A Comprehensive Virulence and Resistance Characteristics of Listeria monocytogenes Isolated from Fish and the Fish Industry Environment.

Listeria monocytogenes is an important pathogen, often associated with fish, that can adapt and survive in products and food processing plants, where it can persist for many years. It is a species characterized by diverse genotypic and phenotypic characteristics. Therefore, in this study, a total of 17 L. monocytogenes strains from fish and fish-processing environments in Poland were characterized for their relatedness, virulence profiles, and resistance genes. The Core Genome Multilocus Sequence Typing (cgMLST) analysis revealed that the most frequent serogroups were IIa and IIb; sequence types (ST) were ST6 and ST121; and clonal complexes (CC) were CC6 and CC121. Core genome multilocus sequence typing (cgMLST) analysis was applied to compare the present isolates with the publicly available genomes of L. monocytogenes strains recovered in Europe from humans with listeriosis. Despite differential genotypic subtypes, most strains had similar antimicrobial resistance profiles; however, some of genes were located on mobile genetic elements that could be transferred to commensal or pathogenic bacteria. The results of this study showed that molecular clones of tested strains were characteristic for L. monocytogenes isolated from similar sources. Nevertheless, it is worth emphasizing that they could present a major public health risk due to their close relation with strains isolated from human listeriosis.

Understanding the main factors that influence consumer quality perception and attitude towards meat and processed meat products.

Meat and meat products consumer behaviour is becoming less predictable. The objective of this review was to determine the attributes associated with the consumer's perception of quality and identify factors influencing the perception and consequent attitude. In conclusion, the findings showed that factors impacting nutritional quality, chemical and biological hazards, animal welfare, beliefs, and fraud could affect consumers' perception of how safe meat products are. Consumers positively perceive sensory attributes and recognize meat's nutritional value, still concerned with fat. Animal welfare and environmental impact have become significant drivers of consumer perception. The presence of chemical additives is a severe concern. Information received by consumers through media strongly influences perception and behaviour. The negative stigmatization of meat and meat products and beliefs often not scientifically-based shapes consumer perception. Their sensory impact and price frame the acceptance of animal production or processing modifications.

Pulsed Light Application for Campylobacter Control on Poultry Meat and Its Effect on Colour and Volatile Profile.

Campylobacter on poultry meat needs to be controlled to reduce the risk of infection caused by the consumption of chicken meat. Pulsed light (PL) application on poultry meat was studied to control Campylobacter spp. The effect of this technology was evaluated regarding poultry meat colour and volatile compound changes. Two breast sample groups were prepared: inoculated with Campylobacter (107 bacteria of Campylobacter jejuni strains) and not inoculated. Samples were submitted to PL, five pulses/s of 300 ms, 1 Hz, and 1 J/cm2 in the apparatus, PL Tecum unit (Claranor). A response surface experimental design was applied regarding the factors of voltage (1828 to 3000 W) and distance to the source UV lamp (2.6 to 5.4 cm). The binomial factorial treatment (voltage and distance) with PL induced different energy doses (fluence J/cm2) received by samples, 2.82 to 9.67 J/cm2. Poultry meat pulsed light treated had a significant decrease of Enterobacteriaceae counts. The treatments applied were unable to reduce 1 log Campylobacter cfu/g of poultry meat. The poultry meat PL treated became slightly light, redder, and yellower than those not treated. PL can decrease the proportion of aldehydes on total volatiles in meat, particularly on those associated with chicken-like, chicken skin-like, and sweet odour notes in fresh poultry meat. Further studies of PL with higher energy doses will be necessary to confirm if there are Campylobacter reductions and about poultry meat treated under storage to evaluate if volatile compounds can affect the flavour of PL-treated meat samples.

High pressure processing, acidic and osmotic stress increased resistance to aminoglycosides and tetracyclines and the frequency of gene transfer among strains from commercial starter and protective cultures.

