance prevalences for several drugs, including erythromycin and gentamicin, were significantly higher among poultry isolates, compared to retail beef or pork isolates. None of the isolates was resistant to the clinically important human drug vancomycin, only 1 isolate was resistant to linezolid, and resistance to tigecycline was below 1%. In contrast, a majority of both E. faecalis (67.5%) and E. faecium (53.7%) isolates were resistant to tetracycline. Overall, the robust NARMS testing system employed consistent sampling practices and methods throughout the testing period, with the only significant trend in resistance prevalence being decreased E. faecium resistance to penicillin. These data provide excellent baseline levels of resistance that can be used to measure future changes in resistance prevalence that may result from alterations in the use of antimicrobials in food animal production.IMPORTANCE Enterococci, including E. faecalis and E. faecium, are present in the guts of food-producing animals and are used as a measure of fecal contamination of meat. We used the large consistent sampling methods of NARMS to assess the prevalence of Enterococcus strains isolated from retail meats, and we found over 90% of meats to be contaminated with enterococci. We also assessed the resistance of the Enterococcus strains, commonly used as a measure of resistance to agents with Gram-positive activity, in foods. Resistance prevalence was over 25% for some antimicrobials and sample sources but was less than 1% for several of the most important therapeutic agents used in human medicine.

Volatile N-nitrosamines in meat products: Potential precursors, influence of processing, and mitigation strategies.

Meat products can be contaminated with carcinogenic N-nitrosamines, which is ascribed to the reaction between a nitrosating agent, originating from nitrite or smoke, and a secondary amine, derived from protein and lipid degradation. Although in model systems it is demonstrated that many amine containing compounds can be converted to N-nitrosamines, the yield is dependent of reaction conditions (e.g., low pH and high temperature). In this article, the influence of the composition of the meat products (e.g., pH, aw, spices) and processing (e.g., ageing, ripening, fermentation, smoking, heat treatment and storage) on the presence and availability of the amine precursors and the N-nitrosamine formation mechanism is discussed. In addition, this article explores the current N-nitrosamine mitigation strategies in order to obtain healthier and more natural meat products.

Survival of Clostridium difficile spores at low temperatures.

Clostridium difficile's presence has been reported in meat products stored typically at low temperatures. This study evaluated the viability in phosphate buffer saline (PBS) of spores from epidemic C. difficile strain R20291 (4.6 log CFU/ml) and M120 (7.8 log CFU/ml). Viability was assessed during 4 months at -80 °C, -20 °C, 4 °C (refrigeration), and 23 °C (room temperature), and after 10 freeze (-20 °C)/thaw (+23 °C) cycles. Although spore viability decreased, significant viability was still observed after 4 months at -20 °C, i.e., 3.5 and 3.9 log CFU/ml and -80 °C, i.e., 6.0 and 6.1 log CFU/ml for strains R20291 and M120, respectively. The same trend was observed for M120 at 4 °C and 23 °C, while for R20291 the viability change was non-significant at 4 °C but increased significantly at 23 °C (p > 0.05). After 10 freeze-thaw cycles, viability of both strains decreased but a significant fraction remained viable (4.3 and 6.3 log CFU/ml for strain R20291 and M120, respectively). Strikingly, both strains showed higher viability in a meat model than in PBS. A small but significant decrease (p < 0.05) from 6.7 to 6.3 log CFU/ml in M120 viability was observed after 2-month storage in the meat model while the decrease from an initial 3.4 log CFU/ml observed for R20291 was non-significant (p = 0.12). In summary, C. difficile spores can survive low-temperature conditions for up to 4 months.