This study analyzed the effect of food-related stresses on the expression of antibiotic resistance of starter and protective strains and resistance gene transfer frequency. After exposure to high-pressure processing, acidic and osmotic stress, the expression of genes encoding resistance to aminoglycosides (aac(6')Ie-aph(2″)Ia and aph(3')-IIIa) and/or tetracyclines (tetM) increased. After cold stress, a decrease in the expression level of all tested genes was observed. The results obtained in the gene expression analysis correlated with the results of the phenotype patterns. After acidic and osmotic stresses, a significant increase in the frequency of each gene transfer was observed. To the best of the authors' knowledge, this is the first study focused on changes in antibiotic resistance associated with a stress response among starter and protective strains. The results suggest that the physicochemical factors prevailing during food production and storage may affect the phenotype of antibiotic resistance and the level of expression of antibiotic resistance genes among microorganisms. As a result, they can contribute to the spread of antibiotic resistance. This points to the need to verify strains used in the food industry for their antibiotic resistance to prevent them from becoming a reservoir for antibiotic resistance genes.

Bacteriocin-Producing Strain Lactiplantibacillus plantarum LP17L/1 Isolated from Traditional Stored Ewe's Milk Cheese and Its Beneficial Potential.

Stored ewe's milk lump cheese is a local product that can be a source of autochthonous beneficial microbiota, especially lactic acid bacteria. The aim of this study was to show the antimicrobial potential of Lactiplantibacillus plantarum LP17L/1 isolated from stored ewe's milk lump cheese. Lpb. plantarum LP17L/1 is a non-hemolytic, non-biofilm-forming strain, susceptible to antibiotics. It contains genes for 10 bacteriocins-plantaricins and exerted active bacteriocin with in vitro anti-staphylococcal and anti-listerial effect. It does not produce damaging enzymes, but it produces ß-galactosidase. It also sufficiently survives in Balb/c mice without side effects which indicate its safety. Moreover, a reduction in coliforms in mice jejunum was noted. LP17L/1 is supposed to be a promising additive for Slovak local dairy products.

Nitrite-Free Implications on Consumer Acceptance and the Behavior of Pathogens in Cured Pork Loins.

Cured pork loins are valued products due to their particular sensory characteristics. These products are usually prepared with nitrite to guarantee adequate color and pathogen control. The use of nitrite in meat products has been criticized due to its potential contribution to carcinogenic N-nitroso-compound formation. The present work aimed to evaluate the effect of eliminating nitrite from the manufacturing of cured loins made with wine- and water-based marinades on the color evaluation of consumers and on the behavior of Clostridium sporogenes, Listeria monocytogenes, and Salmonella. The use of nitrite in processing cured loins resulted in a color considered adequate by more than 50% of the consumers. When nitrite was not used, the color was described mainly as weak. The hedonic evaluation of cured loins did not reflect the color evaluation. The samples with a weak and an adequate color had similar hedonic evaluations. The present work did not allow us to infer the potential interest in injecting S. xylosus into meat to prepare cured loins. The use of nitrite did not affect the survival of Cl. sporogenes, L. monocytogenes, or Salmonella. The reduction in the aw was the primary determinant influencing pathogen survival. The production of nitrite-free cured loins seems possible once the control of pathogens can be achieved. However, the product will have a weaker color. Consumers appreciate sensory aspects other than color, which, combined with the positive impact of the "additive-free" claim, can support the possibility of producing cured loins without nitrite.

Use of Healthy Emulsion Hydrogels to Improve the Quality of Pork Burgers.

The present research evaluated the use of oil mixture emulsion hydrogels as animal fat replacers and their effect on the physicochemical, nutritional and sensory characteristics of pork burgers. Three different types of burgers were manufactured: control (samples elaborated with 100% pork fat), T1 and T2 (pork fat totally replaced by emulsion hydrogels of walnut or pistachio oil and algal oil, respectively). Fat replacement increased the moisture and ash contents and colour parameters (L* and b*) of pork burgers. Modified samples turned out to be firmer and chewier than those in the control group. The addition of oil emulsion hydrogels caused a significant decrease in fat and energy contents and the products obtained can be considered "reduced fat content". Moreover, the content of saturated fatty acids decreased, while mono- and polyunsaturated fatty acids increased, constituting an improvement in health indices. Sensory differences were found between the samples and T2 was the most preferred for flavour and overall. However, both modified burgers had good levels of acceptability. To conclude, the use of the proposed oil mixture emulsion hydrogels as pork backfat substitutes represents a promising strategy to obtain healthier pork burgers without negatively affecting technological or sensory properties.

The Development of Smoked Mackerel with Reduced Sodium Content.

The World Health Organization recommends reducing salt (sodium chloride, NaCl) intake by 30% by 2025. Since smoked fish can deliver up to 4 g NaCl/100 g, the aim of this study was to develop safe, healthy and attractive smoked chub mackerel (Scomber japonicus) with a reduced NaCl content. Two brines (5% and 10%) were used with different ratios of NaCl and potassium chloride (KCl). In each brine, 0%, 25%, 50% and 75% of NaCl was replaced by KCl, resulting in 1.3, 1.1, 0.9 and 0.6 g NaCl (5% brine), and 2.6, 2.0, 1.2 and 0.8 g NaCl (10% brine) per 100 g, respectively. Similar yield, nutritional, safety, texture and colour properties were found in most formulations. The most desirable taste attributes (negligible bitterness and adequate saltiness) were obtained with a 5% brine prepared with 75% NaCl + 25% KCl. Such conditions seemed to allow for obtaining an attractive product for conscious consumers.

The Risk of Salt Reduction in Dry-Cured Sausage Assessed by the Influence on Water Activity and the Survival of Salmonella.

Water activity (aw) is the main hurdle for microbial control in dry-cured sausages. The aw can be influenced by drying or adding electrolytes or humectants. Dry-cured meat products are partially dried, which, together with added salt, results in safe aw values. Currently, there is a trend to reduce salt in meat products, which can compromise the preservation process. The present work aims to evaluate the influences of added salt levels (1% or 3%) and the use or omission of phosphates and wine on the aw of a dry-cured sausage, and to evaluate the possibility of estimating the aw from the moisture loss and the behavior of Salmonella during dry-cured sausage (chouriço) processing. There was a strong relationship between moisture and aw, regardless of the salt level and the presence of phosphates or wine. Predicting aw from moisture loss is possible using the Boltzmann sigmoid function. The salt level strongly influences Salmonella behavior, mainly through aw reduction. An increase in aw by 0.01 units reduced the odds of achieving a 5-log reduction in Salmonella counts to half. Increasing added salt from 1% to 3% increased the odds of achieving a 5-log Salmonella reduction 7.5-fold. The current trend to reduce salt in foods must be carefully approached if applied to cured meat products, as it has substantial consequences on aw evolution and Salmonella survival.

Nitrate Is Nitrate: The Status Quo of Using Nitrate through Vegetable Extracts in Meat Products.

Nitrate and nitrites are used to give the characteristic color to cured meat products and to preserve them. According to the scientific knowledge available at the moment, these compounds are approved as food additives based on a detailed ponderation between the potential risks and benefits. The controversy over nitrites has increased with the release of an IARC Monograph suggesting an association between colorectal cancer and dietary nitrite in processed meats. The trend in "clean label" products reinforced the concern of consumers about nitrates and nitrites in meat products. This review aims to explain the role of nitrates and nitrites used in meat products. The potential chemical hazards and health risks linked to the consumption of cured meat products are described. Different strategies aiming to replace synthetic nitrate and nitrite and obtain green-label meat products are summarized, discussing their impact on various potential hazards. In the light of the present knowledge, the use or not of nitrite is highly dependent on the ponderation of two main risks-the eventual formation of nitrosamines or the eventual out-growth of severe pathogens. It is evident that synthetic nitrite and nitrate alternatives must be researched, but always considering the equilibrium that is the safety of a meat product.

Evidence for the Dissemination to Humans of Methicillin-Resistant Staphylococcus aureus ST398 through the Pork Production Chain: A Study in a Portuguese Slaughterhouse.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) ST398 was recovered from infections in humans exposed to animals, raising public health concerns. However, contact with food producing chain as a means of transmission of LA-MRSA to humans remains poorly understood. We aimed to assess if pork production chain is a source of MRSA ST398 for human colonization and infection. MRSA from live pigs, meat, the environment, and slaughterhouse workers were analyzed by Pulsed-Field Gel Electrophoresis (PFGE), spa, MLST typing, SNPs and for antibiotic resistance and virulence gene profiles. We compared core and accessory genomes of MRSA ST398 isolated from slaughterhouse and hospital. We detected MRSA ST398 (t011, t108, t1451) along the entire pork production chain (live pigs: 60%; equipment: 38%; meat: 23%) and in workers (40%). All MRSA ST398 were multidrug resistant, and the majority carried genes encoding biocide resistance and enterotoxins. We found 23 cross-transmission events between live pigs, meat, and workers (6-55 SNPs). MRSA ST398 from infection and slaughterhouse environment belonged to the same clonal type (ST398, t011, SCCmec V), but differed in 321-378 SNPs. Pork production chain can be a source of MRSA ST398 for colonization of human slaughterhouse workers, which can represent a risk of subsequent meat contamination and human infection.

Optimization of the Effect of Pineapple By-Products Enhanced in Bromelain by Hydrostatic Pressure on the Texture and Overall Quality of Silverside Beef Cut.

Dehydrated pineapple by-products enriched in bromelain using a hydrostatic pressure treatment (225 MPa, 8.5 min) were added in marinades to improve beef properties. The steaks from the silverside cut (2 ± 0.5 cm thickness and weight 270 ± 50 g), characterized as harder and cheaper, were immersed in marinades that were added to dehydrated and pressurized pineapple by-products that corresponded to a bromelain concentration of 0-20 mg tyrosine, 100 g-1 meat, and 0-24 h time, according to the central composite factorial design matrix. Samples were characterized in terms of marination yield, pH, color, and histology. Subsequently, samples were cooked in a water-bath (80 °C, 15 min), stabilized (4 °C, 24 h), and measured for cooking loss, pH, color, hardness, and histology. Marinades (12-24 h) and bromelain concentration (10-20 mg tyrosine.100 g-1 meat) reduced pH and hardness, increased marination yield, and resulted in a lighter color. Although refrigeration was not an optimal temperature for bromelain activity, meat hardness decreased (41%). Thus, the use of pineapple by-products in brine allowed for the valorization of lower commercial value steak cuts.

Application of Edible Alginate Films with Pineapple Peel Active Compounds on Beef Meat Preservation.

Alginate-based edible films containing natural antioxidants from pineapple peel were applied in the microbial spoilage control, color preservation, and barrier to lipid oxidation of beef steaks under storage at 4 °C for five days. Different stabilization methods of pineapple peel compounds were used before incorporation into alginate films, including extracted compounds with an hydroalcoholic solvent encapsulated in microparticles, microparticles produced by spray-drying pineapple peel juice, and particles obtained by milling freeze dried pineapple peel. Bioactive films exhibited higher antioxidant activity (between 0.15 µmol to 0.35 µmol FeSO4.7H2O/g dried film) than the alginate film without these compounds (0.02 µmol FeSO4.7H2O/g dried film). Results showed that control films without active compounds had no significant effect on decreasing the microbial load of aerobic mesophilic and Pseudomonas spp., while the films containing encapsulated hydroalcoholic extract showed a significant inhibitory effect on microbial growth of meat at two days of storage. Alginate films containing peel encapsulated extract were effective for maintaining the color hue and intensity of red beef meat samples. Pineapple peel antioxidants have the potential to retard lipid oxidation in meat samples, and the possibility of incorporation of a higher amount of pineapple peel bioactive compounds in the films should be investigated.

Influence of Meat Spoilage Microbiota Initial Load on the Growth and Survival of Three Pathogens on a Naturally Fermented Sausage.

Meat products are potential vehicles for transmitting foodborne pathogens like Salmonella, S. aureus, and L. monocytogenes. We aimed to evaluate (1) the effect of the meat's initial natural microbiota on Salmonella, S. aureus, and L. monocytogenes growth and survival in a batter to prepare a naturally fermented sausage, made with and without curing salts and wine (2) the effect of a lactic acid bacteria (LAB) starter culture and wine on the survival of the three pathogens during the manufacturing of a naturally fermented sausage made with meat with a low initial microbial load. The results revealed that the reduced contamination that is currently expected in raw meat is favorable for the multiplication of pathogens due to reduced competition. The inhibitory effect of nitrite and nitrate on Salmonella, S. aureus, and L. monocytogenes was confirmed, particularly when competition in meat was low. In any attempt to reduce or eliminate nitrite from naturally fermented sausages, the use of LAB starters should be considered to ensure an unfavorable competition environment for pathogens. In the experiment with naturally fermented sausage, chouriço, the reduction in aw strongly inhibited the challenged pathogens, particularly when a LAB starter culture and wine were used.

Impact of a 25% Salt Reduction on the Microbial Load, Texture, and Sensory Attributes of a Traditional Dry-Cured Sausage.

Catalão is a Portuguese dry-cured traditional sausage, highly appreciated for its distinctive sensory properties. The aim of this study was to evaluate the impact of a 25% salt reduction on Catalão manufactured with either purebred Alentejano (Al) or crossbred Iberian × Duroc (IDr) pork meat, on its physicochemical and microbiological stability, texture parameters, and sensory attributes. No significant effect of salt reduction or genotype was observed for pH, aW, and microbiological parameters. PUFA content was significantly higher for Al Catalão, particularly due to the content in linoleic and linolenic fatty acids. IDr 3% NaCl samples had the highest mean n6/n3 PUFA ratio, and the highest mean values for the atherogenicity and thrombogenicity indices, showing that both genotype and salt content influence these nutritional indices. Texture profile of Catalão was significantly influenced by salt content and genotype. Al samples were less adhesive, cohesive, and easier to chew. Low-salt Catalão was harder, more adhesive, and less cohesive, with lower resilience and higher chewiness values. Regarding sensory attributes, salt content influenced the product aroma, with reduced-salt sausages being evaluated as significantly less aromatic. Overall, a 25% salt reduction did not have a significant impact on the quality, stability, and sensory evaluation of Catalão.

Autochthonous Starter Cultures Are Able to Reduce Biogenic Amines in a Traditional Portuguese Smoked Fermented Sausage.

Traditional smoked fermented sausages are highly appreciated in Portugal and are mostly manufactured according to traditional procedures. The aim of the present work was to evaluate the effect of autochthonous starter cultures on the safety and quality of a smoked fermented sausage, Painho da Beira Baixa (PBB), preserving its sensory quality. Physicochemical parameters, namely pH and water activity (aW), microbiological parameters, biogenic amines, colour, texture profile and sensory attributes were assessed. Different starters were selected based on our previous work. Staphylococcus equorum S2M7, Staphylococcus xylosus CECT7057, Lactobacillus sakei CV3C2, Lactobacillus sakei CECT7056 and a yeast strain (2RB4) were co-inoculated in meat batters at defined concentrations. Starters had a significant effect on the reduction of pH. Enterobacteria and Listeria monocytogenes were not detected in inoculated end-product sausages. Moreover, sausages inoculated with S. equorum S2M7/L. sakei CV3C2/yeast 2RB4 showed a significant reduction in the total content of biogenic amines. No significant differences between treatments were observed for colour and texture parameters, except for adhesiveness. The studied starters did not compromise the sensory characteristics of PBB. To our knowledge, this is the first comprehensive study on the quality and safety of this type of smoked fermented sausage from the central region of Portugal.

Red Wine and Garlic as a Possible Alternative to Minimize the Use of Nitrite for Controlling Clostridium Sporogenes and Salmonella in a Cured Sausage: Safety and Sensory Implications.

The use of nitrite in meat products has been questioned due to its potential association with colon cancer. This work aimed to evaluate the behavior of Clostridium sporogenes (used as a surrogate for Cl. botulinum) and Salmonella in a dry-cured sausage, chouriço, made with and without nitrite and nitrate or with red wine and garlic, and to study the sensory implications through a consumer test. The survival of Cl. sporogenes and Salmonella was determined mainly by the reduction in water activity (aw), but the use of wine or wine and garlic contributed to the control of Salmonella during processing. The challenge test with Cl. sporogenes revealed no effect of the curing salts, wine, or garlic on the population of this microorganism. The use of curing salts resulted in a more reddish color that was recognized by the consumer as over-cured and artificial when compared with chouriço made with wine or wine and garlic, which were better rated in the hedonic test. In cured sausages of small caliber, the use of nitrite might be reconsidered, as the values of aw necessary to inhibit Clostridium toxinogenesis and growth are achieved rapidly.

Dry-Cured Meat Products According to the Smoking Regime: Process Optimization to Control Polycyclic Aromatic Hydrocarbons.

The manufacturing of dry-cured meat products usually includes a smoking step. Polycyclic aromatic hydrocarbons (PAHs) are potentially carcinogenic chemical compounds that may result from smoking. The aim of the present study was to optimize the smoking regime of traditional dry-cured meat products in order to minimize the presence of PAHs. Dry-cured sausages were submitted to different smoking regimes: (A) no smoking; (B) 20 h effective smoking; (C) 60 h effective smoking; (D) effective smoking until reaching 38%-40% weight losses. Three independent batches were produced per smoking regime, and three samples per batch were analyzed. Microbiological, physicochemical, and sensory analyses were performed. The total PAHs content was generally low and did not differ significantly in meat products submitted to the four different smoking regimes. The PAH4 and benzo(α)pyrene levels were below the established legal limits in all analyzed dry-cured sausages. Nevertheless, non-smoked sausages always showed lower PAHs values for all PAHs groups.

What is the Main Processing Factor Influencing Staphylococcus Species Diversity in Different Manufacturing Units?

The microbiota of traditional dry-cured sausages and industrial environment was assessed to characterize the diversity of coagulase-negative staphylococci (CNS), and establish potential relationships with hygiene level or technological characteristics. Eight processing units from South Portugal were audited according to a checklist of requirements. Environmental and products' samples at different production stages were evaluated regarding hygiene and safety criteria. CNS were recovered, characterized, and their potential use as starters evaluated. Low genetic diversity was observed for Staphylococcus xylosus, whereas Staphylococcus equorum showed diverse genetic profiles. Staphylococcus xylosus predominated in products with a long period of cold smoking, Staphylococcus saprophyticus in products with a long period of hot smoking, Staphylococcus epidermidis in products with a short period of cold smoking, and S. equorum in nonsmoked products. Most S. xylosus were resistant to tetracycline, whereas S. equorum were susceptible. Antibioresistance restricted the selection of starters due to safety recommendations. PRACTICAL APPLICATION: The present manuscript highlighted a few staphylococci strains that could potentially be used as starter cultures in fermented meat products. These selected strains do not show resistance to antimicrobials, exhibit adequate technological features, and are well adapted to the industrial environments of meat processing industries using different processing technologies. Therefore, the selected strains ready to be used in the manufacturing of traditional fermented meat products to ensure safety, standardize product properties, and shorten ripening.

Influence of Food Characteristics and Food Additives on the Antimicrobial Effect of Garlic and Oregano Essential Oils.

Utilization of essential oils (EOs) as antimicrobial agents against foodborne disease has gained importance, for their use as natural preservatives. Since potential interactions between EOs and food characteristics may affect their antimicrobial properties, the present work studies the influence of fat, protein, pH, aw and food additives on the antimicrobial effect of oregano and garlic EOs against Salmonella spp. and Listeria monocytogenes. Results showed that protein, pH, aw, presence of beef extract, sodium lactate and nitrates did not influence their antimicrobial effect. In contrast, the presence of pork fat had a negative effect against both EOs associated with their dilution of the lipid content. The addition of food phosphates also exerts a negative effect against EOs probably associated with their emulsification properties as observed with the addition of fat. The results may help the food industry to select more appropriate challenges to guarantee the food safety of foodstuffs.